Investigations on the role of staphylococci in the pathogenesis of German Shepherd dog Pyoderma (GSP) M. A. Wisselinkl, J. P. Koeman2, T. S. G. A. M. van den Ingh2, and A. Willemsel SUMMARY Skin reaction_patterns to intradermal injections ofa Staphylococcus intermedius antigen were examined in physically healthy dogs and in dogs with German Shepherd dog Pyoderma (GSP) at 15 and 30 minutes and at I, 2, 4, 6, 8, 24, 48 and 72 hours after the injection. In both groups the skin histopathology revealed an aspecific inflammatory response of an early polymorphonuclear reaction, followed by a mononuclear cell reaction at 24 and 48 hours. It is concluded that hypersensitivity to staphylococcal antigens does not play a role in the pathogenesis of GSP INTRODUCTION
German Shepherd dog Pyoderma (GSP) is a chronic deep pyoderma in pure-bred and cross-bred German shepherd dogs characterised by multiple deep-seated skin lesions. The most commonly associated bacteria are coagulase-positive staphylococci (CPS) (18). So far the role of CPS in the pathogenesis of GSP is not clear. In general, CPS are considered by far the most important pathogenic organisms in bacterial skin diseases in dogs (1, 10, 11). Recently three coagulase-positive or coagulase-variable species (S. aureus, S. intermedius and S. hyicus) have been determined on the basis of biotyping and DNA-homology studies (8, 9). Among these, S. intermedius has been identified as the major CPS associated with normal and infected canine skin (2, 3). In addition to infection, CPS may also induce hypersensitivity reactions. Such bacterial hypersensitivity has been describel in man and (laboratory) animals (5, 7, 12, 14, 16). In the dog the occurrence of types III and IV hypersensitivity reactions has been suggested (4, 13). In a previous report (19) bacterial hypersensitivity to CPS has been hypothesised as one of the predisposing factors for GSP. This hypothesis was tested by studying the histopathologic changes of skin-test sites at different time intervals, following injection of an extract of S. intermedius. MATERIALS AND METHODS
Animals
Group A consisted of 14 male (two castrated) and 12 female (four castrated) German
shepherd dogs with the clinical features of GSP (18). Their ages ranged from I to 12 years, with a median age of 6 years. Group B consisted of eight male and five female physically healthy German shepherd dogs kept under household conditions. Their ages ranged from 1 to 13 years (median: 8 years). Group C was comprised of six male and four female physically healthy dogs kept in the university clinic, including one bouvier, one beagle, one greyhound, one German shepherd and six mongrel dogs. Their ages ranged from 5 to 9 years (median: 7.5 years). I
2
Department of Clinical Sciences of Companion Animals, University of Utrecht, P.O. Box 80.154, 3508 TD Utrecht, The Netherlands. Department of Veterinary Pathology, University of Utrecht, The Netherlands.
THE VETERINARY QUARTERLY, VOL. 12, No. I, JANUARY 1990
.
29
Bacterial cultures
Duplicate samples of exudate from skin areas with features of GSP were collected with sterile swabs from the dogs of Group A. Following disinfection with methyl alcohol the exudate was squeezed out of the lesions. Duplicate bacterial samples were taken from the dogs of Group B with sterile cotton swabs from the surface of the skin in the neck region, avoiding contact with the haircoat. Bacteria were cultured on 5% horse-blood agar plates. Pure CPS cultures were placed on Dorset media to preserve them for further identification. The bacteria were identified using a commercially available kit (API Staph-Ident System S.A., 38390 Montalieu Vercieu, France) after being reactivated in serum broth or on horseblood (5%) agar plates. Skin tests A commercially available crude extract of S. intermedius (ARTU Biologicals N.V., Lelystad, The Netherlands) was used for the skin tests. To determine the skin threshold concentration of the S. intermedius extract, the following concentrations were used in the dogs of Group C (n = 10): 10 million, 100 million and 1,000 million bacteria per ml (Mb/m1).
Ten dogs of Group A were injected intradermally with the skin threshold concentration of the S. intermedius extract. The test was performed as described by Willemse and Van den Brom (17). The results were interpreted at 15 and 30 minutes and at 1, 2, 4, 6, 8, 24, 48 and 72 hours after the injection. Skin-test reactivity was considered positive if the wheal exceeded the average of the wheal diameter of the histamine solution and the diluent control. Histopathology
From five dogs of Group A and six dogs of Group C punch biopsies (6 mm 0) were collected
from skin-test sites under local anaesthesia, using a 1% lidocain solution. Following injection of the skin threshold concentration of the S. intermedius extract, biopsies were taken at t = 0, 15 and 30 minutes, and at 1, 2, 4, 8, 24, 48 and 72 hours. All biopsies were
fixed in 10% neutral buffered formalin and embedded in paraffin. Six-micrometer sections were cut and stained with haematoxylin-eosin (HE). Some specimens were also stained with Giemsa stain. Each biopsy specimen was interpreted histologically using a grading scale to indicate the degree of cellular infiltration (mild, moderate, or severe) for each cell type observed: neutrophils, eosinophils, histiocytes, lymphocytes, plasma cells and mast cells. RESULTS
Bacterial cultures and further identification
In Group A, CPS were cultured from the exudates of all dogs. However, after being preserved on the Dorset media, cultures from 13 GSP dogs could not be reactivated and as a result were lost for further identification. In the remaining 13 dogs the API Staph-Ident System identified S. aureus three times and S. intermedius five times. Identification was impossible in five cases due to a code not fitting in the API Staph-Ident profile index. In Group B, seven out of 13 cultures were positive. S. intermedius was identified once. Coagulase-negative staphylococci were cultured from the remaining six dogs. S. aureus was not identified. Skin-test reactivity In eight out of 10 dogs of Group C, skin reactivity to the S. intermedius antigen at the concentration of 1,000 Mb/ml was observed at 4 hours after injection. The wheals, ranging from 8 to 13 mm in diameter, disappeared within 8 hours after
the injection. Reactions to 10 and 100 Mb/ml concentrations and the diluent control were invariably negative. Skin reactivity to a 0.01% histamine solution occurred in all dogs after 15 minutes. On the basis of these results 100 Mb/ml (S100) was considered the skin threshold concentration and was therefore used as the test concentration for the GSP dogs. 30
THE VETERINARY QUARTERLY, VOL 12, No, 1, JANUARY 1990
.
Positive reactions to this test concentration were not observed in the 10 tested dogs of Group A. This group's reaction to the diluent control and the 0.01% histamine solution were similar to those observed in Group C. Histopathology I. Healthy dogs (Group C) Beginning at 2 hours, a mild intravascular and perivascular neutrophilic infiltration was observed, peaking at 4 to 8 hours (Table 1). At 24 hours this infiltrate had almost disappeared, while the interstitial reaction was still present. At 24 and 48 hours a mild-to-moderate mixed neutrophilic and mononuclear histiocytic and lymphocytic reaction was present in most dogs, especially in the interstitium. This reaction pattern had waned at 72 hours. A plasma-cell reaction was not observed. Table I. Histologic reaction pattern in six healthy dogs to S100. The dermal reaction at different times (hours) was graded as mild (mi), moderate (mo) or severe (se). Time after injection hours 00.15
Grade
Intravascular gra
Perivascular gra
lym
Interstitial his
gra
lym
his
mi
mo se
00.30
mi
1
mo se
01.00
mi
mo se
02.00
mi
3
2
2
4
3
3
I
1
3
2
mo se 04.00
mi
1
1
3
3
mo se
08.00
mi
3
mo
1
se
24.00
mi
1
6
1
mo se
48.00
mi
1
1
3
mo
I
1
2
2
1
2
2
2
se
72.00
mi
mo
1
1
3
se
(gra) granulocytes, (lym) lymphocytes, (his) histiocytes
THE VETERINARY QUARTERLY. VOL. 12, No. I, JANUARY 1990
31
GSP dogs
2.
A mild-to-moderate intravascular and perivascular granulocytic infiltrate was observed starting between 30 minutes and 1 hour after injection (Table 2); the infiltrate increased between 2 and 8 hours after injection and declined in the following hours (Figure 1). The same pattern was observed in the interstitium. Perivascular mononuclear reactions composed of lymphocytes and histiocytes were all graded as mild and were observed in small numbers from 24 hours till
72 hours (Table 2, Figure 2). The same mononuclear cell infiltration was observed in the interstitium. One dog's results have been placed in brackets because of the acanthosis in the epidermis, which hampered proper grading. . NtddW"""'
"T"'
ifft" -"c.""- V..-- --"-_....
*
A4
,
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.
00
al.°
dote ,14.4 # 0 .., ;
.
:,
'
':*:'''ru.
,
.
-,
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r it?, . ,
'
7
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-
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1
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i
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PIO
"I
I
/ l
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.
Its j
a.
I-
r
'111
4. L
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-,
4
, -44.
411
C.
ma 1
ee 1
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0
,
I.. att
i
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,..e V
S
t
1
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!:.
a
i
J
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S
,
A
O
a
It 1.
Igil b ; - 7 !, .
,
!
a
4,,
t
.
1- .,, ;
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71
-
Figure I. Intravascular and perivasular granulocytic reaction at 2 hours after injection of S100. Margination of intravascular leucocytes. H&E, obj. 40x.
7 --I
6
1
it.
--VT,
.
s
is
,
ff.:.
,,,.
,
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.
,... 4.
-..,
41.
4f
.
..
,
.
.. ,d,
k
Oaio
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g
Nipt: .
.5
4,,,
iv. ipst*.
4,,
Ilw
..
trio
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At 1 1,1s ,
4.
- .** I
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...4% h
,77. %IV A, of 71`,,ct frif.4441 tolp
la* ' Nor "eon.. p,'-i 4.,
OP" ...
[
."41...01.
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Ps
. a
. I i.b's or.
.1,,,:j6,4, r ,...,,,,,
4
411"' qh
T
,
,,,,2
i
1
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i.
, ) °44: *6 04 to$&14.4 .. iik . AI # - .0- .4r. v, Ats 4'; a
-
.-
..
...-.41
Figure 2. Lymphocytic, histiocytic and granulocytic perivascular infiltration at 24 hours after injection of S100. H&E, obj. 40x.
32
THE VETERINARY QUARTERLY, VOL. 12, No. I, JANUARY 1990
Table 2. Histologic reaction pattern in five GSP dogs to S100. The dermal reaction at different times (hours) was graded as mild (mi), moderate (mo) or severe (se).
Time after injection hours 00.15
Grade
Intravascular gra
Perivascular gra
lym
his
Interstitial
gra
lym
his
2
2
3
1
2
1
1
1
(I)
(1)
(1)
(1)
mi
1
mo se
00.30
mi
1
1
1
2
mi
3
3
1
mo
2
2
2
3
3
2
mo se
01.00
mi
mo se
02.00
se
04.00
mi
mo
1
se
08.00
mi
3
4
2
mo
1
se
24.00
1
mi
2
2
mo
1
1
se
48.00
mi
2
I
1
mo se
72.00
mi
1
mo se
1
(gra) granulocytes, (lym) lymphocytes, (his) histiocytes
DISCUSSION
S. intermedius, being the major representative of CPS, is now generally recognised
as one of the most common skin pathogens in the dog (2, 3). Its role in the
pathogenesis of GSP is uncertain. So far the identification of staphylococci in this condition has been confined to the characterisation of the coagulase activity. In this study, the use of the API Staph-Ident System revealed that the skin exudate of GSP dogs contains S. intermedius. This study also confirms the finding of others (2, 3) that S. intermedius can be found on unaffected canine skin. This suggests that S. intermedius itself does not play a crucial role in the pathogenesis of GSP. Bacterial hypersensitivity in man has been associated with recurrent bacterial skin
infections (5, 12). In atopic dermatitis in man, for instance, elevated levels of antistaphylococcal IgE or cell-mediated immune responses to S. aureus may trigger or perpetuate dermatitis (6). THE VETERINARY QUARTERLY, VOL
12, No. 1,
JANUARY 1990
33
Cell-mediated responses to S. aureus, characterised at 48 hours by a mononuclear cell infiltrate, have also been described in mice (7,15). In the dog, type III bacterial hypersensitivity reactions associated with the histologic features of vasculitis have been described (13). This study is the first in which skin threshold concentrations to S. intermedius extracts in the dog have been determined. No immediate or delayed type reactions were observed macroscopically from intradermal tests with the S100 antigen in the GSP group.
The histopathologic studies revealed essentially the same cell patterns in the healthy dogs and in GSP dogs. Mild inflammatory reactions (14) characterised by early polymorphonuclear infiltration followed by mild mononuclear infiltration at 24 and 48 hours were found in both groups of dogs. We consider this
histologic reaction pattern an aspecific inflammatory reaction that might be caused by all kinds of stimuli, including bacterial extracts (14). The microscopic observations of the skin tests in combination with the histopathologic features lead to the conclusion that hypersensitivity to staphylococcal antigens does not play a role in the pathogenesis of GSP. REFERENCES I. Baker E. Staphylococcal disease. Vet Clin North Am 1974; 4: 107-17. 2.
Berg J, Wendell D, Vogelweid C et a/. Identification of the major coagulase-positive Staphylococcus species of dogs as Staphylococcus intermedius. Am J Vet Res 1984; 45: 1307-9. 3. Biberstein E, Jang S, and Hirsch D. Species distribution of coagulase positive staphylococci in animals. J Clin Microbiol 1984; 19: 610-15. 4. Breen PT. Diagnosis and treatment of allergy-induced skin disorders. In: Proc Am Anim Hosp Assoc 1975; 42: 136-41.
5. Champion RH and Parish WE. Allergic diseases in the skin. 1183-1217. In: PGH Gell, RRA Coombs, and PJ Lachmann (Editors). Clinical aspects of immunology, 3rd ed., Blackwell Scientific Publications, Oxford, UK, 1975.
6. Dahl MV. Staphylococcus aureus and Atopic Dermatitis. Arch Dermatol 1983; 119: 840-6. 7. Easmon CSF and Glynn AA. Cell-Mediated immune responses in Staphylococcus aureus
infections in mice. Immunology 1975; 29: 75-85. a new species isolated from animals. Int J Syst Bacteriol
8. Hajek V. Staphylococcus intermedius,
1976; 26: 401-8. 9. Kloos W and Wolfshohl J. Identification of Staphylococcus species with the API Staph-Ident
System. J Clin Microbiol 1982; 16: 509-16.
10. Krogh HV and Kristensen S. A study of skin diseases in dogs and cats. Nord Vet Med 1976; 28: 459-63.
11.
Muller GH, Kirk RW, and Scott DW. Small Animal Dermatology. 3rd. ed., WB Saunders Co, Philadelphia, USA, 1983. 12. Rook A, Wilkinson DS, and Ebling FJG. Textbook of Dermatology. 3rd ed., Blackwell Scientific Publications, Oxford, UK, 1979. 13. Scott DW, MacDonald JM, and Schultz RD. Staphylococcal hypersensitivity in the dog. J Am Anim Hosp Assoc 1978; 14: 766-79.
14.
Slauson DO and Cooper BJ. Mechanisms of disease: A textbook of comparative general pathology. 1st ed., Williams and Wilkins, Baltimore, USA, 1982. 15. Taubler JH. Staphylococcal delayed hypersensitivity in mice. I Induction and in vivo demonstration of delayed hypersensitivity. J Immunol 1968; 101: 546-50. 16. Tizard I. Resistance to bacteria and related organisms. 193-208. In: An introduction to Veterinary Immunology, 2nd ed., WB Saunders Co, Philadelphia, USA, 1982. 17. Willemse A and Van den Brom WE. Evaluation of the intradermal allergy test in normal dogs. Res Vet Sci 1982; 32: 57-61. , 18. Wisselink MA, Willemse A, and Koeman JP. Deep pyoderma in the German shepherd dog. J Am Anim Hosp Assoc 1985; 21: 773-6. 19. Wisselink MA, Bernadina WE, Willemse A, and Noordzij A. Immunologic aspects of German Shepherd dog Pyoderma. J Vet Immunol Immunopathol 1988; 19: 67-77.
34
THE VETERINARY QUARTERLY, VOL 12, No. I, JANUARY 1990
German Shepherd dog Pyoderma (GSP) is a chronic deep pyoderma in pure-bred and cross-bred German shepherd dogs characterised by multiple deep-seated skin lesions. The most commonly associated bacteria are coagulase-positive staphylococci (CPS) (18). So far the role of CPS in the pathogenesis of GSP is not clear. In general, CPS are considered by far the most important pathogenic organisms in bacterial skin diseases in dogs (1, 10, 11). Recently three coagulase-positive or coagulase-variable species (S. aureus, S. intermedius and S. hyicus) have been determined on the basis of biotyping and DNA-homology studies (8, 9). Among these, S. intermedius has been identified as the major CPS associated with normal and infected canine skin (2, 3). In addition to infection, CPS may also induce hypersensitivity reactions. Such bacterial hypersensitivity has been describel in man and (laboratory) animals (5, 7, 12, 14, 16). In the dog the occurrence of types III and IV hypersensitivity reactions has been suggested (4, 13). In a previous report (19) bacterial hypersensitivity to CPS has been hypothesised as one of the predisposing factors for GSP. This hypothesis was tested by studying the histopathologic changes of skin-test sites at different time intervals, following injection of an extract of S. intermedius. MATERIALS AND METHODS
Animals
Group A consisted of 14 male (two castrated) and 12 female (four castrated) German
shepherd dogs with the clinical features of GSP (18). Their ages ranged from I to 12 years, with a median age of 6 years. Group B consisted of eight male and five female physically healthy German shepherd dogs kept under household conditions. Their ages ranged from 1 to 13 years (median: 8 years). Group C was comprised of six male and four female physically healthy dogs kept in the university clinic, including one bouvier, one beagle, one greyhound, one German shepherd and six mongrel dogs. Their ages ranged from 5 to 9 years (median: 7.5 years). I
2
Department of Clinical Sciences of Companion Animals, University of Utrecht, P.O. Box 80.154, 3508 TD Utrecht, The Netherlands. Department of Veterinary Pathology, University of Utrecht, The Netherlands.
THE VETERINARY QUARTERLY, VOL. 12, No. I, JANUARY 1990
.
29
Bacterial cultures
Duplicate samples of exudate from skin areas with features of GSP were collected with sterile swabs from the dogs of Group A. Following disinfection with methyl alcohol the exudate was squeezed out of the lesions. Duplicate bacterial samples were taken from the dogs of Group B with sterile cotton swabs from the surface of the skin in the neck region, avoiding contact with the haircoat. Bacteria were cultured on 5% horse-blood agar plates. Pure CPS cultures were placed on Dorset media to preserve them for further identification. The bacteria were identified using a commercially available kit (API Staph-Ident System S.A., 38390 Montalieu Vercieu, France) after being reactivated in serum broth or on horseblood (5%) agar plates. Skin tests A commercially available crude extract of S. intermedius (ARTU Biologicals N.V., Lelystad, The Netherlands) was used for the skin tests. To determine the skin threshold concentration of the S. intermedius extract, the following concentrations were used in the dogs of Group C (n = 10): 10 million, 100 million and 1,000 million bacteria per ml (Mb/m1).
Ten dogs of Group A were injected intradermally with the skin threshold concentration of the S. intermedius extract. The test was performed as described by Willemse and Van den Brom (17). The results were interpreted at 15 and 30 minutes and at 1, 2, 4, 6, 8, 24, 48 and 72 hours after the injection. Skin-test reactivity was considered positive if the wheal exceeded the average of the wheal diameter of the histamine solution and the diluent control. Histopathology
From five dogs of Group A and six dogs of Group C punch biopsies (6 mm 0) were collected
from skin-test sites under local anaesthesia, using a 1% lidocain solution. Following injection of the skin threshold concentration of the S. intermedius extract, biopsies were taken at t = 0, 15 and 30 minutes, and at 1, 2, 4, 8, 24, 48 and 72 hours. All biopsies were
fixed in 10% neutral buffered formalin and embedded in paraffin. Six-micrometer sections were cut and stained with haematoxylin-eosin (HE). Some specimens were also stained with Giemsa stain. Each biopsy specimen was interpreted histologically using a grading scale to indicate the degree of cellular infiltration (mild, moderate, or severe) for each cell type observed: neutrophils, eosinophils, histiocytes, lymphocytes, plasma cells and mast cells. RESULTS
Bacterial cultures and further identification
In Group A, CPS were cultured from the exudates of all dogs. However, after being preserved on the Dorset media, cultures from 13 GSP dogs could not be reactivated and as a result were lost for further identification. In the remaining 13 dogs the API Staph-Ident System identified S. aureus three times and S. intermedius five times. Identification was impossible in five cases due to a code not fitting in the API Staph-Ident profile index. In Group B, seven out of 13 cultures were positive. S. intermedius was identified once. Coagulase-negative staphylococci were cultured from the remaining six dogs. S. aureus was not identified. Skin-test reactivity In eight out of 10 dogs of Group C, skin reactivity to the S. intermedius antigen at the concentration of 1,000 Mb/ml was observed at 4 hours after injection. The wheals, ranging from 8 to 13 mm in diameter, disappeared within 8 hours after
the injection. Reactions to 10 and 100 Mb/ml concentrations and the diluent control were invariably negative. Skin reactivity to a 0.01% histamine solution occurred in all dogs after 15 minutes. On the basis of these results 100 Mb/ml (S100) was considered the skin threshold concentration and was therefore used as the test concentration for the GSP dogs. 30
THE VETERINARY QUARTERLY, VOL 12, No, 1, JANUARY 1990
.
Positive reactions to this test concentration were not observed in the 10 tested dogs of Group A. This group's reaction to the diluent control and the 0.01% histamine solution were similar to those observed in Group C. Histopathology I. Healthy dogs (Group C) Beginning at 2 hours, a mild intravascular and perivascular neutrophilic infiltration was observed, peaking at 4 to 8 hours (Table 1). At 24 hours this infiltrate had almost disappeared, while the interstitial reaction was still present. At 24 and 48 hours a mild-to-moderate mixed neutrophilic and mononuclear histiocytic and lymphocytic reaction was present in most dogs, especially in the interstitium. This reaction pattern had waned at 72 hours. A plasma-cell reaction was not observed. Table I. Histologic reaction pattern in six healthy dogs to S100. The dermal reaction at different times (hours) was graded as mild (mi), moderate (mo) or severe (se). Time after injection hours 00.15
Grade
Intravascular gra
Perivascular gra
lym
Interstitial his
gra
lym
his
mi
mo se
00.30
mi
1
mo se
01.00
mi
mo se
02.00
mi
3
2
2
4
3
3
I
1
3
2
mo se 04.00
mi
1
1
3
3
mo se
08.00
mi
3
mo
1
se
24.00
mi
1
6
1
mo se
48.00
mi
1
1
3
mo
I
1
2
2
1
2
2
2
se
72.00
mi
mo
1
1
3
se
(gra) granulocytes, (lym) lymphocytes, (his) histiocytes
THE VETERINARY QUARTERLY. VOL. 12, No. I, JANUARY 1990
31
GSP dogs
2.
A mild-to-moderate intravascular and perivascular granulocytic infiltrate was observed starting between 30 minutes and 1 hour after injection (Table 2); the infiltrate increased between 2 and 8 hours after injection and declined in the following hours (Figure 1). The same pattern was observed in the interstitium. Perivascular mononuclear reactions composed of lymphocytes and histiocytes were all graded as mild and were observed in small numbers from 24 hours till
72 hours (Table 2, Figure 2). The same mononuclear cell infiltration was observed in the interstitium. One dog's results have been placed in brackets because of the acanthosis in the epidermis, which hampered proper grading. . NtddW"""'
"T"'
ifft" -"c.""- V..-- --"-_....
*
A4
,
......
.
00
al.°
dote ,14.4 # 0 .., ;
.
:,
'
':*:'''ru.
,
.
-,
*411E,
r it?, . ,
'
7
/Is;
' !It:),
-
. ..!:..16.-z.
....,
1
duo fa . .:*,,.
i
...4
PIO
"I
I
/ l
..
.
Its j
a.
I-
r
'111
4. L
)
-,
4
, -44.
411
C.
ma 1
ee 1
..;:,
.74)...
0
,
I.. att
i
,lei 10
.r7,00 * It.Q.,
,..e V
S
t
1
_ _1:-:
!:.
a
i
J
1
S
,
A
O
a
It 1.
Igil b ; - 7 !, .
,
!
a
4,,
t
.
1- .,, ;
.3
71
-
Figure I. Intravascular and perivasular granulocytic reaction at 2 hours after injection of S100. Margination of intravascular leucocytes. H&E, obj. 40x.
7 --I
6
1
it.
--VT,
.
s
is
,
ff.:.
,,,.
,
I .1 4 I
..
-..' -2%.,....tA.
,
0* .;
,A.
. 4-....,
.
::..
,--'''
s' .
04. evii lv
**
.
,... 4.
-..,
41.
4f
.
..
,
.
.. ,d,
k
Oaio
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g
Nipt: .
.5
4,,,
iv. ipst*.
4,,
Ilw
..
trio
leintiotP ,d, ,,e viti, I
At 1 1,1s ,
4.
- .** I
"*"
...4% h
,77. %IV A, of 71`,,ct frif.4441 tolp
la* ' Nor "eon.. p,'-i 4.,
OP" ...
[
."41...01.
_ to --Ash OF.L
Ps
. a
. I i.b's or.
.1,,,:j6,4, r ,...,,,,,
4
411"' qh
T
,
,,,,2
i
1
0.'' 4/11.-- Tio-l.,4, 10.1. 1.7 Fl4 1,
i.
, ) °44: *6 04 to$&14.4 .. iik . AI # - .0- .4r. v, Ats 4'; a
-
.-
..
...-.41
Figure 2. Lymphocytic, histiocytic and granulocytic perivascular infiltration at 24 hours after injection of S100. H&E, obj. 40x.
32
THE VETERINARY QUARTERLY, VOL. 12, No. I, JANUARY 1990
Table 2. Histologic reaction pattern in five GSP dogs to S100. The dermal reaction at different times (hours) was graded as mild (mi), moderate (mo) or severe (se).
Time after injection hours 00.15
Grade
Intravascular gra
Perivascular gra
lym
his
Interstitial
gra
lym
his
2
2
3
1
2
1
1
1
(I)
(1)
(1)
(1)
mi
1
mo se
00.30
mi
1
1
1
2
mi
3
3
1
mo
2
2
2
3
3
2
mo se
01.00
mi
mo se
02.00
se
04.00
mi
mo
1
se
08.00
mi
3
4
2
mo
1
se
24.00
1
mi
2
2
mo
1
1
se
48.00
mi
2
I
1
mo se
72.00
mi
1
mo se
1
(gra) granulocytes, (lym) lymphocytes, (his) histiocytes
DISCUSSION
S. intermedius, being the major representative of CPS, is now generally recognised
as one of the most common skin pathogens in the dog (2, 3). Its role in the
pathogenesis of GSP is uncertain. So far the identification of staphylococci in this condition has been confined to the characterisation of the coagulase activity. In this study, the use of the API Staph-Ident System revealed that the skin exudate of GSP dogs contains S. intermedius. This study also confirms the finding of others (2, 3) that S. intermedius can be found on unaffected canine skin. This suggests that S. intermedius itself does not play a crucial role in the pathogenesis of GSP. Bacterial hypersensitivity in man has been associated with recurrent bacterial skin
infections (5, 12). In atopic dermatitis in man, for instance, elevated levels of antistaphylococcal IgE or cell-mediated immune responses to S. aureus may trigger or perpetuate dermatitis (6). THE VETERINARY QUARTERLY, VOL
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Cell-mediated responses to S. aureus, characterised at 48 hours by a mononuclear cell infiltrate, have also been described in mice (7,15). In the dog, type III bacterial hypersensitivity reactions associated with the histologic features of vasculitis have been described (13). This study is the first in which skin threshold concentrations to S. intermedius extracts in the dog have been determined. No immediate or delayed type reactions were observed macroscopically from intradermal tests with the S100 antigen in the GSP group.
The histopathologic studies revealed essentially the same cell patterns in the healthy dogs and in GSP dogs. Mild inflammatory reactions (14) characterised by early polymorphonuclear infiltration followed by mild mononuclear infiltration at 24 and 48 hours were found in both groups of dogs. We consider this
histologic reaction pattern an aspecific inflammatory reaction that might be caused by all kinds of stimuli, including bacterial extracts (14). The microscopic observations of the skin tests in combination with the histopathologic features lead to the conclusion that hypersensitivity to staphylococcal antigens does not play a role in the pathogenesis of GSP. REFERENCES I. Baker E. Staphylococcal disease. Vet Clin North Am 1974; 4: 107-17. 2.
Berg J, Wendell D, Vogelweid C et a/. Identification of the major coagulase-positive Staphylococcus species of dogs as Staphylococcus intermedius. Am J Vet Res 1984; 45: 1307-9. 3. Biberstein E, Jang S, and Hirsch D. Species distribution of coagulase positive staphylococci in animals. J Clin Microbiol 1984; 19: 610-15. 4. Breen PT. Diagnosis and treatment of allergy-induced skin disorders. In: Proc Am Anim Hosp Assoc 1975; 42: 136-41.
5. Champion RH and Parish WE. Allergic diseases in the skin. 1183-1217. In: PGH Gell, RRA Coombs, and PJ Lachmann (Editors). Clinical aspects of immunology, 3rd ed., Blackwell Scientific Publications, Oxford, UK, 1975.
6. Dahl MV. Staphylococcus aureus and Atopic Dermatitis. Arch Dermatol 1983; 119: 840-6. 7. Easmon CSF and Glynn AA. Cell-Mediated immune responses in Staphylococcus aureus
infections in mice. Immunology 1975; 29: 75-85. a new species isolated from animals. Int J Syst Bacteriol
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1976; 26: 401-8. 9. Kloos W and Wolfshohl J. Identification of Staphylococcus species with the API Staph-Ident
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Muller GH, Kirk RW, and Scott DW. Small Animal Dermatology. 3rd. ed., WB Saunders Co, Philadelphia, USA, 1983. 12. Rook A, Wilkinson DS, and Ebling FJG. Textbook of Dermatology. 3rd ed., Blackwell Scientific Publications, Oxford, UK, 1979. 13. Scott DW, MacDonald JM, and Schultz RD. Staphylococcal hypersensitivity in the dog. J Am Anim Hosp Assoc 1978; 14: 766-79.
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Slauson DO and Cooper BJ. Mechanisms of disease: A textbook of comparative general pathology. 1st ed., Williams and Wilkins, Baltimore, USA, 1982. 15. Taubler JH. Staphylococcal delayed hypersensitivity in mice. I Induction and in vivo demonstration of delayed hypersensitivity. J Immunol 1968; 101: 546-50. 16. Tizard I. Resistance to bacteria and related organisms. 193-208. In: An introduction to Veterinary Immunology, 2nd ed., WB Saunders Co, Philadelphia, USA, 1982. 17. Willemse A and Van den Brom WE. Evaluation of the intradermal allergy test in normal dogs. Res Vet Sci 1982; 32: 57-61. , 18. Wisselink MA, Willemse A, and Koeman JP. Deep pyoderma in the German shepherd dog. J Am Anim Hosp Assoc 1985; 21: 773-6. 19. Wisselink MA, Bernadina WE, Willemse A, and Noordzij A. Immunologic aspects of German Shepherd dog Pyoderma. J Vet Immunol Immunopathol 1988; 19: 67-77.
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