‘J‘H'sllflk‘l’fl'xl‘Ml

W

W

‘1

fl
\

 

126
984
TH _

THE. OCCURRENCE AND SIGNEFECANCE OF
RENAL LESIONS EN SWENE
PROCE§$ED F352 F909

Thesis for ”19 Degree of M. S.
MICHIGAN STATE UNWERSETY
James Henry Stewart

1962

H 5515

   
     

  

LIBRARY

Michigan State
University

 

 

 

 

ABSTRACT

THE OCCURRENCE AND SIGNIFICANCE OF RENAL
LESIONS IN SWINE PROCESSED FOR FOOD

by James Henry Stewart

A study was made of grayish-white lesions observed
during the post-mortem inspection of swine slaughtered for
food purposes. The study included serological, cultural,
and pathological examinations for determination of the oc-
currence and a possible etiological agent.

A review of the literature revealed a few references
of experimental studies concerning renal lesions resulting
from incidental bacterial infections, and many references
on leptospiral micro-organisms as a causative agent of
nephritis in swine.

The specimens collected for this study were obtained
at a local food processing establishment during slaughter-
ing operations.

The results of the microscopic agglutination—lysis
test for leptospiral antibodies in one hundred swine sera
samples, when compared with the pathological lesions in
the kidneys of these same animals, failed to reveal a sig—
nificant correlation.

Grossly, the renal lesions consisted of grayish—white

James Henry Stewart

foci in the parenchymatous tissues. Microscopically, the
lesions were an inflammatory process characterized by a
leukocytic infiltration consisting principally of lympho-
cytes in the intertubular spaces.

An etiological agent responsible for the grayish-
white renal lesion was not established on the basis of his-
topathological and cultural investigations.

The handling and consumption of swine kidneys ex-
hibiting grayish—white foci may be a possible human health
hazard. It is thought that these foci consisted of abnormal

tissue and should be eliminated from the consumer's food

supply.

THE OCCURRENCE AND SIGNIFICANCE OF RENAL
LESIONS IN SWINE PROCESSED FOR FOOD

by

James Henry Stewart

A THESIS

Submitted to the College of Veterinary Medicine
Michigan State University of Agriculture and
Applied Science in partial fulfillment of
the requirements for the degree of

MASTER OF SCIENCE

Department of Surgery and Medicine

1962

ACKNOWLEDGMENTS

I wish to express my appreciation and gratitude
to the following persons:

Doctor Gabel Conner, Department of Surgery and Med-
icine, my major professor, for his advice, encouragement,
and guidance.

Doctor Ralph Barner, Department of Surgery and Med—
icine, for his advice and guidance during this study.

Doctor Stuart Sleight, Department of Veterinary
Pathology, and Mrs. Athalie Lundberg, Department of Micro-
biology and Public Health, for their assistance on the lepto-
spiral studies.

Mrs. Marian Bennett, Department of Microbiology
and Public Health, for her assistance on the bacteriolog-
ical studies.

Miss Joyce Remsberg, Department of Surgery and Med-
icine, for her assistance on the histopathological studies.

Doctor Rollo Van Pelt, Department of Surgery and
Medicine, for his helpfulness on the photomicrographs.

My appreciation and gratitude is also extended to
others in the College of Veterinary Medicine for encourage—

ment given during this study.

I would like to thank the Meat Inspection Division,
Agricultural Research Service, United States Department of

Agriculture, whose support made possible this study.

Dedicated to my wife,

Katharina

TABLE OF CONTENTS

I 0 INTRODUCTION. 0 O O O O O O O O O O O O 0
II. REVIEW OF THE LITERATURE. . . . . . . . .
A. Introduction. . . . . . . . . . . .

B. Leptospirosis . . . . . . . . . . .

1. Gross Pathology . . . . . . .

2. Histopathology. . . . . . . .

3. serOlogy. O O O O O O O O O O

C. Pyelonephritis. . . . . . . . . . .

1. Gross Pathology . . . . . . .
2O HiStOpathOlogyo O O O 0 O O 0

III. MATERIALS AND METHODS . . . . . . . . . .
A. Collection Procedures . . . . . . .

I. Blood . . . . . . . . . . . .

2. Bacteriological and Histopathol gical

Specimens . . . . . . . . . .
B. Laboratory Procedures . . . . . . .

I Blood Study . . . . . . . . .

. Tissue Studies. . . . . . . .

isms. O O O O O O O O 0 O O O
. Isolation of Leptospirae. . .

2
3
4
IV. RESULTS . . . . . . . . . . . . . . . . .
V. DISCUSSION. . . . . . . . . . . . . . . .
VI. SUMMARY AND CONCLUSIONS . . . . . . . . .

VII. BIBLIOGMPHY. O O O O O O O O O O O O O 0

Isolation of Bacterial Micro-organ—

Page

NON-P- 45- \N \N

10

11
13
13
13
13
14

l4
15

15
l6

17
35

39
4O

Table
1.

LIST OF TABLES

Page
The incidence of grayish—white foci in the kidneys
of swine examined on post—mortem inspection . . . . l9
Serological results of swine serum samples to two
leptospiral serotypes . . . . . . . . . . . . . . . 2O
Serological results of swine sera samples reacting
to two leptospiral serotypes. . . . . . . . . . . . 21
Results of the histopathological examination of
swine kidneys containing grayish-white foci . . . . 22
Results of bacterial cultures of swine kidneys
containing grayish-white foci . . . . . . . . . . . 31
Results of Brown and Breen's staining method for
bacteria in tissues . . . . . . . . . . . . . . . . 32
Results of the Warthin—Starry's stain for lepto-
spirae in tissues . . . . . . . . . . . . . . . . . 33
Results of tissue culture attempts to isolate

leptospirae . . . . . . . . . . . . . . . . . . . . 34

Figure

1.

LIST OF FIGURES

Page

A diffuse infiltration of inflammatory cells

in the cortex of a swine kidney . . . . . . . . .
A higher magnification of Fig. 1 depicts the
diffuse infiltration of the inflammatory exudate
and degeneration of the convoluted tubules. . . .
A higher power of the preceding two photomicro-
graphs reveals a focus of mononuclear cells in
the affected area. The predominant cell types
are lymphocytes and mononuclear phagocytes. . . .
A subcortical infiltration of inflammatory cells
in a swine kidney . . . . . . . . . . . . . . . .
Greater magnification of Figure 4 shows subcap-
sular degenerative changes in the area of in-
flammation. . . . . . . . . . . . . . . . . . . .
A peritubular focal infiltration of inflammatory
cells in a swine kidney cortex. . . . . . . . . .
A subacute, subcapsular focus of inflammatory
cells in a swine kidney. The separation of the
capsule from the kidney tissue is an artifact . .
A higher power of Figure 7 exhibits the focus of

lymphocytes in the subcapsular area . . . . . . .

23

23

25

25

27

27

29

I. INTRODUCTION

The importance of animal health in relation to a
dependable meat supply has been recognized since the earli-
est history of man. The people of the United States are
particular in regard to the quality and character of food
consumed and object to consumption of animal tissues af—
fected with a disease condition. Since an abundance of
healthy animals is available to adequately supply meat for
the diet, this objection should be respected.

Veterinarians engaged in meat hygiene are perform-
ing a basic function in a civilized society by striving to
provide clean, wholesome, disease—free meat as a source
of protein for the diet. A veterinarian who does not pro-
tect the human consumer from meat that might cause disease
or that might otherwise be misrepresented is not perform-
ing an important function.

Veterinarians in meat hygiene must constantly strive
to learn new facts and to re-evaluate known facts about
diseases of animals for the meat consuming public. In the
inspection of swine slaughtered for food purposes, small
(pin-point) grayish-white foci are frequently observed in
the kidneys. The study of these foci was initiated with

two objectives in mind: (I) the occurrence, and (2) to

investigate the etiological, histopathological, and gross

pathological aspects of the foci.

II. REVIEW OF THE LITERATURE

A. Introduction:

Several infectious diseases occur in swine in which
only slight, or perhaps no, symptoms are apparent. Veter-
inarians performing post-mortem examinations of swine may
observe gross lesions in the cortex and medulla of the kid—
ney which apparently did not elicit clinical evidence of
disease. Pathological changes will be more meaningful to
a veterinarian performing a post—mortem examination when
the infectious diseases responsible for lesions in the kid-
neys are more thoroughly studied.

Pathological evidence of an infectious disease pro-
cess in a kidney is characterized by the micro-organisms
creating the lesion. In the kidneys of swine containing
small foci of infection in the cortex and medulla, a collec-
tion of leukocytes comprises an inflammatory exudate. This
is the result of the infectious process and is referred
to as interstitial nephritis.

Interstitial nephritis of swine caused by patho-
genic micro-organisms may be identified by gross patholog-
ical, histopathological, and cultural techniques. A sup—
purative or a non-suppurative interstitial nephritis can
result from an infectious process in the kidneys of swine.

Examples of micro—organisms causing suppurative interstitial

nephritis are CorynebacteriumApyogenes, Corynebacterium

 

renale, Proteus vulgaris, and Escherichia coli. The pre-

 

 

 

vailing leukocyte present in the inflammatory exudate of
suppurative interstitial nephritis is the polymorphonuclear
neutrophil.

Swine infected with leptospirae may have a non-sup-
purative interstitial nephritis as a result of the agent
localizing in the kidneys. The kidney lesions can grossly
resemble suppurative interstitial nephritis, but the com-
position of the inflammatory exudate will differ. The ex-
udate in non-suppurative interstitial nephritis will consist

of lymphocytes and reticulo-endothelial mononuclear cells.
B. Leptospirosis:

1. Gross pathology:

Leptospirae have a preference for the kidneys of
an animal where localization and reproduction occurs in
the tubules. The disease syndrome caused by the leptospirae
is fundamentally the same regardless of the species of lep—
tospirae or the species of animal involved. Only modifica-
tions in the virulence of the micro—organisms and the in-
tensity of the clinical and pathological changes will oc—
cur (28) (14) (8).

Small grayish—white foci observed in the kidneys
of swine examined during post-mortem inspection conducted

at local slaughtering establishments were suggestive of

leptospirosis (5) (8) (14). The lesion may appear singly
or scattered throughout the kidney substance, but the ani-
mal usually has not presented clinical symptoms (5) (8)
(14) (21) (40).

The gross lesions are limited principally to the
kidneys and appear either unilaterally or bilaterally in
an affected animal. The grayish-white foci vary in size
and number. The gross appearance of the lesion will range
from barely discernible to about 5 mm. in diameter, and
from one to many will be apparent in a single kidney. The
site of the lesion is usually the cortex, less frequently
in the medulla, of the kidney. The foci either appear even
with or slightly elevated above the cortical surface (5)
(8) (14) (10) (21) (40).

Adhesions between the capsule and the cortex usu-
ally do not occur and the capsule can be removed without
difficulty (8) (10) (14) (21) (40). It has been reported
in the literature that some kidneys may be roughened with
small depressions and the capsules adhere to the cortical
surface (8). In long standing chronic infections the kid-
neys have appeared shrunken and small bands of fibrous tis-
sue are present on the surface of the cortex extending into
the parenchyma (8).

The lesion is produced as early as seven days after
experimental infection with Leptospira pomona and persists

as an active inflammatory process for ten to fourteen

months (21) (29) (40). The significant features of the
gross renal lesions in swine experimentally infected with

Leptospira pomona are the early appearance and the relative

 

severity of the lesion created by the micro-organism (40).

2. Histopathology:

Histopathological lesions of leptospirosis are
present in the interstitial spaces, tubules, and glomeruli
in the kidneys of affected swine. The pathology produced
by the leptospiral micro-organisms is characterized by in—
flammatory and degenerative changes (8). The consistent
inflammatory reaction is an infiltration of leukocytes into
the interstitial spaces (5) (21) (29) (4C). The leukocytic
infiltration comprises an inflammatory exudate and consists
principally of lymphocytes, plasma cells, and some macro—
phages. The inflammatory exudate also may be present in
the perivascular spaces (21) (29) (40). The degree of in-
flammation in the cortex and medulla of a kidney is depend-
ent on the leptospirae creating the pathological changes (8).

The predominant degenerative pathological change
is hydropic degeneration of the kidney tubules (14) (40).
The earliest change occurring is a slight degree of hydropic
degeneration and occasional pyknosis of the nuclei of the
tubular cells. In more severe hydropic degeneration, the
nuclei are invisible or crowded to the surface and charac-
terized by large, swollen, vacuolated tubular cells (14)

(21) (40). The tubules within a foci of infection may be

destroyed resulting in physiologically inactive glomeruli
(14) (21) (40).

Glomerular changes can be frequent, severe, or only
minor.(5) (14) (21). In minor changes, Bowman's capsule
is surrounded by inflammatory cells and the subcapsular
spaces may contain some proteinaceous material (5) (21).
Hylanization of the glomerular tufts is evident (5) (21).
Glomeruli may appear hypercellular due to the presence of
lymphocytes and plasma cells (5) (21). Bowman's capsule
in the more extensive pathological changes can be completely
filled with a swollen glomerulus and adhesions may form
between the capillary loop‘s and the parietal layer of the
capsule (14). The glomerular tufts may even disappear leav—
ing a distorted Bowman's capsule with an empty lumen (21).
Some glomeruli may show evidence of undergoing fibrosis
with thickening of Bowman's capsule resulting in oblitera-
tion of the capsular space and loss of separation between
the capsule and glomerular tuft (14) (29).

3. Serology:

Serological surveys to reveal the incidence of lep-
tospiral antibodies in the serum of swine have been conducted
in many areas of the world. The interpretation of the sero—
logical findings has been difficult because a correlation
between the actual antibody titers present and its diag-
nostic significance is not fully understood in evaluating
leptospiral infections (19) (35) (5) (8) (9) (13) (24) (28)
(32) (48)-

Swine experimentally infected with Leptospira pomona
2

 

had a serum antibody titer detectable at dilutions of IO-
and 10-3 eight days after inoculation and the maximum anti-

body response (10-7 to 10—8) occurred three to four weeks

post inoculation (2s). The antibody titer declined to 10‘2

to 10—5 four or five months later (28). However, antibody

2

titers (10- to 10-3) have persisted in swine experiment-

ally infected with Leptospira_pomona for one year (28).

 

Swine probably are actively infected with Leptospira

 

pomona when the serum agglutination-lysis titers are present
in dilutions of 10-4 or higher (28). A potential hazard

to livestock and human health exists during an active in-
fection because the affected animal may become a renal car-
rier and transmit the micro-organisms within an environment
(4) (6) (8) (10) (25) (26) (28) (48) (50)-

Agglutination-lysis reactions in the dilutions of

-1 -2

lO , lO , and 10-3 can be interpreted with different diag-
nostic meanings in relation to leptospiral infections in
swine (5) (24) (28) (32) (35) (e) (9) (19) (13) (48). These
low antibody titers may indicate inactive infection in which
the antibody titer has decreased, or a recently acquired
active infection where the antibody titer has not obtained
its maximum level (5) (19) (24) (28) (35) (32).

The epizootiology of leptospirosis is complicated

by the ability of the leptospirae to become localized in

the kidney tubules of swine and rats with a low or absent

serum antibody titer (5) (8) (18). The isolation of Lep-

tospira icterohemorrhagiae from the kidneys of rats whose

 

sera did not contain demonstrable antibody titers has been
accomplished (18). Leptospira pomona have been recovered
from the kidneys of swine whose sera did not react or re-
acted at the lower dilutions when tested with the plate
agglutination or agglutination-lysis tests (5) (8).

Piglets do not contain agglutination-lysis titers
against a leptospiral serotype at birth because the maternal
antibodies are withheld from the feti in utero by the pla-
cental barrier (8) (28). However, antibodies are acquired
from the colostral milk and piglets will reveal, soon after
suckling, approximately the same antibody titer as the sow
(8). In the absence of an active leptospiral infection,

a progressive decline in the piglets acquired antibody titer
occurs (8).

Disappearance of antibody titers in sera of animals
with active leptospiral infections may occur upon adminis-
tration of antibiotic drugs (18). The leptospiral micro-
organisms present in the liver, spleen, and interstitial
kidney tissues may be destroyed without affecting the micro-
organisms localized in the kidney tubules. These animals
can remain carriers capable of transmitting the leptospiral
micro—organisms (18). Other phenomena that may influence
the development of an antibody titer are the age and number

of leptospirae in the infective dose, and neutralization

10

by the micro-organisms of the circulating antibody (28)

(29) (32).
C. Pyelonephritis:

1. Gross pathology:

Veterinary literature describing pyelonephritis
in swine is limited because a greater emphasis has been
directed toward the disease condition in other animals,
especially cattle and horses. The term pyelonephritis sig-
nifies an inflammatory process occurring in the pelvis and
parenchyma of the kidney (44). Pyelonephritis is usually
of an infectious nature and several genera and species of
micro-organisms are implicated as etiological agents re-
sponsible for the pathology produced (23) (44). An extensive
and diffuse necrotizing inflammatory process usually char-
acterizes the disease condition (17) (20) (23) (34) (44)
(45).

The inflammatory process is frequently bilateral
in an affected animal, but an occasional unilateral infec-
tion does occur (17) (23) (45). The size, color, and tex-
ture of an affected kidney will vary in gross appearance
dependent upon the severity, extent, and duration of the
pathological changes resulting from the infection. The
size of a kidney may be normal or greatly enlarged with
very little kidney tissue present (17) (23) (34) (45).

The cortical surface can be mottled with grayish-white foci

11

which contain an inflammatory exudate (34) (44) (45) (49).
The texture of the kidney substance can vary from
normal to firm depending upon the degree of fibrosis that
resulted from the inflammatory process (23) (34) (44) (45).
The pelvis of an affected kidney may contain a non-odorous
gray or yellow purulent material, often mixed with fibrin,
small blood clots, necrotic tissue, and calcareous matter
(17) (23) (34) (44) (45) (49). The purulent material is
tenacious in consistency and will distend the pelvis when
a large enough quantity is present. The papillae of the
renal pyramids can become necrotic in the course of the
inflammatory process (17) (23) (34). The medullary and
cortical areas of an affected kidney may possess abscesses
and hemorrhages that vary in size from micrOSCOpic to macro-
scopic (17) (23) (34) (44) (49). The capsule of an affected
kidney may be adherent to the cortical surface and will be
difficult to remove (23) (54) (49).

2. Histopathology:

A histopathological examination will reveal an in-
flammatory exudate, degenerative, and proliferative path-
ological changes in the kidney tissues affected by the in-
flammatory process (54) (44) (45). Polymorphonuclear leu-
kocytes usually will be the major component of the exudate
and in conjunction with lymphocytes, mononuclear phagocytes,
as well as other inflammatory elements, will constitute

a purulent exudate (34) (44) (45). The exudate may appear

12

to be diffusely located among the kidney tubules in a more
or less radial direction (34) (44).

Degenerative pathological changes of the kidney
tubules are associated with the inflammatory process that
occurs in an affected kidney. MicrOSCOpically, a kidney
infected with pyelonephritis will reveal pathological changes
such as cloudy swelling, fatty degeneration, and necrosis
of the epithelial cells comprising the kidney tubules (34)
(44) (45). These changes observed on histopathological
examination may be due to a reaction between the kidney
cells and a toxin derived from the growth of the micro-
organism responsible for the pathological syndrome (44).

A fibrous proliferative tissue response accompanies
the inflammatory process in the localized infected areas
of the kidney. The tissue reaction is primarily a pericap-
sular proliferation that encircles Bowman's capsules with
one or several layers of fibrotic tissue. The glomerulus
usually is not invaded by fibrous tissue, but a prolifera-
tion may extend into it. An extension of the intertubular
connective tissue of the medulla and thickening of arteries
and arterioles may occur in the immediate vicinity of the

inflammatory process (34) (44) (45).

III. MATERIALS AND METHODS

A. Collection Procedures:

The specimens for this study were obtained from
swine slaughtered at a food processing establishment lo—
cated in East Lansing, Michigan. The swine were selected
at random from 16 separate slaughtering Operations. The
health status, environmental conditions, and the animal
husbandry practices were not known prior to the collection
of the various specimens. Variations in the age, breed,
or sex were not considered a factor in the selection of
the swine. The writer performed an ante-mortem and post-
mortem examination on all of the swine that were slaughtered.
None of the animals presented for slaughter exhibited clin-
ical symptoms of a disease condition.

1. Blood:

All blood samples were obtained from the anterior
vena cava after the swine had been shackled and placed on
an overhead track in the eviscerating room. The blood was
collected in dry vials immediately after severance of the
anterior vena cava and allowed to clot. Humane slaughter-
ing techniques were not employed to produce surgical anes-
thesia before severance of the anterior vena cava.

2. Bacteriological and Histopathological Specimens:

The swine were placed in a scalding vat following

13

14

exsanguination. The hair and scurf became less firmly fixed
which facilitated its removal by a dehairing machine. Each
carcass was suspended on an overhead dressing track after
removal of the hair and scurf for evisceration and inspec-
tion. Carcasses that were not rejected after examination

of the head and viscera continued along the dressing track
and were split and the kidneys removed from their capsules.
The kidneys were inspected and those with grayish—white

foci were removed from the carcass. The affected portions
were excised and placed in a container (8 ounce "dispo-cup"*)
for bacteriological examination. Suitable Specimens were
likewise immediately placed in 4 ounce screw cap jars con-
taining ten per cent neutral formol-saline solution for
fixation for histopathological examination. Approximately
30 minutes elapsed from the time of exsanguination to col—

lection of the specimens.
B. Laboratory Procedures:

1. Blood Study:
The microscopic agglutination-lysis test for lepto-
spiral antibody titers was conducted on the serum from 100

swine. Leptospiragpomona (strain Johnson) and Leptospira

 

 

icterohemorrhagiae live antigens and the serums were mixed

 

 

*Disposable Specimen Cup. Distributed by the Sci—
entific Products. A Division of American Hospital Supply
Corp., Evanston, Ill.

l5

1

in dilutions of 10‘ , 10‘2

, and 10-3. The tubes containing
the serum mixed with live antigen were incubated for two
hours at 37 C. in a thermostatically controlled water bath
before reading the test. A microscope, with an Abbe con-
denser into which a star diaphram was fitted, was employed
to produce a modified dark-field type of illumination.

The degree of agglutination was recorded as (1+ (less than
25%). 1+ (25%). 2+ (50%). and 3+ (75%).

2. Tissue Studies:

The kidneys of all swine were inspected during the
post-mortem examination. When grayish—white foci were ob-
served on the cortex or occasionally in the medulla, tissue
sections were taken for histopathological studies. The
staining procedures used were as follows: hematoxylin and
eosin, Brown and Breen's method for bacteria in tissues
(1), and Warthin—Starry's technique for spirochetes (1).

3. Isolation of Bacterial Micro-organisms:

Tryptose agar (Difco) with defibrinated blood con-
stituting a final concentration of seven per cent was used
in attempts to isolate bacterial micro-organisms from the
grayish-white kidney lesions. Samples from the lesions
were streaked on this medium and incubated for 24 hours at
37 C. The presence or absence of bacterial growth was ob-
served at the end of the incubation period. In the absence
of growth, further bacteriological culturing was not under-

taken.

16

4. Isolation of Leptospirae:

The isolation of leptospirae from the kidneys con-
taining grayish-white foci was attempted by a culture medium
technique (5) (9) (25). This tissue was placed in approx—
imately 20 ml. of 0.85% sodium chloride solution and macer-
ated in a Waring blender. Three or four drops of the emul-
sion produced was inoculated into a culture tube contain-
ing Stuart's Difco medium with rabbit serum added to a final
concentration of ten per cent. The culture tubes were in—
cubated for four weeks at 30 C. and the presence of lepto-
spirae was investigated at weekly intervals by a microscopic

dark—field examination.

IV. RESULTS

During the post-mortem inspection of swine at a
local food processing establishment, it was found that a
number of the kidneys examined contained grayish-white foci
in the cortices. The gross lesions observed resembled the
characteristic lesion produced in the kidneys of swine ex-

perimentally infected with Leptospiragpomona (8) (10) (21)

 

(40). However, under field conditions other etiological
agents may produce a similar type of lesion (8).

The examination of 200 kidneys revealed a relatively
high incidence of grayish—white foci in the cortices. The
percentage of kidneys with foci observed in various slaughter
groups ranged from 50.00 to 11.54. Table 1 presents data
showing that 41 kidneys had 1 to 2 foci and that 13 kidneys
had 10 to 20 foci. The capsules stripped easily from the
cortical surfaces of the kidneys. Some of the lesions were
slightly raised above the cortical surface, others were
not, and ranged in size from barely visible to 4 mm. in
diameter. When sectioned saggitally the grayish-white foci
were seen extending into the medulla of some kidneys.

The results of the micrOSCOpic agglutination-lysis
test conducted on 100 swine sera are presented in Tables

2 and 3. leptospiragpomona and Leptospira icterohemorrhagiae

 

 

live antigens were used in testing the sera for antibody

17

18

titers. Agglutination—lysis reactions were detectable in
19 of the sera. Of these, 12 reacted in a dilution so low
that they were considered insignificant, and 7 reacted in
dilutions indicative of possible exposure to a leptospiral
micro-organism (Table 3) (8) (5) (28) (19) (9) (13) (24)
(52) (55).

The results of the histopathological examination
of the kidneys containing grayish-white foci is presented
in Table 4, and Figures 1-8. The examination consistently
revealed an interstitial nephritis and an inflammatory ex-
udate comprised predominantly of lymphocytic cells. There
was also evidence of degeneration in areas infiltrated by
the inflammatory exudate.

The results of attempts to isolate a microbial
species as an etiological agent are presented in the data
(Tables 5-8). The isolation and identification of a micro-
organism responsible for the inflammatory process was not

accomplished in this study.

Table 1.

19

The incidence of grayish—white foci in the kid-
neys of swine examined on post-mortem inspection

 

 

 

Date Number of Number of Kidneys Kidneys Per Cent
Kidneys Kidneys with with of Kidneys
Examined with 1-2 10-20 with Foci
Foci Foci Foci
5—23-61 52 10 5 5 51.25
6-6-61 16 7 5 2 45.75
7-25-61 52 6 6 0 11.54
9-12-61 24 9 7 2 57.50
9-25-61 6 5 2 l 50.00
10-3—61 8 2 2 0 25.00
10—17—61 24 7 5 2 29.17
10-24-61 24 4 4 O 16.67
12-19-61 14 6 5 l 42.86
Total 200 54 V 41 13 27.00

 

20

Table 2. Serological results of swine serum samples to
two leptospiral serotypes

 

 

Serotype Swine Sera Swine Sera Swine Sera
Tested Negative Reacting
L. pomona 100 92 8

 

L. icterohemorr-
hagiae 100 89 ll

 

21

Table 3. Serological results of swine sera samples react-

ing to two leptospiral serotypes

 

 

 

 

Serotype Number
of Sera Dilutions
Samples
‘ 10’1 10"2 10‘3
(1+ 1+ 2+ 3+ 1+ 2+ 3+ 1+ 2+ 3+
5 + - - - - - - - - -
2 - + — - - - - - - -
L. pomona
2 - - + — — — - - - -
l — - + - + - - - - -

 

7 - + - - - - - - - -
2 - — + - - — - - - -
L. icterohemorr-
hagiae l - - + - - + - - - —
l

 

22

Table 4. Results of the histopathological examination of
swine kidneys containing grayish-white foci

 

 

 

Date Number of Interstitial Predominant Degeneration

Kidneys Nephritis Cell in the of the Kidney

Examined Inflammatory Parenchyma-

Exudate tous Tissues

5-23-61 5 Yes Lymphocyte Yes
6-6-61 3 Yes Lymphocyte Yes
7-25-61 4 Yes Lymphocyte Yes
9-12-61 4 Yes Lymphocyte Yes
9-25-61 3 Yes Lymphocyte Yes
10-3—61 2 Yes Lymphocyte Yes
10—17-61 4 Yes Lymphocyte Yes
10-24-61 3 Yes Lymphocyte Yes
12-19-61 3 Yes Lymphocyte Yes
Total 31

 

25

Figure 1
x 31
A diffuse infiltration of inflammatory cells in the

cortex of a swine kidney. H & E.

Figure 2

x 125
A higher magnification of Fig. 1 depicts the diffuse
infiltration of the inflammatory exudate and degenera-

tion of the convoluted tubules. H & E.

24

 

 

 

 

25

Figure 3
x 500
A higher power of the preceding two photomicrographs
reveals a focus of mononuclear cells in the affected

area. The predominant cell types are lymphocytes (A) and

mononuclear phagocytes (B). H & E.

Figure 4
x 31
A subcortical infiltration of inflammatory cells (A) in

a swine kidney. H & E.

26

 

 

 

27

Figure 5
x 125
Greater magnification of Fig. 4 shows subcapsular degen—

erative changes in the area of inflammation. H & E.

Figure 6
x 31
A peritubular focal infiltration of inflammatory cells

in a swine kidney cortex. H & E.

 

 

 

 

28

 

29

Figure 7

x 31
A subacute, subcapsular focus of inflammatory cells in
a swine kidney. The separation of the capsule from the

kidney tissue is an artifact. H & E.

Figure 8
x 500
A higher power of Fig. 7 exhibits the focus of lympho—

cytes (A) in the subcapsular area. H & E.

 

 

50

 

51

Table 5. Results of bacterial cultures of swine kidneys
containing grayish-white foci

 

 

Date Number of Bacteriological
Kidneys Findings
Cultured
5-23-61 3 Negative
6-6—61 1 Negative
7-25—61 4 Negative
9-12-61 3 Negative
9-25-61 2 Negative
10-3—61 2 Negative
10-17-61 2 Negative
10—24-61 1 Negative
l2-l9—61 2 Negative

 

Total 20

 

52

Table 6. Results of Brown and Breen's staining method for
bacteria in tissues

 

 

 

Date Number of Bacterial
Kidneys Findings
Examined
5-25-61 5 Negative
6-6-61 5 Negative
7-25-61 4 Negative
9-12-61 4 Negative
9+25-6l 5 Negative
10-3-61 2 Negative
10—17-61 4 Negative
10-24-61 3 Negative
l2-l9-6l 3 Negative
Total 31

 

55

Table 7. Results of the Warthin-Starry stain for lepto-
spirae in tissues

 

 

 

Date Number of Leptospirae
Kidneys in the Tissues
Examined
5—23—61 5 Negative
6-6-61 3 Negative
7—25-61 4 Negative
9-12—61 4 Negative
9-25-61 3 Negative
10-3-61 2 Negative
10-17-61 4 Negative
10—24-61 3 Negative
12—19-61 3 Negative
Total 31

 

54

Table 8. Results of tissue culture attempts to isolate

 

 

 

leptospirae
Date Number of Leptospiral
Cultured Kidneys Isolations

Cultured

1-9-62 4 Negative
1-16-62 4 Negative
1-23-62 3 Negative
2—6-62 4 Negative
2-13—62 2. Negative
3-13-62 2 Negative
3-27-62 2 Negative
Total 21

 

V. DISCUSSION

During the post—mortem examination of clinically
normal swine slaughtered in a local food processing estab-
lishment grayish-white renal lesions were observed. The
gross and histopathological lesions resembled an inflamma—
tory process created by leptospiral micro—organisms. Gross-
ly, the lesions observed were small grayish-white foci vary-
ing in size and number which were limited to the parenchy-
matous tissues of the kidneys, especially the cortices.
A suppurative process characterized by a tenacious exudate
was not evident upon gross examination. The microscopic
lesions consisted of a leukocytic infiltration principally
in the intertubular spaces of the cortex and medulla, al-
though it occurred in perivascular and periglomerular areas.
The lesion was usually focal, but in several kidneys it
was rather diffuse. The unit of the nephron in an affected
area was either undergoing degeneration or had entirely
disappeared. The findings were similar to the pathological
processes produced in pigs experimentally infected with
Leptogpiragppmona (8) (10) (21) (40).

A comparison of the histopathological and serolog-

ical results (Tables 2 and 3) obtained using Leptospira

 

pomona and Leptospira icterohemorrhagiae live antigens

 

failed to indicate definite correlation. The interpretation

55

56

of the serological findings is difficult because the diag-
nostic value of antibody titers in the lower dilutions is
not completely understood in leptospiral infections in swine
(5) (8) (9) (13) (19) (24) (28) (32) (35). If the reactions
of 25% agglutination or less at the dilution of 10—1 are
eliminated and only the higher reactions considered several
explanations are possible. Certain factors such as the

age, number, and virulence of the leptospirae in the in-
fective dose may influence the height of the titer in a

pig that becomes infected (32). Antibiotic therapy could
eliminate the leptOSpirae from a pig except in the kidney
tubules and a loss of a demonstrable blood titer might oc-
cur (18). Piglets are born without antibodies against L32-
tospirappomona and can acquire them from the colostral milk
of the sow. However, immunity acquired in this manner will
decline (8). When the disease is endemic most of the pigs
probably become infected during early life and a low anti-
body titer may be present at the time of slaughter (37).

The serological results could be an indication that infec—
tion may have been caused by leptospirae other than.I§ptg-

spiragpomona or Leptospira icterohemorrhggiae (39). The

 

 

comparatively low antibody levels in the pig sera tested
may have been due to any one, or a combination, of the
aforementioned possibilities.

It has been reported in the literature that 20%

of 114 swine sera collected at a slaughtering establishment

57

contained antibodies against Leptospira_pomona, but 50%

 

of these animals revealed histopathological lesions indic—
ative of leptospirosis (8). micrOSCOpic evidence of a lep-
tospiral infection was found in 86% of 338 cattle received
at the Plum Island Animal Disease Laboratory during a one
year period. A correlation between the serological and
histopathological results was not evident (39).

In this study, the kidneys from 100 swine had a
27% incidence of grayish-white foci in the parenchymatous
tissues without significant serological reactions to Lento-

*

§piragpomona and Leptospira icterohemorrhagiae antigens.

 

These observations suggest that the incidence of infection
with leptospirae may be higher than indicated by the micro-
SCOpic agglutination-lysis test.

The results of three procedures presented in the
data (Tables 4-6) suggest that the kidney lesions did not
contain an intact bacterial micro—organism: (l) the per-
sistent absence of suppuration in the tissue sections stained
with hematoxylin and eosin, (2) the failure to obtain growth
of a bacterial species from lesions cultured on tryptose
blood agar, (3) the negative results of the Brown and Breen's
staining method for bacteria in tissues.

The tissue cultures used in this study failed to
isolate a leptospirae from the grayish-white renal foci.
However, it was judged that the abnormal tissue observed
in the swine kidneys should be eliminated from the consumer's

food supply.

58

Veterinarians engaged in meat hygiene are striving

to provide a meat supply free from diseases injurious to

the health of the consumer. Leptospirae in animals intended
for human food have the ability to localize and reproduce

in the kidneys for long periods of time (4) (8) (28) (36)
(37). Research evidence on swine leptospirosis and the
observations herein presented indicates there may be a cor-
relation between the grayish—white foci and the leptospirae

and further research appears necessary.

VI. SUMflARY AND CONCLUSIONS

Grayish—white renal foci were seen during the post—
mortem examination of swine conducted at a local food pro-
cessing establishment. The abnormal tissue was located
in the cortex and medulla of the kidney. Microscopically,
the lesions were a leukocytic infiltration in the inter-
tubular spaces and consisted principally of lymphocytes.

The sera of 100 swine were tested by the micro-
scopic agglutination-lysis test against Leptospiragpomona
and Leptospira icterohemorrhagiae live antigens. The anti-
body titers were relatively low and difficult to interpret
in relation to exposure to leptospirae.

The research evidence appearing in the literature
and the observations presented in this study suggest that
the grayish-white renal foci may indicate a higher incidence
of leptospiral infections than is revealed by the microscopic
agglutination-lysis test.

The procedures used in this study did not confirm
a specific etiological agent responsible for the grayish-
white renal lesion.

The grayish-white foci consist of abnormal tissue
and for this reason such tissue should be eliminated from

the normal channels of food production.

59

VII. BIBLIOGRAPHY

Ambrogi, L. P.: Editor. Manual of Histologic and
Special Staining Techniques. 2nd ed. McGraw-Hill
Co. New York. (l960):l80—l8l; 184-185.

Baker, C. E., Gallian, M. J., and White, E. A.: Lep-
tospirosis. 1. Therapeutic Studies on the Erad-
ication of Renal Carriers of Porcine Leptospirosis.
Vet. Med., 52, (March, l957):lO5-107.

Baker, E. D., Piraino, F., and Ruedy, D.: A Survey
of Swine Brucellosis and Leptospirosis in Wisconsin.
J.A.V.M.A., 134, (May, l959):440-442.

Bohl, E. H., Powers, T. E., and Ferguson, L. C.: Lep-
tospirosis in Swine. Proc. U.S. Livestock San. A.,
58, (1954):565-567.

Boulanger, P., Mitchell, D., Corner, A., and Bourassa,
M.: Observations on Leptospirosis in Swine. Canad.
J. Comp. Med. & Vet. Sci., 23, (Nov., 1959):354-359.

Braun, J. L.: Epidemiology of Leptospirosis in Iowa--
A Study of Sporadic and Epidemic Cases. J.A.V.M.A.,
158, (May, l96l):552-556.

Bryan, H. 8.: Studies on Leptospirosis in Domestic

Animals. VI: Vaccination of Swine with Leptospira

 

pomona Bacterin. Vet. Med., 52, (Feb., l957):51-57.

4o

IO.

11.

12.

15.

14.

15.

41

Burnstein, T., and Baker, J. A.: Leptospirosis in
Swine Caused by Leptospira pomona. J. Infect. Dis.,
94, (1954):55—64.

Coghlan, J. D., and Norval, J.: Canicola Fever in Man
Through Contact with Infected Pigs. Brit. Med. J.,
(Feb., l957):257-26l.

Coghlan, J. D., and Norval, J.: Canicola Fever in Man
from Contact with Infected Pigs. Further Observa-
tions. Brit. Med. J., (Dec., l960):17ll-l713.

DeLay, P. D., Daley, R., and Callahan, 0.: LeptOSpiro-
sis in Relation to Porcine Abortion. Calif. Vet.,
8, (Mar/Apr., l955):36—37.

Ferguson, L. C., and Powers, T. E.: Experimental Lep-
tospirosis in Pregnant Swine. Am. J. Vet. Res.,

17, (July, 1956):47l—477.

Ferguson, L. C., Lococo, S., Smith, H. R., and Hamdy,
A. H.: The Control and Treatment of Swine Lepto-
spirosis During a Naturally Occurring Outbreak.
J.A.V.M.A., 129, (Sept., l956):263~265.

Ferguson, L. C.: Leptospirosis. In Diseases of Swine.
Edit. by H. W. Dunne. lst ed. Iowa State College
Press, Ames, Iowa. 1958:253—266.

Florin, M. 0.: Porcine Nephritis. Thesis. Cornell

University. 1941:1-33.

l6.

17.

18.

19.

20.

21.

22.

Gambrel,

An Outbreak of Porcine Leptospirosis.

Vet.,

Hagan, W. A.,

eases of Domestic Animals. 4th ed.

42

P. T., Gambrel, E. T., and Gambrel, L. E.:

North Am.

57, (Jan., l956):l9-21.

and Bruner,

D. W.: The Infectious Dis-

Comstock Pub-

lishing Associates, Cornell University Press, Ithaca,

1961:149—154.

N. Y.

Howarth, J. A.:

in an Outbreak of Leptospirosis of Swine.

Effect of Aureomycin and Polyotic

J.A.V.M.A.,

129, (Sept., 1956):268-271.

Keast, J.

C., Forbes,

B. R. V., and Wannan, J. S.‘

Serological Survey of Pigs in New South Wales for

Leptospirosis and the Incidence of Swine Abortion.

Austral. Vet. J., 32,
Kenzy, S. G.,
Comparison
uated Live
in Cattle.
Langham, R. E., Morse, E. V.,

perimental Leptospirosis.

Gillespie, R.

J.A.V.MOA. ’

(Jan., l956):ll-17.

W. H., and Lee, J. H.:

of Leptospira_pomona Bacterin and Atten-

Culture Vaccine for Control of Abortion

139. (Aug., 1961):452-454.
and Morter, R. L.: Ex-

V. Pathology of Lepto-

gpira pomona Infection in Swine. Am. J. Vet. Res.,

199 (April, 1958):395-4OO.

Lingard, D. R.,

and Hanson,

spira pomona on the Reproductive Efficiency of

Cattle.

J.A.VOlflOA. ’

159,

(Aug., 1961):450-451.

230

24.

25.

26.

27.

28.

29.

50.

45

The Merck Veterinary Manual. Edit. by 0. H. Siegmund.
2nd ed. Merck and Company, Inc., Rahway, N. J.
1961:1064-1067; 1082-1083.

Michna, S. W.: Antibodies in the Sera of Pigs: A
Preliminary Report. Vet. Rec., 70, (Jan., 1958):
80-82.

Michna, S. W.: The Isolation of Leptospira canicola

 

from the Kidney of a Pig. Vet. Rec., 71, (Jan.,

Michna, S. W.: The Survival of Leptospira canicola

 

in the Renal Tissue of a Pig. Vet. Rec., 71, (Aug.,
l959):549-552.

Monlux, A. W., Siebold, H. R., ShalkOp, W. T., and
Davis, C. L.: Leptospirosis in Hogs. North Am.
Vet., 55, (July, l952):467-469.

Morse, E. V., Bauer, D. C., Langham, R. F., Lang, R. W.,
and Ullrey, D. E.: Experimental Leptospirosis. IV.

Pathogenesis of Porcine Leptospiragpomona Infections.

 

Am. J. Vet. Res., 19, (April, l958):388—594.
Morter, R. L., Morse, E. V., and Langham, R. F.: Ex-
perimental Leptospirosis. VII. Re—exposure of

Pregnant Sows with Leptospira pomona. Am. J. Vet.
Res., 21, (Jan., l960):95-98.
Mullins, J. C.: An Outbreak of Leptospiragpomona in

Swine. Southwest Vet., 8, (Fall, 1954):88.

31.

32.

55.

54.

55.

36.

57.

58.

44

Norval, J.: Leptospirosis in Pigs. Vet. Rec., 70,
(June, l958):470-472.

Powers, T. E., Bohl, E. H., and Ferguson, L. 0.: Clin-
ical Studies on Leptospirosis as a Cause of Abortion
in Swine. J.A.V.M.A., 129, (Dec., 1956):568-572.

Roberts, C. H., Turner, L. W., and Livingston, J. H.:
Bovine Leptospirosis in Alabama-~A Five Year Study.
J.A.V.M.A., 139. (Oct., l96l):877—883.

Runnels, R. A.: Animal Pathology. 5th ed. Iowa State
College Press, Ames, Iowa. 1954:442-463.

Russell, R. R., and Hansen, N. F.: The Incidence of

Leptospira hyos and Leptospiragpomona Infections

 

in Pigs in New Zealand. N. Zeal. Vet. J., 6,
(April, l958):50-51.

Riley, J. W., and Simmons, G. C.: Leptospirosis of
Pigs with Special Reference to Birth of Dead Pigs
and Neo-natal Mortality. Austral. Vet., J., 30,
(July, l954):205-208.

Riley, J. W.: Leptospirosis in Swine. Austral. Vet.
J., 32, (Jan., l956):4—ll.

Schnurrenberger, P. R., Tjalma, R. A., Stegmiller,

H. E., and Wentworth, F. H.: Bovine Leptospirosis—-
A Hazard to Man. J.A.V.M.A., 139, (Oct., 1961):
884—888.

39.

40.

41.

42.

430

44.

450

46.

45

Siebold, H. R., Keech, H., and Bokelman, D. L.: Sub-
clinical Leptospirosis Among Cattle. J.A.V.M.A.,
138, (April, l96l):424-451.

Sleight, S. D., Langham, R. F., and Norter, R. L.:
Experimental Leptospirosis. The Early Pathogenesis
of Leptospiragpomona Infection in Young Swine. J.
Infect. Dis., 106, (May-June, 1960):262-269.

Sleight, S. D., and Williams, J. A.: Transmission of
Bovine Leptospirosis by Coition and Artificial In-
semination--A Preliminary Report. J.A.V.M.A., 138,
(Feb., 1961):151-152.

Sleight, S. D., and Lundberg, A. M.: Persistence of

LeptOSpira pomona in Porcine Tissues. J.A.V.M.A.,

 

159. (Aug., l961):455-456.
Smith, H. C.: Advances Made in Swine Practice. Vet.
Med., 51, (Jan., 1956):55-57.
Smith, H. A., and Jones, T. C.: Veterinary Pathology.
Lea & Febiger. Philadelphia. 1958:782-815.
Soltys, M. A., and Spratling, F. R.: Infectious Cys-
titis and Pyelonephritis of Pigs: A Preliminary
Communication. Vet. Rec., 69, (May, l957):500-504.
Tammemagi, L., and Simmons, G. 0.: Experimental In-

fection of Pigs with Leptospira hyos Savino and

 

Rennella. Queensland J. Agri. Sci., 13, (Sept.,
l956):l69-l74.

47.

48.

49.

50.

46

Tammamagi, L., and Simmons, G. C.: Further Observa-
tions on the Experimental Infection of Pigs with
Lgptospira hyos Savino and Rennella. Queensland
J. Agri. Sci., 15, (Sept., l958):l37-l44.

Tobie, J. E., and McCullough, N. B.: Serological Evi-
dence of Leptospira pomona Infections in Meat In-
spectors. J.A.V.M.A., 158, (April, l96l):454-456.

Udall, D. H.: The Practice of Veterinary Medicine.
5th ed. Ithaca, New York. 1947:254—259.

Van Der Hoeden, J.: Leptospirosis Canicolaris in Pigs
and Its Probable Transfer to Human Beings. J. In-

fect. Dis., 98, (Feb., l956):35-58.

”1144411113111 [1113111111 1111311111“