Carriage of Staphylococcus schleiferi from canine otitis externa: antimicrobial resistance profiles and virulence factors associated with skin infection

Abstract

The recent emergence of Staphylococcus schleiferi in dogs with otitis externa or skin and soft tissue infections has become a significant zoonotic issues. In the current study, we investigated 1) the carriage rates of S. schleiferi among major staphylococci in healthy dogs and dogs with otitis externa, 2) antibiotic susceptibility profiles of S. schleiferi, particularly methicillin resistance (MR), and 3) virulence factors associated with skin and soft tissue infections such as ability to form biofilm, resistance to cationic antimicrobial peptides (CAMPs), and carriage of staphylococcal enterotoxin genes. Among the 21 S. schleiferi isolates, 5 isolates (24%) were determined to be methicillin-resistant (MRSS). Staphylococcal cassette chromosome mec (SCCmec) typing revealed the presence of SCCmec type V in 4 MRSS isolates and type VII in one MRSS. Higher levels of antibiotic resistance, especially multidrug resistance, were observed in MRSS isolates compared to the methicillin-susceptible S. schleiferi (MSSS) isolates. In addition, MRSS isolates exhibited enhanced ability to form biofilm under static condition and all the 5 MRSS isolates carried three or more enterotoxin genes. However, there were no significant differences in resistance to CAMPs between MRSS and MSSS isolates. These findings suggest that coagulase-negative S. schleiferi is becoming more prevalent in canine otitis externa cases. Our results also highlight the presence of multidrug-resistant MRSS isolates with enhanced biofilm production and carriage of multiple enterotoxins.

Keywords: Staphylococcal otitis externa; dogs; methicillin resistance; virulence factors.

Fig. 1. Antimicrobial susceptibility of S. schleiferi isolates from dogs with otitis externa (A) and frequency of multidrug resistance among the S. schleiferi isolates (B). The percentages indicate the rates of resistant isolates among the S. schleiferi isolates. Susceptibility assays were performed using the disc diffusion methods according to the 2017 Clinical and Laboratory Standards Institute guidelines [14].

AMP, ampicillin; CHL, chloramphenicol; ERY, erythromycin; ENR, enrofloxacin; GEN, gentamicin, KAN, kanamycin; RIF, rifampicin; SXT, trimethoprim-sulfamethoxazole; TET, tetracycline; MRSS, methicillin-resistant S. schleiferi; MSSS, methicillin-susceptible S. schleiferi.

Fig. 2. Biofilm formation of S. schleiferi isolates. The data was normalized to the wild-type S. aureus Newman strain. Data represent the means (± standard deviation) from three independent experiments.

MRSS, methicillin-resistant S. schleiferi; MSSS, methicillin-susceptible S. schleiferi; OT, dogs with otitis externa; HT, healthy dogs. ^*p < 0.05, ^†p < 0.01.

Fig. 3. In vitro susceptibility profiles of S. schleiferi strains to K9CATH (A) and LL-37 (B). In vitro cell survival assays were performed with K9CATH (3 μg/mL) and LL-37 (1 μg/mL) as described previously [2627]. The in vitro survival assays were performed at least three times in triplicate. Data represent means (± standard deviation).

MRSS, methicillin-resistant S. schleiferi; MSSS, methicillin-susceptible S. schleiferi; OT, dogs with otitis externa; HT, healthy dogs. ^*p < 0.01; ^†not significant.