veterinary

microbiology
ELSEVIER Veterinary Microbiology 39 (1994) 255-260

Characterization of the most frequently encountered

Staphylococcus sp. in cats.
Shizunobu Igimi a'*, H. Atobe b, Y. Tohya u, A. Inoue c, E. Takahashi b,
S. K o n i s h i b
"Departmentof Biomedical Food Research, National Institute of Health, 1-23-! Toyama, Shinjuku-Ku
Tokyo 162, Japan
bDepartmentof VeterinaryMicrobiology, Faculty of Agriculture, The Universityof Tokyo, 1-1-I Yayoi,
Bunkyo-ku, Tokyo, Japan
CDepartmentof Molecular Biology, Toho UniversitySchool of Medicine, Ota-ku, Tokyo, Japan
(Accepted 26 August 1993)

Abstract

Ninety three staphylococci isolated from clinical specimens from cats were characterized and iden-
tified. Because the biochemical characteristics of Staphylococcusfelis were very similar to those of
Staphylococcus simulans, results were submitted to numerical analysis and DNA homology. Forty-
two isolates (45%) were identified as S. felis, and 4 isolates (4%) as S. simulans. The other species
identified, in order of their frequency were, 12 Staphylococcus aureus (13%), 9 Staphylococcus
intermedius (10), 6 Staphylococcus sciuri (6), 6 Staphylococcus epidermidis (6), 2 Staphylococcus
haemolyticus (2), 2 Staphylococcus xylosus (2), 1 Staphylococcus capitis (1), 1 Staphylococcus
equorum ( 1 ), 1 Staphylococcus gallinarum ( 1 ) and 1 Staphylococcus lentus (1).

Key words."Staphylococcus; Characterization; Cat

1. Materials and methods

Staphylococci are isolated alone or in c o m b i n a t i o n with other bacteria from various

clinical specimens of cats. However, staphylococci do not generally appear to cause any
major specific diseases in cats. Early literature contained a few reports of staphylococci
from cat lesions ( B a b a et al., 1981 ; Biberstein et al., 1984; Cox et al., 1985). The sporadic
nature of staphylococcal infection in cats can perhaps be explained by the fact that the major
pathogenic staphylococci Staphylococcus aureus, Staphylococcus intermedius, and Staph-
ylococcus hyicus are not indigenous to cats. Devriese et al. ( 1 9 8 4 ) found that Staphylococ-

Corresponding author: S. Igimi, Tel.: ( + 81 ) 3-5285- 1111; Fax: ( + 81 ) 3-5285-1150.

0378-1135/94/$07.00 © 1994 Elsevier Science B.V. All rights reserved

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256 s. lgimi et al. / Veterinary.Microbiology 39 (1994) 255-260

cus simulans was the most common Staphylococcus sp. present on both carrier sites and
lesions in cats. Though they had taken into account all staphylococcal species reported
before 1984, since then 8 additional staphylococcal species have been defined (Freney et
al., 1988; Hajek et al., 1992; Igimi et al., 1989; 1990; Schleifer et al., 1984; Varaldo et al.,
1988). Further, Igimi et al. (1989) isolated a new species, Staphylococcusfelis from cats.
As biochemical reactions of S. felis are similar to those of S. simulans, it is difficult to
distinguish between S. felis and S. simulans only by their biochemical properties. In this
paper, we attempt to clarify which species is most frequently isolated from cats by employing
the numerical method and the DNA-DNA hybridization in addition to the conventional
identification method.
All staphylococci which were isolated from cat patients at the Veterinary Hospital, Faculty
of Agriculture, The University of Tokyo, between 1982 and 1990 were identified according
to the Tables of characteristics listed in Bergey's Manual of Systematic Bacteriology
(1986). A total of 93 isolates, all of which showed > 20 colonies on the primary plate,
were examined. Samples of urinary tract infections were obtained from sediments, and of
the other anatomic sites by means of direct swabs. The isolates were cultured in Trypticase
soy agar (BBL Microbiology Systems, Cockeysville, Md.) supplemented with 5% defibri-
nated horse blood. The following biochemical characteristics were determined as previously
described (Igimi et al., 1989); cell and colony characteristics; the production of catalase,
oxidase, coagulase, clumping factor, heat-stable nuclease, hyaluronidase, arginine dehydro-
lase, phosphatase, urease, and acetoin; anaerobic growth in thioglycolate medium; hemol-
ysis; nitrate reduction; acid from carbohydrates; antibiotic susceptibilities; and enzymatic
reactions.
Forty isolates (43%) belonging to 9 species were identified according to the Tables of
characteristics listed in Bergey's Manual. All these isolates displayed the typical character-
istics of each species. Forty six of the remaining 53 isolates coincided with the characteristics
of S. simulans. However, as mentioned by Devriese et al. (1984), most isolates from cat
lesions have positive phosphatase reactions, despite of the fact that S. simulans is normally
phosphatase negative or weakly positive (Kloos and Schleifer, 1975).
In addition to this manual, we considered the following new species descriptions: S.
arlettae, S. equorum, S. kloosii ( Schleifer et al., 1984); S. lugdunensis, S. schleiferi ( Freney
et al., 1988); S. delphini (Varaldo et al., 1988); S.felis (Igimi et al., 1989); S. schleiferi
subsp, coagulans (Igimi et al., 1990); and S. muscae (Hajek et al., 1992). After we added
the 8 new species, we identified 42 isolates as S.felis, 4 as S. simulans and 1 as S. equorum.
As the isolates identified as either S. felis or S. simulans closely resembled each other using
the abore test, we tried to differentiate them by bacitracin resistance, acid production from
mannose, and phosphatase reactions. However, we did not wish to identify the isolates into
either of the two species at present by the following reasons. Bacitracin susceptibility, a
useful property for distinguishing S. felis from S. simulans (Igimi et al., 1989), requires
circumspection due to the possibility of resistant bacteria being present. Secondly, acid
from mannose is not always a suitable characteristic for differentiation of the two species.
The S. felis produced acid from mannose, and the S. simulans for dogs and cats did not
(Igimi et al., 1989). However, Kloos and Schleifer ( 1975 ) reported that many strains of S.
simulans ( > 60%), which were isolated from man, were mannose positive. Finally, while
S. felis is phosphatase positive (Igimi et al., 1989) and the S. simulans is normally phos-
 S. Igimi et al. /Veterinary Microbiology 39 (1994) 255-260 257

phatase negative or weakly positive (Kloos and Schleifer (1975), a phosphatase positive
S. simulans has been described by Sloan et al. (1982).
Because of these uncertainties, four isolates which had been identified as S. simulans and
6 randomly selected isolates of S. felis were verified by numerical analysis. Each isolate
was characterized by 70 characteristics, and analyzed by numerical methods. Tested char-
acteristics are 33 biochemical reactions listed in Bergey's Manual (1986), 19 enzymatic
activities detected by API zyme (API System S.A.), and a futher 18 characteristics as
follows: catalase, bacitracin susceptibility, alpha-hemolysin, beta-hemolysin, glucose, glyc-

(~ Similarity)

Strain 100 90 80 70 60 50

S. simulans JCM 2424T

No.29

No.262

No.87

No.144

S. f e l l s GD521T - [

No.359

No.104

No.121

No.36

No.141

No.106

S. a u r e u s CCM 885T
L l
S. intermedius JCM 2422T-

S. cohnii JCM 2417T

S. xylosus JCM 2418T

100 90 80 70 60 50

Fig. 1. Relatedness among Staphylococcus isolates and type strains by numerical analysis with 70 biochemical
reactions.
258 S. l g i m i et al. / V e t e r i n a r y M i c r o b i o l o g y 39 (1994) 255-260

Table 1
Identification of Staphylococcus sp. isolated from cats

Species No. of strains isolated Others

Total Urinary Otitis Dermatitis Rhinitis Abscesses

tract extema
Infection

S. felis 42 10 15 6 5 3 Conjunctivitis( 3 )
S. aureus 12 3 2 1 1 1 Mastitis(2), Lymphadenitis( 1), Arthritis( 1 )
S. intermedius 9 4 2 1 1 1
S. sciuri 6 3 1 2
S. epidermidis 6 3 3
S. simulans 4 3 1
S. haemolyticus 2 1 l
S. xvlosus 2 1 1
S. cap#is 1 1
S. gallinarum 1 1
S. lentus 1 1
S. equorum 1 1
Unidentified 6 3 2 Colpitis ( 1 )

Totals 93 34 21 13 11 6 8

erol, melibiose, rhamnose, glycogen, sorbitol, erythritol, inositol, adonitol, dulcitol, alpha
methylglucoside, alpha methylmannoside, dextline, amygdalin. All characteristics were
coded as binary data for cluster analysis. The value 1 was assigned if the characteristic was
present or positive. Multistate characteristics, such as a range of tolerance to NaC1 concen-
trations, were regarded as in dividual tests and recorded as individual twostate responses.
Numbers were analyzed by calculating similarity matrices using the simple matching coef-
ficient of Sokal and Michener (1958). Clustering was carried out by single linkage ( Sneath
and Sokal, 1973) using a calculating program made on PC-9801 model VM (NEC, Tokyo,
Japan) by M. Yamada, Faculty of Agriculture, Tokyo University of Agriculture and Tech-
nology, which we modified partly.
As shown in Fig. 1, all isolates (No. 29, No. 87, No. 144 and No. 262) identified as S.
simulans and the type strain of S. simulans shared > 90% similarity values among all strains
in one cluster. The 6 selected isolates (No. 36, No. 104, No. 106, No. 121, No. 141 and No.
359) identified as S. felis, and the type strain of S. felis shared > 88% similarity values
among all strains in the other cluster. Each cluster shared about 78% similarity values.
In addition, DNA-DNA hybridization experiments were performed as described previ-
ously (Igimi et al., 1989). The DNA relatedness results obtained with labeled DNA from
S. felis GD521 v are as follows. All four isolates (No. 36, No. 104, No. 121 and No. 141)
of S. felis tested exhibited high levels of homology ( 81 to 93 %) with type strain of S. felis,
whereas all two isolates (No. 29 and No. 144) of S. simulans tested were rather low (6 and
4%).
These results of numerical analysis and DNA homology supported our previous classi-
fication of these strains. Bacitracin resistance, acid production from mannose, and phos-
 s. lgimi et al. / Veterinary.Microbiology 39 (1994) 255-260 259

phatase reactions are useful properties for distinguishing S. felis from S. simulans. Finally,
we identified 93 staphylococci as shown in Table 1. Three p r e d o m i n a n t species accounted
for 68% of the 93 isolates: S.felis ( 4 5 % of isolates), coagulase-positive S. aureus ( 1 3 % ) ,
and S. intermedius ( 1 0 % ) . According to the report of Devriese et al. ( 1 9 8 4 ) , 38% of 79
isolates from cats' lesions were coagulase positive Staphylococcus and were identified as
24 S. aureus and 6 S. intermedius. C o m p a r e d with their results, the ratios of S. aureus to S.
intermedius and coagulase-positive to coagulase-negative Staphylococcus are rather low.
Devriese et al. also reported that 30% of Staphylococcus sp. from cat carrier sites revealed
the same biochemical patterns as those from lesions and were identified as "S. simulans".
The present study showed that all S. felis are phosphatase positive, and S. simulans from
cats are negative. Because "S. simulans" isolated by Devriese et al. produced phosphatase,
we think most of their isolates were probably S. felis. The frequency of occurrence by
anatomic site are also listed in Table 1. As can be seen Table 1, all isolates originated from
rather superficial lesions, most of which are prone to superficial contamination. In this type
of lesion, the interpretation of the etiological role of the isolates is difficult. In our study,
growth on the primary plates was rated as heavy ( > 100 colonies) with 28 out of the 43 S.
felis, and 19 S.felis were recovered in pure culture.

2. Acknowledgements

The authors thank Prof. T. Mikami, Department of Veterinary Microbiology, Faculty of

Agriculture, The University of Tokyo, for a critical review of the manuscript. W e are also
grateful to the veterinarians who provided samples.

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