Isolation of Staphylococcus schleiferi from dogs with pyoderma

OBJECTIVE: To determine frequency with which Staphylococcus schleiferi could be isolated from dogs with pyoderma and antimicrobial susceptibility patterns of isolates that were obtained. DESIGN: Prospective study. ANIMALS: 54 dogs with a first (n = 14) or recurrent (40) episode of pyoderma. PROCEDURE: Specimens were obtained and submitted for bacterial culture. Isolates were identified as S schleiferi on the basis of growth and biochemical characteristics. Two isolates were submitted for DNA sequencing to confirm identification. Methicillin susceptibility was determined by means of disk diffusion with oxacillin-impregnated disks. RESULTS: 3 of 14 dogs examined because of a first episode of pyoderma and 12 of 40 dogs examined because of a recurrent episode of pyoderma were receiving antimicrobials at the time of specimen collection. Staphylococcus schleiferi was not isolated from any dog with first-time pyoderma but was isolated from 5 dogs with recurrent pyoderma that were not receiving antimicrobials at the time of specimen collection and 10 dogs with recurrent pyoderma that were receiving antimicrobials. Nine isolates were identified as S schleiferi subsp schleiferi, and 6 were identified as S schleiferi subsp coagulans. All S schleiferi subsp schleiferi isolates were resistant to methicillin, but only 2 S schleiferi subsp coagulans isolates were. Two methicillin-resistant isolates were also resistant to fluoroquinolones, and 1 isolate had intermediate susceptibility to fluoroquinolones. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that S schleiferi subsp schleiferi and S schleiferi subsp coagulans may be isolated from dogs with recurrent pyoderma. Although isolates from dogs with pyoderma were frequently resistant to methicillin, multiple drug resistance was uncommon.     

Isolation and characterization of staphylococci from external auditory meatus of dogs with or without otitis externa with special reference to Staphylococcus schleiferi subsp. coagulans isolates

Staphylococci were isolated from the external auditory meatus in 14 (48.3%) of 29 dogs affected with otitis externa (OE dogs) and 28 (68.3%) of 41 dogs without OE (non-OE dogs). Twenty-two OE isolates were identified as belonging to 12 species, and 42 non-OE isolates were identified as belonging to 13 species. The predominant species found in both OE and non-OE isolates were S. intermedius, and S. epidermidis. Thirty-eight (59.4%) of 64 isolates were resistant to one or more of the 17 antimicrobial agents tested. Resistance to PCG and ABPC was most frequent. S. schleiferi subsp. coagulans, a recent etiologic agent of canine OE, was isolated from OE and non-OE dogs. All of the 5 S. schleiferi subsp. coagulans isolates showed typical characteristics. No clear difference in the extracellular enzyme or toxin profiles, nor in the PFGE patterns, was demonstrated between the OE and non-OE isolates of S. schleiferi subsp. coagulans. A new PCR primer set specific for 16S rDNA was designed to identify strains of S. schleiferi subsp. coagulans. The amplified fragment was detected in all of the 5 isolates as well as the type strain GA 211 (=JCM 7470) and a reference strain GA 11, but was not detected in any strains of the related species, S. aureus, S. intermedius and S. hyicus. The PCR may allow a simple, rapid and precise identification of S. schleiferi subsp. coagulans, in addition to the standard tube test for free coagulase.     

Methicillin resistance of staphylococci isolated from the skin of dogs with pyoderma

OBJECTIVE: To determine the methicillin-resistant profile of staphylococcal isolates from the skin of dogs with pyoderma. ANIMALS: 90 dogs with pyoderma. PROCEDURE: Staphylococci isolated from dogs with pyoderma were tested for susceptibility to methicillin by use of a standard disk diffusion test with oxacillin disks. The DNA extracted from the isolates was tested for the mecA gene that encodes the penicillin-binding protein 2a (PBP2a) by use of a polymerase chain reaction (PCR) assay. The expression of PBP2a was determined with a commercial latex agglutination assay. Species of staphylococcal isolates were identified by use of morphologic, biochemical, and enzymatic tests. RESULTS: Most of the isolated staphylococci were methicillin-susceptible, coagulase-positive Staphylococcus intermedius isolates. Whereas only 2 of 57 S. intermedius isolates were resistant to methicillin, approximately half of the isolates had the mecA gene and produced PBP2a. Staphylococcus schleiferi was the second most common isolate. Widespread resistance to methicillin was found among S. schleiferi isolates. More coagulase-negative S. schleiferi isolates were identified with mecA gene-mediated resistance to methicillin, compared with coagulase-positive S. schleiferi isolates. CONCLUSIONS AND CLINICAL RELEVANCE: The latex agglutination assay for the detection of PBP2a expression coupled with the PCR assay for the mecA gene may provide new information about emerging antimicrobial resistance among staphylococcal isolates.     

Microbiological features of Staphylococcus schleiferi subsp. coagulans, isolated from dogs and possible misidentification with other canine coagulase-positive staphylococci

Staphylococcus schleiferi subsp. coagulans has only rarely been isolated and identified from the external auditory meatus of dogs suffering from external otitis. Its morphological and basic biochemical characteristics are of relatively little value for identification, as it phenotypically resembles another coagulase-positive staphylococci (CPS) and, consequently, may be easily misidentified as S. intermedius or even as S. aureus. In the present work, differentiation of S. schleiferi ssp. coagulans was therefore based on specific biochemical and genetic methods. All the strains were evaluated with the following commercial methods: Api Staph System (bioMérieux, Marcy l'Etoil, France), BBL Crystal Identification Systems (Gram-Positive ID Kit and Rapid Gram-Positive ID Kit; Becton Dickinson), and GEN-PROBE AccuProbe, Staphylococcus aureus identification test (bioMérieux). Gram-Positive ID System/GP database includes the broadest range of staphylococcal species and correctly identifies the majority of strains important in veterinary medicine. Therefore, it is an acceptable alternative to conventional methods for identification of canine staphylococcal isolates. Reliable differentiation of S. aureus from S. schleiferi ssp. coagulans and S. intermedius was feasible with AccuProbe for S. aureus, which gave positive results only for S. aureus; all other CPS tested were negative.     

Methicillin-resistant staphylococcal colonization in dogs entering a veterinary teaching hospital

Nasal, axillary and rectal swabs were collected from 193 dogs admitted to the Ontario Veterinary College Veterinary Teaching Hospital. Enrichment culture was performed and coagulase positive staphylococci were identified via standard methods. Methicillin-resistant Staphylococcus pseudintermedius was isolated from 4/193 (2.1%) dogs, and methicillin-resistant Staphylococcus aureus and methicillin-resistant Staphylococcus schleiferi subsp. coagulans were each isolated from 1/193 (0.5%) dogs. Methicillin-resistant Staphylococcus intermedius was not identified. All S. pseudintermedius isolates were unrelated on pulsed-field gel electrophoresis. Evaluation of the epidemiology of methicillin-resistant staphylococcal colonization is necessary to understand the apparent emergence of these strains and to develop appropriate control strategies.     

Screening for skin carriage of methicillin-resistant coagulase-positive staphylococci and Staphylococcus schleiferi in dogs with healthy and inflamed skin

Methicillin resistance rates of 41% for Staphylococcus aureus, 16% for S. intermedius, and 40% for S. schleiferi have recently been reported in canine patients. These were deemed to be reflective of referral and clinician-selection biases, which would imply significantly lower methicillin-resistant staphylococcal carriage rates in less-biased canine populations. In this study, swabs for bacterial culture were collected from five cutaneous sites on each of 50 healthy dogs and 59 dogs with inflammatory skin disease to determine prevalence of carriage and relative frequency of methicillin resistance in coagulase-positive staphylococci and S. schleiferi ssp. schleiferi. These were identified morphologically and by Gram's staining, catalase and coagulase testing, and biochemical speciation. Coagulase-positive staphylococci and S. schleiferi ssp. schleiferi were isolated from 88% (52 of 59) of affected dogs. Species identified in the culture-positive dogs were: S. aureus in 12%, S. intermedius (92%), S. schleiferi ssp. schleiferi (10%), and S. schleiferi ssp. coagulans (10%) with methicillin resistance rates of 17%, 8%, 20% and 20%, respectively. Coagulase-positive staphylococci were isolated from 74% (37 of 50) of healthy dogs: S. aureus (16%), S. intermedius (92%) and S. schleiferi ssp. coagulans (5%). Methicillin resistance rates were 0%, 3% and 50%, respectively. Of total methicillin-resistant isolates, 11 of 13 were positive for PBP2a via latex agglutination. Methicillin-resistant S. intermedius and S. schleiferi ssp. schleiferi isolates were all positive for the mecA gene via PCR. There was no significant difference in staphylococcal isolation or methicillin resistance between study groups. While present, methicillin-resistant coagulase-positive staphylococci are significantly less common in these less-biased populations than in the clinical isolates previously reported from this institution which provided the impetus for this study.     

Evaluation of aerobic bacteriologic culture of epidermal collarette specimens in dogs with superficial pyoderma

OBJECTIVE: To evaluate a method of aerobic bacteriologic culture of epidermal collarette specimens from dogs with superficial pyoderma and compare results with those for aerobic bacteriologic culture of abdominal skin specimens in healthy dogs. DESIGN: Prospective study. ANIMALS: 22 dogs with epidermal collarettes and 24 healthy dogs. PROCEDURE: Dry sterile cotton swabs were rolled across epidermal collarettes or hairless areas of abdominal skin in healthy dogs and submitted for aerobic bacteriologic culture. Hemolytic colonies of gram-positive-staining cocci were tested for catalase production, and if results were positive, a coagulase test was performed. Colonies with coagulase activity were tested for the ability to ferment mannitol. Antimicrobial susceptibility testing was performed on all Staphylococcus spp that were isolated. RESULTS: S. intermedius was isolated from collarettes in 18 of 22 dogs with superficial pyoderma but not from healthy dogs. Estimated sensitivity and specificity of the culture method were 81.8% and 100%, respectively. There were no significant differences in the ability to culture S. intermedius, the number of S. intermedius isolates without resistance to antimicrobials, and the number of S. intermedius isolates resistant to penicillin G when comparing dogs with superficial pyoderma for the first time and dogs with recurrent pyoderma, dogs that did or did not receive concurrent antimicrobials, and dogs with and without underlying allergic disease. CONCLUSIONS AND CLINICAL RELEVANCE: Bacteriologic culture of epidermal collarette specimens was a simple and reliable method for identification of S. intermedius in dogs with superficial pyoderma, regardless of history of pyoderma or current antimicrobial use.     

Surveillance of healthy cats and cats with inflammatory skin disease for colonization of the skin by methicillin-resistant coagulase-positive staphylococci and Staphylococcus schleiferi ssp. schleiferi

In this study, bacterial cultures were collected from five sites on each of 50 healthy cats and 48 cats with inflammatory skin disease (ISD), to determine prevalence of carriage and relative frequency of methicillin resistance in coagulase-positive staphylococci and Staphylococcus schleiferi ssp. schleiferi. Latex agglutination testing for penicillin-binding protein 2a (PBP2a) and pulsed field gel electrophoresis (PFGE) were performed on all methicillin-resistant (MR) isolates. Polymerase chain reaction (PCR) for the mecA gene was performed on MR S. intermedius and S. schleiferi isolates. Staphylococcal chromosomal cassette (SCCmec) typing was performed on all MR S. aureus isolates. Coagulase-positive staphylococci and S. schleiferi ssp. schleiferi were isolated from 24 of 48 cats with ISD: Staphylococcus aureus (14 of 24, 58%), Staphylococcus intermedius (11 of 24, 46%), Staphylococcus schleiferi ssp. schleiferi (1 of 24, 4%), and Staphylococcus hyicus (1 of 24, 4%). Prevalence of MR was 7% for S. aureus, 0% for S. intermedius, 100% for S. schleiferi ssp. schleiferi, and 0% for S. hyicus. Coagulase-positive staphylococci were isolated from 17 of 50 healthy cats: S. aureus (10 of 17, 59%), S. intermedius (11 of 17, 65%), and S. schleiferi ssp. coagulans (1 of 17, 6%). Prevalence of MR was 20% for S. aureus, 18% for S. intermedius, and 0% for S. schleiferi ssp. coagulans. All MR isolates were positive for PBP2a via latex agglutination. Methicillin-resistant S. intermedius and S. schleiferi ssp. schleiferi isolates were also positive for the mecA gene via PCR. Methicillin-resistant S. aureus isolates were identified as SCCmec type II. Results of PFGE indicated heterogeneity among isolates. There was no significant difference in staphylococcal isolation or methicillin resistance between study groups. While present, MR coagulase-positive staphylococci are significantly less common in these study populations.     

Heterogeneity of Staphylococcus pseudintermedius isolates from atopic and healthy dogs

Staphylococcus pseudintermedius is part of the normal canine flora but frequently causes pyoderma in canine atopic dermatitis (AD). This study aimed to determine whether particular S. pseudintermedius strains were associated with AD and/or pyoderma. Ninety‐six S. pseudintermedius isolates from the ear, nares, perineum and lesions of 21 atopic and 16 healthy dogs were lysed with proteinase K and digested with 40 U SmaI. Restriction products were separated using pulsed‐field gel electrophoresis (PFGE) with an Oxford S. aureus control and lambda‐ladder DNA concatomer markers. A dendrogram was constructed by the unweighted pair group method. All isolates showed a ≥56% similarity coefficient. Nine distinct PFGE clusters were identified, as follows: five from both atopic and healthy dogs; three from atopic dogs only; and one from healthy dogs only. Nine clusters were isolated from the nares, eight from the perineum, five from the ears and six from pyoderma lesions. There were no significant differences in the frequency of isolation from atopic or healthy skin, body sites or infected lesions for any of the clusters. Two of six healthy dogs and 18 of 20 atopic dogs with multiple isolates had closely related isolates (less than three band differences) at more than one sampling site. Isolates from pyoderma lesions were closely related to at least one mucosal isolate in 11 of 16 dogs. Staphylococcus pseudintermedius isolates appear to be heterogeneous, and colonization or infection of atopic skin was not associated with any particular strain or cluster of strains.     

A real-time PCR assay to detect the Panton Valentine Leukocidin toxin in staphylococci: screening Staphylococcus schleiferi subspecies coagulans strains from companion animals

Recent reports suggest that methicillin-resistant strains of Staphylococcus schleiferi subspecies coagulans are now commonly isolated from dogs. Given the association of a potentially mobile SCCmec type IV element with lysogenic phage-encoded Panton Valentine Leukocidin (PVL) toxin genes in community-acquired methicillin-resistant Staphylococcus aureus strains we hypothesized that methicillin-resistant S. schleiferi ssp. coagulans strains may also encode PVL toxin genes. Forty S. schleiferi ssp. coagulans strains isolated from companion animals were studied. Susceptibility to oxacillin was determined by broth microdilution and all isolates were screened by PCR for the presence of the mecA gene. SCCmec typing was performed on 14 isolates. A real-time PCR assay was developed for the detection of the PVL genes using a SmartCycler. Pulsed-field gel electrophoresis (PFGE) was performed to determine whether S. schleiferi ssp. coagulans strains were homogeneous. Twenty-eight of the 40 isolates (70%) were resistant to oxacillin and 26/28 possessed the mecA gene by PCR. SCCmec IV was identified in seven strains; the other seven isolates were not typable by this technique. All 40 strains were negative for the PVL toxin gene. PFGE showed a heterogeneous population and 13 different profiles were determined. In conclusion, this study showed that PVL toxin genes were not detected in a heterogeneous population of methicillin-resistant S. schleiferi ssp. coagulans strains isolated from companion animals.     

Antimicrobial resistance of Staphylococcus pseudintermedius isolates from healthy dogs and dogs affected with pyoderma in Japan

Staphylococcus pseudintermedius strains were isolated from healthy dogs and dogs with pyoderma in 2000-2002 and 2009. All the isolates from dogs with pyoderma in 1999-2000 and from healthy dogs in 2000-2002 and 2009 were susceptible to cefalexin and/or other cephalosporins and oxacillin. However, 7.1-12.5 and 11.4% of S. pseudintermedius isolates from dogs with pyoderma in 2009 were resistant to cephalosporins and oxacillin, respectively. All S. pseudintermedius isolates from dogs with pyoderma in 1999-2000 and those from healthy dogs in 2000-2002 were susceptible to fluoroquinolones; however, 50% of the S. pseudintermedius strains isolated from dogs with pyoderma in 2009 and 30% of the S. pseudintermedius strains isolated from healthy dogs in 2009 were resistant to fluoroquinolones. Of the 21 oxacillin-resistant S. pseudintermedius (MRSP) isolates, 11 carried SCCmec type V and 10 carried hybrid SCCmec types II-III. Staphylococcus pseudintermedius strains that were resistant to only one of three fluoroquinolones had a mutation in the quinolone resistance determination region of grlA, whereas S. pseudintermedius strains that were resistant to two or more fluoroquinolones had mutations in the quinolone resistance determination regions of both grlA and gyrA.     

In vitro antimicrobial resistance of staphylococci isolated from canine urinary tract infection

This study determined the diversity of species and antimicrobial resistance of staphylococci isolated from dogs with a presumptive diagnosis of urinary tract infection (UTI). Urine samples from 348 dogs with clinical signs of UTI, according to clinical examination and urinalysis, were processed for isolation of Staphylococcus. Colonies in pure culture were identified by biochemical reactions and tested for susceptibility to 15 antimicrobials. Seventy isolates of staphylococci were obtained (20.1%). Staphylococcus pseudintermedius was the most frequent species (32.8%), followed by S. epidermidis (18.6%), S. simulans (15.7%), S. schleiferi schleiferi (11.4%), S. aureus (11.4%), S. schleiferi coagulans (7.2%) and S. saprophyticus (2.9%). All the isolates were resistant to at least 1 drug and 77.1% were multiresistant. The study reports the alarming antimicrobial resistance of members of the Staphylococcus genus isolated from canine UTI and highlights the importance of coagulase-negative staphylococci in its etiology.     

Antimicrobial Susceptibility and rRNA Gene Restriction Patterns among Staphylococcus intermedins from Healthy Dogs and from Dogs Suffering from Pyoderma or Otitis Externa

A total of 60 Staphylococcus intermedins strains from dogs were investigated by their sensitivity to various antibiotics (50 strains) and by their rRNA gene restriction patterns (ribotyping) (60 strains). Fifteen isolates were from healthy dogs, 9 with otitis externa, and 36 with pyoderma, including 10 strains from a previous study. Sixty per cent of the 50 strains tested for antibiotic susceptibility demonstrated resistance to penicillin, 24% to spiramycin, 20% to tetracycline, 16% to chloramphenicol, and 2% to fucidic acid. All isolates were susceptible to amoxycillin with clavulanic acid, enrofloxacin, and sulphonamides with trimethoprim. There were no significant differences in antimicrobial susceptibility patterns observed among isolates from pyoderma, otitis externa or healthy dogs. Among the 60 strains studied by ribotyping, 10 different ribotypes were identified: 6 different ribotypes among isolates from otitis externa, 8 among isolates from pyoderma, and 5 among isolates from healthy dogs. One ribotype (profile C) was dominant among the isolates from healthy dogs while another ribotype (profile A) was dominant among strains from dogs suffering from pyoderma. This profile was not demonstrated in any of the strains from healthy dogs. From 5 different dogs suffering from pyoderma, 2 different clones were demonstrated based on their plasmid profile and antibiogram. In these dogs 1 of the clones always belonged to ribotype A. The results concerning strains of S. intermedins isolated from furunculosis suggest the existence of distinct subpopulations with different pathogenicity to dogs.     

Prevalence of oxacillin- and multidrug-resistant staphylococci in clinical samples from dogs: 1,772 samples (2001-2005)

OBJECTIVE: To determine whether resistance to oxacillin and other antimicrobials in 3 Staphylococcus spp commonly isolated from dogs increased from 2001 to 2005. DESIGN: Retrospective case series. SAMPLE POPULATION: 1,772 clinical samples of various types obtained from dogs examined at the University of Tennessee Veterinary Teaching Hospital or at regional veterinary hospitals and submitted to the bacteriology and mycology laboratories associated with the teaching hospital. PROCEDURES: Samples were submitted by attending veterinarians to the bacteriology and mycology laboratories for routine aerobic microbial culture. Identification and antimicrobial susceptibility procedures were performed on all isolates. Susceptibility reports for each antimicrobial and Staphylococcus spp were determined from aggregate electronically archived test results. Oxacillin and multidrug resistance for Staphylococcus intermedius was analyzed by reviewing disk diffusion zone measurements. RESULTS: Oxacillin resistance increased among S. intermedius isolates during the past 5 years, and the increase was associated with multidrug resistance. In 2005, 1 in 5 Staphylococcus spp isolates from canine clinical samples was resistant to oxacillin. The most common staphylococcal species isolated were S. intermedius (n = 37), Staphylococcus schleiferi (21), and Staphylococcus aureus (4), and frequencies of oxacillin resistance in isolates of these species were 15.6%, 46.6%, and 23.5%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Veterinarians should be aware of the potential for empiric drug treatment failures in instances where Staphylococcus spp infections are common (eg, pyoderma). Judicious use of bacterial culture and susceptibility testing is recommended.     

Identification and antimicrobial susceptibility of bacteria and yeasts isolated from healthy dogs and dogs with otitis externa

The bacterial and fungal flora of the external ear canal of dogs with otitis externa and of healthy dogs were studied. The most frequently isolated microorganism from otitic ears was Staphylococcus intermedius (58.8%), followed by Malassezia pachydermatis (30.9%), Streptococcus canis (29.9%), Proteus spp. (14.4%) and Escherichia coli (10.3%). A statistical analysis of our results showed that the prevalence of these microorganisms is significant in dogs with otitis externa. Furthermore, the antimicrobial susceptibility patterns of isolated strains were determined. Majority of all bacterial isolates were most susceptible to gentamicin. Malassezia pachydermatis, the most prevalent yeast in this study, showed an excellent level of susceptibility to all antifungal agents tested.     

Malassezia pachydermatis isolated from normal and diseased external ear canals in dogs: a comparative analysis

To investigate the role of Malassezia pachydermatis as a pathogenic agent in canine otitis, a comparative analysis of isolates from normal and diseased external ear canals in dogs was undertaken. Specimens were collected from the ears of dogs with unilateral or bilateral otitis and from healthy dogs. Mycological analysis was by direct microscopy and fungal culture on Sabouraud's dextrose agar and Dixon's agar. Of the otitis specimens, 63.7% showed typical Malassezia cells on cytological examination. In samples taken from the healthy ears of dogs with unilateral otitis, only 21.43% (P<0.05) showed evidence of Malassezia. M. pachydermatis was identified cytologically and culturally in 57.53% (P<0.05), 14.29% and 30.0% of samples from the ears of dogs with otitis, from the healthy ears of dogs with unilateral otitis and from the ears of healthy dogs with no otitis. In the group with otitis associated with M. pachydermatis, the poodle was the most common breed (39.29%; P<0.05), whereas in the group without otitis, the German Shepherd breed was prominent (although this observation was not statistically significant). In both groups, the majority of dogs with M. pachydermatis were aged between 1 and 3 years (P<0.05). The higher incidence of M. pachydermatis isolated from the ears of dogs with otitis externa suggests a putative pathogenic role of this yeast in this condition.     

Microbial flora and antimicrobial susceptibility patterns from dogs with otitis media

This study evaluated the middle ear microbial flora and antimicrobial susceptibility patterns from dogs with otitis media. Antimicrobial susceptibility patterns were obtained on 164 micro-organisms from 107 ears of 82 dogs with otitis media that presented to Louisiana State University over a 12-year period. The most common micro-organisms isolated were Staphylococcus intermedius (26.8%), Pseudomonas aeruginosa (23.2%), β-haemolytic streptococcus (12.8%), Proteus spp. (11.0%) and Staphylococcus epidermidis (8.5%). Antimicrobial susceptibility testing was performed on all isolates employing the disk diffusion method (Kirby-Bauer). Minimum inhibitory concentration (MIC) testing for enrofloxacin and ciprofloxacin was performed on 10 of the 18 P. aeruginosa isolates found to be resistant to enrofloxacin on Kirby-Bauer. Of these isolates, 6 (60%) were susceptible, 1 (10%) was intermediate and 3 (30%) were resistant to enrofloxacin. Eight of these isolates (80%) were susceptible to ciprofloxacin, 1 (10%) showed intermediate susceptibility and 1 (10%) was resistant.     

Antimicrobial susceptibility and mechanism of resistance to fluoroquinolones in Staphylococcus intermedius and Staphylococcus schleiferi

The objective of the present study was to determine the antimicrobial susceptibility of 136 canine isolates of Staphylococcus intermedius and 10 canine isolates of S. schleiferi subspecies coagulans to 16 fluoroquinolones (FQs), and to investigate the mechanisms of resistance in the nonsusceptible isolates. Of the 136 of S. intermedius tested 98.5% were susceptible to all 16 FQs whereas only 40% of the 10 isolates of S. schleiferi subspecies coagulans were susceptible. Two isolates of S. intermedius and six isolates of S. schleiferi, were found to be resistant to 13 out of 16 FQs, while they retained their susceptibility to fourth generation FQs such as gatifloxacin, moxifloxacin and trovafloxacin. Sequencing of the quinolone-resistance determining regions of gyrA and grlA genes showed that in S. intermedius, dichotomous resistance to FQs was associated with the occurrence of one alteration in GyrA-84 and one in GrlA-80, while in S. schleiferi the same pattern of resistance was observed in isolates showing these changes only in gyrA. This study is the first to screen FQs of the second, third and fourth generation for antimicrobial resistance in clinical isolates of S. intermedius and S. schleiferi of canine origin, and to describe mutations in gyrA and grlA associated with FQ resistance in these bacterial species.     

Adherence of Staphylococcus intermedius to canine corneocytes in vitro

This study investigated the in vitro adherence of Staphylococcus intermedius to canine corneocytes, collected from a healthy dog using double-sided adhesive tape. Adherence was shown to depend on duration (P < 0.001) and temperature of incubation (P < 0.001) and the concentration of bacteria (P < 0.001). Isolates of S. intermedius from lesions of pyoderma were not generally more adherent to healthy canine skin than were isolates from healthy dogs. Significant differences in adherence were demonstrated between individual isolates within both groups (P < 0.001). The study suggests that among S. intermedius there is no correlation between virulence and adherence to canine corneocytes in vitro. The finding may be important for the potential use of avirulent variants of S. intermedius as antagonistic strains against canine pyoderma. However, more studies are needed to compare the adherence of the isolates to skin cells obtained from dogs with diseases predisposed to pyoderma.     

Genotypic versus phenotypic identification of staphylococcal species of canine origin with special reference to Staphylococcus schleiferi subsp. coagulans

A comparative study was performed to examine the respective accuracy of 16S rDNA sequencing and of the commercial biochemical assay ID32 STAPH (bioMérieux, Marcy l'Etoile, France) in the identification of 232 staphylococcal samples representing 20 species and subspecies isolated from 367 dogs. Notable differences in species distribution were observed by comparing genotypic and phenotypic data. Partial sequencing of 16S rDNA resulted in an unambiguous identification of 226 (97.4%) of the isolates, whereas the phenotypic approach resulted in a correct diagnosis of 162 (69.8%) of the isolates. Statistical agreement between genotypic and phenotypic identification of staphylococci was substantial (Kappa coefficient of 0.6-0.8) for Staphylococcus aureus, S. hominis, S. warneri, S. cohnii subsp. urealyticus, and S. simulans, and "almost perfect" (Kappa coefficient of 0.8-1) for S. intermedius, S. epidermidis, S. equorum, S. haemolyticus, S. sciuri, and S. kloosi. No agreement above that expected by chance (Kappa coefficient=0) was observed for S. schleiferi subsp. coagulans, which was either confounded with S. intermedius and S. capitis, or categorized as unacceptable by the biochemical assay. Given the growing importance of this pathogen in veterinary medicine and its frequent misidentification with related staphylococci, a PCR-RFLP approach producing a S. schleiferi-specific restriction profile was developed. This fast and reliable assay represents a valuable tool in assisting in the monitoring of this pathogen.