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USE OF ANTIMICROBIAL AGENTS IN DAIRY CALVES AND
THE PUBLIC HEALTH CONCERN

presented by

James John Averill, DVM

has been accepted towards fulfillment
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USE OF ANTIMICROBIAL AGENTS IN DAIRY CALVES AND THE PUBLIC
HEALTH CONCERN

By

James John Averill, DVM

A DISSERTATION
Submitted to
Michigan State University
in partial fulfillment of requirements
for the degree of
DOCTOR OF PHILOSOPHY
LARGE ANIMAL CLINICAL SCIENCES

2009

ABSTRACT

USE OF ANTIMICROBIAL AGENTS IN DAIRY CALVES AND THE PUBLIC
HEALTH CONCERN

By
James J. Averill, DVM
Since their inception, antimicrobial agents have been used in multiple ways to

enhance the health and well-being of humans and animals. Today, in developed I'T

countries, we presume that antimicrobial agents will cure the vast majority of bacterial
diseases. However, bacterial resistance to antimicrobial agents is an emerging problem
that challenges the biomedical professions and public health. Additionally, the use of
antimicrobial agents in animal agriculture has been implicated as contributing to bacterial
resistance in human medicine. Veterinary medicine can play a key role in helping
mitigate the potential increase of antimicrobial resistance.

This study was developed with the overall goal to enhance knowledge on the
appropriate use of antimicrobial agents in veterinary medicine. This overall goal was
addressed through the four following objectives: 1) Evaluate growth, morbidity, and
mortality of Holstein heifer calves fed milk replacer with or without oxytetracycline and
neomycin add, 2) Determine the steady state pharmacokinetics of oxytetracycline
administered in milk replacer, 3) Compare minimum inhibitory concentrations of
Escherichia coli and Pasteurella multocida isolated from calves fed milk replacer with or
without antimicrobial agents, and 4) Develop a computer-aided learning program on the
appropriate use of antimicrobial agents in veterinary medicine to be used in veterinary

school curricula.

Findings are briefly summarized below. Calves fed medicated milk replacer
(MMR) were heavier at 42 and 150 days of age than calves fed non-medicated milk
replacer. Risk factors that influenced weight gain were treatment group, birth weight,
season of birth, and having an episode of respiratory disease. Though there was no
difference in morbidity between treatment groups. There was a numeric difference with
mortality, as the medicated group had lower mortality rates.

Plasma concentrations of oxytetracycline in dairy calves fed medicated milk
replacer were below minimum inhibitory concentration (MIC) values for Escherichia coli
and Pasteurella multocida. For Pasteurella multocida, the medicated group did have a
higher MIC9O compared to the non-supplemented calves, while for Escherichia coli there
was no difference.

A website was developed which contains a ‘Principle’ module that goes over the
basics of antimicrobial resistance and the role of antimicrobial use in animal and human
health. There are also multiple ‘Case Study’ modules that apply these principles to
specific clinical situations involving dairy and beef cattle, small animals, pocket pets and
swine. Each module is about how a veterinary student or veterinarian investigates an
issue concerning the appropriate use of antimicrobial agents. Within each module there
are videos, animations, and questions to engage the learner.

In conclusion, MMR increases growth in dairy calves and plasma levels of
oxytetracycline do not reach MIC values for either pathogen tested. Feeding MMR
during the first few weeks of life may be beneficial to reduce mortality but given the
concern regarding potential development of resistance, MMR not be feed for the entire

suckling period.

To my parents David and Sandra.
Without your unconditional love and guidance
this would never have been possible.

iv

ACKNOWLEDGEMENTS

I am sincerely appreciative of Dr. Ronald Erskine, my major advisor, for his
guidance, insight and patience throughout my research. I’d also like to thank my
graduate committee - Drs. Paul Bartlett, Theresa Bernardo, Carole Bolin and Patricia
Somsel -- for their time, valuable suggestions, and comments regarding my research.
This research would not have been possible without the cooperation of Cary Dairy, who
allowed me to use their facilities and employees’ time during my research.

I owe the College of Veterinary Medicine, especially the Department of Large
Animal Clinical Sciences, faculty and staff my gratitude for the opportunity to pursue my
doctoral degree and provide me with an enjoyable learning environment. The Michigan
Department of Community Health, Bureau of Laboratories, provided me the opportunity
to grow professionally as an educator and researcher. Also among those who contributed
in helping with the challenge of my work and sanity are my fellow graduate students:
Amanda Fine, RoseAnn Miller, and Roxanne Pillars. I am also indebted to a few key
individuals who assisted much of my research: Anne Brockett, Elizabeth Ritchie, Katie
May, and Steve Haskell.

None of this would be possible without the support of my family. Thanks goes to
my parents (David and Sandra) and my brothers (Travis, Duncan and Kevin) for always
being there and showing interest even if you didn’t know what I was talking about.
Finally, I will always be indebted to Donna Letavish for her patience and fortitude in
seeing me through this process. Her support, willingness to help, and love has meant so

much to me. I could never thank her enough.

TABLE OF CONTENTS

LIST OF TABLES ...........................................................................
LIST OF FIGURES .........................................................................
INTRODUCTION ...........................................................................

CHAPTER 1

USE OF AN TIMICROBIALS IN DAIRY CALVES AND PUBLIC HEALTH
CONCERNS: REVIEW .....................................................................

Introduction .....................................................................
Discovery of Antimicrobial Agents .........................................
Development of Antimicrobial Resistance .................................
Development of Public Health Concerns ..................................
Types and Mechanisms of Antimicrobial Resistance .....................
Use of Antimicrobials in Animal Feed .....................................
Pharmacokinetics of Oxytetracycline in Milk Replacer ..................
Factors Affecting Calf Growth ...............................................
Conclusion .....................................................................

Cited Literature .................................................................

CHAPTER 2

EFFECTS OF FEEDING MEDICATED OR NON-MEDICATED MILK
REPLACER TO HOLSTEIN HEIFER CALVES ON GROWTH, MORBIDITY
AND MORTALITY .........................................................................

Abstract .........................................................................
Introduction .....................................................................
Materials and Methods .......................................................

Results ..........................................................................

Discussion ......................................................................
Conclusion ......................................................................
Cited Literature ................................................................

CHAPTER 3

PHARMACOKINETICS OF OXYTETRACYCLINE IN MILK REPLACER
FED TO HOLSTEIN CALVES ............................................................

Abstract .........................................................................
Introduction .....................................................................
Materials and Methods .......................................................

Results ..........................................................................

Discussion ......................................................................
Conclusion ......................................................................
Cited Literature ................................................................

vi

viii

ix

1

5

6

7

10
13
17
23
26
30
38
39

50
51
52
53
57
59
63
70

73
74
75
76
80
81
85
91

CHAPTER 4
TEACHING ANTIMICROBIAL RESISTANCE VIA COMPUTER AIDED

LEARNING ................................................................................... 93
Abstract ......................................................................... 94
Introduction ..................................................................... 94
Materials and Methods ....................................................... 97
Description of Computer Aided Learning Tool ........................... 100
Usability Test Results .......................................................... 103
Discussion ...................................................................... 104
Conclusion ...................................................................... 106
Cited Literature ................................................................ 109

CONCLUSIONS .............................................................................. 1 11

APPENDIX .................................................................................... 116
Appendix A (Appropriate Use of Antimicrobial Agents in Veterinary
Medicine) ........................................................................ 1 17

vii

LIST OF TABLES

Table 1.1 Historical Timeline of Antimicrobial Development, Introduction, and
Important Events ............................................................... 11

Table 1.2 Pharrnacokinetic Studies of the Tetracycline Family in Dairy Calves
and Pigs .......................................................................... 28

Table 1.3 Recommended Milk-Replacer Ingredients for Replacement Calves... 32

Table 1.4 Recommended Calf-Starter Composition .................................. 32
Table 2.1 Mean Growth for Calves ...................................................... 65
Table 2.2 Calf Morbidity Data ........................................................... 65
Table 2.3 Number of Cases Per Animal ................................................ 66
Table 2.4 Type and Number of Mortalities Per Treatment Group .................. 66

Table 2.5 Fixed Effects Values for Repeated Measures Model With Weight as
Outcome ......................................................................... 66

Table 3.1 Plasma Concentration of Oxytetracycline in Calves at Each

Collection Time Point ......................................................... 86
Table 3.2 Pharmacokinetic Values for Oxytetracycline .............................. 86
Table 3.3 MICSO and MIC90 Values of Oxytetracycline for P. multacida and

E. coli ............................................................................ 90
Table 4.1 Usability Testing Task List for the Website ................................ 108

viii

Figure 1.1

Figure 1.2
Figure 2.1

Figure 2.2

Figure 2.3

Figure 3.1

Figure 3.2
Figure 3.3

Figure 4.1

Figure A]
Figure A.2
Figure A.3
Figure A.4

Figure A.5

LIST OF FIGURES

Methods of Transferable Resistance for Antimicrobial Agents;
Conjugation, Transformation, Transduction .............................

Mechanisms of Resistance for Antimicrobial Agents ..................
Calf Mortalities by Treatment Group .....................................

Weight Gained in the First 8-Weeks for Calves With or Without a
Respiratory Case ..............................................................

Weight Gained at 42 Days of Age ........................................

Steady State Kinetics, Mean Plasma Concentration for
Oxytetracycline ..............................................................

MIC Values of Oxytetracycline for Pasteurella multocida ............
MIC Values of Oxytetracycline for Escherichia coli ...................

Intended Flow of the Website and Themes Covered in Each
Module ........................................................................

The Principles Module ......................................................
Medicated Milk Replacer Module ........................................
Neonatal Scours Module ...................................................
Contagious Mastitis Module ...............................................

Farm Based Mastitis Module ..............................................

ix

19

22

67

68

69

87

88

89

107

119

120

121

122

123

L

INTRODUCTION

RATIONALE

The discovery of antimicrobial agents advanced medicine from the inability to
treat complications fiom a simple scratch to the ability to cure life-threatening diseases
such as syphilis, cholera, tuberculosis and pneumonia. Since their inception,
antimicrobial agents have been used in multiple ways to enhance the health and well-
being of humans and animals. Today, in developed countries, we presume that
antimicrobial agents will cure the vast majority of bacterial diseases. However, bacterial
resistance to antimicrobial agents is an emerging problem that challenges the biomedical
professions and public health. Additionally, the use of antimicrobial agents in animal
agriculture has been implicated as contributing to bacterial resistance in human medicine.

Antimicrobial agents were added to animal feed as early as 1948. Soon after,
antimicrobial agents were added to milk replacer to improve the growth of dairy calves
and for disease prevention. By the mid 1950s research demonstrated that medicated milk
replacer increased growth 10 to 30 percent in the first eight weeks of life. As animal
husbandry and preventive management improved over the last half century, the literature
has offered paradoxical results as to the benefit of feeding medicated milk replacer.
National Animal Health Monitoring System (NAHMS) surveys of the dairy industry in
2002 and 2007 reported that supplementing milk replacer with antimicrobial agents is a
still a common practice as over fifty percent of all milk replacers include antimicrobial

agents. The most common drugs added are oxytetracycline and neomycin.

As antimicrobial resistance has increased, questions have arisen regarding the
need to add antimicrobial agents to milk replacer and other animal feeds. Scrutiny over
the use of antimicrobial agents began in 1969 when the British government released the
Swan report, calling for antimicrobial agents in animal agriculture to be used by
prescription only and for drugs used as feed additives be prohibited if used in human
medicine. The World Health Organization released several reports in the late 1990s and
into the 21St century stating concern regarding the use of antimicrobial agents in animal
feed and potential transfer of resistance to humans. In 2006, the European Union banned
the use of all antimicrobial agents in animal agriculture for growth promotion, including

ionophores.

PROBLEM STATEMENT

With the emergence of antimicrobial resistance as a public health concern,
veterinary medicine plays a critical role in mitigating further development of resistance.
Although there have been advancements in animal husbandry and preventive medicine
practices (vaccines, nutrition and biosecurity) over the past fifty years, medicated milk
replacer is still commonly used in the dairy industry. There is a need to investigate the
necessity of adding antimicrobial agents to milk replacer for aid in growth and/or prevent

disease given the concern regarding the development of antimicrobial resistance.

RESEARCH QUESTIONS TO BE ADDRESSED
The overall aim of this dissertation is to enhance scientific knowledge on the

appropriate use of antimicrobial agents in dairy cattle. Specifically, to address the use of

antimicrobial agents in dairy calves. Key research questions that should be answered by
the studies conducted;

1) Do calves fed milk replacer supplemented with antimicrobial agents grow
faster and have a lower disease burden?

2) Do plasma-steady state pharmacokinetic values for oxytetracycline as fed in
medicated milk replacer achieve minimum inhibitory concentrations for
Escherichia coli and Pasteurella multocida?

3) Will a computer-aided learning module be useful to teach veterinary students

concepts about appropriate use of antimicrobial agents?

HYPOTHESES TO BE TESTED

To address the above overall aim and research questions the following hypotheses

were developed. They are:

1) Holstein heifer calves fed a milk replacer supplemented with oxytetracycline
and neomycin will gain more weight and have a lower morbidity and
mortality rate than calves fed the same milk replacer without antimicrobial
agents.

2) Plasma concentration of oxytetracycline in calves fed a milk replacer
supplemented with oxytetracycline will reach minimum inhibitory
concentrations for Escherichia coli from fece and Pasteurella multocida from

nasal swab.

OVERVIEW OF RESEARCH

A literature review regarding discovery and development of antimicrobial
resistance and the use of antimicrobial agents in milk replacer are provided in chapter
one. Chapter two addresses hypothesis 1 via a field trial where calves’ growth and
health records were monitored weekly until weaning and then again at five months of
age. Hypothesis 2 is addressed in chapter three which describes an investigation of the
steady-state pharmacokinetics of oxytetracycline in six calves and compares minimum
inhibitory concentrations of oxytetracycline to two common bacterial pathogens to
plasma drug levels in calves fed milk replacer with or without oxytetracycline and
neomycin. Chapter four focuses on hypothesis 3, which describes a website was on the
appropriate use of antimicrobial agents in veterinary medicine to be used as an adjunct to

a veterinary students education.

CHAPTER 1

USE OF ANTIMICROBIALS IN DAIRY CALVES AND PUBLIC HEALTH
CONCERNS: A REVIEW
INTRODUCTION:

The discovery of antimicrobial agents advanced medicine from the inability to
treat complications fiom a simple scratch to the ability to cure life-threatening diseases
such as syphilis, cholera, tuberculosis and pneumonia. Since their inception,
antimicrobial agents have been used in multiple ways to enhance the health and well-
being of humans and animals. Today, in developed countries, we presume that
antimicrobial agents will cure the vast majority of bacterial diseases. However, bacterial
resistance to antimicrobial agents is an emerging problem that challenges the medical
professions and endangers public health. The use of antimicrobial agents in animal
agriculture has been implicated as contributing to bacterial resistance in human medicine.

Antimicrobial agents were added to animal feed as early as 1948 (J ukes and
Williams, 1953). Soon after, antimicrobial agents were added to milk replacer to
improve the growth of dairy calves and for disease prevention. As animal husbandry and
preventive management improved over the last half century, questions arose regarding
the need to add antimicrobial agents to milk replacer, and other animal feeds.

The purpose of this literature review is to impart a historical and current
understanding of the use of antimicrobial agents in animal feed. In particular, topics will
include: 1) discovery of antimicrobial agents, 2) the mechanisms of bacterial resistance,
3) public health concerns regarding the use of antimicrobial agents in animal agriculture,
4) the practice of using medicated milk replacer, 5) pharmacokinetics of oxytetracycline

in milk replacer, and 6) factors associated with dairy calf growth.

DISCOVERY OF ANTIMICROBIAL AGENTS:

Antimicrobial agents are substances that inhibit microorganism viability, and
derive their name from the Greek words anti, meaning “against,” micros, meaning
“little,” and bios, meaning “life.” Substances used to treat bacterial infections in animals
and humans are typically referred to as antibiotics. The terms antimicrobial agents and
antibiotics are not interchangeable; antibiotics refer to substances of microbial origin that
act upon other microorganisms, while the broader term, antimicrobial agents, also
includes synthetic compounds.

In recalling the discovery of penicillin, Sir Ernest Chain stated that it “is
practically impossible for anyone growing bacteria not to come across chance
contaminants with antagonistic properties” (Chain, 1980). During the 1870s, scientific
emphasis on the germ theory derived the term “abiogenesis.” Roberts documented this in
his “Studies of Abiogenesis” in 1874, where he noted antagonism between fungi and
bacteria (Roberts, 1874). The use of such an antagonist for therapy was not suggested
until 1885 by Cantani’s paper in Medical News (Cantani, 1885). In 1889, a French
biologist, P. Vuillemin, described the destruction of one organism by another with the
adjective “antibiotic” (Chain, 1980). Later in that century, Paul Ehrlich, who was later
proclaimed by some as the father of antibacterial therapy, determined that the chemical
dye salvarsen had potential antimicrobial effects in patients with syphilis. However,
salvarsen was later found to have unpleasant side effects as the compound contained an
arsenic derivative (Amyes, 2001). In his early work, Ehrlich suggested that dyes might
be the “magic bullet” for treating bacterial infections. Ehrlich’s work was rediscovered

in 1929 by Gerhard Domagk who took a more systematic approach to discovery of

chemicals to treat bacterial infections by using a mouse model (Amyes, 2001).

Domagk’s vigilance paid off in 1932 when a new drug, the red dye Prontosil, was
successfiill in treating Streptococcus in mice (Amyes, 2001 ). Prontosil was tested against
a variety of bacteria and was shortly discovered not to be effective against gram-negative
bacteria. Domagk’s discovery was not well accepted by the medical community until
1936, when the son of Franklin D. Rooosevelt was cured from tonsillitis by Prontosil
(Amyes, 2001). The press hailed the miracle drug. Later research by a French laboratory
discovered that the active compound in Prontosil was a sulphanilarnide, the first of this
class of antimicrobial agents (Amyes, 2001).

In his St. Mary’s laboratory during the 1920’s, Alexander Fleming was
investigating the antibacterial properties of body secretions (Chain, 1980). In these
experiments he demonstrated that a teardrop on a culture caused rapid lysis of certain
bacterial organisms. This lytic enzyme was named lysozyme, and was Fleming’s first
major scientific finding (Amyes, 2001). Further studies found that lysozyme did not
readily kill pathogenic bacteria; this did not deter Fleming fi'om continuing to look for
new antiseptics. Fleming was not known for being a tidy, well-organized bacteriologist,
as any visitor to his laboratory would see clutter and colonized petri dishes on the bench
tops. One day in early September 1928, Fleming was going through some old petri
dishes inoculated with Staphylococcus when he noticed mold contaminating one area of a
particular plate (Wainwright, 1990). Around the area of contamination was a wide circle
of inhibited growth of Staphylococci. Fleming named this inhibitory substance
penicillin, after the mold Penicillium notatum that was growing on the plate (Wainwright,

1990). Fleming was able to reproduce this phenomenon for other pathogenic gram-

positive bacteria, but not for gram-negative bacteria (Wainwright, 1990). Further studies
not only demonstrated that penicillin was effective against staphylococci but also non-
toxic to mice and rabbits (Chain, 1980). One question that Fleming did not answer with
his studies was the potential chemotherapeutic effect of penicillin.

Ironically, his discovery went unnoticed by the medical community for almost 10
years until 1936 when Florey and Chain attempted to determine the structure and
chemotherapeutic benefit of Fleming’s discovery (Chain, 1980). In 1940, they succeeded
with the help of Heatley to purify penicillin and later tested it in mice infected with
streptococci. After receiving penicillin doses every three hours, all but one survived, as
compared to all control mice dying within 16 hours (Chain, 1980). Fleming was pleased
with these findings and was quoted as saying, “they have turned out to be the successful
chemist I should have liked to have with me in 1929.” (Amyes, 2001). The next step was
to test the compound on humans and mass-produce penicillin via fermentation. F lorey
struggled to get pharmaceutical companies to explore large vat fermentation even after
several successful treatments in humans. Finally, John L. Smith, president of Chas Pfizer
Company was intrigued by the success of the compound and Chas Pfizer became the first
company to begin large-scale production of the drug in 1942 (Amyes, 2001).

Penicillin’s breakthrough in the United States took place when a devastating fire
at the Coconut Grove Nightclub in Boston occurred on November 29, 1942 (Levy, 2002).
Merck and Company sent a small supply of penicillin to Massachusetts General Hospital
for patients receiving skin grafts. This became one of the most important clinical trials
demonstrating the safety and efficacy of penicillin to the United States government. The

media touted penicillin as the “miracle drug” due to its ability to control infectious

bacteria that previously were not treatable (Levy, 2002). Thus, the discovery of clinical
applications for sulfonamides and penicillin revived the quest for what Paul Ehrlich
called the “magic bullet”, a drug that could kill bacteria without harming humans (Levy,
2002). For the discovery of penicillin, mass production, and success in treating soldiers’
wounds during World War II, Fleming, Florey, and Chain received the Nobel Prize for
Medicine in 1945 (Amyes, 2001).

In 1941, SA. Waksman introduced the term antibiotics, a noun, as “chemical
substances that are produced by microorganisms and that have the capacity, in dilute
solution, to selectively inhibit the growth of and even destroy other microorganisms”
(Aarestrup, 2006). This term was commonly used from this time forward when referring
to a compound used to treat bacterial infection(s). Through the mid 1940s into the early
1960s, many antimicrobial agents were discovered and brought into clinical practice

(Table 1).

DEVELOPMENT OF ANTIMICROBIAL RESISTANCE:

Alexander Fleming, in a 1945 interview with The New York Times, warned that
misuse of penicillin could lead to bacterial resistance (Levy, 2002). Fleming had
observed this phenomenon in his laboratory; strains of previously susceptible organisms,
in the presence of low concentrations, were no longer inhibited by penicillin (Levy,
2002). To avoid such development, Fleming spoke out saying that complete courses of
therapy were necessary to avoid initiating resistance. Little was done to address

Fleming’s concern due to the overwhelming benefit antimicrobial agents had on

10

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medicine. However, by 1946, resistance to penicillin was noted in a London hospital
where 14% of the isolated strains were resistant to the drug, and by the end of the decade,
the frequency of resistant strains had increased to 59 percent in the same hospital (Levy,
2002)

The proportion of penicillin-resistant organisms continued to increase into the
1950’s, as the drug was sold over-the-counter and incorporated into numerous products to
treat various ailments. By 1955, most countries had documented an increase in penicillin i
resistance, which prompted many countries to require a prescription for human use
(Levy, 2002). During this period, emerging resistance among pathogens in animal
agriculture was also documented, and most believed this evolution was strictly
chromosomally mediated (Elam et al., 1951a, b; Jones and Ricket, 2003). Barnes
confirmed these earlier studies (mentioned above), and determined that antimicrobial
agents in animal feed altered the microflora and increased the proportion of organisms
resistant to antimicrobial agents (Barnes, 195 8). The concern over potential transfer of
antimicrobial resistance to zoonotic pathogens came to a head during an outbreak of
chloramphenicol-resistant Salmonella Typhimurium in dairy calves, which was

implicated to cause infections in humans (Anderson, 1968).

DEVELOPMENT OF PUBLIC HEALTH CONCERNS:

As antimicrobial agents were being rapidly discovered, they were being employed
therapeutically or non-therapeutically, for animals and humans, and via a variety of
routes of administration and therapeutic regimens. In 1969, the “Swarm Report,” was

released by the British government regarding animal husbandry and the use of

13

antimicrobial agents by veterinary medicine in animal agriculture (Swann, 1969). This
report was prompted by: 1) an outbreak of resistant Salmonella in calves that infected
humans (Anderson, 1968), 2) an outbreak of chloramphenicol-resistant Salmonella typhi
in Central America (Randall, 1969), 3) evidence that antimicrobial agents used in animal
feed for growth promotion were causing the development of resistant bacteria isolated
from pigs and chickens (Barnes, 1958; Smith, 1975, 1977) and 4) demonstration of
transferable resistance (Anderson, 1968). This committee concluded that therapeutic use
of antimicrobial agents in farm animals should be used by prescription only and that feed
additive use of antimicrobial agents should only be permitted if the drug is not used in
humans (Swann, 1969). Tetracycline and penicillin did not meet the feed additive use
guidelines and were banned in the United Kingdom from such use (Aarestrup, 2006).
The Swann report received mixed reviews, as few believed such
recommendations were warranted (Freeman, 1970; Prescott et al., 2000), but it did
initiate debate as to how antimicrobial agents should be used in animal agriculture.
However, as time passed, scrutiny of non-therapeutic use of antimicrobial agents in
animal agriculture from public health and human medical communities increased. A
report from the US. Food and Drug Administration (FDA) in 1972 concluded that use of
antimicrobial agents in food animals could promote resistance in Salmonella and required
manufacturers to demonstrate that their product did not increase the prevalence of
resistant Salmonella in animals (FDA, 1972). This report was the first in the United
States to suggest that there was enough scientific information to justify a ban on non-
therapeutic use of penicillin and chlortetracycline, except under veterinary prescription.

However, this recommendation was not accepted (Force, 1972).

14

In 1980, the National Research Council of the US. released “The Effects on
Human Health of Subtherapeutic Use of Antimicrobials in Animals” (IOM, 1980). This
report stated that there was not adequate information to prove or disprove a relationship
between the use of antimicrobial agents in animal feed and increased bacterial resistance
in human medicine. This report further commented that the United Kingdom had seen
little benefit from changes recommended in the Swann Report of 1969 (IOM, 1980),
though the authors did recognize that non-therapeutic use of antimicrobial agents
increased isolation of resistant E. coli and Salmonella. Similar results were found in the
Institute of Medicine report in 1989, which determined a lack of substantial evidence
linking the use of penicillin and tetracycline at non-therapeutic levels in animal feed to
any impact on human health (IOM, 1989). Both of these studies did state that there was a
need for more information, as was reiterated in 1995 by the Office of Technology
Assessment (OTA, 1995).

A report in 1981 stated that banning the use of antimicrobial agents in animal feed
at non-therapeutic levels would cost the United States economy 3 .5 billion dollars
(Technology, 1981). The authors went on to say that in their opinion, banning non-
therapeutic antimicrobials would only be beneficial if therapeutic use was banned, too.
This might result in unethical and inhumane treatment of livestock.

After considerable debate, Sweden in 1985, prohibited non-therapeutic use of
antimicrobials in animal feed (SOU, 1997). This had economic ramifications, as the
mortality rate increased for piglets at weaning and necrotic enteritis emerged in broiler

chickens (Wierup, 2001). A self-study in 1997 concluded that the benefits out-weighed

15

the risks of banning the use of antimicrobial agents for growth promotion, and non-
therapeutic use (SOU, 1997).

The World Health Organization recommended the discontinuation of
antimicrobial agents in animal feed for growth promotion in 1997 (WHO, 1997).
Specifically, antimicrobials used in human medicine that were also used in animal feed
for growth promotion and evolved towards cross-resistance were to be prohibited from
animal use. The following year the Ministry of Agriculture, Fisheries and Food (U .K.)
released a report stating that resistance in animal pathogens is due to antimicrobial use.
Specifically Campylobacter and Salmonella with regards to certain antimicrobial agents,
can reach humans through the food supply and cause disease or colonize humans
allowing resistance to be transferred (Aarestrup, 2006).

Public health concerns continued to develop over the use of antimicrobials in
animal feed at non-therapeutic levels, and numerous reports were released stating that
such practice was leading to increased resistance in human medicine and decreasing
therapeutic success (Barza and Gorbach, 2002; JETACAR, 1999; WHO, 2003b). These
findings were strengthened by the Danish experience on banning antimicrobials in animal
feed for growth promotion and disease prevention; that ban resulted in a greater than 50
percent reduction in the use of antimicrobial agents in animal agriculture and decreased
prevalence of resistant organisms (WHO, 2003a). However, there was an increased
morbidity in weaned pigs and decreased feed efficiency that lead to a 1% increase in
production cost (WHO, 2003b). Over a five-year period, the FDA and pharmaceutical

industries debated the use of a fluoroquinolone drug in poultry because of concerns that

16

feeding this drug resulted in antimicrobial resistance in bacteria that cause infections in
humans, and the license for this use was withdrawn in September 2005 (FDA, 2005).

Given the overwhelming public health concern and regulatory response, the
scientific community continues to seek an objective viewpoint on the ban of
antimicrobials in animal feed for growth promotion and disease prevention. The National
Research Council and Institute of Medicine concluded that there was not enough
information to demonstrate an immediate public health concern (IOM, 1989; NRC and
IOM, 1999). The NRC/IOM report did acknowledge that there were many gaps and
further information may change their conclusions. It was also estimated that banning
non-therapeutic use of antimicrobial agents in animal feed would add $5-$10 3 year to the
cost of food for each United States citizen (NRC and IOM, 1999). Bywater et al reported
in 2000 a qualitative assessment of antimicrobial resistance and animal agricultures role.
Results of a questionnaire sent to experts in the United Kingdom found that use of
antimicrobial agents in animal agriculture contributed only 3.88 percent of the resistance
in major bacterial pathogens affecting humans, while the rest was due to use of such

drugs in human medicine (Bywater and Casewell, 2000).

TYPES AND MECHANISMS OF ANTIMICROBIAL RESISTANCE:

Bacteria can be intrinsically resistant to antimicrobial agents from a lack of targets
for drug activity, the inability of the drug to enter the bacteria, genetically coded pumps
to export the drug, or the presence of an enzyme that inactivates the drug (Sefton, 2002).
Intrinsic resistance occurs in many bacteria to various antimicrobial agents, for example,

glycopeptides are not able to enter the outer membrane of gram-negative bacteria

17

(Aarestrup, 2006). Perhaps of greater concern is acquired resistance, as this causes the
emergence and spread of resistance in what was once considered a susceptible organism.
Acquired resistance arises from a genetic mutation of an organism or via
acquisition of genetic material through horizontal transmission (Sefton, 2002; Tenover,
2006). Mutations that confer resistance spontaneously (endogenous resistance) occur at

varying rates among species and among strains within a species. As an example,
Staphylococcus aureus is approximately lOO-fold more likely to develop resistance to
rifarnpin than Escherichia coli, making the treatment of staphylococci with rifarnpin
inappropriate (Aarestrup, 2006).

Exogenous resistance, or transferable resistance, can occur by one of three
possible mechanisms (Figure 1): transformation, transduction, and conjugation (cell to
cell transfer of genetic material) (Aarestrup, 2006; Sefton, 2002; Tenover, 2006).
Transformation and transduction do not require contact between organisms, however
acquisition of antimicrobial resistance by these mechanisms is limited to closely related
species or genus, as homology is required between donor and recipient (Aarestrup, 2006).
Transduction occurs via plasmid DNA incorporated into a bacteriophage that attaches to
bacteria and transfers DNA into the bacteria, which potentially carries genes coding for
antimicrobial resistance, although this is believed to be a rare event (Tenover, 2006).
Transformation is the transfer of naked DNA from one cell to another (Prescott et al.,
2000). This form of transferable resistance is becoming recognized as an important
source of emerging resistance, e.g. Slrepotococcus transferring genes for penicillin-

binding-protein 2 (Aarestrup, 2006; Prescott et al., 2000).

18

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Conjugation is likely to play the biggest role in acquired resistance as genes for
antimicrobial resistance are commonly found on conjugative genetic elements; plasmids,
transposons, and integrons (Prescott et al., 2000; Sefton, 2002). Plasmids are
extrachromosomal circular DNA in bacteria, which replicate independently of
chromosomal DNA, but at the same time (Prescott et al., 2000). Plasmids can confer
resistance fi'om 1 to 10 different antimicrobial agents. Transposons are short sequences
of DNA, which readily transfer between plasmids, or between plasmids and
chromosomes (Prescott et al., 2000). In order for transposons to integrate into foreign
DNA, insertion genes are vital; these gene sequences flank both ends of the transposon
(Salyers and Whitt, 2002). Frequency of transposition is highly dependent on the
transposon and bacterial strain. Integrons are mobile genetic elements that are often
found on plasmids that are associated with antimicrobial resistance and other bacterial
changes (Prescott et al., 2000). In order for integrons to function and transfer resistance
they must contain an integrase enzyme for site-specific recombination, a gene-capture
site, and a captured gene (mobile element that contains the gene for antimicrobial
resistance) (Salyers and Whitt, 2002). Plasmids develop multi-drug resistance by
incorporation of multiple integrons conferring antimicrobial resistance (Prescott et al.,
2000; Salyers and Whitt, 2002).

Cross-resistance is the final type of antimicrobial resistance, which occurs when a
bacterium becomes resistance to an antimicrobial agent, and in doing so becomes
resistant to another (Aarestrup, 2006). For example, macrolides, lincosamides, and

streptogramins act on ribosomes, and bacterial adaptation of the 50s ribosomal RNA

20

confers resistance not just to one of the antimicrobial agents, but to all three (Aarestrup,
2006).

Mechanisms of antimicrobial resistance can be classified on a biochemical basis
into the following classifications (figure 2); modifying enzyme, impermeability, active
expulsion of drug, and modification of target. Modifying enzymes that inactivate the
drug can occur naturally and/or be plasmid mediated (Aarestrup, 2006; Sefton, 2002).
This mechanism is commonly seen with resistance to B-lactams and aminoglycosides; an
example is B-lactarnase that binds to penicillin to deactivate the drug. Resistance to B-
lactams and fluoroquinolones can also arise from reduced permeability of the drug into
the bacteria because membrane porins are too small or the drug is unable to diffuse
through the cytoplasmic membrane (Aarestrup, 2006; Sefton, 2002). An example of this
is the lack expression of OmpF porin in E. coli, which reduces susceptibility to B-
lactams, tetracyclines, and quinolones (Jensen et al., 1999). An efflux pump is a
transmembrane protein that works by active expulsion of the drug and is typically
plasmid mediated (Aarestrup, 2006; Sefion, 2002). This mechanism is often seen with
tetracycline where the drug enters the cytoplasm of resistant bacteria and is pumped back
out via the efflux pump (Sefton, 2002). The final major mechanism is modification of
the drug target from structural changes, replacement or protection (Prescott et al., 2000).
This mechanism is ofien seen in penicillin, macrolide, lincomycin, streptomycin, and
quinolone families. As an example, methicillin-resistant S. aureus expressing the mecA
gene synthesize a penicillin-binding-protein that has a lower affinity for methicillin

(Walsh, 2003).

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USE OF ANTIMICROBIALS IN ANIMAL FEED:

The idea of using antimicrobial agents in animal feed was presented in a report by
Moore et al in 1946 (Moore et al., 1946) in a study where chicks fed streptomycin and
sulfasuxidine in purified diets had accelerated growth. This idea was further supportedby
another study in 1948 that found a similar result (Stokstad et al., 1949). During this time,
research was focused on an “animal protein factor” (APF) as a growth promtant, the
activity of which was derived from vitamin B12. Stokstad’s study found that an APF
fermented from Steptomyces aureofaciens contained an unidentified growth factor, which
caused greater growth than in chicks fed diets only supplemented with vitamin B12
(Stokstad et al., 1949). Similar results were observed in pigs and turkeys when fed the
same fermented material used by Stokstad et al (Cunha et al., 1949; McGinnis et al.,
1949). The fermented product was later found to contain aureomycin, otherwise known
as chlortetracycline (McGinnis et al., 1949; Stokstad and Jukes, 1950). Finding that
antimicrobials increased the growth rate in animals was somewhat surprising to scientists,
as work with sulfonamides in rats lead to vitamin deficiencies and decreased growth rates
(Jukes, 1971). It was eventually determined that diets containing sulfonamides fed to rats
suppress normal intestinal bacterial flora that synthesize vitamins. Although numerous
studies with chickens and pigs determined the beneficial effect of antimicrobial agents in
animal feed on growth, it remains unclear what mechanisms are involved.

Although early studies in dairy calves found no benefit of antimicrobial agents in
calf feed (Rusoff and Haq, 1950; Williams and Knodt, 1950), Loosli and Wallace
determined APF, or aureomycin, in milk fed to dairy calves increased weight gain and

decreased the frequency of diarrhea when compared to the controls (Loosli and Wallace,

23

1950). Subsequent trials determined dairy calf feed supplemented with either APF or
aureomycin increased weight gain (Bartley et al., 1950a; Bartley et al., 1950b; Hogue et
al., 1957b; Lassiter, 1955; Loosli and Wallace, 1950; Rushoff, 1950; Rusoff et al., 1951).

These findings led to multiple studies of antimicrobial agents in feed of young
dairy calves. In 1955, Lassiter conducted a review of the published literature regarding
antimicrobial agents in dairy calves and as a growth promotant (Lassiter, 1955). In this
review, there was an attempt to discuss all antimicrobials being studied in dairy calves
(mainly aureomycin and terramycin), the effective antibiotic level, the effect of route of
administration on growth, and the effect of combinations of antimicrobial agents.
Lassiter concluded that when antimicrobial agents were added to feed, especially in milk,
that growth benefits were likely. The growth benefits ranged from a 10 to 30 percent
increase over the controls in the first 16 weeks of life. Most of the benefit was observed
in the first 8 weeks. Addition of antimicrobial agents to feed also led to decreased
frequency of diarrhea and mortality. These benefits were reported when the
antimicrobial agents where fed at 0.33 to 0.44 mg/kg or 15-20 mg per 45 kilograms of
body weight daily; and higher levels had no additional benefit (Lassiter, 1955).

Studies investigating antimicrobial agents in milk replacer continued to
demonstrate benefits in growth (Brown et al., 1960; Everett et al., 1958; Felsman et al.,
1973; Hogue et al., 1957a; Hvidsten, 1959; Jorgensen et al., 1964; Morrill et al., 1977;
Radisson et al., 1956; Rusoff et al., 1959; Swanson, 1963; Thomas et al., 1959).
However, over this period of time there were also reports that demonstrated no
statistically significant benefit of antimicrobial agents in milk fed to dairy calves (Bush et

al., 1959; Edwards, 1962; Gabrilidis, 1972; Lassiter et al., 1958; Preston et al., 1959). As

24

is seen in a report with mixed review on the benefit of antimicrobial agents based on
studies conducted in 1956 and 1960, in which a statistically significant higher rate of
growth was observed in a single group for both years when compared to controls, while
all other groups and treatments showed no increased growth (Edwards, 1962). Preston et
a1 and Bush et al found a benefit in growth once the calves were consuming adequate
grain that contained antimicrobial agents (Bush et al., 1959; Preston et al., 1959).

Preston et al proposed that the addition of chlortetracycline to milk did not benefit growth
because of reduced effectiveness of the drug when added to milk versus grain.

More recent literature has also offered paradoxical results as to the effects of
antimicrobial agents in milk replacer. Tomkins reported in 1991 that antimicrobial agents
improved performance and decreased the incidence of diarrhea in dairy calves (Tomkins
and Jaster, 1991). Quigley et al found a trend towards increased growth when feeding
medicated milk replacer at 57 mg/day of oxytetracycline (OTC), but there was no
statistically significant difference over the controls (Quigley et al., 1997). Additionally,
two other studies found no differences in growth between antibiotic or control groups
(Donovan et al., 2002; Heinrichs et al., 2003) when fed at 64mg and 403 mg per day.

The authors offered no hypothesis for why there was no difference between antibiotic and
control groups. However, there are multiple factors that affect calf growth; such as sex,
stress, environment, nutrition, and it is unknown if a higher proportion of gut flora in the
calves in the later studies may have been resistant to the antimicrobial agent(s) selected

for the study.

25

PHARMACOKINETICS OF OXYTETRACYCLINE IN MILK REPLACER:

The National Animal Health Monitoring System (NAHMS) Dairy Study of 2002
and 2007 reported that oxytetracycline and neomycin are the most frequently used
antimicrobial agents in milk replacer for dairy calves (USDA, 2005, 2008a).
Oxytetracycline is active against susceptible Gram-positive and Gram-negative bacteria
and is readily absorbed after oral administration to fasting animals (Plumb, 2002).
However, the presence of food or dairy products can significantly reduce the ability of
oxytetracycline to be absorbed by an animal fi'om the gastrointestinal tract (Plumb, 2002).
Because of their lipophilic nature, tetracyclines have wide distribution throughout the
body except for cerebrospinal fluid, and are eliminated unchanged primarily via
glomerular filtration (Plumb, 2002). This drug is bacteriostatic and inhibits protein
synthesis by binding to the 308 ribosomal subunit of susceptible organisms.

Phannacokinetic studies of oxytetracycline in dairy calves are limited, especially
those including administration in milk replacer (Palmer et al., 1983; Schifferli et al.,
1982). Red Holstein- Simmental crossed calves, administered oral oxytetracycline in
milk at a dose of 50 mg/kg of body weight, attained peak serum concentrations of 3.10 to
6.39 ug/ml between 6 and 12 hours after administration (Schifferli et al., 1982).
Bioavailability of oxytetracycline via oral administration was 46% and the elimination
half-life ranged from 7.95 to 15.20 hours (Schifferli et al., 1982).

A study of calves fasted overnight and fed 9 mg/kg of oxytetracycline in milk
replacer had statistically significantly lower serum concentrations than calves
administered the drug in water or an electrolyte solution (Palmer et al., 1983). Palmer et

al. also determined the area under curve (AUC) was statistically different for milk

26

replacer, water and electrolyte solution: 570, 754, and 1306 ug/ml‘min respectively.
Sixty-three percent of oxytetracycline was bound to the milk replacer, and therefore not
available for absorption (Palmer et al., 1983). Luthman et al reported similar results to
Palmer et al. with a higher dose of oxytetracycline; 50 mg/kg in cows milk, milk replacer
or water (Luthman and J acobsson, 1983). In a later study, Luthman et al administered
tetracycline chloride at 25 mg/kg in milk replacer twice a day into dairy calves resulting
in serum concentrations above lug/ml for most of the day, these values are similar to
Palmer et a1 (Luthman et al., 1989). This study also evaluated bolus administration of
tetracycline chloride at 50 mg/kg in milk replacer or 4 hours post feeding in water and
found that serum concentrations peaked at four hours in both groups, although the milk
replacer group had statistically higher serum concentrations, which contradicts the reports
cited above. Luthman did hypothesize that the lower serum concentrations attained with
the water bolus may be in part due to grain and hay consumption during the 4-hour time
frame, which may have reduced absorption of tetracycline chloride.

Young pigs also have been used as a model for oral administration of
oxytetracycline pharmacokinetics and are similar to pre-ruminant neonatal calves both
are essentially physiological monogastrics. F asted pigs, tube fed oxytetracycline at a
dose of 45 mg/kg, had increased plasma concentrations and areas under the curves than
pigs fed before administration. However, bioavailability was low for both fasted and fed
pigs; 18 and 5 percent respectively (Nielsen and Gyrd-Hansen, 1996). These
pharmacokinetic values are lower than those reported by Schifferli et al who fed the
calves at a similar level, 50 mg/kg (Schifferli et al., 1982). Similar results were seen in

earlier studies by Mevis et al and Hall et al, in which drug levels after oral administration

27

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of oxytetracycline in feed peaked at 0.2 and 0.4 ug/ml in serum respectively (Hall et al.,
1989; Mevius et al., 1986). The three studies mentioned above did have varied time to
peak concentration, ranging from 4 to 48 hours. A steady state kinetic study from the
Netherlands determined that pigs administered a dose of 54.5 mg/kg of body weight in
the feed attained plasma concentrations from 0.39-1.14 ug/ml, which is similar to studies
in pigs and calves fed at comparable levels (Pijpers et al., 1991a).

In diseased animals, the administration of antimicrobial agents through feed or
water is typically not recommended as consumption of the drug may be reduced. Pijpers
et a] determined that administration of oxytetracycline in feed to pigs at 50mg/kg that
were challenged with respiratory pathogens had shorter times to peak plasma
concentration and higher volumes of distribution and AUC when compared to the non-
challenged pigs (Pijpers et al., 1991b). A similar finding was reported following
intravenous administration of oxytetracycline in feedlot cattle that were challenged with a
respiratory pathogen when compared to healthy cattle (Ames et al., 1983).

A few studies have proposed that serum or plasma concentrations of
oxytetracycline following feeding of calves or pigs may be high enough to treat some
bacterial pathogens, based on typical minimum inhibitory concentrations (Hall et al.,
1989; Mevius et al., 1986; Schifferli et al., 1982). Schifferli et a1 looked at serum drug
concentrations to determine if they were high enough to therapeutically treat an
Escherichia coli infection in gastrointestinal tract of calves fed at 50 mg/kg.
Oxytetracycline via oral administration did attain and maintain serum drug
concentrations, peak concentration between 6 to 12 hours was 3.10 to 6.39 jig/ml and

concentration remained above 2.0 ug/ml at 24 hours; these levels were above the

29

minimum inhibitory concentration (Morrill et al., 1977) of E. coli (0.5 ug/ml) for 24
hours (Schifferli et al., 1982). However, this study was conducted in the early 19803. In
2004 the National Antimicrobial Resistance Monitoring System in the United States
reported that 65 percent of bovine E. coli isolates from feces were resistant with MIC __>_16
ug/ml for tetracycline (NARMS, 2004). Mevis et al reported that feeding oxytetracycline
to pigs at 400 ppm resulted in plasma concentrations that may be effective for some
bacterial pathogens whose MIC ranged from 0.1 to 0.5 jig/ml, such as Streptococci spp,
and F usibacterium necrophorum, but not for Pasteurella and Bordetella spp whose
MIC’s ranged from 0.2 to 3.0 jig/ml (Mevius et al., 1986). Hall et al made a similar
conclusion as plasma oxytetracycline concentration did not exceed 0.4 ug/ml when fed at

0.55 mg/kg (Hall et al., 1989).

FACTORS AFFECTING CALF GROWTH:

Holstein heifers should attain a weight of 84 kg and height of 87 cm at the withers
by 60 days of age (Heinrichs and Hargrove, 1987). Optimal grth is positively
associated with reproduction and lactation performance once a heifer enters the lactating
herd (Hoffman and Funk, 1992). Additionally, the cost of raising replacement heifers
ranges from 15 to 20 percent of the cost of producing milk in United States dairy herds
(Heinrichs, 1993). Several factors impact optimal grth of dairy replacement heifers,
including nutrition, passive transfer of immunoglobulins, disease, housing, environment,
dam, parity, and season.

Dairy replacement heifers in the United States are generally fed 8 to 10 percent of

their body weight in milk on a daily basis until they are able to consume calf-starter grain

30

at approximately 0.75 kg per day (Jurgens, 1993; NRC, 2001). This diet allows calves to
be weaned as early as 4 weeks of age. A milk replacer should contain a minimum of 20
percent crude protein, 10 percent crude fat, and a maximum of 1 percent crude fiber, with
adequate vitamins and minerals (Table 2) (Adams et al., 1995; Heinrichs et al., 2003;
NRC, 2001). However, calf growth is enhanced when they are fed milk replacers that
contain 26-30 and 20 percent protein and fat, respectively (Nonnecke et al., 2003). Calf-
starter grain should contain 16 to 20 percent crude protein and three percent crude fat
with balanced vitamins and minerals (Table 3) (Heinrichs and Jones, 2003).
Milk-based products; dried skim milk, buttermilk, whey, and casein are the preferred
protein ingredients of a milk replacer (Adams et al., 1995; Heinrichs and Jones, 2003).
Plant proteins and other sources of protein are inferior ingredients because of the
immature digestive system of a newborn calf that has limited ability to digest non-milk
proteins until after 3 weeks of age (NRC, 2001). The primary source of energy in milk
replacer is tallow, as either white grease or lard (NRC, 2001).

Dry matter intake of milk, milk replacer and/or grain affects calf growth (Bar-
Peled et al., 1997; Brown et al., 2005; Jasper and Weary, 2002; Jenny et al., 1982; Place
et al., 1998; Quigley et al., 2006; Thomas and Tinnimit, 1976). An Israeli study allowed
calves to suckle on a cow three times a day for 42 days while the control calves were fed
a limited amount of milk replacer daily. Suckled calves had a 0.85 kg average daily gain
for the first 6 weeks, which was almost 0.2 kg higher then the control calves (Bar-Peled
et al., 1997). Milk replacer with different crude protein and energy levels fed at either 1.1
percent or 2.0 percent of body weight, resulted in calves with an average daily gain of

0.668 kg for high intake diet and 0.379 kg for the low intake diet at 8 weeks of age

31

Table 1.3 Recommended milk-replacer ingredients for replacement calves

 

 

 

 

 

 

 

 

 

 

 

 

Nutrient Amount
Crude protein, min (%) 20 - 28
Fat, min (%) 10 — 22
Crude fiber, max (%) 1
Vitamin A, (IU/lb) 4091
Vitamin D, (IU/IQ 273
Vitamin E, QU/lb) 22.7
Iron (ppm) 100
Selenium (ppm) 0.3
Calcium (%) 1.0
Phosphorus (%) 0.7
Magnesium (%) 0.07

 

Source: Adapted fi'om Heinrichs, AJ and Jones CM. “Feeding the Newborn Calf”.
Extension Circular. www.pubs.cas.psu.edu

Table 1.4 Recommended calf-starter composition

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Nutrient Amount
Crude protein @1) 18 — 20
Fat (%) 3
ADF (%) 11.6
NDF (%) 12.8
MB (Mcal/lb) 1.49
Vitamin A, (IU/ lb) 1 818
Vitamin D, (IU/lb) 273
Vitamin E, (IU/lb) 11.4
Manganese (ppm) 40.0
Iron (ppm) 50.0
Copper (ppm) 10.0
Zinc (ppm) 40.0
Cobalt (ppm) 0.10
Iodine (ppm) 0.25
Selenium (ppm) 0.30
Calcium (%) 0.70
Phosphorus (%) 0.45
Magnesium (%) O. 10
Sulfur (%) 0.20
Potasium (%) 0.65

 

Source: Adapted from Heinrichs, AJ and Jones CM. “Feeding the Newborn Calf”.
Extension Circular. www.pubs.cas.psu.edu

32

(Brown et al., 2005). A Pennsylvania study investigating factors that affect dairy heifer
growth included dry matter intake as part of their final model (Place et al., 1998). Jasper
and Weary demonstrated that calves fed milk replacer ad libitum were 10.5 kg heavier at
35 days of age than calves fed conventionally, and that this difference in weight
continued until the study ended on day 63 (Jasper and Weary, 2002). However, other
studies question the benefits of higher dry matter intake on calf performance. Higher dry
matter intake was associated with increased fi'equency of diarrhea and treatment
compared to feed-limited controls (Quigley et al., 2006). This agreed with an earlier
report that found increased dry matter intake prior to weaning resulted in higher body
weight initially, but was followed by decreased average daily gains after weaning (at 4
weeks of age), and ultimately, no difference in body weight gained by 6 weeks of age
(Jenny et al., 1982).

Calves rely on passive transfer of immunoglobulins in colostrum for maternal
antibodies due to a lack of in-utero transfer of these antibodies in the bovine. Calves
should receive immunoglobulins via colostrum within the first 24 hours of life to allow
absorption of macromolecules, such as immunoglobulins, before the gut wall becomes
impermeable. After 24 hours of age, the passive transfer of antibodies across the gut wall
ceases or is limited (Franklin et al., 1998; Stott et al., 1979). Serum immunoglobulin G
(IgG) concentration in calves is positively correlated to calf health and grth (Berge et
al., 2005; Davidson et al., 1981; DeNise et al., 1989; Donovan et al., 1998a; Nocek et al.,
1984; Pare. et al., 1993; Robison et al., 1988; Van Donkersgoed et al., 1993; Virtala et al.,
1996a; Wittum and Perino, 1995). Nocek et a1 determined calves fed high quality

colostrum, (_>_ 60 mg/ml of immunoglobulin in milk) had higher growth rates in the first

33

 

four days of life compared to those fed low quality colostrum (Nocek et al., 1984). A
study of 1000 Holstein calves found that those high serum concentration of
immunoglobulins (>8 mg/ml) had higher average daily gains than calves with low serum
immunoglobulin concentration (Robison et al., 1988). High concentrations of
immunoglobulins in serum were also associated with a decreased number of days that
calves were affected with diarrhea (Pare. et al., 1993). More recently, a California study
determined decreased morbidity and mortality in calves with serum IgG 2 1000 mg/dL
(Berge et al., 2005). However, while positively impacting calf health, a Florida study
suggested that colostral immunoglobulins did not impact calf growth (Donovan et al.,
1998b). A possible reason for this discrepancy was that earlier studies did not control for
disease in their analysis, and if included, immunoglobulin concentrations were not
associated with calf growth. Additionally, growth was only monitored for a short period
of time in some of the studies (Davidson et al., 1981; Nocek et al., 1984), and if measured
for longer durations (greater age), the benefit of colostral immunoglobulins on calf
growth may diminish.

Infectious diseases also impairs calf growth; diarrhea typically is the primary
concern during the first two weeks of life, and pneumonia is a concern for the remainder
of their growing phase until first calving (Roy, 1980). In review, Simensen and Norheirn
emphasized the impact of enteric and respiratory disorders on calf health and grth
(Simensen and Norheim, 1983a). Multiple studies have demonstrated the impact of
disease on calf health and growth (Berge et al., 2005; Curtis et al., 1989; Donovan et al.,
1998a; Ganaba et al., 1995; Heinrichs et al., 2005; Lundborg et al., 2005; Lundborg et al.,

2003; Place et al., 1998; Van Donkersgoed et al., 1993; Virtala et al., 1996a; Waltner-

34

Toews et al., 1986). A study of Holstein calves in Florida documented the negative effect
of diarrhea on weight gain through six months of age (Donovan et al., 1998b). Calves
with diarrhea in the first 90 days of life are 2.5 times more likely to leave the herd, and if
remaining, are three times more likely to calve after 900 days of age (Waltner-Toews et
al., 1986). Contrary to the above study, Curtis et a1 (Curtis et al., 1989) found that
diarrhea had no effect on calves leaving the herd, which may have resulted from
differences in recording morbidity between the studies. Similar findings were observed
in a Quebec study; calves that were ill from diarrhea during the first two weeks of life
compensated for lost weight gain by weaning age (Ganaba et al., 1995). Lundborg et al
found diarrhea and pneumonia were both risk factors for decreased calf growth
(Lundborg et al., 2003). Van Donkersgoed et al also determined pneumonia affected
chest girth, but diarrhea did not (Van Donkersgoed et al., 1993). A New York study
reported that pneumonia verified by a veterinarian was associated with decreased weight
gain of 3.8 kg for the first 90 days, while producer identified pneumonia was not (V irtala
et al., 1996a). The above studies suggest that both the diagnosis and severity of diarrhea
and pneumonia is subjective, and the impact of these diseases on calf growth is difficult
to assess without standard clinical benchmarks.

Calves in the US. are traditionally raised in individual hutches, and most studies
support the concept that this type of housing increases weight gain, while decreasing
disease and cross-sucking (Maatje et al., 1993; Simensen, 1982; Tomkins, 1991; Van
Putten, 1982). In review, Simensen and Norehim summarized that individual housing in

hutches improved grth compared to calves housed within a stanchion barn with cows

(Simensen, 1982), although some studies suggested group housing was not detrimental to

35

calf health. A Canadian study reported that housing calves in pairs did not affect calf
health, performance and behavior (Chua et al., 2002; Kung et al., 1997). Additionally,
Kung et a1, determined that suckling calves raised in groups performed as well as calves
in individual hutches (Kung et al., 1997). Several of the above mentioned authors
suggested that more research is needed to determine the importance of behavioral
patterns and social interactions when comparing group and individual housing impact on
growth (Chua et al., 2002; Simensen, 1982; Wilson et al., 1999).

Evidence for environmental factors such as temperature, relative humidity,
ammonia content in air, and air movement on calf growth is mainly anecdotal. The
optimum environmental temperature for raising calves is between 15 to 25°C, with a
relative humidity of 0.6 to 0.8, though these ranges vary with age of animal (Davis and
Drackley, 1998; Roy, 1990). Environmental temperatures outside the thermoneutral zone
(5 to 20°C) and especially below the lower critical temperature require more energy for
maintenance (Davis and Drackley, 1998; Roy, 1990). A Wisconsin study demonstrated
that calves housed at 21°C versus 3°C on the same diet would gain 586g and 20g daily,
respectively (Gebremedhin et al., 1981). Similar findings where also observed in a study
from Pennsylvania (Scibilia et al., 1987). Calves maintained at a relative humidity of
0.95 on wooden slats had an increased incidence of diarrhea compared to calves
maintained at a relative humidity of 0.75 (Roy, 1990). Others did not find an impact on
calf growth with varying relative humidity and temperatures (Place et al., 1998; Roy,
1990). What may be more detrimental to calf health are large variations in enviroMental
temperature and relative humidity in short periods of time (Roy, 1990). Proper

ventilation, of four complete air exchanges per hour, is believed to decrease the exposure

36

of the calf’s respiratory tract to ammonia, pathogens, and dust (Davis and Drackley,
1998). Natural ventilation, as with hutches, promotes healthier calves (Davis and
Drackley, 1998), although drafty conditions should be avoided (Roy, 1990).

A common practice in the United States is to use wheat straw for dairy calves as
bedding. An Arkansas study found no difference in calf growth when other bedding
material was used (Panivivat et al., 2004), although calves bedded with sand and granite
fines were treated more during the first two weeks of life for diarrhea. Simensen and
Norheirn cited that calves housed above liquid manure pits had increased morbidity and
decreased grth (Simensen and Norheim, 1983a). A study of 51 herds in Norway
demonstrated that calves between 31 to 90 days of age raised on solid floor with limited
bedding, or with deep bedding (sawdust/wood shavings/straw), grew slower compared to
calves raised on slotted floors with or without litter (Simensen and Norheim, 1983a).

Limited literature is available regarding the growth of dairy calves that are born as
twins. There is general agreement that twin calves are smaller at birth and will grow
slower (Ganaba et al., 1995; Lundborg et al., 2003). A study in beef cattle demonstrated
that single calves had a 15 percent greater chance of surviving to 200 days of age then
twin calves (Gregory et al., 1996). The single calves were 8.8 kg heavier at birth and 28
kg heavier at 200 days of age.

Other risk factors impacting dairy calf growth have been investigated, although
reports are limited. A Florida study determined that there was no association between
season of birth (summer, winter) and weight gain (Donovan et al., 1998b). A Norwegian
study of 51 dairy herds found chest girth size increased for the first 90 days of life for

calves born during late summer and early fall which contradicts the findings of 795

37

Holstein calves in Pennsylvania that had improved growth when born in winter and
spring (Place et al., 1998; Simensen and Norheim, 1983b). These same two studies also
reported an association between parity of the dam (first lactation) and decreased growth
of the calf (Place et al., 1998; Simensen and Norheim, 1983b). Reports have suggested
the benefit of family members as compared to hired staff in caring for calves (Hartman et
al., 1974; Jenney et al., 1981; Martin et al., 1975; Simensen, 1982; Speicher and Hepp,
1973) while others have reported no benefit (Hagstad et al., 1984; James et al., 1984;

Lundborg et al., 2005; Oxender et al., 1973).

CONCLUSION:

Research over the past 50 plus years has generally shown that when antimicrobial
agents are added to milk replacer, dairy calf growth is increased. However, there are
limited studies in the past 20 years showing thisbenefit, especially since preventive
medicine practices have been implemented (quality colostrum, proper nutrition, housing
and use of vaccines) to improve the health and well being of dairy calves. Secondly, drug
concentrations of oxytetracycline in milk replacer have varied over time and there are no
pharrnacokinetic studies with the current dose used by the dairy cattle industry in milk
replacer to benefit growth and disease prevention. Finally, the public health community
scrutinizes the use of antimicrobial agents in animal feed, as antimicrobial resistance is a
public health concern. Using antimicrobial agents at non-therapeutic doses, as is
practiced in animal feed, there is a potential that resistance can develop due to survival of

the fittest, as not all bacteria will die when exposed to drugs at low levels.

38

Cited Literature

Aarestrup, F. (2006). Antimicrobial Resistance In Bacteria of Animal Origin. Washington
DC, American Society for Microbiology.

Adams, R. S., J. W. Comerford, et al. (1995). Dairy Reference Manual. Ithaca NY,
Northeast Regional Agricultural Engineering Service.

Ames, T. R., V. L. Larson, et al. (1983). "Oxytetracycline concentrations in healthy and
diseased calves." Am J Vet Res 44(7): 1354-7.

Amyes, S. G. (2001). Magic Bullets Lost Horizon The Rise and Fall of Antibiotics. New
York, Taylor and Fracis Inc.

Anderson, E. (1968). "Drug resistance in Salmonella typhimurium and its implications."
Br Med J 3: 333-339.

Bar-Peled, U., B. Robinzon, et al. (1997). "Increased weight gain and effects on
production parameters of Holstein heifer calves that were allowed to suckle fiom
birth to six weeks of age." J Dairy Sci 80(10): 2523-8.

Barnes, E. (1958). "The effect of antibiotic supplements on the faecal streptococci
(Lancefield group D) of poultry." Br Vet J 114: 333-344.

Bartley, E., F. Fountaine, et al. (1950). "The effects of APF concentrate containing
aureomycin on the growth and well-being of young dairy calves." J Animal Sci 9:
646.

Bartley, E., K. Wheatcroft, et al. (1950). "Effects of feeding aureomycin to dairy calves."
J Animal Sci 10: 1036.

Barza, M. and S. Gorbach (2002). "The need to improve antimicrobial use in agriculture:
Ecological and human health consequences." Clinical InfectiousDiseasgs
34(supplement 3): 871-143.

Berge, A. C., P. Lindeque, et al. (2005). "A clinical trial evaluating prophylactic and
therapeutic antibiotic use on health and performance of preweaned calves." _J_
Daig Sci 88(6): 2166-77.

Brown, E. G., M. J. Vandehaar, et al. (2005). "Effect of increasing energy and protein
intake on body growth and carcass composition of heifer calves." J Daig Sci
88(2): 585-94.

Brown, L., D. Jacobson, et al. (1960). "Urea utilization by young dairy calves as affected
by chlortetracycline supplementation." J Daig Sci 43: 1313-1321.

39

 

Bush, L., R. Allen, et al. (1959). "Effect of chlortetracycline on nutrient utilization by
dairy calves." J Daig Sci 42(4): 671-678.

Bywater, R. J. and M. W. Casewell (2000). "An assessment of the impact of antibiotic
resistance in different bacterial species and of the contribution of animal sources
to resistance in human infections." J Antimicrob Chemother 46(4): 643-5.

Cantani, A. (1885). "Un Tentativo di Batterioterapia." G Int Sci Med 7: 493-496.

Chain, S. E. (1980). A Short Histog of the Penicillin Discoveg From Fleming's Early
Observations in 1929 to the Present Time. The History of Antibiotics: A

Symposium, Madison, WI, American Institute of the History of Pharmacy.

Chua, B., E. Coenen, et al. (2002). "Effects of pair versus individual housing on the
behavior and performance of dairy calves." J Dairy Sci 85(2): 360-4.

Cunha, T., J. Burnside, et al. (1949). Arch Biochem 23: 324.

Curtis, C., M. White, et al. (1989). "Effects of calfhood morbidity on long-term survival
in New York Holstein herds." Prev Vet Med 7: 173-186.

Davidson, J ., S. Yancey, et al. (1981). "Relationship between serum immunoglobulin
values and incidence of respiratory disaes in calves." J Am Vet Med Assoc 179:
708-71 0.

Davis, C. L. and J. K. Drackley (1998). The Development, Nutrition. and Managment of
the Young Calf. Ames IA, Iowa State University Press.

DeNise, S. K., J. D. Robison, et al. (1989). "Effects of passive immunity on subsequent
production in dairy heifers." J Daig Sci 72: 552-554.

Donovan, D. C., S. T. Franklin, et al. (2002). "Growth and health of Holstein calves fed
milk replacers supplemented with antibiotics or Enteroguard." J Dg'g Sci 85(4):
947-50.

Donovan, G. A., I. R. Dohoo, et al. (1998). "Associations between passive immunity and
morbidity and mortality in dairy heifers in Florida, USA." Prev Vet Med 34(1):
3 1-46.

Donovan, G. A., I. R. Dohoo, et al. (1998). "Calf and disease factors affecting growth in
female Holstein calves in Florida, USA." Prev Vet Med 33(1-4): 1-10.

Edwards, S. (1962). "Effect of antibiotics on the growth rate and intestinal flora
(Escherichia coli) of calves." J Comp Path 72: 420-432.

40

Elam, J ., L. Gee, et al. (1951). "Effect of feeding penicillin on the life cycle of the chick."
Proc Soc Exp Biol Med 77: 209-213.

Elam, J ., L. Gee, et al. (1951). "Function and metabolic significance of penicillin and
bacitracin in the chick." Proc Soc Exp Biol Med 78: 832-836.

Everett, J. J ., L. Brown, et al. (1958). "Aureomycin as a protein-sparing agent and its
influence on minimum starter protein level satisfactory for nomal growth of dairy
calves." J Daigy Sci 41(10): 1407-1416.

FDA (1972). Report to the Commisioner of the Food and Drug Administration by the
FDA Task Force on the Use of Antibiotics in Animal Feeds. Rockville, MD,
Bureau of Veterinary Medicine, Food and Drug Administration,

Department of Health, Education, and Welfare: 22.

FDA. (2005). "Announces Final Decision About Veterinary Medicine." Retrieved
December 29, 2006, from
http://www.fda.gov/bbs/topics/news/2005/new01212.htmlFDA

Felsman, R., M. Wise, et al. (1973). "Effect of added dietary levels of copper sulfate and
an antibiotic on performance and certain blood constituents of calves." J Animal
_S_c_i 36(1): 157-160.

Force, F. T. (1972). Report to the Commissioner of the Food and Drug Administration,
The use of antibiotics in animal feeds. Rocksville, MD, US. Government Printing
Office.

Franklin, 8., C. Sorenson, et al. (1998). "Influence of vitamin A supplementation in milk
on growth, health, concentrations of vitamins in plasma, and immune parameters
of calves." J Da_ig Sci 81: 2623-2632.

Freeman, A. (1970). "The Swarm Report." J Am Vet Med Assoc 157(1): 13-16.

Gabrilidis, T. (1972). "Antibiotics in dairy calves' rations (abs)." J Daig Sci 55(5): 703.

Ganaba, R., M. Bigras-Poulin, et al. (1995). "Description of cow-calf productivity in
northwestern Quebec and path models for calf mortality and growth." Prev Vet
Med 24: 31-42.

Gebremedhin, K. G., C. O. Cramer, et al. (1981). "Predictions and measurements of heat

production and food and water requirements of Holstein calves in different
environments." Trans Am Soc Agg'c Eng 24: 715-720.

41

Gregory, K., S. Echtemkamp, et al. (1996). "Effects of twinning on dystocia, calf
survival, calf growth, carcass traits, and cow productivity." J Anim Sci 74: 1223-
1233.

Hagstad, H., S. Nicholson, et al. (1984). "Influence of management practices on dairy
calf mortality." Trop Vet 2: 123-127.

Hall, W. F., T. S. Kniffen, et al. (1989). "Plasma concentrations of oxytetracycline in
swine after administration of the drug intrarnuscularly and orally in feed. " J Am
Vet Med Assoc 194(9): 1265-8.

Hartman, D., R. Everett, et al. (1974). "Calf Mortality." J Daig Sci 57: 576-578.

Heinrichs, A. J. (1993). "Raising dairy replacements to meet the needs of the let
century." J Dag Sci 76(10): 3179-87.

Heinrichs, A. J. and G. L. Hargrove (1987). "Standards of weight and height for Holstein
heifers." J Daig Sci 70(3): 653-60.

Heinrichs, A. J ., B. S. Heinrichs, et al. (2005). "A prospective study of calf factors
affecting age, body size, and body condition score at first calving of holstein dairy
heifers." J Daig Sci 88(8): 2828-35.

Heinrichs, A. J. and C. M. Jones (2003) "Feeding the Newborn Dairy Calf." Extension
Circular 311 Volume, DOI:

Heinrichs, A. J ., C. M. Jones, et al. (2003). "Effects of mannan oligosaccharide or
antibiotics in neonatal diets on health and growth of dairy calves." J Dg'ry Sci
86(12): 4064-9.

Hoffman, P. C. and D. A. Funk (1992). "Applied Dynamics of Dairy Replacement
Growth and Management." J. Daig Sci. 75(9): 2504-2516.

Hogue, D., R. Warner, et al. (1957). "Comparison of antibiotics for dairy calves on two
levels of milk feeding." J Dairy Sci 40: 1072.

Hogue, D., R. Warner, et al. (1957). "Comparison of antibiotics for dairy calves on tow
levels of milk feeding." J Da_iry Sci 40: 1072-1078.

Hvidsten, H. (1959). "Studies on chlortetracycline and penicillin in the nurtrition of
young calves." Acg Agric Scand 9: 3-22.

IOM (1980). The effects on human health of subtherapeutic use of antimicrobials in
animal feeds. N. R. Council. Washington DC, National Academy of Sciences.

42

 

IOM (1989). Human Health Risks with the Subtherapeutic Use of Penicillin or
Tetracyclines in Animal Feed. 1. 0. Medicine. Washington DC, National Academy
Press.

James, R., M. McGilliard, et al. (1984). "Calf mortality of Virginia dairy herd
impovement herds." J Daig Sci 67: 908-911.

Jasper, J. and D. M. Weary (2002). "Effects of ad libitum milk intake on dairy calves." J
Daig Sci 85(11): 3054-8.

J enney, B., G. Gramling, et al. (1981). "Management factors associated with calf
mortality in South Carolina dairy herds." J Dag Sci 64: 2284-2289.

Jenny, B. F., H. J. Van Dijk, et al. (1982). "Performance of calves fed milk replacer once
daily at various fluid intakes and dry matter concentrations." J Dg'ry Sci 65(12):
2345-50.

Jensen, L., A. Harnmerum, et al. (1999). "Vancomycin-resistant Enterococcusfaecium
strains with highly similar pulsed-field gel electrophoresis patterns containing
similar Tn1546-like elements isolated fi'om a hospitalized patient and pigs in
Denmark." Antimicrob Agents Chemother 43(724-725).

J ETACAR (1999). The use of antibiotics in food-producing animals: antibiotic resistant
bacteria in animals and humans. J. Turnidge. Canberra, Biotext.

Jones, F. and S. Ricket (2003). "Observations on the History of the Development of
Antimicrobials and Their Use in Poultry Feeds." Poultry Science 82: 613-617.

Jorgensen, N., M. Owens, et al. (1964). "Effect on growth rate and health...adding
antibiotics to milk for dairy calves raised in outdoor hutches." South Dakota Farm
and Home Resear_ch 19(3): 10-13.

Jukes, T. (1971). "The present status and background of antibiotics in the feeding of
domestic animals." Ann NY Agd Sci 182: 362-379.

Jukes, T. and W. Williams (1953). "Nutritional effects of antibiotics." Pharrnacol Rev
5(4): 381-420.

Jurgens, M. (1993). Animal Feeding and Nutrition. Dubuque, IA, Kendall/Hunt
Publishing Company.

Kung, L., Jr., S. Demarco, et a1. (1997). "An evaluation of two management systems for
rearing calves fed milk replacer." J Dairy Sci 80(10): 2529-33.

Lassiter, C. (1955). "Antibiotics as growth stimulants for dairy cattle: A review. " J Dairy
_S_c_i 38: 1102.

43

Lassiter, C., R. Grimes, et al. (1958). "Influence of antibiotics on the growth and protein
metabolism of young dairy calves." J Daig Sci 41(10): 1417-1424.

Levy, S. (2002). The Antibiotic Paradmg How the Misu_se of Antibiotics Destroys Their
Curative Powers. Cambridge, Perseus Books Group.

Loosli, J. and H. Wallace (1950). "Influence of APF and aureomycin on the growth of
dairy calves." Proc Soc Exptl Biol Med 75: 531.

Lundborg, G., E. Svensson, et al. (2005). "Herd-level risk factors for infectious diseases
in Swedish dairy calves aged 0-90 days." Preventive Veterinary Medicine 68:
123-143.

Lundborg, G. K., P. A. Oltenacu, et al. (2003). "Dam-related effects on heart girth at
birth, morbidity and growth rate fi'om birth to 90 days of age in Swedish dairy
calves." Prev Vet Med 60(2): 175-90.

Luthman, J. and S. O. Jacobsson (1983). "The availability of tetracyclines in calves."
Nord Vet Med 35(7-9): 292-9.

Luthman, J ., S. O. J acobsson, et al. (1989). "Studies on the bioavailability of tetracycline
chloride after oral administration to calves and pigs." Journal of Vetering
Medicine. Series A 36(4): 261-268.

Maatje, K., J. Verhoeff, et al. (1993). "Automated feeding of milk replacer and health
control of group-housed veal calves." VEt Rec 133: 266-270.

Martin, S., C. Schwabe, et al. (1975). "Dairy calf mortality rate: influence of management
and housing factors on calf mortality rate in Tulare County, California." Am J Vet
_Rfi36: 1111-1114.

McGinnis, J ., E. Stephenson, et al. (1949). Response of chicks and turkey poults to
vitamin B12 supplements produced by fermentation with different organisms. Am
Chem Soc MeetingAtlantic City, New Jersey: 42A.

Mevius, D. J ., L. Vellenga, et al. (1986). "Pharmacokinetics and renal clearance of
oxytetracycline in piglets following intravenous and oral administration." Vet Q
8(4): 274-84.

Moore, P., A. Evenson, et al. (1946). "Use of sulfasuxidine, streptothricin, and
streptomycin in nutritional sutdies with the chick." J Biol Chem 165: 437-441.

Morrill, J ., A. Dayton, et al. (1977). "Cultured milk and antibiotics for young calves." J
Dg'ry Sci 60(7): 1105-1109.

44

 

NARMS (2004). NARMS Annual Veterinary Isolate Data. Athens, GA, United States
Department of Agriculture, Agriculture Research Services.

Nielsen, P. and N. Gyrd-Hansen (1996). "Bioavailability of oxytetracycline, tetracycline
and chlortetracycline after oral administration to fed and fasted pigs." J Vet
Pharmacol Ther 19(4): 305-11.

Nocek, J. E., D. G. Braund, et al. (1984). "Influence of neonatal colostrum
administration, immunoglobulin, and continued feeding of colostrum on calf gain,
health, and serum protein." J Dairy Sci 67(2): 319-33.

Nonnecke, B. J ., M. R. F oote, et al. (2003). "Composition and functional capacity of
blood mononuclear leukocyte populations from neonatal calves on standard and
intensified milk replacer diets." J Dg'ry Sci 86(11): 3592-604.

NRC and IOM (1999). The us_LOf dggs in food animals: benefits and risks. Washington
DC, National Academy Press.

NRC. (2001). Nutrient Requirements of Dairy Cattle. Washington, D.C., National
Academy Press.

OTA (1979). Drugs in Livestock Feed. Vol 1. T. R. Office of Technology. Washington
DC, US. Government Printing Office.

OTA (1995). Impacts of Antibiotic-Resistant bacteria. O. o. T. Assessment. Washington,
DC, US. Government Printing Office.

Oxender, W., L. Newman, et al. (1973). "Factors influencing dairy calf mortality in
Michigan." J Am Vet Med Assoc 162: 458-460.

Palmer, G. H., R. J. Bywater, et a1. (1983). "Absorption in calves of amoxicillin,
ampicillin, and oxytetracycline given in milk replacer, water, or an oral
rehydration formulation. " Am J Vet Res 44(1): 68-71.

Panivivat, R., E. Kegley, et al. (2004). "Growth Performance and Health of Dairy Calves
Bedded with Different Types of Materials." J Dm’ Sci 87: 3736-3745.

Pare" J ., M. Thurmond, et al. (1993). "Effect of birthweight, total protein, serum IgG and
packed cell volume on risk of neonatal diarrhea in calves on two California
daires." Can J Vet Res 57: 241-246.

Pijpers, A., E. J. Schoevers, et al. (1991). "Plasma levels of oxytetracycline, doxycycline,

and minocycline in pigs after oral administration in feed. " J Anim Sci 69(11):
45 12-22.

45

 

Pijpers, A., E. J. Schoevers, et al. (1991). "The influence of disease on feed and water
consumption and on pharmacokinetics of orally administered oxytetracycline in
pigs." J Anim Sci 69(7): 2947-54.

Place, N. T., A. J. Heinrichs, et al. (1998). "The effects of disease, management, and
nutrition on average daily gain of dairy heifers from birth to four months." J Daig
_Sii 81(4): 1004-9.

Plumb, D. C. (2002). Veterinary Drug Handbook. Ames IA, Iowa State Press.

Prescott, J ., J. Baggot, et al. (2000). Antimicrobial Therapy in Veterifinary Medicine.
Ames, Iowa, Iowa State Press.

Preston, T., N. McLeod, et al. (1959). "The effect of chlortetracycline on growth of early-
weaned calves." Animal Production 1.: 13-19.

Quigley, J. D., 3rd, J. J. Drewry, et al. (1997). "Body weight gain, feed efficiency, and
fecal scores of dairy calves in response to galactosyl-lactose or antibiotics in milk
replacers." J De_u_ry' Sci 80(8): 1751-4.

Quigley, J. D., T. A. Wolfe, et al. (2006). "Effects of additional milk replacer feeding on
calf health, growth, and selected blood metabolites in calves." J Daig Sci 89(1):
207-16.

Radisson, J ., C. Smith, et al. (1956). "The mode of action of antibiotics in the nutrition of
the dairy calf. I. Effect of terramycin administered orally on the performance and
intestinal flora of young dairy calves." J Daria Sci 39: 1260-1267.

Randall, C. (1969). "The Swann Committee." Vet Rec 85: 616-621.

Roberts, W. (1874). "Studies of Abiogenesis." Phil Trans 104: 466.

Robison, J. D., G. H. Stott, et al. (1988). "Effects of passive immunity on growth and
survival in the dairy heifer." J Dairy Sci 71(5): 1283-7.

Roy, J. H. (1980). "Symposium: disease prevention in calves. Factors affecting
susceptibility of calves to disease." J Dg'g Sci 63(4): 650-64.

Roy, J. H. (1990). The Calf Vol. 1: Management of health. London, Butterworths.
Rushoff, L. (1950). "A.P.F. supplements for calves." J Animal Sci 9: 666.
Rusoff, L., A. Cummings, et al. (1959). "Effect of high-level administration of

chlortetracycline at birth on the health and grth of young dairy calves." J Dairy
S91 42: 856-862.

46

Rusoff, L., A. Davies, et al. (1951). "Growth-promoting effect of aureomycin on young
claves weaned from milk at an early age." J Nutrition 45: 289.

Rusoff, L. and M. Haq (1950). J Dg'ry Sci 33: 379.

Salyers, A. and D. Whitt (2002). Bacterial Pathogenesis A Molecular ApproaLh.
Washington DC, ASM Press.

Schifferli, D., R. L. Galeazzi, et al. (1982). "Pharmacokinetics of oxytetracycline and
therapeutic implications in veal calves." J Vet Pharmacol Ther 5(4): 247-57.

Scibilia, L. S., L. D. Muller, et al. (1987). "Effect of environmental temperature and
dietary fat on growth and physiological responses of newborn calves." J Daig Sci
70: 1426-1433.

Sefion, A. (2002). "Mechanisms of Antimicrobial Resistance, Their clinical relevance in
the new millennium." Drugs 62(4): 557-566.

Simensen, E. (1982). "An Epidemiological Study of Calf Health and Performance in
Norweigan Dairy Herds 1. Mortality: Literature Review, Rates and
Characteristics." Acta Agric Scand 32: 411-419.

Simensen, E. and K. Norheim (1983). "An Epidemiological Study of Calf Health and
Perfomance in Norwegian Dairy Herds III. Morbidity and Permformance:
Literature Review, Characteristics." Acta Agfi: chd 33: 57-64.

Simensen, E. and K. Norheim (1983). "An Epidemiological Study of Calf Health and
Performance in Norweigan Dairy Herds V. Changes in Performance with Season
and Age." Acta Agric Scand 33: 129-135.

Smith, W. (1975). "Persistence of tetracycline resistance in pig E.coli." Nature 258: 628.

Smith, W. (1977). Antibiotic resistance in bacteria_and associated probelms in farm
animals before and after the 1969 Swarm Report. London, Butterworths.

SOU (1997). Antimicrobial feed additives. . Report from the Commission on
Antimicrobial Feed Additives. Stockholm. 1997:132.

 

Speicher, J. and R. Hepp (1973). "Factors associated with calf mortality in Michigan
dairy herds." J Am Vet Med Assoc 162: 463—466.

Stokstad, E. and T. Jukes (1950). "Growth-promoting effect of aureomycin on turkey
poults." Poultry Science 29: 611-612.

Stokstad, E., T. Jukes, et al. (1949). "The multiple nature of the animal protein factor." J
Biol Chem 180: 647-654.

47

Stott, G., D. Marx, et al. (1979). "Colostral immunoglobulin transfer in calves. I. Period
of absorption." J Dairy Sci 62: 1632-1638.

Swann, M. (1969). Report of Joint Committee on the Use of Antibiotics in Animal
Husbandry and Veterinary Medicine. London, Her Majesty's Stationary Office.

Swanson, E. (1963). "Effects of chlortetracyline in calf starter and milk." J Dag Sci 46:
955-958.

Technology, C. o. A. S. a. (1981). Report 88, Antibiotics in animal feeds. C. f. A. S. a.
Technology. Ames, IA.

Tenover, F. (2006). "Mechanisms of Antirncirobial Resistance in Bacteria." Ih_e
American Journal of Medicine 119(6A): S3-SlO.

Thomas, J ., R. McDowell, et al. (1959). "Effects of feeding aureomycin to dairy calves."
J Dg'ry Sci 42(4): 658-665.

Thomas, J. W. and P. Tinnimit (1976). "Amounts and sources of protein for dairy
calves." J Dm‘ Sci 59(11): 1967-84.

Tomkins, T. (1991). Loose-housing experience in North America Proc. of the Int. Symp.
on Veal Calf Production, Wageningen, Netherlands, EEAP.

Tomkins, T. and E. Jaster (1991). Preruminant calf nutrition. Vet Clin North Am Food
Anim Pract. 7: 557.

USDA (2005). Part IV: Antimicrobial Use in U. S. Dairy Operations, 2002. National
Animal Health Monitoring System. USDA:APHIS:VS:CEAH. Fort Collins, CO:
61.

Van Donkersgoed, J ., C. Ribble, et al. (1993). "Epidmeiological study of enzootic
pneumonia in dairy calves in Saskatchewan." Can J Vet Res 57: 247-254.

 

Van Putten, G. (1982). "Welfare in veal calf units." Vet Rec 111: 437-440.

Virtala, A.-M., G. Mechor, et al. (1996). "The effect of calfliood diseases on growth of
female dairy calves during the first 3 months of life in New York State." J Dairy
§c_i 79: 1040-1049.

Wainwright, M. (1990). Miracle Cure. Cambridge, Basil Blackwell Ltd.

Walsh, C. (2003). Antibiotics: Actiorrs, OfiginaLResistance. Washington DC, ASM Press.

48

Waltner-Toews, D., S. W. Martin, et al. (1986). "The effect of early calflrood health
status on survivorship and age at first calving." Can J Vet Res 50(3): 314-7.

WHO. (1997). "The Medical Impact of the Use of Antimicrobials in Food Animals.
Report of a WHO meeting. Berlin, Germany, 13-17 October, 1997."
WHO/EMC/ZOO/97.4 Retrieved December 19, 2006, from
http://whqlibdoc.who.int/hq/1997/WHO EMC ZOO_97.4.pdf.

WHO (2003). Impacts of antimicrobial growth promotion termination in Denmark.
Geneva, World Health Organization.

WHO (2003). Joint FAO/OIE/WHO Expert workshop on non-human antimicrobial usage
and antimicrobial resistance: Scientific assessment. Geneva.

Wierup, M. (2001). "The Swedish experience of the 1986 year ban of antimicrobial
growth promoters, with special reference to animal health, disease prevention,
productivity, and usage of antimicrobials." Microb Drag Resist 7(2): 183-90.

Williams, J. and C. Knodt (1950). J Dg'g Sci 33: 380.

Wilson, L. L., T. L. Terosky, et al. (1999). "Effects of individual housing design and size
on behavior and stress indicators of special-fed Holstein veal calves." J Anim Sci
77(6): 1341-7. '

Wittum, T. and L. Perino (1995). "Passive immune status at postpartum hour 24 and
long-terrn helath and performance of calves." Am J Vet Res 9(1149-1154).

49

CHAPTER 2

50

EFFECTS OF FEEDING MEDICATED OR NON-MEDICATED MILK
REPLACER TO HOLSTEIN CALVES ON GROWTH, MORBIDITY AND
MORTALITY

Averill, JJ‘, Erskine, RJ', and Bartlett, PC’
1Michigan State University, College of Veterinary Medicine. East Lansing MI 48824.
ABSTRACT

Three hundred and one Holstein heifer calves were randomly assigned to be fed a
milk replacer with 440 mg/kg of neomycin and 220 mg/kg of oxytetracycline, or a milk
replacer with no antimicrobial agents added. Calves were fed approximately 220g of milk
replacer twice a day starting at three days of age and were provided ad libitum access to
water and grain until weaning at 7 weeks of age. Occurrence of health events and weekly
gains in height and weight were recorded until weaning and were again obtained at 150
days of age. Height at birth, 42- and ISO-days of age did not differ between treatment
groups (P>0.4). However, the medicated treatment group had higher weights at 42- and
ISO-days of age (P<0.05), and weeks 5 through 8 (P<0.01). Calves with a pre-weaning
episode of respiratory disease gained less weight than calves without respiratory disease
(P<0.01). Calves born in the fall and winter months had higher weights than calves born
in the summer and spring (P5 0.05). There was no difference in morbidity rates for
enteritis, respiratory, and other diseases but there was a trend towards overall lower
mortality rates in the medicated group (P= 0.099). Our results suggest that calves fed

medicated milk replacer grow faster than calves fed non-medicated milk replacer,

although there was no difference in morbidity for any particular disease.

51

INTRODUCTION

In 1955, a review of the published literature concluded that antimicrobial agents
added to feed, especially in milk, would likely increase growth rates in dairy calves
(Lassiter, 1955). Growth rates increased from 10 to 30 percent in the first 8 weeks of life.
Over the next 20 years, additional studies reported improved growth from the feeding of
antimicrobial agents in milk replacer (F elsman et al., 1973; Hvidsten, 1959; Morrill et al.,
1977; Radisson et al., 1956; Swanson, 1963). However, other studies demonstrated no
statistically significant benefit of antimicrobial agents in milk fed to dairy calves (Bush et
al., 1959; Edwards, 1962; Gabrilidis, 1972; Lassiter et al., 1958; Preston et al., 1959).

More recent literature has also offered paradoxical results as to the effects of
antimicrobial agents in milk replacer. Tomkins and J aster reported that antimicrobial
agents improved performance and decreased the incidence of diarrhea in dairy calves
(Tomkins and J aster, 1991). Quigley et al found a trend towards increased growth in
feeding medicated milk replacer, but there was no statistically significant difference
compared to the controls (Quigley et al., 1997). Two additional reports have found no
differences in growth between antibiotic or control groups (Donovan et al., 2002;
Heinrichs et al., 2003).

Animal agriculture has received scrutiny from the feeding of antimicrobial agents
at subtherapeutic levels because of the potential for development of antimicrobial
resistance (WHO, 2003b). The European Union has banned all antimicrobial agents for
growth promotion (EUROPA, 2002). Thus, the use of antimicrobials in milk replacer fed

to dairy calves, especially if benefits are equivocal, may be cause for concern. Over the

52

years the concentration of antimicrobial agents in milk replacer have varied. Industry
standards today use 440 mg/kg of neomycin and 220 mg/kg of oxytetracycline in milk
replacer fed to dairy calves, as medicated milk replacers are used in over 50% of dairy
farms in the United States. Studies demonstrating a benefit in growth promotion for
feeding medicated milk replacer have generally only followed the calves to weaning, so
there is no literature demonstrating if the difference in growth is maintained further into
the growing heifers life.

The objectives for this study was to determine the effect of feeding milk replacer
supplemented with or without antimicrobial agents on the height, weight, morbidity, and
mortality of dairy calves from birth to five months of age between the two treatment
groups. Secondly, apply a repeated measures model to determine risk factors affecting

growth in these calves and determine age at which two treatment groups differ in growth.

MATERIALS AND METHODS

The Institutional Animal Care and Use Committee of Michigan State University
approved the experimental protocol of this study.

m A commercial 600-cow dairy farm was selected based on willingness to
participate and their ability to keep records. At birth, all calves had navels dipped with
iodine, weight recorded, were placed in an individual hutch bedded with straw, and then
received one gallon of quality colostrum, immunoglobulin level >50 mg/ml.

Three hundred and one heifer calves were enrolled over a 13-month period. Each animal
was enrolled from three days until five months of age. Inclusion criteria for the study

were: Holstein heifer, non-twin, no signs of birth defects, and over 250 days in-utero.

53

The farm was blinded as to which milk replacer was medicated; all feeding equipment,
milk replacer bags and hutches were color coded to maintain consistent feeding of the
correct treatment assignment.

Milk Replacer: At birth, calves were randomly assigned to one of the two study
groups: milk replacer without antimicrobial agents (n = 151) or milk replacer with
antimicrobial agents (11 = 149). The medicated milk replacer had 440 mg/kg of
neomycin and 220 mg/kg of oxytetracycline. Milk replacer was fed at approximately
220g, dissolved in 2 liters of water, twice a day until one week prior to weaning when
they were reduced to once a day feeding. Calves received a total dose of about 45 mg of
oxytetracycline (1mg/kg twice a day) and 90 mg of neomycin (2 mg/kg twice a day) at
each feeding. The composition of the milk replacer included 20 percent crude protein, 20
percent crude fat and a crude fiber less then 0.15 percent. The protein was entirely
derived from milk derivatives.

Calf Management: Within the first day, the calves received injections of Vitamin
A, D and E, selenium, and an intranasal modified-live Infectious Bovine Rhinotracheitis
and Parainfluenza-3 vaccine. All calves in the study received vaccines per the farm’s
vaccine management schedule for replacement heifers. Calves received two liters of
colostrum twice a day until being placed on milk replacer at three days of age. Calves
were offered fresh water and a calf-starter, ad libitum, beginning on day 3. Once weaned,
calves were moved into group housing with other heifers of the same age and size.
Calves remained in this transition group until approximately 5 months of age. At
weaning, they were initially fed calf starter and hay and slowly transitioned to a com-

silage-based total mixed ration (TMR). .

54

Data Collection: Height and weight data were collected weekly until weaning and
once at five months of age before leaving the transition barn. Health records of
individual animals were also monitored during weekly visits to the farm. Body weight
was obtained with a digital scale that was calibrated each week. Wither height was
obtained with a sliding ruler; measurements were taken three times and averaged.

Assess_ment of Passive Tran__s_f_‘e_r_: In order to assess serum total protein, a blood
sample was collected from each calf via jugular venipuncture with a 20G needle into a 10
ml vacutainer during the first week of birth. The sample was placed on ice, and allowed
to clot. The vacutainers were centrifuged at 1,000 x g for 10 minutes. Serum was
harvested and a total solid was determined with a refi'actometer (Reichert TS-Meter,
Model 1310400A, Depew NY). The remaining serum was frozen at -80°C until radial
immunodiffusion was performed by a commercial kit (V MRD, Inc., Pullman WA).
Briefly, 3 ul of serum was placed into each well and held at room temperature for 18 to
24 hours. Controls and specimen diameters were measured and plotted on a semi-log
graph to determine immunoglobulin G concentration.

Case definitions: We used the following case definitions of clinical disease in
calves for this study.

An ENTERIC CASE at least exhibited loose stool and rectal temperature >39.5°C, and
may also show signs of depression, anorexia, off feed and/or dehydrated.

A RESPIRATORY CASE presented with coughing or abnormal thoracic sounds, mucous
discharge, and rectal temperature >39.5°C, and may also shown signs of depression.
OTHER INFECTIOUS CASES included: calves that did not match the enteric or

respiratory definition but had an elevated temperature or keratoconj unctivitis.

55

OTHER NON-INFECTIOUS CASES included calves that were anorexic, lame, or
bloated.

Statistical analysis: Weekly data (weight, height, and health records), passive
transfer of antibodies and management practices were entered into a Microsoft Access
database (2000, Microsoft Corporation, Redmond, WA). Separate investigators reviewed
the data to detect data entry errors. Microsoft Excel (2000, Microsoft Corporation,
Redmond, WA) was used to calculate morbidity and mortality rates. Average daily gain,
and adjusted 42-day and ISO-day weight and height were calculated with SAS v.9.1
(Statistical Analysis System Institute, Cary, NC). PROC ANOVA procedure was
performed with SAS v.9.1 and used to make comparisons in growth variables between
the two treatment groups at birth, weaning and 5 months of age. Weekly weight was
analyzed as a repeated measure by PROC MIXED procedure with first-order
autoregressive covariance structure. Effect variables treatment and week were forced
into the model while all others were included if they initially had a P-value of 0.20 or
less. The final model included effect variables with a P-value less than or equal to 0.05.
All values from each calf were used in the modeling, even if the calf died. Categorical
data, such as morbidity and mortality, was compared using EPI INFO v.3.4.3 (Centers for
Disease Control and Prevention, Atlanta GA). Results were expressed as relative risk
with confidence intervals. Statistically significant differences were determined when the

confidence interval did not include 1.0.

56

RESULTS

There was no difference in starting wither height of 76.41 and 76.52 cm (P=0.72)
and birth weight of 42.45 and 42.35 kg (P= 0.85) for the medicated and non-medicated
groups, respectively (Table 2.1). Birth weight of thirty-seven calves were not recorded,
19 for the medicated group and 18 for non-medicated group. A mean weight of 42.4 kg
was used for these calves to calculate their growth from birth. The first recorded height,
either at birth or within week born, was used as the starting height for calculating change
in height for each calf.

Sixty-six calves did not receive any treatments dming the study, thirty-three from
each treatment group (Table 2.3). There was no statistical difference between the two
groups for the number of respiratory, enteric, other infectious, or other non-infectious
cases, as the relative risk confidence intervals included 1.0 (Table 2.2). A majority
(135/ l 39) of the enteric cases occurred prior to weaning, and a total of four cases
occurred post-weaning. Conversely, respiratory cases (110/147) tended to occur more
frequently after 42 days. Calves had fewer other cases of disease during the post-
weaning period, compared to the pre-weaning period. Non-infectious cases were
predominantly bloat and lameness while cases in the other infectious category included
elevated temperature of unknown origin, infectious bovine keratoconjunctivitis and/or
otitis. The medicated milk replacer treatment had 50 calves with two or more disease
cases during the study, while the non-medicated treatment had 37, relative risk of 1.14
(CI=0.98 to 1.32) (Table 2.3).

A numeric difference in mortality rate occurred by eight weeks of age, 7.4% and

1 3.2 % for the medicated and non-medicated groups, respectively, Mantel-Haenszel Chi-

57

Square (MB) of 2.72 (P=0.099). At five months of age, mortality rate for the medicated
group was 8.7% and 15.1% for non-medicated group, respectively MH of 2.92
(P=0.087). Sixty-nine percent of deaths occurred during the first two weeks of life
(Figure 2.1). Twenty-three deaths (Table 2.4) occurred in the non-medicated group and
thirteen in the medicated group. For the medicated group, the case fatality rates for
respiratory, enteric, and other cases are; 1.32, 12.9 and 10.7 % respectively, while the
non-medicated group had similar results; 1.41 , 23.2, and 16.2 % for respiratory, enteric,
and other cases, respectively.

Initial wither height measurements were not different among treatment groups
(Table 2.1). There was no statistical difference in wither height between treatment
groups at 42— and ISO-days of age (Table 2.1). Initial body weight did not differ between
treatment groups (Table 2.1). However, calves fed the medicated milk replacer had
higher mean body weights at 42 days (P= 0.011), and at 150 days (P= 0.0015) (Table
2.1). Calves in both treatment groups experienced a positive weight gain throughout the
first 8 weeks of age except for the second week. Modeling weight of calves at weekly
intervals from birth to 8 weeks of age by repeated measures determined that the
medicated group had higher weights fi'om week five through eight than the non-
medicated group (P<0.01). The final model had a Bayesian Information Criteria (BIC) of
11707.5 and included the following variables; treatment, week, weight at birth (wtb),
season born (season), respiratory case (resp), and interaction terms: treatrnent*week and
week‘resp (Table 2.5).

Since the occurrence of a respiratory case was included in the model, weights

were analyzed for those with respiratory cases (n=59) or no respiratory cases (n=242)

58

pre-weaning. Weights of calves with a pre-weaning respiratory case were lower
compared to calves without a pre-weaning respiratory case (P=0.008) at 8-weeks of age
when treatment groups were combined (Figure 2.2). There also was a difference in
weight between treatment groups at 8 weeks of age for calves with a pre-weaning
respiratory case as the medicated group was 5 kg (P=0.036) heavier, and for the non-
respiratory cases as the medicated group gained an additional 1.4 kg (P=0.01).

Weight gained at 42 days of age was stratified by season of birth. There was no
difference in weight gained between treatments groups within season born (P>0.05).
However, there were statistical differences in weight gained between seasons (Figure
2.3), as spring born calves were lightest, followed by summer and fall born calves and
winter born calves gained the most weight (P<0.05). Mean total protein for the
medicated group was 5.59 g/dl (range: 4.3 to 7.2) and did not differ (P=0.39) from the
non-medicated group (5.65g/dl; range: 4.2 to 7.4). Thirty-three and 40 calves were below
5.2 g/dl for the medicated and non-medicated groups respectively. There is an

association between a total protein less than 5.2 g/dl to death (RR=1.99 (1 07-368)).

DISCUSSION

The judicious use of antimicrobial agents has been a topic of increasing concern
for animal agriculture. The American Veterinary Medical Association (AVMA) and the
Food and Drug Administration (FDA) have developed guidelines on the judicious use of
antimicrobial agents in veterinary medicine to help mitigate antimicrobial resistance
(FDA, 2001). In particular, the use of antimicrobial agents in a non-therapeutic manner

in farm animals has been criticized (WHO, 1998, 2003b). The American College of

59

Veterinary Internal Medicine has developed a position statement that voluntary actions
should be taken by the veterinary profession to conservatively use antimicrobial agents to
minimize adverse effects on animal or human health (Morley et al., 2005)

Our study is the largest conducted to date investigating the relationship between
dairy calf growth and health, and milk replacer with or without antimicrobial agents
added. This study was conducted on a well-managed 600—cow commercial dairy farm in
Michigan and there was no difference in crude morbidity between the two treatment
groups. This is contrary to previous studies that showed a difference in morbidity in
calves fed milk replacer with antimicrobial agents compared to the controls (Berge et al.,
2005; Braidwood and Henry, 1990; Heinrichs et al., 2003). Lundborg et a1 2005 found
morbidity on Swedish farms ranged fi'om 0 to 57.6% with a median of 21 .6% (Lundborg
et al., 2005). Our overall morbidity rate (78%) was also higher than the NAHMS 2007
Dairy study (38.5% in unweaned calves and 9.5% in weaned calves) (USDA, 2008c).
The mortality rate in the present study of 11.9% is higher than previously reported in
other studies as they ranged from 5.6 to 9.4% (Losinger and Heinrichs, 1997; Tyler et al.,
1998; Virtala et al., 1996b; Waltrrer-Toews et al., 1986; Wells et al., 1997). The higher
overall morbidity and mortality rates in our trial can be attributed to calves that had a
serum total protein below 5.2 g/dl and two disease outbreaks; a week-long respiratory
outbreak during the fall in weaned calves, and an enteritis outbreak in suckling calves
that occurred late fall into early winter. Increased mortality rate in this study is
suggestive of a more severe infection, though diagnostics was not conclusive on

causative agent for enteric cases. Morbidity rates may also be higher in this study as they

60

were producer diagnosed and are subjective based on the caretaker’s interpretation of
clinical signs.

Calf weight in the first eight weeks was below standard weight curves, compared
to a Pennsylvania (Heinrichs and Hargrove, 1987) and a national study (Heinrichs and
Losinger, 1998). However, calves in both treatment groups exceeded the standard
growth curves by 150 days of age. Standard height was attained for calves in this study at
8 weeks and 150 days of age (Heinrichs and Hargrove, 1987; Heinrichs and Losinger,
1998). Suckling calves were being fed approximately 220g of milk replacer twice a day
prior to weaning, which is below the standard recommendation of 250g per feeding
(Tomkins and J aster, 1991) for a 50-kg calf, which in part may explain the sub-standard
weight gain.

Medicated calves had a higher weight at 42 days and 150 days of age. This
confirmed previous studies that determined antimicrobial agents improve growth of
suckling calves (Berge et al., 2005; Morrill et al., 1977; Quigley et al., 1997). Other
studies found no statistical increase in weight gain following feeding of medicated milk
replacer, although numerical differences were observed (Donovan et al., 2002; Heinrichs
et al., 2003). Today, the dairy cattle industry adds oxytetracycline and neomycin at 220
mg/kg and 440 mg/kg to milk replacer and when feed according to the label calves
receive 1 mg/kg and 2 mg/kg twice a day for oxytetracycline and neomycin respectively.
Caution should be applied in comparing earlier studies to our results as all studies used
higher concentrations of oxytetracycline in the milk replacer. With studies using
different drug concentrations may explain the previous paradoxical results regarding the

benefit of antimicrobial agents in milk replacer. To the best of our knowledge, there are

61

no previous studies that compared weight between calves fed medicated versus non-
medicated milk replacers post-weaning.

In our repeated measures model all data was included until the calf left the study
(including data from calves prior to their death). There was no difference between
treatment groups in weight until after four weeks of age in the repeated measures model.
This is similar to findings reported by Morrill et al, which reported no difference until 3
weeks of age (Morrill et al., 1977). Other significant variables that affected weight at
weaning included birth weight, respiratory cases, and season of birth, which have been
shown to impact growth in previous studies (Place et al., 1998; Virtala et al., 1996a).
Virtala et al demonstrated that for each week a calf had a respiratory case, weight was
decreased by 0.8 kg (V irtala et al., 1996a). Dairy calves treated by producers for
pneumonia had an average weekly growth (chest girth) of 2.00 cm versus 2.47 cm in non-
treated calves (Van Donkersgoed et al., 1993).

Virtala et a] reported that calves born during the winter had higher weight gain
than calves born in the summer (V irtala et al., 1996a), which was similar to our findings.
Place et al hypothesized that differences in growth relative to season of birth may be due
to the availability of the caretaker to spend time with the calves (Place et al., 1998).
Seasonal differences were also reported by Donovan et al, but saw the inverse of the
reports stated above, with higher gains in summer rather than winter-bom calves
(Donovan et al., 2002). A likely factor in season being included in our final model may

have been due to the enteritis outbreak that started in October and extended into

February.

62

Total serum protein was not included in our final model as a variable affecting
weight gain. Studies have adequately shown that the failure of passive transfer increases
the risk of calves becoming ill and therefore decreasing growth (Jarmuz et al., 2001;
Vann and Baker, 2001; Virtala et al., 1996a). We did see that low serum total protein of
less than 5.2 g/dl is associated with death in agreement with previous studies. The non-
significant effect of an enteritis case is in agreement with previous studies (Sivula et al.,
1996; Van Putten, 1982). However, other studies have demonstrated that morbidity due
to enteritis does impact calf growth (Donovan et al., 1998b; Walther-Toews et al., 1986).
Additionally, Waltner-Toews reported that calves with enteritis were 2.5 times more
likely leave the herd in first 90 days (W altner-Toews et al., 1986). Enteritis may have not
impacted growth in our study due to almost identical number of cases between two
treatment groups in the first 8 weeks of life or the fact that calves were followed for more

than four weeks and were able to compensate for weight lost in first few weeks of life.

CONCLUSION

Calves fed milk replacer with antimicrobial agents gained more weight by 42 and
15 0 days of age than calves fed non-medicated milk replacer, with numerical differences
being seen starting at 4 weeks of age. Risk factors that influenced weight gain were
treatment group, birth weight, season of birth, and having a respiratory case. Though
there was no difference in morbidity between treatment groups, there was a non-
significant association with mortality. The benefit of feeding medicated milk replacer
may have resulted, in part, fi'om the occurrence of an enteric outbreak during the trial as

more calves survived and/or consumed more feed. Calves receiving the non-medicated

63

milk replacer had higher mortality and decreased weight gain while having equal
morbidity rate compared to the medicated milk replacer group. This suggests that the
medicated milk replacer calves recover better from enteric disease, especially since a
majority of mortalities were of enteric disease.

Given the benefit in growth for calves receiving medicated milk replacer, is a 6 kg
difference at 150-days of age biologically significant. This study is not able to answer
that question, as the calves were not followed through puberty and into their first
lactation. However, one could argue that the difference is not large enough for the added
cost of feeding medicated milk replacer. More importantly, this study shows that calves
fed medicated milk replacer have a lower mortality rate, which is very important to a
producer. With the increased scrutiny over the use of antimicrobial agents in animal feed
this study would show the benefit in growth and prevention of death. If one was to
conclude that the growth differences are not biologically significant than using medicated
milk replacer throughout the pre-weaning period may not be necessary. This would
decrease the use of antimicrobial agents and may decrease the chance for antimicrobial

resistance to develop, as the drugs would be used less fiequently.

64

Table 2.1. Mean growth for calves

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Medicated N on-Medicated P-value
# Calves Enrolled 149 152
# Calves Weaned 139 131
# Calves Finished Trial 136 129
Birth Weight (kgL 42.35 42.45 0.85
Adj. 42-Day Weight (kg) 64.18 61.99 0.011
42-Day ADG' (kg) 0.513 0.464 0.012
Adj. 150-Day Weight (kg) 181.4 175.7 0.0015
150-Day ADG‘ (kg) 0.925 0.888 0.0018
Startingfleight (cm) 76.41 76.52 0.72
Adj. 42-Day Height (cm) 82.99 82.74 0.43
42-Day ADHG2 (cm) 0.129 0.125 0.60
Adj. 150-Day Height (cm) 104.6 104.6 0.85
150-Day ADHGZ (cm) 0.188 0.187 0.93
IAverage Daily Gain
2Average Daily Height Gain
Table 2.2 Calf Morbidity Data
Medicated Non-Medicated Relative Risk (CI)
# Total Illnesses 174 177 1.00 (0.92-1.08)
Respiratory 76 71 1.10 (0.87-1.40)
Enteric 70 69 1.05 (0.82-1.34)
Other_Nl 15 20 0.86 (0.45-1.66)
Other; I 13 17 0.66 (0.34-1.27)
# Cases 542 days 116 125 0.98 (0.85-1.13)
Respiratory 27 32 0.86 (0.54-1.36)
Enteric 67 66 1.03 (0.80-1.70)
Other_NI 11 14 0.80 (0.37-1.70)
Other_I 11 11 1.02 (0.46-2.28)
# Cases > 42 days 58 52 1.13 (0.84-1.52)
Respiratory 49 39 1.19 (0.84-1.68)
Enteric 3 1 2.85 (0.30-27.01)
Other_Nl 4 6 1.81 (0.34-9.72)
Other; I 2 6 0.94 (0.31-2.85)

 

 

 

 

 

Other_NI is Other_Non-Infectious case classification

Other_I is Other_Infectious case classification

65

 

Table 2.3 Number of cases per animal

 

 

 

 

 

 

 

 

# Cases/Animal Medicated Non-Medicated
0 33 33
l 66 82
2 44 22
3 4 10
4 2 4
5 0 l

 

 

 

 

Table 2.4 Tye and number of mortalities per treatment group

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Medicated Non-medicated Relative Risk (CI)
# Total Mortalities 13 0.58 (0.30-1.10)
Enteric 9 0.57 (0.26-2.00)
Respiratory 1 1 1.02 (0.06-16.16)
Other_Infectious 0 0 --
Other_Non-infectious 3 6 0.51 (0.13-2.00)
# Mortalities 542 Days 10 20 0.51 (0.25-1.05)
Enteric 9 16 0.51 (0.26-1.26)
Respiratory 0 0 -
Other_Infectious 0 0 --
Other_pNon-infectious 1 4 0.26 (0.03-2.26)
# Mortalities >42 Days 3 3 0.97 (0.20-4.72)
Enteric 0 0 --
Respiratory 1 l 0.98 (006-1548)
Other_Infectious 0 0 -
Other_Non-infectious 2 2 0.95 (0.14-6.63)

 

 

 

 

Table 2.5. Fixed Effects values for Repeated Measure Model with Weight as Outcome

 

Type 3 Tests of Fixed Eflects

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Num Den

Effect DF DF F Value Pr>F
Treatment 1 999 2.75 0.0973
Week 8 1799 372.31 <.0001
Month 12 443 6.74 <.0001
WTB 1 779 46.68 <.0001
_ Resp 1 2226 0.50 0.4748
Treatrnent*Week 8 1797 1.79 0.0755
Week*Resp 8 1797 2.25 0.0217

 

 

66

 

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Cited Literature

Berge, A. C., P. Lindeque, et a]. (2005). "A clinical trial evaluating prophylactic and
therapeutic antibiotic use on health and performance of preweaned calves." J
Dg'ry Sci 88(6): 2166-77.

Braidwood, J. and N. Henry (1990). "Clinical efficacy of chlortetracycline hydrochloride
administered in milk replacer to calves." Vet Rec 127: 297-301 .

Bush, L., R. Allen, et al. (1959). "Effect of chlortetracycline on nutrient utilization by
dairy calves." Dg'g Sci 42(4): 671-678.

Curtis, C., M. White, et al. (1989). "Effects of calfhood morbidity on long-term survival
in New York Holstein herds." Prev Vet Med 7: 173-186.

Donovan, D. C., S. T. Franklin, et al. (2002). "Growth and health of Holstein calves fed
milk replacers supplemented with antibiotics or Enteroguard." J Daig Sci 85(4):
947-50.

Donovan, G. A., I. R. Dohoo, et al. (1998). "Calf and disease factors affecting growth in
female Holstein calves in Florida, USA." Prev Vet Med 33(1-4): 1-10.

Edwards, S. (1962). "Effect of antibiotics on the growth rate and intestinal flora
(Escherichia coli) of calves." J Comp Path 72: 420-432.

EUROPA (2002). Commission proposes new safety rules for feed additives and to
prohibit antibiotics as growth promoters.

FDA. (2001, October 2008). "CVM and Judicious Use of Antimicrobials." Retrieved
November, 2008, from http://www.fda.gov/cvm/JudUse.htm.

Felsman, R., M. Wise, et al. (1973). "Effect of added dietary levels of copper sulfate and
an antibiotic on performance and certain blood constituents of calves." J Animal
S_ci 36(1): 157-160.

Gabrilidis, T. (1972). "Antibiotics in dairy calves' rations (abs)." J Dairy Sci 55(5): 703.

Heinrichs, A. J. and G. L. Hargrove (1987). "Standards of weight and height for Holstein
heifers." J Daifl Sci 70(3): 653-60.

Heinrichs, A. J ., C. M. Jones, et al. (2003). "Effects of mannan oligosaccharide or
antibiotics in neonatal diets on health and growth of dairy calves." J Dairy Sci
86(12): 4064-9.

Heinrichs, A. J. and W. C. Losinger (1998). "Growth of Holstein Dairy Heifers in the
United States." J Anim Sci 76: 1254-1260.

70

Hvidsten, H. (1959). "Studies on chlortetracycline and penicillin in the nurtrition of
young calves." Acta Agric Scand 9: 3-22.

James, R., M. McGilliard, et a]. (1984). "Calf mortality of Virginia dairy herd
impovement herds." J Dag Sci 67 : 908-911.

Jarmuz, W., I. Szelag, et al. (2001). "Relationships between concentration of serum
immunoglobulins and growth rate of dairy heifers, abstract 1792." J Dag Sci
84(Suppl. 1): 432.

Lassiter, C., R. Grimes, et al. (1958). "Influence of antibiotics on the growth and protein
metabolism of young dairy calves." J Dg'g Sci 41(10): 1417-1424.

Lundborg, G., E. Svensson, et al. (2005). "Herd-level risk factors for infectious diseases
in Swedish dairy calves aged 0-90 days." Preventive Veterim Medicine 68:
123-143.

Morrill, J ., A. Dayton, et al. (1977). "Cultured milk and antibiotics for young calves." 1
Day Sci 60(7): 1105-1109.

Place, N. T., A. J. Heinrichs, et al. (1998). "The effects of disease, management, and
nutrition on average daily gain of dairy heifers from birth to four months." J Dg'g
_S_cj 81(4): 1004-9.

Preston, T., N. McLeod, et al. (1959). "The effect of chlortetracycline on growth of early-
weaned calves." Animal Production 1: 13-19.

Quigley, J. D., 3rd, J. J. Drewry, et al. (1997). "Body weight gain, feed efficiency, and
fecal scores of dairy calves in response to galactosyl-lactose or antibiotics in milk
replacers." J Dairy Sci 80(8): 1751-4.

Radisson, J ., C. Smith, et al. (1956). "The mode of action of antibiotics in the nutrition of
the dairy calf. I. Effect of terramycin administered orally on the performance and
intestinal flora of young dairy calves." J Dag Sci 39: 1260-1267.

Sivula, N., T. R. Ames, et al. (1996). "Descriptive epidemiology of morbidity and
mortality in Minnesota dairy heifer calves." Prev Vet Med 27: 155-171.

Swanson, E. (1963). "Effects of chlortetracyline in calf starter and milk." J Dag Sci 46:
955-958.

Tomkins, T. and E. Jaster (1991). Preruminant calf nutrition. Vet Clin North Am Food
Anim Pract. 7: 557.

71

USDA (2008). Dairy, 2007, Part III: Reference of Dairy Cattle Health and Management
Practices in the United States, 2007. Fort Collins, CO, USDA-APHIS-VS,
CEAH: 160.

Van Donkersgoed, J ., C. Ribble, et al. (1993). "Epidmeiological study of enzootic
pneumonia in dairy calves in Saskatchewan." Can J Vet Res 57: 247-254.

Van Putten, G. (1982). "Welfare in veal calf units." Vet Rec 111: 437-440.

 

Vann, R. and J. Baker (2001). "Calf serum IgG concentrations affects weaning
performance, abstract 926." J Anim Sci 79(Suppl. 1): 223.

Virtala, A.-M., G. Mechor, et al. (1996). "The effect of calfliood diseases on growth of
female dairy calves during the f1rst3 months of life in New York State." J D '
Sci 79: 1040-1049.

Waltner-Toews, D., S. W. Martin, et al. (1986). "The effect of early calfhood health
status on survivorship and age at first calving." Can J Vet Res 50(3): 314-7.

WHO. (1998). "The Medical Impact of the Use of Antimicrobials in Food Animals.
Report of a WHO meeting. Berlin, Germany, 13-17 October, 1997."
WHO/EMC/ZOO/97.4 Retrieved December 19, 2006, from
http://whcfladocwhoint/hq/ 1997/WHO_EMC ZOO 97.4.pdf.

 

WHO (2003). Joint FAO/OIE/WHO Expert workshop on non-human antimicrobial usage
and antimicrobial resistance: Scientific assessment. Geneva.

72

CHAPTER 3

73

PHARMACOKINETICS OF OXYTETRACYCLINE IN MILK REPLACER FED
TO HOLSTEIN CALVES
Averill, 11‘, Wagner, SAZ, Imerman, PM3, and Erskine, RJ‘
1 Michigan State University, College of Veterinary Medicine. East Lansing MI 48823.
2 North Dakota State University, Department of Animal Sciences. Fargo ND 58108.
3 Iowa State University, Veterinary Diagnostic Laboratory. Ames IA 50011.
ABSTRACT
Six healthy Holstein heifer calves were fed approximately 45 mg of
oxytetracycline in milk replacer twice a day beginning at 3 days of age in order to
determine steady state pharmacokinetics for the drug. At 10 days of age, plasma samples
were collected over a 9-hour period (0, 60, 90, 120, 150, 180, 210, 300, 390, 480, 540

minutes) after the morning feeding. Liquid chromatography was used to determine
oxytetracycline concentration in plasma. Mean Cmax of oxytetracycline was 0.178 ug/ml
and mean Tmax was 290 minutes post feeding. Additionally we compared minimum

inhibitory concentrations of oxytetracycline for Pasteurella multocida and Escherichia
coli isolated from calves that were fed medicated or non-medicated milk replacer. E. coli
and P. multocida were isolated from fecal samples and nasal swabs of calves at 2 and 8
weeks of age, respectively. Samples were ollected from 15 calves fed medicated milk
replacer (220 mg/kg oxytetracycline and 440 mg/kg neomycin) and 15 calves fed non-

medicated milk replacer at each age group, with susceptibility to oxytetracycline

evaluated using microbroth dilution. The MIC50 and MIC90 were identical for E. coli in
both groups, >8 pig/ml, while for P. multocida the MIC50 of 2 ug/ml was identical in

both groups. The P. multocida MIC90 for the non-medicated group was 2 ug/ml, while

the medicated group had a one-fold higher dilution of 4 ug/ml (P < 0.05). The steady-

74

state pharmacokinetic values in this study are lower than previously reported for

oxytetracycline in milk replacer. Peak plasma concentrations of oxytetracycline were

below the MIC50 for E. coli and P. multocida. Higher concentration of the drug in milk

replacer may be necessary for therapeutic success.

INTRODUCTION

The National Animal Health Monitoring System (NAHMS) Dairy Study of 2002
and 2007 reported that oxytetracycline and neomycin are the most frequently used
antimicrobial agents in milk replacer for dairy calves (USDA, 2005, 2008b).
Tetracyclines are also the most commonly used drug to treat enteritis in unweaned dairy
calves, while florfenicol is for respiratory disease (USDA, 2008c). Oxytetracycline is
effective against susceptible Gram-positive and Gram-negative bacteria and is readily
absorbed after oral administration to fasting animals (Plumb, 2002). However, the
presence of food or dairy products can reduce the ability of oxytetracycline to be
absorbed by an animal in the gastrointestinal tract (Plumb, 2002). Because of their
lipophilic nature, tetracyclines have wide distribution throughout the body, and they are
eliminated unchanged primarily via glomerular filtration (Plumb, 2002).

Pharmacokinetic studies of oxytetracycline in dairy calves are limited, especially
those studying administration via milk replacer (Palmer et al., 1983; Schifferli et al.,
1982). Studies that have been conducted were administered at doses not in use by the
dairy industry today, as manufacturers of milk replacer commonly supplement their
product with 220 mg/kg of oxytetracycline and 440 mg/kg of neomycin. Since neonatal

calves are physiologically monogastric, young pigs are another animal model that can be

75

 

used to assess pharmacokinetic values of oxytetracycline after oral administration (Hall et
al., 1989; Mevius et al., 1986; Nielsen and Gyrd-Hansen, 1996; Pijpers et al., 1991a).

Animal agriculture has received scrutiny from the feeding of antimicrobial agents
at subtherapeutic levels because of the potential for development of antimicrobial
resistance (WHO, 2003b). Studies in calves and pigs have hypothesized that serum or
plasma concentrations of oxytetracycline administered per as in feed may be high enough
to treat some common bacterial pathogens, based on typical minimum inhibitory
concentrations (MIC) (Hall et al., 1989; Mevius et al., 1986; Schifferli et al., 1982). With
antimicrobial resistance emerging, this may no longer be true.

The objectives for the current investigation are to evaluate the steady state
pharmacokinetics of oxytetracycline administered to dairy calves in milk replacer and
determine if the plasma concentration of the drug is greater than the in vitro minimum
inhibitory concentration (MIC). The second objective will be based on oxytetracycline
MIC for Escherichia coli from fecal samples and Pasteurella multocida from nasal swabs

as both pathogens are commensal organisms within their respective body organs.

MATERIALS AND METHODS
The Institutional Animal Care and Use Committee (IACUC) at Michigan State
University approved all procedures involving live animals, as well as the trial protocol.
Pharmacokinetics:

M; Six healthy Holstein heifer calves aged 9 to 14 days and of similar

weight (42.5 to 47.3 kg) were used in a steady state pharmacokinetic study.

76

Milk Replacer: Calves received quality colostrum (immunogloban level >50
mg/ml) at birth and until 3 days of age. At three days of age, calves were switched to a
commercial medicated milk replacer with 220 mg/kg of oxytetracycline and 440mg/kg of
neomycin (Nutrenam Snowflakes 20-20 All-Milk Medicated Milk Replacer). The milk
replacer was labeled to contain 20 percent crude protein (milk based) and crude fat, and
0.15 percent crude fiber. Milk replacer was fed as approximately 220 g dissolved in 2
liters of water twice a day. Thus, at each feeding medicated calves received
approximately 45 mg of oxytetracycline (1 mg/kg twice daily) and 90 mg of neomycin (2
mg/kg twice daily).

Specimen Collection and Handling: Whole blood samples were collected via
jugular venipuncture from each calf prior to the morning feeding and at 60, 90, 120, 150,
180, 210, 300, 390, 480, and 540 minutes post feeding. Specimens were transported to
the laboratory at Michigan State University on ice and were spun at 1000 x g for 10
minutes. Plasma was separated and frozen in SmL polypropylenes tubes at -80°C.

Liquid Chromatography: Plasma oxytetracycline levels were determined by

liquid chromatography (LC). A modification of the Association of Analytical

Committees (AOAC® Official Methods“) method 995.09 for chlortetracycline,
oxytetracycline and tetracycline in edible animal tissues was used (AOAC International,
2000). One ml of plasma was mixed with 10ml of McIlvaine buffer and passed through a
C18 solid-phase extraction column for cleanup. The oxytetracycline was eluted fiom the
solid phase extraction column with ethyl acetate and 5:95, methanolzethyl acetate. The
elution fi'actions were concentrated to dryness under nitrogen and resolvated in 100 ul of

methanol and sent to LC. A set of standards (50, 100, 200, 500, 1000, and 2000 ppb

77

oxytetracycline) in a blank plasma matrix were also solid phased extracted and used as a
standard curve. Standards and samples sent to LC were chromatographed using 3x3
Perkin Elmer C18 guard column and an Axiom ODS 5 micron 150mm x 3mm
analytical column. Detection of oxytetracycline was at 350 nm by UV light. Results
were quantified against the plasma matrix standard curve. The retention time of the
oxytetracycline was 6.5 minutes. For this method the oxytetracycline limit of detection
(LOD) was 10 ppb and the limit of quantification (LOQ) was 25 ppb.
Antimicrobial Susceptibility Testing

gigs; Thirty healthy calves with no previous illnesses were selected from two
different age groups, suckling (about 2 weeks of age) and just-weaned heifer calves
(about 8 weeks of age). These calves were from the same farm mentioned above. Within
each age group 15 calves fed medicated milk replacer and 15 fed an identical milk
replacer, but without antimicrobial agents were randomly selected for specimen
collection. Calves in the suckling group were used to collect fecal specimens for culture
and isolation of Escherichia coli while those calves in just-weaned group were used for
nasal swab specimens to culture and isolate Pasturella multocida.

chimen Collection: Approximately 10 grams of fecal matter were collected
from each calf via digital rectal palpation; using a separate glove for each animal.
Approximately 1 gram of fecal matter was placed in Cary-Blair transport media for
transport to the laboratory on ice. Nasal specimens were collected using a sterile guarded
swab. Swab and guard was inserted into the right nostril approximately 10 cm, then the
swab was inserted another 5 cm, rotated 360°, retracted back into the guard and removed.

After sampling, each swab was inserted into transport media (BBL Culture Swab media,

78

Becton, Dickinson and Company, Sparks, MD) and placed on ice for transport to the
laboratory at Michigan State University.

Bacterial Culturing: We cultured all specimens within 6 hours of collection.
Fecal specimens were inoculated on a MacConkey agar gel plate, triple sugar iron agar
slant, and urea slant and incubated at 37°C with 5% C02 overnight. Suspect E. coli
colonies underwent further biochemical tests for confirmation; indole, methyl red,
vogues-proskauer, citrate, sorbitol, and motility. Quality control of biochemical tests
were performed using Salmonella Java (CDC control), Enterococcus aerogenes ATCC®
13048, and Escherichia coli ATCC® 25922. Nasal swab specimens were inoculated on a
blood agar plate and incubated at 37°C with 5% C02 overnight. Suspect colonies of
Pasteurella multocida were then gram stained to confirm a small Gram-negative rod.
Confirmatory biochemical testing was performed using APIZONE strips following the
manufacture’s instructions (bioMerieux, Inc, Durham, NC). Quality control of APIZONE
strips was performed using Sphingobacterium multivorum ATCC® 35656, Aeromonas
hydrophilia ATTCC® 35654, Pseudomonas aeruginosa ATCC® 27853 and Alcaligenes
faecalis ATCC® 35655. Isolates were frozen at -80°C in skim milk until susceptibility
testing was performed.

S_usc_eptibility Testirg; MIC values were determined using a microbroth dilution
system. A sterile inoculating loop was used to obtain a sample of E. coli or P. multocida
from hand thawed skim milk and streaked onto MacConkey agar for E. coli or blood agar
for P. multocida. Plates were incubated at 37°C for 24 hours. A single colony from each
plate was isolated then streaked onto a Mueller Hinton agar and incubated at 37°C for 24

hours. Minimum inhibitory concentration was determined for the bacteria using the

79

Sensititre semi-automated antimicrobial susceptibility testing system following the
manufacturer’s instructions (Trek Diagnostic Systems, Westlake OH). Sensititre
antimicrobial susceptibility plates used for E. coli and P. multocida were BOPOlF and
CMV lABPF respectively. The minimum dilution of antimicrobial agent that inhibited
growth was recorded as the MIC. Quality control was conducted using Escherichia coli
ATCC® 25922. Results were entered into Microsoft Access database (2000, Microsoft
Corporation, Redmond, WA) and data entry reviewed for errors by separate individuals.
Statistical Analysis

Microsoft Excel (2000, Microsoft Corporation, Redmond, WA) was used to

calculate pharmacokinetic values, and the. mean MIC values to inhibit growth for 50

percent of the isolates (MIC 50) or for 90 percent of the isolates (MIC90) of P. multocida

and E. coli to oxytetracycline. When exact MICSO’s and 90’s could not be calculated, a
conservative approach was followed, by rounding up to the next isolate in the order.
Mantel-Haenszel Chi-square analysis for trend (SAS 9.1, Cary, NC) was used to compare
MIC values between the medicated and non-medicated milk replacer groups. Statistical

significance was designated a priori as a P-value of 0.05 or less.

RESULTS

Mean weight was 44 kg (range 42 to 47) and mean age 11 days (range 9 to 14),
for the six calves used for the pharmacokinetic portion of this study. All calves were
healthy and had not received any medications prior to enrollment. Individual and mean
QSEM) plasma concentrations at each time point following feeding of milk replacer with

oxytetracycline is presented in Table 3.1. Plasma concentrations generally increased

80

until 300 minutes post feeding and then declined to 0.135 ug/ml at 540 minutes (Fig.
3.1). Mean peak serum concentration occurred at 290 minutes (range 90 to 540). The
mean plasma concentration for oxytetracycline over the nine-hour period was 0.178
rig/ml and ranged from 0.098 to 0.275 rig/ml (Table 3.2).

Antimicrobial susceptibility results were obtained on bacterial isolates fi'om 60
calves; 15 fed medicated milk replacer and 15 fed non-medicated milk replacer within the
suckling and just-weaned groups. For the 13 P. multocida isolates from the medicated
group their MIC ranged from 1.0 —— 4.0 ug/ml while 11 isolates from the non-medicated
calves ranged from 0.5 — 2.0 rig/ml (Fig. 3.2). The difference in MIC of isolates

collected from the medicated group compare to non-medicated was significant (Mantel-

Haenszel Chi-Square of 4.83; p < 0.028). The MIC50 was 2 rig/ml and the MIC90 was 4

rig/ml for isolates from the medicated calves, while isolates from the non-medicated
calves were 2 ug/ml and 2 rig/ml, respectively (Table 3.3). All E. coli isolates had an
MIC >8 rig/ml except for one isolate in the non-medicated group (Fig. 3.3).

MIC values for oxytetracycline against P. multocida and E. coli exceeded the

plasma concentration that this drug attained in calves receiving the medicated milk

replacer at all collection times for all calves. MIC50 was 2.0 rig/ml for P. multocida

while the highest individual calf plasma concentration only reached 0.275 ug/ml.

DISCUSSION

Oxytetracycline is approved by the US. Food and Drug Administration as a

supplement to feed for growth promotion, to improve feed efficiency, and to aid/control

81

in treatment of bacterial pathogens when administered at 0.11-0.22 mg/kg/day. For the
treatment of bacterial enteritis and pneumonia due to an infection with pathogens
susceptible to oxytetracycline, the dose to be administered is 22 mg/kg/day (Bayley,
2006). For this study, the calves received approximately 45 mg at each feeding in the
milk replacer, thus receiving approximately 1 mg/kg per feeding. On the manufacturer’s
label, it states that this product is to “aid in the treatment of bacterial enteritis (scours) ”
(Nutrenam Snowflakes 20—20 All-Milk Medicated Milk Replacer).

Previous pharmacokinetic studies of oxytetracycline in calves were conducted
with doses ranging from 5 to 50 mg/kg in milk replacer (Luthman and Jacobsson, 1983;
Palmer et al., 1983; Schifferli et al., 1982). The steady state plasma concentrations of
oxytetracycline obtained in our study (Table 3.1) were low compared to previous work.
Schifferli et al reported serum levels of oxytetracycline ranged from 0.75 to 1.2 rig/ml in

calves administered 5 mg/kg in milk replacer twice a day for five days (Schifferli et al.,
1982). Curve simulation in this study approximated a Tmax post-feeding of

approximately 6 hours, which is comparable to our results (Schifferli et al., 1982). In our
study, the average concentration for oxytetracycline was 0.125 ug/ml. A steady-state

kinetic study in feeder pigs fed oxytetracycline in feed at 13 mg/kg of body weight

(Pijpers et al., 1991a), resulted in a Cmax of 0.13 to 0.22 rig/ml, which is similar to our

findings. Pijpers et al did not calculate Tmam as plasma samples were only collected prior

to feeding and 3 hours post feeding (Pijpers et al., 1991a).
A study in calves fasted overnight and administered a single dose of

oxytetracycline at 9 mg/kg in milk replacer reported results similar to ours (Palmer et al.,

1983), TmX of 6 hours and a Cmax of 0.5 rig/ml. As one would expect, studies

82

 

administering 50 mg/kg of oxytetracycline for one dose in milk replacer fed to calves

achieved higher Cmax and longer Tm. Schifferli et a1 and Luthman et a1 published

results with a Cmax of 4.99 and 1.2 rig/ml and a Tmax of 9.16 and 4 hours in the serum,

respectively (Luthman and Jacobsson, 1983; Schifferli et al., 1982).

The high level of resistance in our E. coli specimens to oxytetracycline correlates
with previous studies (Catry et al., 2007; Khachatryan et al., 2004; NARMS, 2004; Sato
et al., 2005). However, previous reports used tetracycline for their susceptibility testing,
while we used oxytetracycline. We reported, at a breakpoint of 16 ug/ml, 100 percent
resistance as did Catry et a1 (Catry et al., 2007). Data from the Michigan State University
Diagnostic Center for Population and Animal Health (DCPAH) from 1998 to 2002
reported a high level of resistance in E. coli to tetracycline, as 23 percent of isolates were
susceptible (Averill, 2005). These results are higher than those reported by Sato et a1 and
Khachatryan et al, 55 and 79 percent, respectively (Khachatryan et al., 2004; Sato et al.,
2005). The age in which calves were sampled may have been a factor between these
studies. Khachatryan et al demonstrated that as calves aged, the resistance to tetracycline
of E. coli isolates collected from them decreased, so that by 6 months only 17% of
isolates were resistant to tetracycline (Khachatryan et al., 2004).

The mean MIC for oxytetracycline to P. multocida in calves previously reported

by Catry et al in two separate studies were 0.25 and 0.5 rig/ml respectively, values lower

than our MIC5OS (Catry et al., 2006; Catry et al., 2005). Though our MIC90 values were

lower than what Catry reported in 2005 and 2006, which were 64 and 32 ug/ml
respectively (Catry et al., 2006; Catry et al., 2005). DCPAH data from 1998 to 2002 had

a high level of susceptibility, 80 percent, to tetracycline for P. multocida, which is similar

83

to our results as all isolates were _<_ 4 ug/ml. A study reporting the MIC values of P.

multocida to oxytetracycline in ill calves also had higher values than observed in our

study, as the MIC50 and MIC90 were both _>_l6 rig/ml (Mevius et al., 1990). The higher

MIC values reported in the above studies may be due to multiple reasons; including
unknown drug use on farm and treatment of calves before bacterial sampling.

In our study, oxytetracycline MIC values for P. multocida were higher for isolates
collected from medicated calves. This difference may be due to the fact that the
medicated calves were receiving milk replacer containing oxytetracycline since three
days of age. However, the sample size was limited, as isolates were only obtained from
11 and 13 calves from the control and medicated groups, respectively. At the time of
study design, the NCCLS recommended that 10 isolates for an antibiogram were
sufficient to make valid comparisons between two groups. Since then, the NCCLS, now
the CLSI (Clinical and Laboratory Standards Institute), has changed their
recommendation to state that 30 isolates are necessary to report susceptibility results of
an organism to an antimicrobial agent in an antibiogram (CLSI, 2005). There is a
possibility of type 1 error where the MICs are truly similar between the two groups.

Previous studies have looked at serum or plasma concentrations of
oxytetracycline administered in the feed to determine if therapeutic levels are achieved to
treat various bacterial pathogens (Hall et al., 1989; Mevius et al., 1986; Schifferli et al.,
1982). Schifferli et al reported that feeding 50 mg/kg of oxytetracycline in milk replacer
to calves could achieve and maintain a serum concentration above 1.0 ug/ml for 24
hours, and exceeded the MIC of 0.5-1 .0 rig/ml for respiratory pathogens (Schifferli et al.,

1982). Within the same study, steady-state kinetic values from feeding calves

84

oxytetracycline at 5 mg/kg achieved serum peak concentrations around 1 rig/ml, which is
above the reported MIC of 0.5 ug /ml. However, it was demonstrated in our study, that
feeding oxytetracycline at the levels typically added to milk replacer are inadequate to
maintain effective concentrations in plasma for therapeutic efficacy of pneumonia.

In treating animals with antimicrobial agents per as there is no literature, to the
investigators knowledge, that states the concentration that is obtained and maintained in
the lumen of the gastrointestinal tract. Nor is there information stating how plasma drug
concentration relates to gastrointestinal drug concentration. With the lack of literature to
demonstrate what drug concentration is needed to treat bacterial enteritis, it is speculative
to predict whether or not medicated milk replacers offer an effective treatment regimen

for enteritis.

CONCLUSION:

Plasma concentration of oxytetracycline in milk replacer fed to calves does not
reach MIC values for Escherichia coli from feces and Pasteurella multocida from nasal
swabs. Therefore, supplementing milk replacer with oxytetracycline at low levels will
likely not be effective in treating bacterial enteritis or pneumonia caused by the
pathogens tested in this study. Higher concentrations of the drug in milk replacer, or by

separate administration or feeding, may be necessary for therapeutic success.

85

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Cited Literature

Barber, H. E., T. N. Calvey, et al. (1974). "Bilogoical bioavailability and in vitro
dissolution of oxytetracycline dihydrate tablets." Brit J Clin Phgm 1: 405-408.

Barnett, D. B., R. N. Smith, et al. (1974). "Bioavailability of commercial tetracycline
products." Brit J Clin Phgm 1: 319-323.

Bayley, A. (2006). Compendium of Vetering Products, Ninth Edition. Port Huron, MI,
NOrth American Compendiums, Inc.

Blair, D. C., R. W. Barnes, et al. (1971). "Biological availablity of oxytetracycline HCl
capsules." J Amer Med Ass 215: 251-254.

Brice, D. W. and H. F. Hammar (1969). "Therapeutic nonequivalence of oxytetracycline
capsules." J Amer Med Ass 208: 1189-1190.

Catry, B., A. Decostere, et al. (2006). "Detection of tetracycline-resistant and susceptible
pasteurellaceae in the nasopharynx of loose group-housed calves." Vet Res
Commun 30(7): 707-15.

Catry, B., J. Dewulf, et al. (2007). "Antimicrobial resistance patterns of Escherichia coli
through the digestive tract of veal calves." Microb DrugRe_s_i_s_t_ 13(2): 147-50.

Catry, B., F. Haesebrouck, et al. (2005). "Variability in acquired resistance of Pasteurella
and Mannheirnia isolates from the nasopharynx of calves, with particular
reference to different herd types." Microb Drug Resist 11(4): 387-94.

CLSI (2002). Performance Standards for Antimicrobial Disk and Dilution Susceptibility
Tests for BaLcteri_a Isolated from Animals: Approved Sta_ng_ard--Second Edition,
M3 1 -A2. Wayne, PA, NCCLS.

CLSI (2005). Analysisj and Presentation of Cumulative Antimcrobial Susceptibility Test
Data; Approved Guideline--Second Edition. M39-A2. Wayne, Pennsylvania,
Clinical and Laboratory Standards Institute.

Hall, W. F., T. S. Knifi‘en, et a1. (1989). "Plasma concentrations of oxytetracycline in
swine afier administration of the drug intramuscularly and orally in feed." J Am
Vet Med Assoc 194(9): 1265-8.

Khachatryan, A. R., D. D. Hancock, et al. (2004). "Role of calf-adapted Escherichia coli
in maintenance of antimicrobial drug resistance in dairy calves." Appl Environ
Microbiol 70(2): 752-7.

Luthman, J. and S. O. Jacobsson (1983). "The availability of tetracyclines in calves."
Nord Vet Med 35(7-9): 292-9.

91

Luthman, J ., S. O. Jacobsson, et al. (1989). "Studies on the bioavailability of tetracycline
chloride after oral administration to calves and pigs." Journal of Veterinafi
Medicine. Series A 36(4): 261-268.

Mevius, D. J ., H. J. Breukink, et al. (1990). "In vitro activity of flumequine in
comparison with several other antimicrobial agents against five pathogens
isolated in calves in The Netherlands." Vet Q 12(4): 212-20.

Mevius, D. J ., L. Vellenga, et a1. (1986). "Pharmacokinetics and renal clearance of
oxytetracycline in piglets following intravenous and oral administration." Vet Q
8(4): 274-84.

NARMS (2004). NARMS Annual Veterinary Isolate Data. Athens, GA, United States
Department of Agriculture, Agriculture Research Services.

Nielsen, P. and N. Gyrd-Hansen (1996). "Bioavailability of oxytetracycline, tetracycline
and chlortetracycline after oral administration to fed and fasted pigs." J Vet
Pharmacol Ther 19(4): 305-11.

Palmer, G. H., R. J. Bywater, et al. (1983). "Absorption in calves of amoxicillin,
ampicillin, and oxytetracycline given in milk replacer, water, or an oral
rehydration formulation." Am J Vet Res 44(1): 68-71.

Pijpers, A., E. J. Schoevers, et al. (1991). "Plasma levels of oxytetracycline, doxycycline,
and minocycline in pigs after oral administration in feed." J Anim Sci 69(11):
4512-22.

Plumb, D. C. (2002). Veterinary Drug Handbook. Ames IA, Iowa State Press.

Sato, K., P. C. Bartlett, et al. (2005). "Antimicrobial susceptibility of Escherichia coli
isolates from dairy farms using organic versus conventional production methods."

J Am Vet Med Assoc 226(4): 589-94.

Schifferli, D., R. L. Galeazzi, et al. (1982). "Pharmacokinetics of oxytetracycline and
therapeutic implications in veal calves." J Vet Pharmacol Ther 5(4): 247-57.

92

Chapter 4

93

TEACHING ANTIMICROBIAL RESISTANCE VIA COMPUTER AIDED
LEARNING
James J Averill, Paul C Bartlett, Theresa M Bernardo, Robert P Malinowski and Ronald J
Erskine
ABSTRACT
Veterinary medicine plays a key role in helping mitigate antimicrobial resistance.

A computer aided learning tool was developed at the following website:
http://old.cvm.msu.edu/cdc, as an aid for teaching veterinary students about antimicrobial
resistance. This CAL allows for active learning that is student driven and interactive.
Usability testing was conducted to ensure learners could easily navigate the CAL and
follow the logical flow of the teaching objectives and information being presented. The
CAL contains a module regarding the basic ‘Principles’ of antimicrobial resistance,
which summarizes the microbiology, pharmacology and epidemiology concepts in
regards to antimicrobial resistance, and the role of antimicrobial use in animal and human
health. There are also multiple ‘Case Study’ modules that apply these principles to
specific clinical situations involving dairy and beef cattle, small animals, and swine.
These modules relate to a specific scenario regarding antimicrobial usage in veterinary
medicine/animal agriculture. Within each module, there are videos, animations, and

questions to engage the learner. The materials within this CAL can be used as an adjunct

to traditional styles of learning in pre-clinical or clinical settings.

INTRODUCTION
Veterinary school curricula are over-extended as the breadth and depth of

veterinary knowledge continues to expand. Little opportunity exists to add new courses

94

regarding emerging issues, such as antimicrobial resistance (AMR) (Hird et al., 2002).
With the reemergence and interest in the ‘One Health, One Medicine’ concept, AMR is
also an excellent example of how such a concept can be used to address a medical issue
that impacts animals, humans and the environment. If AMR is to be taught in veterinary
curricula, information must be integrated into existing veterinary courses and course
moderators may not know who is teaching specific issue(s). Computer-Aided Learning
(CAL) is one potential tool that can help educate students on AMR.

The American Veterinary Medical Association (AVMA) and American Medical
Association (AMA) are working together towards a common goal ‘One Health, One
Medicine’ (AVMA, 2008). The ‘One Health, One Medicine’ concept brings animal
health, human health and environmental health together as one. AMR is an excellent
example of how this concept can be used to mitigate drug resistance, as physicians,
veterinarians, microbiologist, ecologist and others are working to understand the
development, spread and control of AMR. Antimicrobial agents, since their inception,
have been used in multiple ways (therapeutically or sub-therapeutically via injections, per
os, lotions, and other formats) to enhance the health and well-being of animals and
humans. However, bacterial resistance to antimicrobial agents is an emerging problem
that challenges the biomedical professions and public health (WHO, 1999). Additionally,
the use of antimicrobial agents in animal agriculture has been implicated as contributing
to bacterial resistance in human medicine (J ETACAR, 1999; WHO, 2003a). The time is
now for the veterinary profession to educate all veterinarians about the prudent use of

antimicrobial agents and antimicrobial resistance (Morley et al., 2005). Similar steps are

95

also being in the United Kingdom by the human medical community (Davey and Garner,
2007)

Advancement of information and technology is likely to continue to impact
veterinary education into the future not only in how information is delivered but also the
location of the learner (Short, 2002). These developments are opportunities to enhance
learning and will effect how we teach and learn in the future. Computer-Aided Learning
(CAL) also known as computer-assisted learning, computer-assisted instruction, or
computer-based learning, is a form of self-instruction with no direct interaction with the
instructor. It relies on computer-based lessons that include text, images, video, three-
dirnensional photos and simulated (virtual) reality. A simple lesson in CAL may consist
of text or visual information, such as radiographs, while more complex CAL lessons have
a greater level of interactive learning. For example, students are given a clinical case
where they choose their own pathway to solve the scenario and later explain why they
proceeded in such a manner.

The effectiveness of CAL in teaching is still being debated, but there is general
agreement that it can serve as an adjunct to traditional didactic learning (N erlich, 1995;
Rosenberg et al., 2005; Valcke and De Wever, 2006). The downside of CAL is the up-
front cost of development, programming and training of faculty on how to use such tools
effectively for teaching (Childs et al., 2005; Short, 2002). Advantages of CAL include
the ability of the student to learn at their own pace, the learning can take place anywhere
around the world, many students can be taught simultaneously, and collaboration among
experts can easily be facilitated to design lessons (Childs et al., 2005; Short, 2002). At

least five principles must be met in‘order for CAL or the use of information technology to

96

be effective in teaching, 1) just-in time, personalized learning, 2) student centered
learning, 3) self-paced learning, 4) learning anytime, anywhere and; 5) experimental,
discovery learning (Smith, 2003).

The purpose of this project was to enhance veterinary education on the
appropriate use of antimicrobial agents in veterinary medicine and thereby mitigate the
further development and spread of antimicrobial resistance via a CAL tool based on the
five principles mentioned above. The following objectives were developed for the
learner: 1) Understand the need to appropriately use antimicrobial agents, 2) Learn to
formulate strategic choices regarding the use of antimicrobial agents, 3) Explore the
relationship between animal and human health with respect to use of antimicrobial

agents.

MATERIALS AND METHODS

The CAL titled “Appropriate Use of Antimicrobial Agents” was a collaborative
effort between Michigan State University College of Veterinary Medicine (MSU CVM),
the Centers for Disease Control and Prevention (CDC), and Michigan Department of
Community Health through an Epidemiology and Laboratory Capacity grant from CDC.
This CAL also included participants from several other colleges of veterinary medicine in
United States as authors or editors of modules.

This CAL is intended to be used as an adjunct to didactic learning, such as in
courses of public health, pharmacology and microbiology, and also in clinical settings
such as production medicine clerkships. Individual modules or the entire CAL can be

used by instructors as an adjunct to learning specific topics for students through videos,

97

animations and questions about microbiology, pharmacology, public health and
management of livestock.

In the process of developing this CAL initial discussions centered on what
delivery format should be used, ie. a website or a course management/learning
management system (CMS/LMS) such as Blackboard, Web CT, or Angel. We chose to
use a website, instead of a CMS/LMS, since the tool could be used by anyone and
accessed around the world. The downside to using a website is that an individual whom
knew HTML pro grarnming had to enter the content; this was overcome by using trained
personnel at MSU CVM Information Technology Center. A standard template was
developed for the CAL with three panes within the monitor screen; navigation and a
‘Tools’ tab which contain a glossary, links, references, and credits are in the left pane, the
right pane was for additional information/links, and the middle pane contains the text,
video, animation and questions about the lesson. The CAL was constructed using
Adobe® Dreamweaver®.

In constructing this tool, a team (veterinarians, instructional designer and web
designers) approach was used to develop key themes to be covered within the ‘Principle’
and ‘Case Study’ modules to help with flow and reduce replication. Intended flow of this
tool is for the learner to proceed through the ‘Principles’ module first to obtain the key
concepts regarding antimicrobial resistance then move into species specific topics in the
‘Case Study’ modules (Fig 4.1). Each module or lesson was scripted and storyboarded
by the main author and designers before recording video, creating animations and
finalizing the lesson. This allowed for the authors and designers to work together on the

lesson and decide on the appropriate technology for incorporation into the module. Also

98

included within the storyline were questions for students to answer and evaluate their
understanding of the issue in various formats ranging from multiple-choice to drag-and-
drop using Adobe® Flash? Once a working draft was agreed upon a shot list/storyboard
was developed. After footage was captured, the author(s) reviewed it, and all shots were
cataloged to expedite editing. Video was edited using Adobe® Premiere®, and
compressed for the CAL using either Windows Media® Player format or Adobe® Flash®
Player format. Adobe® Flash® was also used to develop animations to explain or
reinforce a specific topic. Still photos were also used in the CAL and were edited with
Adobe® Photoshop®. Once a module was completed the estimated time to complete the
module was placed at the beginning.

Six veterinary students participated in Usability Testing of the CAL using a
method similar to HincMifie et al (Hinchliffe and Mummerv, 2008). The Institutional
Review Board approved the use and participation of human subjects to test the CAL.
Student selection was based on their willingness to participate in the process and their
comfort with using the computer/intemet. There were two students for each of the
following categories; novice (can turn computer on and run basic programs), intermediate
(novice skills plus ability to browse intemet and download programs), experienced
(intermediate skills plus ability to design websites/programs). Combinations of
quantitative (time taken) and qualitative (administrator’s observations and subjective user
preferences) techniques were used to collect data fiom students while completing eleven
pre-defined tasks (Table 4.1). These tasks focused on the ease of navigating through the
CAL and flow of the material. The administrator explained the process to each student at

the beginning; once the student began the tasks the administrator could not answer any

99

questions to eliminate any potential bias. To record their responses, students were
videotaped with a digital camcorder and by a screen capture program Carntasia StudioTM.
Students were instructed to express verbally their thoughts and explain what they were
doing as they completed the eleven tasks. Each student’s video was evaluated by the
development team to identify any difficulties they had in navigating the CAL. Areas of
difficulty were recorded, along with comments fiom the participants, and used to ilnprove

navigation of the CAL.

DESCRIPTION OF COMPUTER AIDED LEARNING TOOL

This CAL is accessible at http://old.cvm.msu.edu/cdc. A broadband intemet
connection is recommended, due to the number of embedded video and audio clips. Most
modules are based on a storyline with a veterinary student or veterinarian addressing an
antimicrobial resistance issue. Within the lesson are videos, cartoons, and animations to
further explain a topic. As learners proceed through the CAL they have opportunities to
answer questions to determine their comprehension of the topic with immediate feedback
as to what is the correct answer. The main module is called ‘Principles’, which includes
an overview of key concepts in microbiology, pharmacology and epidemiology/public
health regarding AMR. Other lessons specific to one species are contained within ‘Case
Study’ section. The case studies offer more detailed information about antimicrobial
resistance topics pertinent to dairy cattle, beef cattle, swine and small animal. Each
module also underwent peer review by veterinarians who are experts in the given topic to

ensure accuracy of the material.

100

Principles: This lesson follows Jeff, a veterinary student and his personal
experience with a Salmonella infection and how he collaborated with the State Public
Health Veterinarian to determine the source of his infection. Jeff’s journey leads him to
reflect on and investigate his past food consumption along with small and large animal
exposures. Along the way, he learns about the microbiology, pharmacology and
epidemiology/public health of antimicrobial resistance. To complete the investigation,
Jeff visits the State Diagnostic Laboratory to review culture and susceptibility testing and
also visits with his pharmacology professor to discuss antimicrobial agents, and how to
use MICs (minimum inhibitory concentrations). Finally, Jeff explores the use of
antimicrobial agents within animal agriculture and their role in developing resistance in
human pathogens.

Case Studies: This section contains case studies in dairy cattle, beef cattle, swine,
pocket pets and companion animals. Each module covers a specific topic within that
specie in greater detail as compared to the ‘Principles’. Below is a brief description of
each of the dairy cattle modules.

Medicated Milk Replacer: This module regards a veterinary student, Gretchen,
who is riding with a veterinarian, Dr. J essup, and visits a farm with an enteritis problem
in suckling calves. Objectives of this module are to understand what medicated milk
replacer is, the different antimicrobial agents added to milk replacer, the advantages and
disadvantages of medicated milk replacer and the importance of a colostrum management
program. During Gretchen’s history taking, she discovers a bag of (milk replacer with a
label. She notices that oxytetracycline and neomycin are included in the milk replacer.

Gretchen is surprised by this and asks the farmer how long and why they have been using

101

the product. Gretchen then questions Dr. J essup about the practice and purpose of
supplementing antimicrobial agents in milk replacer. Dr. J essup attempts to answer all of
Gretchen’s questions regarding the use of antimicrobial agents in feed.

Neonatal Scour-s: Pertains to Dr. Karl’s response to a farm call where the farmer,
Chuck Erby, has tried numerous drugs to treat enteritis in his dairy calves while
experiencing increased mortality. This module focuses on when antimicrobial agents
should or should not be used to treat enteritis and on management practices to reduce
drug use on a farm. Learners are able to view the farm operation via video, interpret
physical exam findings of healthy and ill calves, determine which specimens to collect
for submission to the laboratory, review herd health records and discuss treatment options
with the farmer. Discussions with Mr. Erby are centered on his desire to give the ill
calves medication while Dr. Karl explains the need to know the pathogen(s) before
determining if antimicrobial agents are necessary, especially in calves not demonstrating
signs of systemic disease. Dr. Karl explains to Mr. Erby that antimicrobial agents are not
always appropriate for treating enteritis in calves. In fact, management practices can play
a major role in preventing neonatal calf diseases. A suggestion that Dr. Karl makes to
Mr. Erby is to improve husbandry practices to minimize antimicrobial use.

Contagious Mastitis: This module profiles a veterinarian, Dr. Susan Keller,
working with a client, Mr. Oliver McCormick, who is concerned about a high somatic
cell count (SCC). Main topics of discussion are preventive measures to mitigate mastitis,
importance of culturing mastitis cases, the proper use of antimicrobials in treating
mastitis and how treatment duration is important for effective therapy. During the herd

visit, the last milk test and SCC reports are reviewed and analyzed. Dr. Keller and Mr.

102

McCormick discuss options and agree to a return visit to monitor the milking procedure
and collect milk samples for culture from cows with high SCC. Once culture results are
reported, they discuss treatment options with antimicrobial agents ranging from
intramammary infusions to systemic therapy. Dr. Keller also emphasizes the importance
of management practices to mitigate disease transmission of contagious mammary
pathogens from cow to cow.

Farm Based Mastitis Program: This is a 22-minute video written and filmed in
the style of a local news story regarding how one large dairy farm uses an evidence-based
approach to preventing and controlling mastitis. Learner objectives for this module
include the role of a veterinarian in developing standard operating procedures, how to use
a farm antibiogram to determine treatment regimens, the five key topics to consider when
selecting a drug and the importance of preventive measures to mitigate mastitis. The
news reporter, Ms. Becky Dewitt, interviews the farm veterinarian, about their mastitis
program. An evidence-based approach is taken for each case: identify causal pathogen,
determine appropriate drug to use based from the farm antibiogram, treat the cow and
monitor. This is the standard operating procedure for treating mastitis at this operation.
Antimicrobial agents are only a part of the mastitis program on this farm. This evidence-
based plan that is presented, decreased antimicrobial agent use by 74 percent and the

number of cows treated and duration in the mastitis pen.

USABILITY TEST RESULTS
Six veterinary students completed the 11 tasks in a reasonable amount of time.

Overall feedback was that the CAL could be navigated and material flowed smoothly.

103

However there were a few suggestions or areas of difficulty that were identified by the
administrators. Three key issues were identified: students looked for content only within
the middle pane, a need for page advancement buttons and change the format of
interactive questions. Participants frequently used the left pane to navigate throughout
the CAL and looked for content within the middle pane. Only two participants, in their
first attempt, completed a question that required the participant to go to the right-side
pane and click on a link for more information, while two never answered the question. To
correct this, the right pane of the CAL is reserved only for remedial, advanced or
additional information that is not critical for the learner. The usability testing also
demonstrated a need for easier navigation with ‘page forward’ and ‘page backward’
buttons on each screen, and to limit the amount of material within each screen to reduce
scrolling. The final change was in regard to question formatting, as there was a need to
give more clear directions to ‘drag and drop’ or just ‘click and point’. Additionally,
questions with two parts were removed as all six of the participants were confused with
them. Conducting usability testing early in the development process allowed for a
standard template to by created which saved time in creating further modules, as

author(s) knew the appropriate layout and placement of content.

DISCUSSION

This CAL was designed using the five principles of information technology in
teaching; 1) just-in time (detailed information about a current topic that is not
incorporated in the classroom or clinical teaching), personalized learning, 2) student

centered learning, 3) self-paced learning, 4) learning anytime, anywhere and; 5)

104

experimental, discovery learning (Smith, 2003). Antimicrobial resistance is a current
issue within the medical fields, and is an excellent example of how a ‘One Medicine’
concept can be used to address and resolve this issue. Veterinary medicine plays an
important role in this topic, as there is a potential for the use of antimicrobial agents in
animals to impact the ecosystem and human health (WHO, 1998, 2003b).

This CAL allows learners to expand upon what they have learned in the
classroom. Learners are likely to have some prior knowledge regarding antimicrobial
agents and/or AMR, and will have an idea of what information he/she needs to learn.
Also, the learners are able to work at their own pace, choose which modules they wish to
view, and the location where they prefer to learn. These parameters are what make the
CAL student-centered and allow it to function as an active learning environment
(Arseneau and Rodenburg, 2004; Smith, 2003). This is in great contrast to a teacher-
centered, passive learning atmosphere, such as lectures in a large classroom.

Active learning is a major component of this CAL. Incorporating multiple forms
of media, variety in content delivery, and use of different question formats allows the
learners to be more engaged, compared to traditional formats of learning (Arseneau and
Rodenburg, 2004). In addition, learners receive immediate feedback from the questions
they answer, allowing for self-evaluation and reinforcement of learning objectives. There
is a chance that learners may only view this CAL at a superficial level; if true, the learner
is still dictating what is viewed and the direction taken through the CAL, and is thus
engaged in an active learning process. .

Experimental learning is another key factor for this site. Learners are able to

discover unknown facts and enhance their knowledge throughout each module’s

105

storyline, by the questions asked, or investigating the various links and references for
more information. Experimental learning is the primary way in which adult learners

further their knowledge (Kolb, 1984).

CONCLUSION

This CAL is meant to be an adjunct to traditional formats of learning in the
classroom or clinical settings. The exact role of CAL in veterinary education is still
being developed, as there are several complicating factors: a sharp learning curve,
unwillingness of faculty to develop material, and the start-up costs for implementing the
technology (Childs et al., 2005; Dale et al., 2005; Short, 2002). At a minimum, CAL is
an excellent addition to traditional learning (Nerlich, 1995; Rosenberg et al., 2005). At
the onset of this project, the intention was for this CAL to be used in veterinary school
curriculums, but there is potential for this to be used in continuing education for graduate
veterinarians too. Further evaluation of this tool is needed to determine its effectiveness
as a teaching tool.

This CAL tool continues to be enhanced and evolve and the most recent version is
now located at the following website: http://arls.cvm.msu.edu. Additional modules have
been added regarding basic concepts of microbiology, pharmacology and public health
beyond what is mentioned in the ‘Principles’ section. Another change that has been
made is that the left-side pane has been moved to the top of the CAL, allowing the
storyline to contain more of the screen. These changes were made by the design team,
which includes Michigan State University and CDC personnel, to enable authors of the

modules to more easily edit content in the future.

106

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Table 4.1 Usability Testing Task List for the CAL

 

Go to the following website http://www.cvm.msu.edu/cdc the usemame and
password is “cdcmsu”.

1. Find objectives for this program
2. Start the Investigation
3. Use the side bar to list some superbugs

4. In the next page locate and define 3 methods for resistant genes to be
transferred

5. Go to “Use in Agriculture” and answer the first question

6. On the next page work through the first question, determine if
Tetracycline is an over the counter drug

7. Answer the first two questions in the “Small Animal Clinic” section

8. In the “Microbiology” section review the video on “Bacterial Culturing”
9. In the “Pharmacology” section locate the dose formula for an antibiotic
10. Use the glossary to define pharmacokinetics

11. Go to the “Milk Replacer” case study

 

 

 

108

Cited Literature

Arseneau, R., Rodenburg, D., 2004, The Develomental Perspective, In: Pratt, D. (Ed.)
Five Perspectives on Teaching in Adult and Higher Education. Krieger
Publishing, Malabar, FL, p. 304.

AVMA 2008. One Health: A New Professional Imperative (Schaumburg, IL, American
Veterinary Medical Association).

Childs, S., Blenkinsopp, E., Hall, A., Walton, G., 2005, Effective e-leaming for health
professionals and students—barriers and their solutions. A systemic review of the
literature--fmdings from the HeXL project. Health Information and Libraries
Journal 22, 20-32.

Dale, V., McConnell, G., Short, A., Sullivan, M., 2005, Ten Years of CLIVE (Computer-
Aided Leanring in Veterinary Education) in the United Kingdom. J Vet Med Educ
32, 47-50.

Davey, P., Garner, S., 2007, Professional education on antimicrobial prescribing: a report
from the Specialist Advisory Committee on Antimicrobial Resistance (SACAR)
Professional Education Subgroup. J Antimcirob Chemother 60, i27-32.

Hird, D., King, L., Salman, M., Werge, R., 2002, A Crisis of Lost Opportunity-
Conclusions from a Symposium on Challenges for Animal Population Health
Education. J Vet Med Educ 29, 205-209.

J ETACAR 1999. The use of antibiotics in food-producing animals: antibiotic resistant
bacteria in animals and humans, Tumidge, J ., ed. (Canberra, Biotext).

Kolb, D., 1984, Experiential Learning. Prentice Hall, Englewood Cliffs, NJ, 256 p.

Morley, P., Apley, M., Besser, T.E., Burney, D., Fedorka-Cray, P., Papich, M., Traub-
Dargatz, J ., Weese, J ., 2005, Antimicrobial drug use in veterinary medicine. J Vet
Intern Med 19, 617-629.

Nerlich, S., 1995, Computer-Assisted Learning (CAL) for General and Specialist Nursing
Education. Australian Critical Care 8, 23-27.

Rosenberg, H., Sander, M., Posluns, J ., 2005, The effectiveness of computer-aided
learning in teaching orthodontics: A review of the literature. American Journal of
Orthodontics and Dentofacial Orthopedics 127, 599-605.

Short, N., 2002, The use of information and communication technology in veterinary
education. Research in Veterinary Science 72, 1-6.

109

Smith, R., 2003, The Application of Information Technology in the Teaching of
Veterinary Epidemiology and Public Health. J Vet Med Educ 30, 344-350.

Valcke, M., De Wever, B., 2006, Information and communication technologies in higher
education: evidence-based practices in medical education. Medical Teacher 28,
40-48.

WHO 1998. The Medical Impact of the Use of Antimicrobials in Food Animals. Report
of a WHO meeting. Berlin, Germany, 13-17 October, 1997. In
WHO/EMC/ZOO/97.4 (Geneva, World Health Organization).

WHO 1999. Containing Antimicrobial Resistance: Review of the Literature and Report H
of a WHO Workshop on the Development of a Global Strategy for the
Containment of Antimicrobial Resistance. In WHO/CDS/CSR/DRS/99.2
(Geneva, Switzerland, World Health Organization), p. 60. L

 

WHO 2003a. Impacts of antimicrobial growth promotion termination in Denmark
(Geneva, World Health Organization).

WHO 2003b. Joint FAO/OIE/WHO Expert workshop on non-human antimicrobial usage
and antimicrobial resistance: Scientific assessment (Geneva).

110

CONCLUSIONS

111

CONCLUSIONS

The overall aim of this dissertation was to enhance scientific knowledge regarding
the appropriate use of antimicrobial agents in dairy cattle. To meet this aim there were a
few questions; 1) Do calves fed milk replacer with antimicrobial agents grow faster and
have a lower disease burden, 2) Do plasma-steady state pharmacokinetic values for
oxytetracycline as fed in medicated milk replacer (MMR) achieve minimum inhibitory
concentrations for Escherichia coli and Pasteurella multocida, and 3) Will a computer-
aided learning module be useful to teach veterinary students concepts about appropriate
use of antimicrobial agents? All of these questions have been answered in the previous
chapters.

In thinking about the results of my research, the first two studies (chapters 2 and
3) compliment each other well. The milk replacer study evaluates the on farm benefit of
antimicrobials in feed, while the pharmacokinetic study investigates the benefit of
oxytetracycline in milk replacer. What is interesting to me is that there are paradoxical
results and differences that are either biologically but not statistically significant or vice
versa. For example, oxytetracycline and neomycin benefits growth yet there is no
difference in morbidity between treatment groups. While there is a ntuneric difference in
mortality rates between treatments but was not statistically significant.

From a production standpoint there is a benefit to feeding MMR but is it
biologically significant with a 6 kg difference at 150 days of age? To me this is not a
large enough difference for the economic investment of an additional 5 dollars per calf
from feeding MMR. This statistical difference is in part likely due to being the largest

study to date as I calculated a sample size to determine a 10-gram difference in average

112

daily gain, instead it was 37 g difference. The difference in weight could have been
higher if there was no enteric disease outbreak, but in looking at the data prior to and
afier the outbreak the difference in weight between treatments was almost the same.

In my opinion the most important finding in these studies is that there is no
difference in morbidity but there were 28 deaths in the first two weeks of life with almost
all of them during the enteric outbreak. This is almost 80% of the deaths for the whole
year, of which 19 were in the non-medicated milk replacer group. Though there is no
statistically significant difference between treatments, there is a definite biological
significance. For me to walk onto a farm and tell them not to feed MMR, when there is a
2 fold increase in the number of deaths in calves fed non-medicated milk replacer, would
be hard to do.

When you look at the pharmacokinetic values of oxytetracycline there clearly is
no benefit for the treatment of systemic infections, such as pneumonia. Nor did it reduce
the morbidity rate in MMR calves. Yet there may be some benefit to feeding MMR as it
may mitigate the severity of illness, shown by the decrease in mortality between
treatment groups. In this study, we only know the plasma concentrations that were
attained for oxytetracycline. However, we were not able to evaluate what was going on
in the gastrointestinal tract with oxytetracycline. Secondly, we only looked at one drug,
as neomycin may also be playing a major role in reducing mortality. We know that
aminoglycosides, like neomycin, are not well absorbed from the gut lumen. Therefore
drug concentrations may be high enough in the gastrointestinal tract to minimize the
severity of enteric disease. Neomycin may also be playing more of a role as there was

increased growth in pre-weaning calves with no respiratory disease compared to calves

113

with respiratory disease pre-weaning, while oxytetracycline plasma concentrations in
calves fed the MMR did not reach MIC values for P. multocida.

. If I was to walk onto a farm and was asked if feeding MMR is of any benefit, I
would have to say YES but with some careful considerations. Results from my research
are only from one farm and extrapolating these results to other farms is hard to do, but in
combination with previous studies you can make some decisions. From an animal
welfare point it would be hard to argue against the use of MMR during an enteric disease
outbreak. Now do calves need to be on MMR for the entire suckling period? This is
where veterinary medicine has to look at the pros and cons of such a practice. There is
likely to be few or no new classes of antimicrobial agents coming to market and therefore
the profession needs to preserve and prolong efficacy of the current drugs for treating ill
animals. Given that I would recommend only using MMR during the first few weeks and
switch over to a product that contains no antimicrobial agents.

Now switching gears, developing the CAL tool (chapter 4) was a great learning
experience in respect to working in a team environment, but also exciting to be part of
educating students in a non-traditional way. This tool can be very beneficial to a
veterinary curriculum as it can be used in different ways: to supplement class material for
various courses (microbiology, pharmacology and public health), to clinical settings as a
review or learn about specific issues. There is also a possibility that veterinarians and/or
producers could use the tool for continuing education.

A concern of mine is that I left this project over two years ago and the tool has yet
to be released to the general public. This is mainly due to the bureaucratic process at the

Centers for Disease Control and Prevention (CDC) to review and approved material.

114

 

Given this, this leads to my biggest concern as to who is going to ensure information
stays current. For instance in the ‘Principles’ module there is a question regarding the
likely source of Salmonella infection and peanut butter sandwich is a possible answer that
is labeled unlikely, which is no longer true given a recent Salmonella Typhimurium
outbreak in peanut butter. There will be a need for an administrator to be identified to
ensure content is kept current either by themselves or through a team. Overall, I look
forward to the day the tool is released for use by veterinary curricula, as I believe it will

be an effective way for students to learn about the prudent use of antimicrobial agents.

115

 

APPENDIX

ll6

 

APPENDIX A

Appropriate Use of Antimicrobial Agents in Veterinary Medicine

117

Appropriate Use of Antimicrobial Agents in Veterinary Medicine

This computer-aided learning module can be viewed at the following website:
http://old.cvm.msu.edu/cdc. At this time the CAL is password protected as both the
usemame and password are cdcmsu. In the near future the CAL will be moving to the
following location: http://arls.cvm.msu.edu. If the CAL is not working feel free to
contact Dr. Paul Bartlett at Michigan State University via phone or email. His contact
information is: phone 517-432-3100 email bartlett@cvm.msu.edu

The following pages contain screen shots to show you what the CAL looks like.
Modules that are included are: Principles and Dairy Cattle, which include medicated
milk replacer, neonatal scours, contagious mastitis, and farm based mastitis program.

118

 

 

 

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