Antimicrobial treatment of methicillin-resistant Staphylococcus pseudintermedius associated with canine wounds represents an important challenge. The aim of this study was to create a canine wound infection model, Lubbock Chronic Wound Biofilm (LCWB), with a focus on S. pseudintermedius, drawing inspiration from the established human model involving S. aureus. Methicillin-resistant S. pseudintermedius 115 (MRSP) and Pseudomonas aeruginosa 700 strains, isolated from dog wounds, were used to set up the LCWB at 24, 48 and 72h. The LCWBs were evaluated in terms of volume, weight, and microbial CFU/mg. The microbial spatial distribution in the LCWBs was assessed by SEM and CLSM imaging. The best incubation time for the LCWB production in terms of volume (3.38 cm3 ± 0.13), weight (0.86 gr ± 0.02) and CFU/mg (up to 7.05 x 106 CFU/mg ± 2.89 x 105) was 48h. The SEM and CLSM images showed a major viable microbial colonization at 48h with a non-mixed bacteria with a prevalence of MRSP on the surface and P. aeruginosa 700 in the depth of the wound. The obtained findings demonstrate the capability of S. pseudintermedius to grow together P. aeruginosa in the LCWB model, representing the suitable model to reproduce the animal chronic wound in vitro.

BACKGROUND: Piper betle L. has potent of antimicrobial activity and is widely used as a traditional remedy to treat skin infections. However, no clear evidence exists concerning antimicrobial and antibiofilm activity against Staphylococcus pseudintermedius and methicillin-resistant S. pseudintermedius (MRSP) opportunistic pathogens that cause wound infections and pyoderma in canines and zoonotic disease.
OBJECTIVE: The antimicrobial and antibiofilm activities of P. betle extract were assessed against S. pseudintermedius and MRSP strains.
METHODS: Ethanol leaf extract of P. betle was investigated for its antibacterial effect on S. pseudintermedius and MRSP by broth microdilution and time-kill assays. Biofilm inhibition and production assays were performed to evaluate antibiofilm and biofilm eradication effects, respectively. Biofilm-associated gene expression was further studied using real-time polymerase chain reaction (PCR). The possible interaction between IcaA and major compounds in P. betle was analyzed by molecular docking.
RESULTS: The extract showed minimum inhibitory concentration (MIC) at 250 μg/mL. Growth inhibition of P. betle at 1 MIC against the bacteria was initially observed after treatment for 4 h. All isolates were completely killed after 18 h exposure to the extract. Minimum biofilm inhibitory concentrations (MBICs) of the extract against the tested isolates ranged 1/2 MIC to 1 MIC, while minimum biofilm eradication concentration (MBEC) of P. betle was initialed at 8 MIC. Quantitative inhibition and eradication effects were observed in representative strains. The extract at 1/2 MIC and 1 MIC values inhibited biofilm formation up to 100%, with bacterial biofilm removed at up to 94.21% by 4 MIC of the extract. The extract downregulated the expression of the icaA gene among biofilm-producing isolates. The most abundant compounds, 4-allyl-1,2-diacetoxybenzene and eugenol showed a strong affinity with IcaA protein at -5.65 and -5.31 kcal/mol, respectively.
CONCLUSIONS: P. betle extract demonstrated the antibacterial, antibiofilm, and biofilm-removal activity against S. pseudintermedius and MRSP. Downregulation of the icaA gene expression and protein interaction were possible modes of action of the extract that impacted biofilm production. This extract showed promise as an alternative treatment for S. pseudintermedius infection, especially drug-resistant and biofilm-associated cases.

UNASSIGNED: Methicillin-resistant Staphylococcus (MRS) has originated, spread extensively, and become a prominent source of bacterial infections in both human and animal.
UNASSIGNED: We report the prevalence, genetic diversity, and antimicrobial resistance pattern of Staphylococcus pseudintermedius and Staphylococcus aureus strains isolated from dogs and cats with eye discharges.
UNASSIGNED: A total of 12 (6.0%) coagulase-positives staphylococci were identified as (6/200, 3%) S. aureus and (6/200, 3%) S. pseudintermedius. The phenotypic methicillin resistance of S. aureus and S. pseudintermedius were 50.0% (3/6) and 16.7% (1/6), respectively. None of the isolates showed biofilm formation in the microtiter plate assay. The highest resistance (50.0%) for S. pseudintermedius strains was detected against clindamycin and tetracycline. 67.0% of S. aureus isolates were resistant to penicillin-G. The PCR analysis conducted for detection of mecA gene indicated that only one S. aureus isolated from a cat was mecA gene positive. Phylogenetic analysis based on repetitive sequence-based PCR (rep-PCR) showed that all strains were typable and generated PCR products ranging from 800 bp to 4,400 bp. The lineages ST241 and the novel ST2361 in multi-locus sequence typing (MLST) analysis were detected in one methicillin-susceptible S. pseudintermedius and methicillin-resistant S. pseudintermedius of dogs, respectively. In addition, the lineages ST4155 and ST7217 of two methicillin-resistant S. aureus strains of cats were connected epidemiologically to previously reported cases.
UNASSIGNED: These results indicate epidemiologically related strains (ST241, ST4155, and ST7217) transferring between animals and humans. Therefore, the strategies to combat the widespread MRS should be based on collaboration between human and veterinary medicine under the One Health concept.

To determine the antibiotypes and frequency of toxin genes in methicillin-resistant Staphylococcus pseudintermedius (MRSP), 281 nasal swab samples were collected from dogs and dog guardians in Abakaliki, Southeastern Nigeria. Antimicrobial susceptibility testing was determined by disc diffusion technique while detection of toxin genes was carried out by PCR. Exactly 41 (28.7 %) and 6 (4.3 %) MRSP were obtained from dogs and dog guardians respectively. Isolates exhibited resistance (100-16.7 %) to amoxicillin-clavulanic acid, cephalosporins, fluoroquinolones, and carbapenems. Seccanine, lukD, siet, and exi toxin genes were harboured by 42 (89.4 %), 47 (100 %), 37 (78.7 %), and 2 (4.3 %) MRSP isolates respectively. This study has shown that dogs and dog guardians in Abakaliki, Southeastern Nigeria are colonized by multiple drug-resistant MRSP which harbour toxin genes. This represents a significant public health problem in veterinary and human medicine.

Staphylococcus pseudintermedius (SP) and methicillin-resistant SP (MRSP) is one of the most important veterinary pathogens in the dog. Herein, from a total of 126 S. pseudintermediusstrains, 23 MRSP (18%) were identified. Multi-Locus Sequence Typing (MLST) revealed that most of MRSP strains belonged to ST71 (26%), which have been already reported in Italy and other countries. Interestingly, nine new sequence types (39%), from 1053 up to 1061, were described for the first time. Moreover, the isolated MRSP strains showed relevant antibiotic resistance profiles. This report highlights the circulation of new sequence types of MRSP in Italy and underlines the need of a global epidemiological surveillance to limit the increasing spread of multidrug-resistant MRSPstrains worldwide, since they may represent a considerable concern for dog\'s health.

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