THE EFFEC‘? 0F CER‘EMN DEETARY INGREDENTS
UPON THE INCIDENCE (3F BLGOL‘SPOTS IN
CHICKEN EGGS

Thai: 50: the Degree of Ph. D.
MICHIGAN STATE UNWERSITY
James Eritrean Wald
19:52

‘thrj‘fé

This is to certify that the

thesis entitled

The Effect of Certain Dietary Ingredients

ion the Incidence of Bloodspots in Chicken Eggs.

presented by

James Britten Ward

has been accepted towards fulfillment
of the requirements for

Ph.D. degree in Poultry Science

r Mfic/ professor

Dat@ 4/ /7é)/

 

0-169

 

LIBRARY

Michigan State
University

 

ABSTRACT

THE EFFECT OF CERTAIN DIETARY INGREDIENTS UPON THE
INCIDENCE OF BLOODSPOTS IN CHICKEN EGGS

by James Britton Ward

Experiments were conducted with three strains of Single Comb
White Leghorn hens to study the effects of certain dietary ingredients
on the incidence of bloodspots -- particularly those concerned with
internal hemorrhaging. The number and size of bloodspots in eggs on an
individual bird basis occurring before, during and after feeding of the
experimental diets were used as criteria to evaluate the effects of the
dietary treatments.

Additions of vitamin 312, selenium, crude cottonseed oil, crude
corn oil, a refined soybean-corn oil blend, beta-aminopropionitrile
fumarate (BAPN), vitamin K (menadione sodium bisulfite), reserpine (an
antihypertensive agent), and combinations of reserpine with sulfa-
quinoxaline, sulfaquinoxaline with soybean-corn oil blend, vitamin K
with soybean-corn oil blend and dehydrated alfalfa leaf meal plus
animal fat were without influence on number or size of bloodspots
produced. Differences in bloodspot incidence between strains of birds
were noted.

The percentage of red blood cells (hematocrit value) was different
for strains, periods and treatment while prothrombin times (blood
clotting potential) were not different for strains or treatments.
Prothrombin times were not determined for each period.

Hematocrit values were significantly depressed by a high level of

cobalt while a low level of cobalt caused a non-significant increase in

hematocrit values. Values determined in July were lower than those in
March or May. \
The highest level of selenium (5 ppm), reserpine (2mg/1b) plus
sulfaquinoxaline (0.125%) and soybean-corn oil blend plus sulfaqui-
noxaline (0.1%) caused a significant depression in egg production.

The highest level of vitamin B12 resulted in a non-significant increase

in production as did the addition of 1 ppm of selenium.

The Effect of Certain Dietary Ingredients

Upon the Incidence of Bloodspots in Chicken Eggs

BY

James Britton Ward

A Thesis
Submitted to
Michigan State University
in partial fulfilment of the requirements
for the degree of
Doctor of Philosophy
Department of Poultry Science

1962

ACKNOWLEDGMENTS
The author wishes to express his sincere thanks to Dr. Philip J.
’ /

Schaible for his excellent guidance and personal interest during this
study.

The author also wishes to express his appreciation to Dr. Howard

. C. Zindel, Chairman of the Department of Poultry Science, and to Michigan

State University for making available Poultry Science Department labor-
atory and farm facilities for the conduct of this project.

The writer is indebted to Drs. Robert K. Ringer, Theo H. Coleman
and L. E. Dawson for their construCtive review of this thesis.

He is expecially grateful to Jerome Yates for his generous assist-

ance, personal interest and helpful suggestions with respect to pro-

cedures employed and collection of the data.

ii

TABLE OF CONTENTS

Introduction

Review of Literature

Experimental Procedure and Results
General Procedure
Introduction to Experiments A and B
Experiment A: Procedure and Results
Discussion of Experiment A
Experiment B: Procedure and Results
Discussion of Experiment B

General Discussion

Conclusions

Literature Cited

Appendix

iii

Page

23
27
39
41
48
50

56

LIST OF TABLES

Table
No.

 

1 Composition of the Basal Ration Fed in Experiment A
2 Changes in Basal Ration for Experiment A

3 Change in Bloodspot Incidence Between Periods 1 and 3
When Calculated on an Individual Bird Basis

4 Change in Bloodspot Incidence Between Periods 3 and 5
When Calculated on an Individual Bird Basis

5 Percentage Egg Production for Periods l and 3
6 Percentage Egg Production for Periods 3 and 5

7 Analysis of Variance of Egg Production Data for Birds in
Group 1 for Periods 1, 3 and 5

8 Analysis of Variance of Egg Production Data for Birds in
Group 2 for Periods l, 3 and 5

9 Analysis of Variance of Egg Production Data for Birds in
Group 3 for Periods 1, 3 and 5

10 Hematocrit Values for Birds in Group 1.

11 Analysis of Variance for Hematocrit Values for Birds in
Group 1 for Periods l, 3 and 5

12 Results of Grading of Eggs by a Commercial Grader
l3 Composition of the Basal Ration Fed in Experiment B
14 Changes in Basal Ration for Experiment B

15 Change in Bloodspot Incidence Between Periods 1 and 3 When
Calculated on an Individual Bird Basis

16 Change in Bloodspot Incidence Between Periods 3 and 5
When Calculated on an Individual Bird Basis

17 The Effect of Handling of Birds for Collection of Blood
for Hematocrit Determinations on the Incidence of Bloodspots

18 The Effect of Handling Birds and Treating for Mites on
the Incidence of Bloodspots

19 Analysis of Variance of Prothrombin Times Obtained During
Period 5

iv

222$
11

12

13

14

15
16

17

18

19

20

21

22
29

3O

31

32

33

33

34

Table

No. Page
20 Analysis of Variance of Hematocrit Values ‘ 35
21 Percentage Egg Production for Periods 1 and 3 36
22 Percentage Egg Production for Periods 3 and 5 37
23 Analysis of Variance of Egg Production Data for 38 '

Periods 1, 3 and 5

 

Bloodspot Incidence

Periods

Bloodspot Incidence

Periods

Percentage of Bloods

1 and 3

Percentage of Bloods

3 and 5

Percentage of Bloods

LIST OF APPENDIX TABLES

Expressed as the Percentage of Eggs
1 and 3

Expressed as the Percentage of Eggs
3 and 5

pots by Strains Within Treatments
for Birds in Group 1

pots by Strains Within Treatments
for Birds in Group 1

pots Falling Into Each Size Category

During Periods 1 and 3

Percentage of Bloodspots Falling Into Each Size Category
During Periods 3 and 5

Percentage of Bloodspots Falling Into Each Size Category
by Strains Within Treatments During Periods l and 3 for

Variance
Period 1

Variance
Period 3

Variance
Period 5

Variance
Period 1

Variance
Period 3

Variance
Period 5

Bloodspot Incidence

Periods

Bloodspot Incidence

Periods

Table
No.
1
Laid During
2
Laid During
3
for Periods
4
for Periods
5
6
7
Birds in GrOup 1
8 Analysis of
Group 2 for
9 Analysis of
Group 2 for
10 Analysis of
Group 2 for
11 Analysis of
Group 1 for
12 Analysis of
Group 1 for
13 Analysis of
Group 1 for
14
Laid During
15
Laid During
16

of Egg Production Data for Birds in

of Egg Production Data for Birds in

of Egg Production Data for Birds in

for Hematocrit Values for Birds in

for Hematocrit Values for Birds in

for Hematocrit Values for Birdsin

Expressed as the Percentage of Eggs

1 and 3

Expressed as the Percentage of Eggs
3 and 5

Percentage of Bloodspots by Strains Within Treatments

for Periods

l and 3

vi

Page

56'
57
58
59
60
61

62

63.
64
65
66
67
68
69
7O

71

Table
No.

 

17
18
19

2O
21
22

23

Percentage of Bloodspots by Strains Within Treatments
for Periods 3 and 5

Percentage of Bloodspots Falling Into Each Size Category

.During Periods l and 3

Percentage of Bloodspots Falling Into Each Size Category
During Periods 3 and 5

Analysis of Variance of Egg Production Data for Period 1
Analysis of Variance of Egg Production Data for Period 3
Analysis of Variance of Egg Production Data for Period 5

Bloodspot Incidence for Birds at Two Levels of Production
Without Regard for Treatment

vii

Page
72

73

74

75
76
77

78

INTRODUCTION

From one-half to three percent of all eggs entering primary market
channels are rejected because of bloodspots. Another five to seven
percent of the eggs actually contain bloodspots which are too small to
be detected by the candling process and thus are sold to the consumer.
Bloodspots cause a substantial loss to poultrymen but the exact amount
is difficult to estimate because of the difficulty in ascertaining what
effect they may have on the consumer.

It has been shown that the heritability of bloodspot incidence is
quite high. However, while some strains of birds have a very low
incidence, it has been virtually impossible to eliminate bloodspots
completely by selection and breeding.

For the purposes of experimentation, it would be desirable to find
some means of increasing the incidence of bloodspots. From a practical
standpoint, of course, it would be desirable to find some means of
reducing the incidence of bloodspots. With these thoughts in mind the
hypothesis was develOped that perhaps bloodspot incidence could be in-
fluenced through nutrition by means of including or deleting certain
dietary ingredients.

The studies reported herein involved dietary supplementation with
various levels of vitamin 812, selenium, vegetable and animal fats,
vitamin K, sulfaquinoxaline, a tranquilizer, and beta-aminopropionitrile
fumarate (BAPN). Of particular interest were treatments affecting ‘

hemorrhaging and blood clotting.

REVIEW OF LITERATURE

Nalbandov and Card (1944) reasoned that bloodspots were the result
of intrafollicular hemorrhaging prior to ovulation and that hemorrhaging
may have occurred several days before the release of the yolk. Later,
Stiles et a1. (1958) injected birds with radioactive phosphorus (P32)

3 1/2 hours after ovulation and could detect no radioactivity in a
bloodspot present at oviposition. When the radioactiye phosphorus

was injected nine hours prior to ovulation, radioactivity was detected
in a bloodspot present in the egg at oviposition. These workers concluded-
that the formation of the bloodspot occurred sometime within the nine-
hour period prior to ovulation.

Quinn and Godfrey (1940) reported that there were breed and family
differences in bloodspot incidence. The heritability of bloodspots was
estimated to be 0.32 by Farnsworth and Nordskog (1955), who based their
estimation on a sample of 15 eggs per hen. Lerner et a1. (1959) in-
creased the incidence of bloodspots from between 0.5 and 1.3 up to 23
percent through selection. The heritability of bloodspotting, expressed
as percent of all eggs laid which contained bloodspots, was found to be
0.5.

The effect of the absence or presence of certain dietary ingredients
linked with capillary fragility, general subcutaneous bleeding and aortic
rupture on the incidence of bloodspots has been investigated. A de-
ficiency of vitamin P, a crude mixture of glycosides of eriodictin and
herperidin, increases capillary fragility and permeability (Titus, 1955).
Rutin, a crystalline glycoside of quercitin, effective in mending capil-
lary fragility and preventing petechial bleeding in humans, has been

studied wlth respect to bloodspot incidence as has vitamin C which causes

2

 

a decrease in capillary fragility in some cases but not in others (Tey,
cited by Houssay, 1955). Carver and Henderson (1948) investigated the
effect of rutin and vitamin C in the diet of laying hens but found no
reduction in the incidence of bloodspots.

According to Yacowitz e£_al. (1955), 0.1 percent sulfaquinoxaline
caused hemorrhaging at four to five weeks of age and decreased hematocrit
values at two to three weeks of age. .Cuckler and Ott (1955) reported
that a level of at least 0.4 percent sulfaquinoxaline in the diet of
broilers was required to produce a prolonged increase in blood clotting
time and even this gave only a slight increase in prothrombin time.
Anderson et a1. (1954) stated that broilers developed hemorrhages when
raised on a wire-floored battery and fed a simplified corn-soybean oil
meal ration. This hemorrhagic condition could be overcome by the
addition of vitamin K. Data presented by Hare et a1. (1953) indicated
that birds fed a corn-soybean oil meal ration from one to six weeks of
age developed a condition similar to the so-called hemorrhagic disease.
Balloun and Johnson (1952) increased blood clotting time by feeding
underheated soybean oil meal to chicks. Additions of vitamin K did not
restore blood clotting time to normal.

When Stephens and Tugwell (1960) fed a "low” vitamin K diet to
chicks, blood clotting time was increased. Blood clotting time was de-
creased when 2.5 percent autoclaved and dried feces or three percent
alfalfa leaf meal was added to the ration. Menhaden fish meal added
to the diet did not influence blood clotting time. However, dicumarol
in the ration of chicks increased blood clotting time (Harms and Tarver,
1957).

By feeding .036 percent beta-aminopropionitrile fumarate (BAPN),

Roy and Bird (1959) produced leg deformities and aortic rupture in chicks
prior to six weeks of age. When Barnett and Morgan (1959) fed BAPN at
.06 percent in both low and high fat diets, incidence of internal
hemorrhaging was 9.5 percent and 15 percent, respectively. Waibel and
Pomeroy (1958) produced aortic rupture in turkeys by feeding .02 percent
BAPN. Vitamin K added to the diet did not consistently reduce BAPN-
induced hemorrhaging (Barnett et a1., 1958).

Card and Nalbandov (1944) reported that allowing birds to run on
range reduced bloodspot incidence but that fresh-cut grass fed inside the
poultry house was not as effective in reducing the incidence. However,
Denton (1947) found no effect from the feeding of green grass on the
incidence of bloodspots. Nalbandov and Card (1947) found no influence
from feeding defatted wheat germ, wheat germ oil, liver meal, condensed
fish solubles, yeast, a "mixture of trace minerals” or folic acid on
bloodspot occurrence.

According to Sauter et al. (1952) ten percent of alfalfa leaf meal
in the ration lowered the incidence of bloodspots. By feeding hens vit-
amin A at levers of 200, 300, and 400 IU per hundred grams of diet,
Bearse et al. (l953)found that a decrease in bloodspots occurred as the
vitamin A level increased. Later, Bearse and Berg (1958) reported that
vitamin A levels of 3,000 and 6,000 IU/lb. lessened the bloodspot problem.
The incidence of bloodspots was greater on high energy diets than on
low energy diets. The authors attributed this higher incidence to a
twenty percent reduction in feed intake with a consequent reduction in
vitamin A intake for birds on the high energy diets. Bearse et al.
(1960) found that vitamin A levels of 1200 to 1600 IU/lb. produced a

minimum of bloodspots and that no further reduction occurred from in-

creasing the vitamin A level up to 15,000 IU/lb. Munro (1952) reported
that the percentage of bloodspots, as determined by candling, was con-
sistently lower on a low energy ration as compared to a high energy
ration (1.77 vs. 2.91 percent).

Using various levels of sodium chloride, potassium chloride, sodium
carbonate and their combinations, Pope et a1. (1961) found no influence
on the incidence of bloodspots. Vitamins E and K, corn fermentation
products and Vigofac were also without influence while "low" levels of
vitamin A caused an increase in bloodspot incidence.

When Pope (1959) fed high levels (ten to twenty times the normal
amount) of vitamin B12, there was an increase in the number of bloodspots.'
When these high levels were fed with vitamin A, the adverse effect of
vitamin 312 was reduced somewhat. Depleting hens of vitamin B12 or
supplementing B12 depleted birds had no effect on the incidence of
bloodspots (Bearse, 1961). Three grams of vitamin K and two pounds of
methionine per ton in the ration of caged layers dki not influence the
incidence of bloodSpots.

Erythrocytosis (abnormal production of red blood cells) can be in-
duced by the administration of "excessive" doses of vitamin 812 or other
cobalt compounds with a resultant increase in peripheral resistance which
may in turn lead to hypertension (Kugelmass, 1959). Weiss (1958) reported
that incidence of bloodspotting was higher in a high blood pressure line
of birds than in a low blood pressure line, but not significantly so.
Speckmann and Ringer (1961) reduced the blood pressure in turkeys by
feeding reserpine. Sturkie (1959) reported that reserpine causes hypo-

tensive changes in chickens.

EXPERIMENTAL PRODECURE AND RESULTS

General Procedure

 

The hens were housed in individual double-tiered, offset-type
cages. Cages were arranged in groups of eight and feed and water were

provided ad libitum. Water troughs extended the full lenght of each

 

row of cages and were cleaned daily. Manure was removed once each
week with an automatic pit cleaner.

Eggs were collected daily and examined twice weekly on the same
days of each week. All eggs were broken out on a glass plate with
mirrors both underneath and behind. This permitted full observation of
the egg for bloodspots. Bloodspots were recorded in one of three
groups according to size: l/l6th inch (1.6 millimeters) or less in
diameter; greater than l/l6th inch (1.6 millimeters) but less than
3/l6th inch (4.8 millimeters) in diameter; 3/16th inch (4.8 millimeters)
or greater in diameter. Bloodspot data from hens laying in only one of
two periods being compared were not included.

The effect of treatment on bloodspot incidence was analyzed
statistically on the basis of the number of birds influenced in'a like
manner. Treatments were group as follows: 1 through 6, 7 through 9 and
11 through 16. The groups will be referred to as Groups 1, 2 and 3
respectively. Treatment 10 was not considered in the analysis because
the birds were not kept on the experimental diet the allotted time.

The Chi-square method (Snedecor, 1956) was employed because it is de-
signed to measure the deviation of frequency of sample numbers in
different categories from the expected frequency. In this case, birds
were placed in one of three categories based on change in bloodspot
incidence for each bird between anv two periods.

6

For the analysis of egg production data hematocrit values and
prothrombin times, the analysis of variance (Snedecor, 1956) was
employed. Tests for significant differences between means were con-
ducted according to the method of Duncan (1955).

Hematocrit values were determined by the micro technique as
described by McGovern g£_al. (1955). Blood from the brachial vein in
the wing was collected directly into capillary tubes. Centrifugation
was carried out in'a micro-capillary centrifuge and readings were made
on a micro-capillary reader according to the method outlined by Jones

(1956).

INTRODUCTION TO EXPERIMENTS A AND B

Conflicting reports with reference to the influence of vitamin 312
on bloodspot incidence exist in the literature. There are indications
that excessively high levels of vitamin B12 increases the number of
bloodspots, but this influence may be modified by factors such as
various levels of vitamin A. Since vitamin B12 and cobalt have been
reported to stimulate erythrocyte production, hematocrit values (per-
centage of red blood cells) were determined.

Pope (1959) obtained a slight decrease in bloodspot incidence by
feeding crude cottonseed oil. On the basis of Pope's work the decision
was made to compare the effect of vegetable oils and animal fat on the
incidence of bloodspots in chicken eggs.

In recent years, vitamin K has received considerable attention in
the popular press as being important in decreasing bloodspot incidence.
In addition, claims are being made that the vitamin K currently being
used is more stable. Although earlier experimental work had indicated
no effect from vitamin K on bloodspot incidence, it seemed advisable to
conduct new tests in light of the greater stability claims and reports
of reduced bloodspot incidence as a result of feeding the more stable
vitamin K. Beta;aminopropionitrile and sulfaquinoxaline, were included
beCause of their association with internal hemorrhaging.

Since reserpine is well known as a hypotensive agent it was
theorized that if there was a correlation between high blood pressure
and bloodspot incidence as suggested by Weiss (1958) then reserpine

should cause a reduction in incidence of bloodSpots.

.Experiment A: Procedure and Results

 

The effects of vitamin 312, cobalt, selenium and fat or oil were
studied with respect to bloodspot incidence. The experiment was di-
vided into five intervals denoted as Periods 1 thrOugh 5. During Periods
l, 4 and 5 all birds were fed the same basal diet; during Periods 2 and 3,
birds on treatments 1, 7 and 11 were on basal; all others were on
experimental diets. Periods 2 and 4 were 14 days each in length and
were considered "switch over” periods. Observations for bloodspots
were made during these periods but were not included in the data.

Periods l and 5 were each 60 days in length for all treatments.
Period 3 was 60 days in length for treatments 1 through 6 and 68 days
in length for treatments 7 through 16. At the end of 60 days, all of
the experimental diets for treatments 7 through 16 had not been consumed,
thus Period 3 was extended for these treatments in order to utilize the
experimental diets completely.

The calendar dates for this experiment were as follows:

Group 1 Group 2 Group 3
Period 1 Dec. 29 to Feb. 26 Nov. 19 to Jan. 17 Same
Period 2 I , Feb. 27 to Mar. 11 Jan. 18 to Jan. 31 as
Period 3 Mar. 12 to May 10 Feb. 1 to Apr. 18 group
Period 4 May 11 to May 24 Apr. 9 to Apr. 22 2
Period 5 May 25 to July 23 Apr. 23 to June 21

Three strains of birds were used. One had a history of "high”
incidence §Strain A), another a history of "medium" incidence (Strain B)
and the third a history of "low" incidence (Strain C). There were 24
birds per treatment in Group 1 and 16 birds-per treatment in Groups 2

and 3. Group 1 contained birds from Strains A, B and C; Group 2 from

10

strains B and C; Group 3 from strain C only.

Group 1 was fed various levels of vitamin B12 or cobalt (the
lowest level of cobalt was the equivalent of that found in 30‘micro-
grams of vitamin B12; the highest level of cobalt was the equivalent of
that found in a commercial trace mineral mix). Group 2 was fed
different levels of selenium and Group 3 different levels of vegetable
oils and animal fat.

A total of 34,155 eggs were examined in this experiment and the
data are summarized in Tables 3 through 12 and Appendix Tables 1

through 13.

11

Table 1. Composition of the Basal Ration Fed in Experiment A

 

 

 

 

Ingredient Amount per th.
Corn, yellow No. 2, ground 66.70 ‘lbs.
Soybean oil meal, 44% protein 19.00 "
Meat and bone scrap, 50% protein 3.00 "
Fish meal (with solubles), 60% protein 1.25 ”
Alfalfa leaf meal, dehyd., 20% protein 1.30 "
Fat, animal, No. 2 yellow grease 1.50 ”
Dicalcium phosphate, feed grade 2.00 “
Limestone, ground 5.00 "
Salt, iodized .25 ”
Manganese sulfate, 70% feed grade .025 "
Riboflavin suppl. (l6 gm/lb) ' 1 gram
Vitamin A (10,000 IU/gm) ~ 20 "
Vitamin D3 (3,000 ICU/gm) 10 ”
Total 100.00 + lbs.

 

Calculated analysis:

Crude protein % 16.9 Vitamin 812 (meg/1b) 1
Crude fat Z 4.5 Riboflavin (mg/lb) 1.
Crude fiber % 3.1 Niacin (mg/lb) 10.
Calcium % 2.8 Pantothenic acid (mg/1b) 2
Phosphorus % .8 Choline (mg/lb) 406
Vitamin D (ICU/lb) 300 Energy (Cal/1b) 945

Vitamin A (IU/lb) 3563

 

.oHumu Hus m CH HHo :Hoo paw HHo cmwn>0m meHmmH wchHmucoo UGmHH <

 

 

 

 

 

 

 

.HHo Ho umw mo mcoHuumHDLSm N
no wcoHqupm HoH mummcmmsoo Cu mums mumB Chou Ho mowmucmoumm msu :H mmwcmno
H
o.m uu nu nn nu Damonom HHo cuoo mpouu
nu o.m nu nu un ucoouma NHHo snooncmmchom
nu nn o.m nu nu ucmowoa HHo pwmmCODDOU mpsuu
nu un uu m.m nu unmoumm umH HmEHc<
mH qH mH NH HH
mumHQ
H
OH m H uu Ema AONmm.mo mm Nmz mmv EoHconm
OH a w m
mumHQ II.

Nm.H nn nn nu uu HH\on Amoo oo mmv uHmnoo
-- oa cm ma _ -- aa\wue . Nam caeaua>
m q m N H much . mucmHUmpwcH

mumHQ

 

 

< u:¢Ewumuum new comumm Hemmm cH mmmcmzo .N mHeme

13

Table 3. Change in Bloodspot Incidence Between Periods 1 and 3
When Calculated on an Individual Bird Basis

A.

 

 

 

 

Percentage

Average Percentage Percentage of birds Chi
percent of birds of birds showing square
Treatment change increasing decreasing no change values
1 - 2 43 57 0 2.15

2 + 1 44 44 12 2-71

3 - 13 p 11 78 11 3.97

4 - 5 35 60 5 .11

5 - 10 27 68 5 .53

6 - 2 35 55 ' 10 .42
Total 9.89

Chi-square value 10 degrees of freedom (P<:.01) = 23.21

7 + 3 60 20 20 1.23

8 + 1 42 50 8 1.59

9 + l 50 p 33 17 .02
Total 2.84

Chi-square value 4 degrees of freedom (P<f.01) = 13.28

11 + 4 28 36 36 3.23

12 - 3 31 46 23 .26

13 - .3 25 56 19 .98

14 - 2 19 62 19 1.19

15 - 2 47 47 '6 2.79

16 - 2 38 54 8 1.22
Total 9.67

Chi-square value 10 degrees of freedom (P<;.01) = 23.21

 

14

liable 4. Change in Bloodspot Incidence Between Periods 3 and 5 When
Calculated on an Individual Bird Basis

 

 

 

 

 

 

'Percentage
Average Percentage Percentage of birds Chi
percent of birds of birds showing square
'Ifireatment change increasing decreasing no change values
1 + 4 53 42 5 .49
2 . - 7 31 62 6 7.45
3 + 3 53 29 18 1.46
4 + 8. 62 38 0 1.86
5 '+ 6 59 ‘35 6 .32
6 + 5 61 11 28 9.82
Total ' 21.40
Chi-square value 10 degrees of freedom (P (.01) = 23.21
7 - 8 15 69 16 .41
8 - 2 33 67 0 2.36
9 - 4 20 50 30 2.11
Total 4.88
Chi-square value 4 degrees of freedom (P.<.01) = 13.28
11 - l 17‘ 58 25 1.83
12 - l. 33 59 8 .46
13 - 2 30 67 13 1.39
14 '- 1 43 36 21 1.54
15 + l 36 50 14 .04
16 + 3 50 42 8 1.72
Total ‘ 6.98

Chi-square value 10 degrees of freedom (P (:01) = 23.21

 

15

Table 5. Percentage Egg Production for Periods l and 3

 

 

 

Treatment , Period 1 Period 3 Amount of change
Z Z
1 64 51 - l3
2 62 54 - 8
3 59 59 0
4 58 67 + 9
5 69 6O ’ - 9
6 58 55 - 3
7 57 65 + 8
8 59 75 + 16
9 64 36 - 28
11 7O 71 + 1
12 69 69 0
13 79 68 - 11
14 72 69 - 3
15 63 60 - 3

16 68 69 + l

16

tfaible 6. Percentage Egg Production for Periods 3 and 5

__-

 

Treatment Period 3 Period 5 Amount of change
% ' %
1 51 54 + 3
2 h 54 66 + 12
3 59 57 - 2
4 67 67 0
5 60 64 + 4
6 55 48 - 7
7 65 68 + 3
8 75 65 - 10
9 36 53 + 17
11 71 53 - 18
12 69 51 - 18
13 68 72 + 4
14 69 68 - 1
15 6O 56 f - 4

16 69 66 - 3

l7

irable 7. Analysis of Variance of Egg Production Data for Birds in
Group 1 for Periods 1, 3 and 5

 

 

Mean

 

 

Means not underscored by the same line are

 

Source of Degrees of
\rariation freedom square F .01
.12reatments 5 387.8 1.89 .06
EStrains 2 1087.0 5.29 .66
I?eriods 2 119.5 .58 .66
T? X P 10 215.7 1.05 .46
'T X S 10 286.6 1.31 .46
P X S 4 347.8 1.70 .36
T X P X S 20 220.0 1.07 .61
Error 311 205.1
Strains A B
Av. eggs/hen .32;_ 37.2

significantly different

18

Table 8. Analysis of Variance of Egg Production Data for Birds in
Group 2 for Periods 1, 3 and 5

 

 

Source of Degrees of Mean

variation freedom square F F .01
Treatments 2 1018.5 5.19 4.82
Strains 1 1118.0 5.70 6.90
Periods 2 148.0 .75 4.82
T x P 4 836.7 4.27" 3.51
T X S 2 517.5 2.64 4.82
P X S 2 542.5 2.77 4.82
T X P X S 4 165.2 .84 3.51
Error . ‘ 103 196.1

 

Since the T X P interaction was significant for this group, an analysis
of variance was conducted on data of all periods individually in order
to determine if treatments had a significant effect within each period.
The analysis of variance tables for individual periods are in the

Appendix.

19

Table 9. Analysis of Variance of Egg Production Data for Birds in
Group 3 for Periods 1, 3 and 5

 

 

 

Source of Degrees of Mean
variation ' freedom square F F .01
Treatments 5 264.0 1.47 3.11
Periods 2 1413.0 7.87 4.71
T X P 10 211.1 1.17 2.41
Error 243 179.4

Periods 5 1 3

Av eggs/hen 36.9 42.2 45.0

 

 

20

Table 10. Hematocrit Values for Birds in Group 1

 

 

 

Treatment - Period 1 Period 3 Period 5
1 28.4 28.5 26.7
2 28.1 29.7 26.8
3 28.4 29.4 26.8
4 29.4 28.6 26.7
5‘ 27.1 28.1 28.0
6 29.8 27.3 26.5
Strains:
A ' 27.2 27.6 25.4
B 29.0 28.8 27.4

 

21

Table 11. Analysis of Variance for Hematocrit Values for Birds in
Group 1 for Periods 1, 3 and 5

 

 

 

 

Source of I Degrees of Mean

variation freedom square F F .01
Treatments 5 3.50 .63 3.09
Periods 2 95.26 17.91 4.69
Strains 2 98.03 17.69 4.69
T X P 10 17.88 3.22 2.39
T X S . 10 4.03 .72 2.39
P X S 4 5.33 .96 3.39
T X P X S 20 6.37 1.15 1.95
Error 298 5.54

Periods 5 l 3

Hct. values ‘26;9 28.5 28.6

Strains A B C

Hct. values 26.9 ’28.5 28.6

 

 

Means not underscored by the same line are significantly different

(P (.01).

Since the T X P interaction was significant an analysis of variance

was conducted on data of all periods individually in order to determine
if treatments had a significant effect within each period. The
analysis of variance tables for the individual periods are in the

Appendix.

22

Table 12. Results of Grading of Eggs by a Commercial Grader

 

 

 

Number Percent
of of
Grade eggs total
A 317 88.2
B 23 a 6.3
C and Checks 12 3.3

Rejects (bloodspots) 8 2.2

 

23

Discussion of Experiment A:

 

For the control ration and treatments involving various levels of
vitamin B12 (15, 30 and 60 micrograms per pound in Treatments 2, 3 and
4, respectively), and cobalt at the lowest level (treatment 5) there was
a non-significant decrease in bloodspot incidence when expressed as
percent of all eggs laid (Appendix Table 1). There was a slight increase
in bloodspot incidence with the highest level of cobalt. When expressed
as average percent change on an individual bird basis, the lowest level
of vitamin 812 (treatment 2) showed a slight increase (Table 3). All
other treatments showed a slight decrease. This is in agreement with
the work of Bearse (1961), who found that including vitamin BIZ in the
diet of depleted birds or depleting birds of vitamin B12 had no in-
fluence on number of bloodspots.

Selenium at any of the levels fed was without influence on the
number of bloodspots (treatments 7 - 9). Birds fed the highest level
of selenium (treatment 10) dropped from 50 percent production in
Period 1 to 25 percent production in Period 2. Carlson g£_gl. (1954)
reported that sodium arsenite or arsanilic acid was partially effective
in counteracting the toxic effects of sodium selenite in a corn-soybean
type ration. Consequently, at the end of the first week in Period 3,
arsanilic acid (90 gms/ton) was added to diet 10. At the end of a
three-week period the birds showed no signs of coming back into pro-
duction and the experimental feed was discontinued.

Addition of fat or oil was without significant influence on
bloodspot incidence (treatments 11 - 16). There was a consistent de-

<2rease in bloodspots with treatments 12 through 16, but it is doubtful

if it can be attributed to the type of fat or oil in the diet since the

24

birds on the ration containing no added fat (treatment 16) also showed
a slight decrease. Pope (1959) obtained a non-significant decrease in
bloodspot incidence when crude tottonseed oil was included in the
ration.

For treatments 1 through 9, a greater percentage of the bloodspots
were of the larger size (over 4.8 mm) in Period 3 than in Period 1.
This was not true for treatments 11 through 16 (Appendix Table 5).

When the data were examined for each strain in Group 1 (Appendix Table
7), all strains were approximately equal in the tendency to produce a
greater percentage of bloodspots over 4.8 mm in Period 3 as compared
with Period 1.

The percentage production for Group 1 for Periods 1 and 3 for all-
birds laying in either Period is given in Table 5. Strain differences
were noted but dietary treatments were without significant influence on
production. It was noted, however, that the decline in production from
Period 1 to Period 3 was not as great for the treatments containing
vitamin 312.

In comparing treatments in Group 2, strain differences in egg
production (Appendix Table 8) were noted in Period 1; however, these
differences did not continue throughout the entire experiment. An
analysis of variance (Appendix Table 9) indicated that treatment effects
were significant during Period 3. Egg production during Period 3 for
the birds on the highest level of selenium (treatment 9) was signifi-
cantly depressed. There was a non-significant increase in production
for the birds on the control ration and the ration containing a low
level of selenium with the latter resulting in the greatest increase.

Effects of treatment on production were not significant (Table 9)

25

for the birds in Group 3. Production was significantly lower in Period
5 than in Period 1 and 3.

Table 10 contains the hematocrit values for birds in Group 1. An
analysis of variance was conducted on data of all periods individually.
It was noted that there were strain and treatment differences during
Period 1 (Appendix Table 11). Treatment differences were not to be
expected since all birds were on the basal ration during this period,
and treatments were actually replicates.

Treatment and strain differences were not significant at the
p.< .01 level-during Period 3 but were significant at the P'<;.05 level.
During Period 5 only strain differences were evident. The strain with
the highest incidence of bloodspots (Strain A) had the lowest hematocrits.
However, examination of the data did not indicate a correlation between
bloodspot incidence and hematocrit values.

Since the hematocrit values during Period 3 decreased, but not
significantly, as the vitamin 312 level was increased, it suggests that
continued use of high levels of vitamin B12 may cause a decrease in
erythrocyte formation.- Hematocrit values for birds on the high-level
of cobalt (treatment 6) were significantly depressed during Period 3
when Periods 1 and 3 were compared for treatment 6 alone. The low level
of cobalt (treatment 5) did not cause a significant increase in he-
matocrit values. Davis et a1. (1945) reported that continued admin-
istration of a "high” level of cobalt to ducks depressed erythropoietic
activity.

The fact that hematocrit values for all treatments except
treatment 5 were lower during Period 5 is not readily explainable.

Cohn and D'Amour (1951) reported that an increase in altitude stimulated

26

erythrOpoietic activity. For this reason, hourly station barometric
pressure values were obtained from the weather bureau during each
period on the days that blood was collected for hematocrit value de-
terminations. Barometric pressure values were not different between
periods.

Doupe g£_al. (1957) reported an increase in plasma volume in
humans in summer months and hematocrit values fluctuated less than
plasma volume.but were generally lower in summer than in winter.

In order to determine the size of bloodspots and percent of blood-
spots that are normally detected by a professional grader, a total of
360 eggs was taken to a commercial processing station. The grades ob-
tained are included in Table 12.

After the eggs had been graded, they were broken out and evaluated.
Twenty-five bloodspots (6.9%) were observed when broken out as opposed
to 8 (2.2%) observed by the grader. Two of the eggs rejected by the
grader because of bloodspots did not have bloodspots but had other
abnormalities that could be mistaken for bloodspots. Of the remaining
6 eggs rejected by the grader, 5 of the eggs had bloodspots over 4.8 mm

in size and 1 egg had a bloodSpot between 1.6 and 4.8 mm.

27

Experiment B: Procedure and Results

 

The effects of BAPN (beta-aminopropionitrile), vitamin K, sulfa-
quinoxaline, reserpine, soybean-corn oil blend, reserpine plus sulfa-'
quinoxaline, sulfaquinoxaline plus soybean-corn oil, vitamin K plus
sulfaquinoxaline, and alfalfa leaf meal plus animal fat were studied
with respect to bloodspot incidence in an experimental plan similar to
that in Experiment A. Periods l, 3 and 5 were 28 days in length while
Periods 2 and 4 were 14 days in length. Eggs were collected five days
each week.

There were 24 birds per treatment; 8 birds from each of the three
strains previously described. Bloodspot incidence was measured for 28
days and birds were then allotted to the experimental treatments so as
to balance initial bloodspot incidence.

Data from treatment 3 are not presented because the birds were rot
kept on the experimental diet the allotted time.

During Periods 4 and 5 the vitamin K level was increased from 4 to
40 mg/lb in treatment 5 and the sulfaquinoxaline from 0.0125 to 0.1
percent in treatments 6 and 11. A total of 17, 242 eggs were examined
in this experiment and the data is summarized in Tables 15 through 23
and Appendix Tables 14 through 23.

The calendar dates for this experiment were as follows:

Period 1 - January 9 to February 5
Period 2 - February 6 to February 19
Period 3 - February 20 to March 18

Period 4 - March 19 to April 2

Period 5 April 3 to April 29

Hematocrit values were determined in only one period of this

28

experiment because the birds began to decrease in production after being
handled for the collection of blood. Since egg production is essential
in order that bloodspot incidence can be determined, collection of

blood for hematocrit determinations was discontinued.

29

Table 13. Composition of the Basal Ration Fed in Experiment B

 

 

 

 

Ingredient Amount per th.

Corn yellow No. 2, ground 70.00 lbs.

Soybean oil meal, 44% protein 19.00 "

Meat and bone scrap, 50% protein 3.00 "

Fish meal with solubles, 60% protein 1.25 "

Dicalcium phosphate, feed grade 2.00 ”

Limestone, ground 5.00 "

Salt, iodized .25 ”

Manganese sulfate, 70% feed grade .025 "

Riboflavin suppl. (16 gm/lb) 1 gram

Vitamin A (10:000 IU/gm) 20 ”

Vitamin D (3,000 ICU/gm) 10 "

Total 100.00 lbs. +

CaIcII lEtEd—ahaIy-s- i-s- : _________________________
Crude protein % 17 0 Vitamin B12 (mcg.1b) 1.3
Crude fat % 3.1 Riboflavin (mg/1b) 1.1
Crude fiber % 3.0 Niacin (mg/lb) 10.0
Calcium % 2 8 Pantothenic acid (mg/lb) 2.8
Phosphorus % .8 Choline (mg/1b) 407
Vitamin D (ICU/lb) 300 Energy (Cal/1b) 945

Vitamin A (IU/lb) 3050

.oHumu Hue m CH

HHo Chou cam cmmHNOm pmcmeH mo mcoHn <

 

 

 

 

 

N

,. .HmmE wmmH mHHmHHm
Ho umm .HHo Ho mcoHUprm How mummcmano cu meme mama :Hoo Ho mowmucmoumm ecu cw mmwcmnu H
m.H nu nn nn nn nn nn nu nu nn nn unmoumm umw HmEHc<
m.H nn nu nn nn nn nn nn nn nu nn ucmona HmoE HmmH mHHmHH<
nn o.m o.m o.m nn nn nn nn nn nn nn osmonma NHHo cuooncmmczom
nn nn nu nn o.N o.N nn nu nn nu nn nH\wE mcHauwmmm
un mNHo. nu nn mNHo. nu mNHo. nn nn nu nn ucmouom mCHmeocHsvmesm
nn nu o.N uu un nn nn 0.6 o.N nu nn nu nH\wE M GHEmuH>
un nn nu nn nn nu nn nu m0. m0. nn ucmonwm zm<m
NH HH OH m m N o d m N H wuHGD usmempwcH

mumHn

m unmEHHmmxm uoH :oHumm Hmmmm :H mowcmno .qH mHan

31

Table 15. Change in Bloodspot Incidence Between Periods 1 and 3 When
Calculated on an Individual Bird Basis

 

 

Percentage
Average Percentage Percentage of birds Chi
percent of birds of birds showing square
Treatment change increasing decreasing no change values
1 + 6 57 A 19 24 3.14
2 - 5 38 38 24 1.10
3 -- __ _- -- -_
4 - l 45 45 ., 10 1.11
5 + l 50 45 5 .50
6 - 6 33 48 I 19 1.06
7 + 5 45 40 15 .32
8 + 8 59 35 6 1 73
9 - 4 48 33 19 19
10 + 1 48 30 22 3.45
11 0 48 38 14 .06
12 1+ 2 43 38 19 .19
Total 12.85

 

Chi-square value for 20 degrees of freedom (P<f.01) = 37.57

32

Table 16. Change in Bloodspot Incidence Between Periods 3 and 5
Calculated on an Individual Bird Basis

 

 

(Percentage

 

 

Average Percentage Percentage of birds Chi
percent of birds of birds showing square
Treatment change increasing decreasing no change values
1 - 5 26 53 21 1.57
2 + 2 43 38 19 .12
3 -- -- -- -- --
4 + 9 57 33 10 2.59
53 - 3 4o 55_ 5 3.33
6b 0 38 37 25 .62
7 + 1 37 42 21 17
8 - 7 29 53 18 l 03
9 + 6 52 29 19 1 65
10 + 7 50 32 18 1.03
11b - 1 32 58 10 2.34
12 + 8 35 35 30 2.17.
Total -I3762_
Chi-square value for 20 degrees of freedom (Pn(.01) = 37.57

Vitamin K increased to 40 mg/lb during Period 5

Sulfaquinoxaline increased to 0.1% during Period 5

33

 

 

 

 

 

Table 17. The Effect of Handling of Birds for Collection of Blood for
Hematocrit Determinations on the Incidence of Bloodspots
Three days before Three days after
No. of No. of No. of No. of
eggs bloodspots % eggs bloodspots %
Handled 61 13 21 64 12 19
Not handled 72 20 28 71 ' 15 21

 

Table 18.

 

 

 

 

The Effect of Handling Birds and Treating for Mites on
the Incidence of Bloodspots
Three days before Three days after
No. of No. of No. of No. of
eggs bloodspots % eggs bloodspots %
Handled 254 57 22 216 36 17
Not handled 245 56 23 242 62 26
Handled 246 50 20 246 52 21
Not handled 217 40 18 213 36 17

 

34

Table 19. Analysis of Variance of Prothrombin Times Obtained
During Period 5

 

 

 

Source of Degrees of Mean
variation freedom square F .01
Treatments 3 9.3 .42 5.95
Strain 2 10.0 .44 6.93
T X S 6 14.8 .65 4.82
Error 12 22.5

Average prothrombin times:

Treatment Seconds
1 28.8
5 31.7
6 31.1
11 31.1
Strains:
A 30.0
B 31.9

 

35

Table 20. Analysis of Variance of Hematocrit Values

 

 

Source of , Degrees of Mean
variation freedom square F F .01
Strains 2 304.37 32.34 4.71
Within 174 9.41
Strains: , A B C

25.1 27.9 29.6

 

Means not underscored by the same line are significantly
different (P (.01).

36

Table 21. Percentage Egg Production for Periods l and 3

 

 

Amount of

 

Treatment ' Period 1 Period 3 change
% % ,
1 73 70 - 3
2 71 62 - 9
3 -- -- --
4 , 72 70 - 2
5 1 69 61 - 8
6 72 62 - 10
7 74 56 p - 18
8 73 52 -.21
9 7O 63 - 7
1o 72 ' 65 - 7
ll 78 62 - 16

12 75 73 - 2

 

37

Table 22.: Percentage Egg Production for Periods 3 and 5

 

 

Amount of

 

Treatment Period 3 Period 5 change
1 7% 6T - 9
2 62 66 + 4
3 -- -- --
4 70 59 - 11
5 61 60 - l
6 62 50 - 12
7 56 65 + 9
8 52 60 + 8
9 63 64 + l

10 ' 65 66 + 1
ll 62 45 - 17

12 73 '67 - 6

 

 

38

Table 23. Analysis of Variance of Egg Production Data for Periods

 

 

 

l, 3 and 5

Source of Degrees of Mean

variation freedom square F F .01
Treatments 10 26.1 2.14 2.35
Periods 2 366.0 29.97 4.65
Strains 2 132.0 10.81 4.65
T X P 20 26.3 ‘ 2.15 1.91
T X S 20 18.3 1.50 1.91
P X S 4 26.7. 2 l9 3 35
T X S X P 40 16.2 11.33 1.63
Error 607 12.2

 

Since T X P interaction was significant, an analysis of variance
was conducted on data of all periods individually in order to
determine if treatments had a significant effect within each period.
Analysis of variance tables for individual periods are in the

Appendix.

 

39

Discussion of Experiment B

The various experimental diets were without significant effect on
the percentage of eggs containing bloodspots (Appendix Table 14) or on
the number of birds increasing or decreasing in the production of blood-
spot eggs (Table 15). Pope g£_al. (1961) also failed to affect bloodspot
incidence when vitamin K was included in the diet. Pope (1959),
however, obtained a non-significant decrease in bloodspots when crude
cottonseed oil was fed. In Experiment A, of the present study, the-
inclusion of crude cottonseed oil or a refined soybean-corn oil blend
produced a non-significant decrease in incidence of bloodspots but the
effect was not as great when repeated in Experiment B (treatment 9).

Between Periods 1 and 3, in general, Strain A showed a decrease
regardless of treatment, Strain B showed an increase, and Strain C was
inconsistent in bloodspot incidence (Appendix Table 32). Between
Periods 3 and 5 no consistent tendency of strains to increase or decrease
was apparent (Appendix Table 17).

In comparing Periods l and 3, or 3 and 5, the percentage of
bloodspots falling in each size category (Appendix Table 18) was
approximately the same for each period.

During Periods 1 and 3, a slightly higher percentage of the eggs
from birds laying at a rate of 50 percent or more contained bloodspots
than the eggs from birds laying at a rate less than 50 percent. The
percent bloodspots for the less than 50 percent group declined slightly
from Periods l to 5. This can be explained on the basis that in Period
5 a smaller percentage of the eggs were from the birds with high blood-
spot incidence (Strain A).

From Tables 17 and 18, it can be seen that handling birds for

 

40

collection of blood or for treatment of mites had no effect on the in-
cidence of bloodspots within a three-day period following handling.
Frightening hens at different hours throughout the day (Jeffrey and
Pino, 1943) or trapnesting (Stiles and Dawson, 1959) had no influence
on the incidence of bloodspots.

During Period 5, prothrombin times, determined for six birds from
each of four treatments (1, 5, 6 and 11), were not different due to
strain or treatments (Table 19).

Hematocrit differences between strains were significantly different

(Table 20).

Significant strain differences in egg production were noted during
Period 1 (Appendix Table 20). During Period 1, treatments were actually
replicates since all birds were on the basal ration. No differences
between treatments existed.

There were differences in egg production due to strain and treat-
ment during Period 3 (Appendix Table 21). Birds on treatment 8, which
contained sulfaquinoxaline and reserpine, produced significantly less
eggs than birds on the control ration. Birds on treatment 12, which
contained dehydrated alfalfa meal and animal fat, produced significantly
more eggs than those birds on treatments 8 or 7 (reserpine alone).

During Period 5, egg production for the birds on the soybean-corn
oil blend plus sulfaquinoxaline (treatment 11) was lower than egg
production for the birds on the control ration but not significantly so
(Appendix Table 22). Egg production for treatments 7, 9, 10, 2 and 12
was significantly higher than treatment 11. This was not true for the

birds on sulfaquinoxaline alone (treatment 6).

 

GENERAL DISCUSSION

As previously mentioned, the incidence of bloodspots in chicken eggs
is influenced considerably by heredity and much progress has been made in
overcoming this problem by the selection of proper breeding stock.
Nevertheless, thus far, genetics has not completely solved this problem
because a small percentage of the eggs being marketed today still contain
this defect. .This indicates that other factors may also be involved.

How should the effect of treatment on incidence of bloodspots be
measured? Numerous workers in the past have used the technique of com-
paring the pre-experimental period with the experimental period but
generally on,a group or pen basis. By maintaining birds in individual
cages it was possible to observe the effect of treatment on the individual
bird and determine the number of birds that were influenced in a like
manner. When group comparisons are made with respect to bloodspot in-
cidence, the change in incidence between pre-experimental and ex-
perimental period may be caused by one or a few birds -- but the change
is attributed to the entire group. Thus, it is possible for a few
individuals to bias the data. In addition, when individual comparisons
are made, this automatically takes into consideration only those birds
that lay in both the pre-experimental and experimental period, thus
eliminating the influence of any particular bird that lays in only one
period.

Bearse (1958) reported an increase in bloodspot incidence when
reserpine was included in the diet. He based his conclusions on a
comparison of the birds on the reserpine-supplemented diet with that of
the birds on the control ration with respect to change in incidence for

both groups between the pre-experimental and experimental period. For

41

42

example, the birds on the reserpine-supplemented diet showed an increase
of two percent while the birds on the control ration showed no change.
(Presumably this was on a group basis).

If using this type of comparison as the sole criterion for eval-
.uation is valid, then reserpine in this study had a beneficial effect
because the birds fed the reserpine did not show as great an increase in
bloodspot incidence as did those fed the control ration (when pre-experi-
mental and experimental periods were compared). 0n the basis of the
same type comparison, BAPN and sulfaquinoxaline were even more effective
in reducing the incidence of bloodspots. Admittedly, experimental
rations must be compared with the control rations but not without also
comparing the pre-experimental period with the experimental period for
any particular ration and determining if the difference between periods
is significant. It would appear that the most valid test for determining
if the difference between periods is significant is to determine the
number of birds influenced in a similar manner.

Another type of comparison that could be made would be to adjust the
values for the control ration to zero and establish comparsions of the
experimental rations with the control ration on this basis. For example,
birds in Experiment A on the control ration showed an average increase
in bloodspot incidence of 6 percent while the birds on treatment 2
showed an average decrease of 3 percent, or a difference of 9 percent
when treatment 2 is compared with the control ration. This type of
comparison, however, has the disadvantage in that it does not take into
consideration the fact that if the pre-experimental period is not com-
pared with the experimental period for any particular ration than those

birds do not act as their own controls.

 

43

.Examination of the data for individual birds in this study indicated
that the birds could be divided into three types on the basis of blood-
spot incidence. The first type laid eggs that contained no bloodspots;
the second type produced eggs infrequently that contained bloodspots
and the third type produced eggs that contained a high percentage of
bloodspots. By determining bloodspot incidence at stated intervals for-
a short period of time, it is conceivable, that at the first "break-
out", a portion of the eggs examined would be from hens of the third
type while at the second "break-out", not any of the eggs would be
from birds of this type. .Consequently, it would be erroneous to draw
the conclusionnthat treatment caused a decrease in bloodspot incidence
on the basis of such experimental procedure. An example of determining
bloodspot incidence at stated intervals is the worh of Berruti and
Dedrick (1960). Bloodspot incidence was determined at 2- and 4-week
intervals after initiation of experimental diets.

Since vitamin K is necessary for prothrombin formation which in
turn is directly involved in blood clotting, much attention has been
given to vitamin K as being a factor in reducing the incidence of_
bloodspots. Blood clotting does not normally occur until the blood has
been released from the blood vessel. Consequently, it would appear
that any influence of vitamin K wodld be on size, rather than on
number, of bloodspots. Nalbandov and Card (1944), Denton (1947) and
Pope et al. (1961) found no effect on number of bloodspots produced
when vitamin K was included in the diet.

Why, then, study the influence of vitamin K on bloodspot incidence
when numerous reports have indicated no effect? Day (1960) reported

that vitamin K (menadione sodium bisulfite) was not stable when present

44

in a calcite carrier and Pope and others used vitamin K in such a
carrier. In addition, Berruti and Dedrick reported that by feeding a
more stable vitamin K they were able to cause a considerable reduction
in the incidence of bloodspots.

Since sulfaquinoxaline had been suggested by Finkel (1961) to be
an antagonist of\vitamin K, it was included in this study to accentuate
the vitamin K-deficient effect, if any.

The findings of this study, which do not agree with those of
Berruti and Dedrick, are supported by the fact that sulfaquinoxaline
had no influence on bloodspot incidence when included in a vitamin
K-low diet. Thus, it is evident that vitamin K per se is not the
solution to the bloodspot problem.

The fact that sulfaquinoxaline did not have any influence on pro~
thrombin times is probably due to the low level used. Cuckler and Ott
(1955) reported that at least 0.4 percent was required to produce an
increase in prothrombin time. The use of higher levels by this author
resulted in cessation of egg production. Consequently, a correlation
between prothrombin time and bloodspot incidence could not be determined.

Does hemorrhaging other than that caused by a vitamin K deficiency
occur, and, if so, what would be the effect of this type of hemorrhaging
on the incidence of bloodspots? Waibel and Pomeroy (1958) reported that
IMXPN produced ”dissecting aneurysm of the posterior aorta and consequent
death due to internal hemorrhage“ in growing turkeys. Barnett ££_al.

613958) reported that supplemental vitamin K did not consistently reduce
BAFQV-induced hemorrhaging in chickens, indicating that the BAPN effect
is riot mediated by causing a vitamin K deficiency.

Since BAPN did not affect the number or size of bloodspots produced

1

45

it is possihle that the effect of BAPN on the small veins and capillaries
of theovary differs from the effect on larger vessels.

High levels of vitamin 812 had no influence on the incidence of
bloodspots. Pope (1959) obtained a significant increase in bloodspot
incidence by feeding high levels of vitamin B12 (30 mcgs/lb). However,
he also noted that the feeding of vitamin A at levels of 3,575 or 10,215
USP units/lb counteracted the detrimental effects of this high level
of vitamin B12. The vitamin A level of the basal ration used in the
work discussed here was 3,563 units/lb which may explain why the use of
higher levels of vitamin B12 than Pope fed did not increase bloodspot
incidence.

Hematocrit values for the birds on the high levels of vitamin B12
were normal which indicated that a condition of erythrocytosis did not
exist. Yet, the literature states that erythrocytosis can be induced
by the administration of "excessive” doses of vitamin B12 or other
cobalt compounds with a resultant increase in peripheral resistance which
may in turn lead to hypertension (Kugelmass, 1959). Weiss (1958) re-
ported that incidence of bloodspotting was higher in a high blood
pressure line of birds than in a low blood pressure line, but not sig-
nificantly so.

If hypertension is associated with bloodspot incidence as suggested
by Weiss, would the feeding of vegetable oils and animal fats result in
different effects on the number of bloodspots produced? Daghir gp_al.
(:1960) reported that soybean oil caused a significant decrease in serum

efliolesterol levels while white grease was without effect. Since the
ctlaracteristic lesion of the atherosclerotic artery may contain up to

“3 percent cholesterol, it is believed that hypercholesterolemia and

46

atherosclerosis are associated in some way (Best and Taylor, 1961).

In humans, a narrowing of the peripheral vessels, such as would
happen when a sclerotic lesion occurs in an artery, can result in hyper-
tension. Although narrowing of the larger peripheral vessels may result
in hypertension, the pressure in the small veins and capillaries does not
change materially. Nalbandov and Card (1944) suggested that bloodspots
may be due to rupture of small blood vessels or capillaries. However,
it does not appear that the rupture would be the result of increased
blood pressure resulting from a condition such as atherosclerosis.

Thus, influencinglblood cholesterol would not be expected to influence

the number of bloodspots produced. In addition, the feeding of reserpine,
an anti-hypertensive agent, had no influence on number or size of
bloodspots produced.

There is a positive correlation between whole blood specific
gravity and hematocrit values (Kugelmass, 1959), therefore, any dis-
turbance that would affect the “corpusclezplasma volume ratio" would
influence hematocrit values. For example, after hemorrhage there is a
fall in the corpusclezplasma volume ratio due to an increase in the
return of fluid from the tissue spaces. The blood loss from hens with
high bloodspot incidence was possibly greater than the loss from hens
iflith low bloodspot incidence but it is doubtful if this loss was
enough to affect the corpuscle:plasma volume ratio, and consequently,
toould not explain why hematocrit values were lowest for the birds with
Elie highest incidence of bloodspots.

Acute disease increases whole blood specific gravity, however,
mC>rtality was highest for the strain of birds with the lowest hematocrit

Values.

47

That the high level of cobalt appeared to depress erythrocyte
formation as evidenced by significantly lower hematocrit values, is in
agreement with Davis E£_al.(l945) who stated that continued administration
of "excessive” amounts of cobalt decreased the erythrocyte count in
ducks.

The influence of Vitamin B12 on egg production suggests re-
evaluation of the vitamin B12 requirement of the laying hen for
optimum egg production. The levels used were considerably higher than
the present recommendations of the National Research Council.

As evidenced by a depression in egg production, the higher levels
of selenium were toxic. On the other hand, the addition of a low level
of selenium appeared to cause an increase in production. This suggests
further study on the selenium requirement of laying hens. Poley et a1.
(1940) reported that better growth was obtained in chicks when 2 or 5

ppm of selenium was supplied in selenium-containing grains.

CONCLUSIONS
Two experiments were conducted using 600 Single Comb White Leghorn
pullets from three different strains to determine the effect of certain
(dietary ingredients on the incidence of bloodspots in chicken eggs.
I3loodspot incidence or size of bloodspots of individual birds before,
(diaring and after feeding the experimental rations was the criteria used
CC) evaluate the dietary treatments employed..
The following dietary additives did not affect the number or size
0 f bloodspots .
Vitamin B12 (15, 30 and 60 meg/lb)
Cobalt, as CoCO3 (1.32 and 90 mcg/lb)
Selenium as Na2303-5H20 (1 and 5 ppm)
Crude cottonseed oil (5.0%)
Crude corn oil (5.0%)
Refined soybean-corn oil in a 4:1 ratio (5.0%)
Fat, animal, No. 2 yellow grease (none or 5.0%)
Beta-aminopropionitrile fumarate (BAPN) (0.03%)
Vitamin K as menadione sodium bisulfite (2,4 and 40 mg/lb)
Sulfaquinoxaline (0.0125% and 0.1%)
Reserpine (2 mg/lb)
Reserpine plus sulfaquinoxaline (2 mg/lb plus 0.0125%)
Refined soybean-corn oil plus vitamin K (5.0% plus 2 mg/lb)

Refined soybean-corn oil plus sulfaquinoxaline (5.0% plus
0.0125 or 0.1%)

Alfalfa leaf meal plus animal fat (1.3% plus 1.5%)
Subtraction from the diet of all the above had no effect on
inciidence of bloodspots.
Thus the hypothesis that the presence or absence of certain

48

49

dietary ingredients might influence the incidence of bloodspots in
chicken eggs was not confirmed.
Egg production was significantly depressed by:
Selenium (5 ppm)
Reserpine (2mg/1b) plus sulfaquinoxaline (0.0125%)

Sulfaquinoxaline (0.1%) plus refined soybean-corn oil
blend (5.0%)

Egg production was depressed, but not significantly, by:
Beta-aminopropionitrile fumarate (0.03%)
Reserpine (2 mg/lb)
Sulfaquinoxaline (0.1%)

Egg production was increased, but not significantly, by:
Vitamin B12 (60 mcg/lb)
Selenium (1 ppm)

Hematocrit values were significantly different between strains of
E>isrds with Strain A being lower than Strains B and C in Experiment A.
ilri Experiment B, all strains were significantly different from each
C>tlner (ranking low to high: Strain A, Strain B and Strain C).

In Experiment A, hematocrit values for July were significantly lower
Flian for March and May. The hematocrit values for all treatments were
lcrwer during July, except for the treatment that had contained the low

level of cobalt during May.

50

LITERATURE CITED
.Anderson, C. C-, J- H. Hare, J. K. Bletner, C. E.-Weakley, Jr. and J. A.
Mason, 1954. A hemorrhagic condition in chicks fed simplified
rations. Poultry Sci. 33: 120-126.

ISalloun, S. L. and E. L. Johnson, 1952. Underheated soybean oil meal
increases blood clotting time of chicks. Poultry Sci. 31:905 (Abs.)

I3airnett, B. D., 1960. The effect of reserpine on artificially produced
and spontaneously appearing aortic rupture. The Second Conference
on the use of reserpine in poultry production. pp. 9-14.

ISairnett, B. D. and C. L..Morgan, 1959. The effect of high levels of
dietary fat on beta-aminopropionitrile induced internal hemorrhage
in chicks. Poultry Sci. 38:589-593.

Isaarnett, B. D., D. J. Richey and C. L. Morgan, 1958. The effect of anti-
coagulants on toxicity of beta-aminopropionitrile. Poultry Sci.37:
1124-1128.

IBeearse, G. E., 1958. How nutrition and management affect bloodspots in
eggs. 11th Washington State College Animal Industries Conference
Abstracts.

ISeaarse, G. E. and L. R. Berg, 1958. The fat soluble vitamins and bloodspot
incidence. Poultry Sci. 37:1184 (Abs.)

Bearse, G. E., C. F. McClary and H. C. Saxena, 1953. Bloodspot incidence
and the vitamin A level of the diet. Poultry Sci. ‘32:888 (Abs.)

‘B‘Eérrse, G. E., C. F. McClary and H. C. Saxena, 1960. Bloodspot incidence
in chicken eggs and vitamin A level of the diet. Poultry Sci. 39:
860-865.

BGEITruti, R. and G. T. Dedrick, 1960. Evidence on the value of stabi-

lized Heterogen K in reducing the incidence of bloodspots. Hetero-

51

chemical Corp. Unpub.
Best, C. H. and N. B. Taylor, 1961. The Physiological Basis of Medical
Practice, 7th ed. The Williams and Wilkins Co., Baltimore, Md.

(33rd, L. E. and A. Nalbandov, 1944. Controlling blood and meat spots.
Poultry Sci. 23:551 (Abs.)

(Sairlson, C. W., E. Guenthner, W. Kohlmeyer and O. E. Olson, 1954.

Some effects of selenium, arsenicals and vitamin 312 on chick
growth. Poultry Sci. 33:768-774.

(Zairver, J. S. and W. Henderson, 1941. The effect Of rutin and ascorbic
acid and of alfalfa on blood and meat spots in hens' eggs. Poultry
Sci. 27:656 (Abs.)

Chahn, E. W. and F. E. D'Amour, 1951. Effects of high altitudes on
growth and polycythemia in rats. Am. J. of Physiol. 161:394-399.

(Zuckler, A. C. and W. H. Ott, 1955. Tolerance studies on sulfaquinbx-
aline in poultry. Poultry Sci. 34:867-879.

Imaghir, N. J., W. W. Marion and S. L. Balloun, 1960. Influence of dietary
fat and choline on serum and egg yolk cholesterol in the laying
chicken. Poultry Sci. 39:1459tl466.

IDEIVIS, J. A., A. W. McCullough and R. H. Rigdon, 1945. Polycythemia
produced by cobalt in the duck. J. of Lab. and Clin. Med. 30:327-
336.

IDEty, E. J. and B. Glick, 1960. Chemical and biological assays of
vitamin K premixes. Presented at the Informal Poultry Nutrition
Conference in Chicago.

I3en1ton, C. A., 1947. Observations on the incidence and characteristics
of blood and meat spots in hens' eggs. Poultry Sci. 26:272-276.

thbupe, J., M. H. Ferguson, and J. A. Hildes, 1957. Seasonal fluctuations

52

in blood volume. Can. J. Biochem. and Physiol. 35:203-213.
jDuncan, D. B., 1955. Multiple range and multiple F tests. Biometrics
11:1-42.
Fearnsworth, G. M. and A. W. Nordskog, 1955. Estimates of genetic param-
’ eters influencing blood spots and other economic traits of the
fowl. Poultry Sci. 34:1192 (Abs.)

Ffiirikel, M. J., 1961. Vitamin K1 and the vitamin K analogues. Clin.
Pharmacol. and Therap. 2:794-814.

Ffirxast, D. V., H. S. Perdue and H. C. Spruth, 1956.

Vitamin K activity of menadione sodium bisulfite in chickens. J.
Nutrition 59:181-196.

Pierre, J. H., G. C. Anderson, C. E. Weakley, Jr. and J. K. Bletner, 1953.
Factors contributing to a hemorrhagic condition in experimental
chicks fed simplified rations. Poultry Sci. 32:904 (Abs.)

lizarms, R. H. and F. R. Tarver, Jr., 1957. The influence of dicumarol
upon blood clotting time and blood loss of young chickens.

Poultry Sci. 36:76-79.

11c>ussay, B. A., 1955. Human Physiology, 2nd ed. McGraw-Hill Book Co.,
Inc., New York.

£T€2ffrey, F. P. and J. Pino, 1943. The effects of heredity and of certain
environmental factors on the incidence of bloodspots in chicken eggs.
Poultry Sci. 22:230-234.

JOnes, A. R., 1956. A device for rapidly deriving the hematocrit of
blood centrifuged in ungraduated tubes. New England Jour. of Med.
254:172-174.

Kl~1gelmass, N. I., 1959. Biochemistry of Blood in Health and Disease.

Chas. C. Thomas Co., Springfield, Ill.

53

Lerner, M. I., L. W. Taylor and D. C. Lowry, 1951. Selection for in-
creased incidence of bloodspots in White Leghorns. Poultry Sci.
30:748-757.

bicGovern, J. J., A. R. Jones and A. G. Steinberg, 1955. The hematocrit
of capillary blood. New England Jour. Of Med. 253:308-312.

ldiinro, S. S., 1952. Effect of different feeding formulas on blood-
spotting. Poultry Sci. 31:929 (Abs.)

lQealbandov, A. V. and L. E. Card, 1944. The problem of blood clots and
meat spots in chicken eggs. Poultry Sci. 23:170-180. .

PQEIIbandov, A. V. and L. E. Card, 1947. The problem of blood and meat
spots in chicken eggs. II. Its importance in poultry flocks, and
a study of the nutritional factors involved. Poultry Sci. 26:400-
409.

I?€2rdue, H. 8., J. A. Kolar and D. V. Frost, 1961. Non-correlation of

I vitamin K status and production of eggs with bloodspots. Poultry

Sci. 40:1443 (Abs.)

I?c>ley, W. E., wfo. Wilsbn and A. L. Moxon, 1940. The effect of selenized
grains on the rate of growth of chicks.7 Poultry Sci. 19:358 (Abs.)

I?cxpe, C. W., 1959. The effect of certain nutrients and other materials
upon the incidence of bloodspots in chicken eggs. Ph. D. Thesis.
Michigan State University.

EVDIJe, C. W., P. J. Schaible and L. E. Dawson, 1961. Effects of certain
nutrients upon bloodspots in chicken eggs. Poultry Sci. 40:377-
382.

CQKIiJnn, J. P. and A. B. Godfrey, 1940. Inheritance and variation of blood
spots in chicken eggs. Poultry Sci. 19:359 (Abs.)

jROY, D. B. and H. R. Bird, 1959. Stimulation of chick growth by proline.

 

 

I i I'll ‘58.! q. I 1

54

Poultry Sci. 38:192-196.
ESauter, E. A., W. J. Stadelman and J. S. Carver, 1952. Factors

affecting the incidence of bloodspots and their detection in hens'
eggs. Poultry Sci. 31:1042-1049.

Sr1eudecor, G. W., 1956. Statistical Methods. 5th ed. Iowa State
College Press, Ames, Iowa.

Sraeeckmann, E. W. and R. K. Ringer, 1961. Hemodynamic responses following
reserpine feeding to turkeys. Poultry Sci. 40:1292-1298.

St:eephens, J. F. and R. L. Tugwell, 1960. Sources and levels of vitamin
K in relation to cecal coccidiosis. Poultry Sci. 39:1183-1187.

EStZiles, P. G. and L. E. Dawson, 1959. The effect of physical disturb-
ance, sound and light on the incidence of blood and meat spots and
other egg quality factors. Poultry Sci. 38:1250 (Abs.)

$5t:iles, P. G., R. K. Ringer and L. F. Wolterink, 1958. A procedure for
labeling bloodspots in chicken eggs with radioactive phosphorus.
Poultry SCi. 37:600-601.

ESt;urkie, P. D., 1954. Avian Physiology. Comstock Publishing Associates,
Ithaca, New York.

53t:urkie, P. D., 1959. Cardiovascular effects of reserpine on the fowl.
Conference on the use of the tranquilizing and antihypertensive agent
Serpasil in animal and poultry production. pp. 18-20.

Titus, H. W., 1955. The Scientific Feeding of Chickens. 3rd. ed. The
Interstate, Danville, Ill.

TI‘avis, H. F., 1960. Nutritional studies on ranch-raised mink. Ph. D.
Thesis, Michigan State University.

‘qéiibel, P. E. and B. S. Pomeroy, 1958. Studies on the production of

aortic hemorrhage in growing turkeys with beta-aminopropionitrile.

55

Poultry Sci. 37:934-938.

‘Weiss, H. S., 1958. Blood pressure and egg formation. Poultry Sci. 37:
33-36. ‘

Tfacowitz, H., R.D. Carter and E. Ross, 1955. Further studies on

hemorrhagic syndrome induced by feeding high levels of sulfaquinox-

aline to chicks. Poultry Sci. 34:1229 (Abs.)

56

Appendix Table 1. Bloodspot Incidence Expressed as the Percentage
of Eggs Laid During Periods 1 and 3

 

 

 

Period Period AmOunt "t"
Treatment 1 3 of change values
% % Z

1 . 21 16 - 5 .42
2 20 19 - l .08.
3 24 17 - 7 ' .52
4 23 18 - 5 .39
5 32 23 - 9 .67
6 ; 19 21 + 2 .16
7 7 10 + 3 30
8 6 4 - 2 22
9 8 9 + l 09
11 8 10 + 2 18
12 11 7 - 4 ll
13 8 5 - 3 34
-14 12 9 - 3 .27
15 11 8 - 3 .30
16 6 4 - 2 23

57

Appendix Table 2. Bloodspot Incidence Expressed as the Percentage of
Eggs Laid During Periods 3 and 5

 

 

Period Period Amount . "t"
Treatment . 3 5 of change values
Z Z Z
l 16 17 + 1 .08
2 17 21 + 4 .29
3 17 16 - l .08
4 14 21 + 7 .52
5 21 24 + 3 .21
6 20 25 + 5 36
7 10 6 - 4 37
8 4 3 - l 13
9 9 7 - 2 l6
3 11 5 4 - 1 12
12 8 6 - 2 19
13 5 3 - 2 .28
14 9 8 - 1 . .10
15 8 9 + 1 .10

16 4 6 - 2 .23

58

Akrapendix Table 3. Percentage of Bloodspots by Strains Within Treatments
for Periods l and 3 for Birds in Group 1

 

 

 

Period Period Amount
'Ifireatment Strain 1 3 of change

Z Z Z

l A 40 32 - 8

‘ B 12 10 - 2 ‘

C 12 4 - 8

2 A 39 39 0
B 18 20 + 2

C ‘ 10 6 - 4

3 A 61 51 -10
B 10 5 - 5

C 8 5 - 3

4 A 48 44 - 4
B 17 15 - 2

C 8 5 - 3

5 A 66 53 -13
B 23 20 - 3

C 8 6 - 2

6 A 52 50 - 2
B 15 20 + 5

C 7 l - 6

 

Appendix Table 4. Percentage of Bloodspots by Strains Within Treatments
for Periods 3 and 5 for Birds in Group 1

 

 

 

Treatment Strain Period Period Amount
3 5 of change
Z Z Z
l A 33 34 + 2
B 10 21 +11
C 4 6 + 2
2 A 37 52 +15
B 21 20 - l
C 6 10 + 4
3 A 54 52 - 2
B 5 5 7 + 2
C 5 11 + 6
4 A 43 71 +28
B 15 22 + 7
C 5 9 + 4
5 A 52 66 +14
B 22 15 - 7
C 6 13 + 7
6 A 50 54 + 4
B 19 23 + 4
C 1 4 + 3

60

Appendix Table 5. Percentage of BloodSpots Falling Into Each Size
Category During Periods l and 3

 

 

 

 

 

 

Period 1 Period 3
Size ___ Size
Under Between Over Under Between Over
Treatment 1.6 mm 1.6-4.8 mm 4.8 mm 1.6 mm 1.6-4.8 mm 4.8 mm
Z Z Z Z Z Z
1 49 22 29 31 26 43
2 56 17 27 31 21 48
3 43 19 38 27 18 55
4 45 22 33 38 26 36
5 37 21 42 37 16 47
6 35 29 36 29 22 49
7 56 21 23 50 25 25
8 78 15 7 63 4 33
9 66 17 17 46 18 36
11 68 16 16 58 14 28
12 64 14 22 64 14 22
13 61 17 22 56 25 19
14 58 17 25 54 17 29
15 65 21 14 60 ’ 21 19

16 65 18 17 70 9 21

 

61

Appendix Table 6. Percentage of Bloodspots Falling Into Each Size
Category During Periods 3 and 5

 

 

 

 

 

Perigd 3 Period 5
___ __.i_‘#_§ize __ Size
Under Between Over Under Between Over

Treatment 1.6 mm 1.6-4.8 mm 4.8 mm 1.6 mm 1.6-4.8 mm 4.8 mm

A— _4_ ‘4flflw‘

 

% z 2 2 2 2

1 31 26 43 4o 23 37

2 31 22 h 48 37 19 44

3 27 18 55 33 26 41

4 38 26 36 36 20 44

5 37 16 47 39 20 41

6 29 23 49 36 18 46

7 5o 25 25 47 22 31

8 63 4 33 38 48 14

9 46 18 36 41 32 27

11 58 14 28 50 14 36
12 64 14 22 74 22 4
13 56 25 19 41 14 45
14 54 17 29 39 18 43
15 6O 21 19 55 19 26

16 70 9 21 50 22 28

 

62

>pwm5de Hmva u. wmenmsnmmm om wHOOQmwOnm meHHsm Hsno men: mHNm omnmmoaw 6% mnHmHsm SHnrH:
Hummnamsnm UCHHDN meHomm H mad w mom wHHam Hp onocm H

 

 

 

 

 

meHOQ H ill wmnHoa w
20. 0m comma wmnSmm: o<mn 20. em canon wmnamm: o<mn
Hammnamsn mnHmH: mv0nm H.o Ba H.onb.m Ba b.m 38 vanm H.@ as H.oub.m Ba b.m as

N N N N N N

H > How mo Hm wN No No No we
w Nm no No Nu Nb No no , Nm

n we be No Nm 0 km NN Nu

N > 8 . as No Nd 8 Na 8 E
N am bb Ho Nu mm uN Nm pm

a Nm mN N HH Hm bu HN so

u > HHN Nu Hm bm mo NH Hm 0H
w No mo 0 No HH mm e we

0 Nu bu we HN Hm so Nw NH

b >. HHN Nb NH _ em NO NO No DH
w, by mu No HN mN wm Nu um

o NN mN He 9 Hm mo N Hw

m > Now Na Ho mu Hoo No H» mu
m oN mu NN HH mH bH Nu NN

n Nm mu H» No NN ob Hb Nu

o > me No No es mo Nb Ho mu
w em , no Nu ww mw No No up

0 Ho mu NH Ho w Nu 0 mm

 

 

 

 

63

Appendix Table 8. Analysis of Variance of Egg Production Data for

Birds in Group 2 for Period 1

 

 

 

Source of Degrees of Mean

variation freedom square .01
Treatments I 2 68.0 .39 5.20
Strains 1 1633.0 9.24 7.33
T X S 2 52.0 .29 5.20
Error 39 176.0

Strains B C

Av. eggs/ hen 30.1 42.2

 

Appendix Table 9.

64

Analysis of Variance of Egg Production Data for
Birds in Group 2 for Period 3

 

Source of Degrees of Mean

variation freedom square F F .01
Treatments 2 2368.0 11.91 5.29
Strains 2 516.0 2.58 7.44
T X S 2 105.5 .53 5.29
Error 34 199.4

Treatments 9 7 8

Av. eggs/hen

24.5 44.2 50.7

 

 

 

Means not underscored by the same line are significantly different

(P < .01).

 

65

Appendix Table 10. Analysis of Variance of Egg Production Data for
Birds in Group 2 for Period 5

 

 

Source of Degrees of Mean

variation freedom square F F .01
Treatments 2 291.5 1.34 5.39
Strains 1 54.0 .25 7.56
T X S ’ 2 655.0 3.02 5.39

Error 30 216.9

 

 

Appendix Table 11.

66

Analysis of Variance for Hematocrit Values for

Birds in Group 1 for Period 1

 

 

 

 

 

Source of Degrees of Mean

variation freedom square F F 01
Treatments 5 21.44 3.42 3.20
Strains 2 44.06 7.05 4.82
T X S 10 3.89 .62 2.51
Error 109 6.25

Treatments 1 3 4 6
Hct. values 27.1 28.1 28.4 28.4 29.4 29.8
Strains C

Hct. values 27 2 29.0 29.1

 

 

Means not underscored by the same line are significantly different

(p<.01)

 

 

Illlltmllllllllnlnlitll'luilll’llllill

 

67

Appendix Table 12. Analysis of Variance for Hematocrit Values for

Birds in Group 1 for Period 3

 

 

 

Source of Degrees of Mean

variation freedom square F F .05
Treatments 5 13.91 2.59 2.30
Strains 2 25.79 4.79 3.09
T X S 10 5.53 1.02 1.92
Error 105 5.38

Treatments 6 5 l 4 3 2

Hct. values

Strains

Hct. values

27.3 28.1 28.5 28.6 29.4 29.7

 

 

27.6 28.8 29.1

 

 

Means not underscored by the same line are significantly different

(P< .05) .

68

Appendix Table 13. Analysis of Variance for Hematocrit Values for
Birds in Group 1 for Period 5

 

 

 

Source of Degrees of Mean

variation freedom square F F .01
Treatments . 5 4.95 1.02 3.25
Strains 2 39.11 8.11 4.88
T X S 10 6.79 1.41 2.55
Error 84 4.82

Strains A B C

Hct. values , 25.4 27.4 27.5

 

 

Means not underscored by the same line are significantly
different (P < .01) .

 

69

Appendix Table 14. Bloodspot Incidence Expressed as the Percentage
of Eggs Laid During Periods l and 3

 

 

 

Period Period Amount ”t"
Treatment 1 3 of change values
Z Z Z -
1 l3 19 + 6 .54
2 29 24 ' - 5 .37
3 -- -- -- --
4 26 25 - l .08
5 22 20 - 2 .16
6 24 18 - 6 .48
7 19 20 + l .08
8 20 25 + 5 .35
9 14 13 - 1 10
10 17 ‘19 + 2 .17
ll 14 15 + l .09

12 21 22 + 1 _ .08

 

70

Appendix Table 15. Bloodspot Incidence Expressed as the Percentage

of Eggs Laid During Periods 3 and 5

 

 

 

Period Period Amount ”t"
Treatment '3 5 of change values
Z Z 2
1 17 13 - 4 .35
2 24 26 + 2 .15
3 -- __ -_ --
4 24 28 + 4 .30
5a 21 19 - 2 .17
6b 19 21 + 2 .14
7 17 23 + 4 .31
8 25 19 - 6 .43
9 14 17 + 3 .29
10 19 24 + 5 .40
11b 15 9 - 6 .57
12 22 26 .30

 

a Vitamin K increased to 40 mg/lb during Period 5

b

Sulfaquinoxaline increased to 0.1% during Period 5

 

ll. infill III {III II in
.I-II' 8
II! II!!! I

71

Appendix Table 16. Percentage of Bloodspots by Strains Within
Treatments for Periods l and 3

 

 

 

Period Period Amount of
Treatment Strain l 3 change

Z Z Z
l A 27 26 - l
B 7 15 + 8
C 8 16 + 8
2 A 54 39 - 13
B 31 34 + 3
C 9 5 — 4
4 A 55 43 - 12
B 19 20 + 1
C l4 17 + 3
5 A 36 35 - 1
B 13 19 + 6
C 16 13 - 3
6 A 48 36 - 12
B 18 17 - 1
C 9 6 - 3
7 A 36 33 - 3
B 7 18 + 11
C l3 l3 0
8 A 51 39 - 12
B 11 32 - + 21
C 6 12 + 6
9 A 31 30 - 1
B 4 11 + 7
C 9 2 - 7
10 A 36 37 + 1
‘ B 12 9 - 3
C 5 12 + 7
11 A 33 21 - 12
B 7 13 + 6
C 6 12 + 6
12 A 39 29 - 10
B 16 25 + 9
C 12 12 0

 

 

litiulll'll’lrilll

72

Appendix Table 17. Percentage of Bloodspots by Strains Within
Treatments for Periods 3 and 5

 

 

 

Period Period Amount of
Treatment . Strain 3 5 change
Z Z Z

l A 22 15 - 7
B 15 16 + 1

C 16 8 - 8

2 A 39 51 + 12
B 34 31 - 3

C 5 5 0

4 A 43 63 + 20
B 20 23 + 3

C 16 17 + l

5‘31 A 37 29 - 8
B 19 18 - l

C 13 12 - l

b

6 A 35 40 + 5
B ' l7 l3 - 4

C 5 16 + 11

7 A 24 37 + 13
B 18 19 + l

C 13 18 + 5

8 A 39 ' 49 + 10
B 32 22 - 10

C 12 1 - 11

9 A 33 38 + 5
B 14 15 + l

C 2 9 + 7

10 A 37 47 + 10
B 9 17 + 8

C 12 6 - 6

11b A 21 7 - 14
B 13 10 - 3

C 12 ll - l

12 A 33 35 + 2
B 25 35 + 10

C 12 8 - 4

 

a Vitamin K increased to 40 mg/lb during Period 5
b Sulfaquinoxaline increased to 0.1% during Period 5

 

73

Appendix Table 18. Percentage of Bloodspots Falling Into Each
Size Category During Periods 1 and 3

1‘

_L‘

 

 

 

 

Period 1 Period 3
Size_ __ Size‘
Under Between Over Under Between Over

Treatment 1.6 mm 1.6-4.8 mm 4.8 mm 1.6 mm 1.6-4.8 mm 4.8 mm

Z Z Z Z 2 2

1 56 24 20 56 18 25

2 46 24 ‘3o 41 13 .46

3 -_ -- -_ -_ -- --

4 46 18 36 44 14 42

5 52 17 32 50 16 34

6 6O 23 .17 43 17 4o

7 a 47 22 31 61 12 27

8 52 17 31 53 18 29

9 58 5 37 55 21 24

10 59 16 26 44 20 36
11 . 49 19 32 49 18 33

12 57 14 . 29 56 20 24

 

74

Appendix Table 19. Percentage of Bloodspots Falling Into Each Size
Category for Periods 3 and 5

 

 

 

 

 

 

Period 3 Period 5
A4:_ Size Size ‘£__
Under Between Over Under Between Over

Treatment 1.6 mm 1.6-4.8 mm 4.8 mm 1.6 mm 1.6-4.8 mm 4.8 mm

 

2 2 2 2 2 2

1 62 15 23 6O 23 17

2 4o , 13 48 49 18 33

3 -- -_ -- -- -- --

4 42 15 43 48 17 36

5a 49 15 36 49 20 31

6b 41 16 43 56 3 41

7 70 11 19 52 14 34

8 53 18 29 46 13 41

9 56 19 25 44 20 36

10 4o 20 36 48 16 36
11b 50 17 33 44 37 19
12 55 20 25 44 15 41

 

a Vitamin K increased to 40 mg/lb during Period 5

Sulfaquinoxaline increased to 0.1% during Period 5

 

75

Appendix Table 20. Analysis of Variance of Egg Production Data
for Period 1

 

 

 

Source of Degrees of Mean
variation freedom square F F .01
Treatments 10 4.9 .47. 2.40
Strains 2 65.5 6.35 4.70
T X S 20 9.4 .92 1.96
Error 222 10.3

Strains C A B

Av. eggs/hen 13.8 14.3 15.5 '

 

 

Means not underscored by the same line are significantly different

(p < .01)

 

76

Appendix Table 21. Analysis of Variance of Egg Production Data for

 

 

 

Period 3
Source of Degrees of Mean
variation freedom square F F .01
Treatments 10 34.5 2.67 ' 2.41
Strains 2 66.0 5.12 4.71
T X S 20 25.1 1.95 1.97
Error 198 12.9
8 7 5 11 2 6 9 10 1 4 12

10.5 11.2 12.2 12.3 12.4 12

.5 12.6 13.0 13.9 13.9 15.3

 

 

Strains A

Av. eggs/hen 11.6

12.7 13.4

 

 

 

Means not underscored by the same line are significantly different

(p (.01)

77

‘ Appendix Table 22.. Analysis of Variance of Egg Production Data for

Period 5

 

Source of Degrees of Mean

variation freedom square F F .01
Treatments 10 39.3 2.60 2.28
Strains 2 54.0 3.58 4.71

T X S 20 16.2 1.07 1.97
Error 196 15.1
Treatments and Av. eggs/hen

11 6 4 8 5 1 9 7 10 2 12

9.0 10.0 11.9 12.0 12.1 12.2 12.8 13.1 13.2 13.3 13.5

 

 

A

Means not underscored by the same line are significantly different

(P < .01)

78

>vwmsde Hmon Nw. wHooamuOn HsoHamonm mou mHHam mn H€o rm<mHm Om wnoacnnHo: an305n wmmmwd
mow anmmnamsn

 

 

 

 

mox wHOQCOnHos omlmnmmnmn fimmm arm: mON vnoaconHo:

zo. om 20. cm wmnnmsn zo. om zo. 0m menmSn

meHOQ mnHmH: mmmm mnOnm mwOnm mmmm muOnm chnm
H .8: do .2 :3 . 80 N»
w Noon mwN No NbH mm Nu
m NmbN mbN NH wa mu NN
H > HHwb boo DH mo NH bN
m Hon HmH H» H» w NH
o HHbN How Ho 00 0 Ho
w > ouu NNm uw 00 No wN
w HONH Hoo Ho 00 Hm Nm
a mom HHo HH 0H HH HN
m > 8» ”ma 3 3 No um
w Hon Now No bu HH No
a muN mw Ho HNw Hm HH

 

IIHHHH llHllllfl

78 0350