Methicillin-resistant Staphylococcus Pseudintermedius Research Articles

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Pyoderma in various manifestations is one of the most common problems in veterinary practice. Multiple studies on the treatment and prevention of pyoderma, combined with an alarming increase in the incidence of methicillin-resistant Staphylococcus pseudintermedius (MRSP) infections, have led to a shift from previous recommendations for the use of systemic antibiotics to the use of topical therapy. Antiseptics are primarily used in the treatment of bacterial infections. The most common component is chlorhexidine, which is available in various dosage forms. Benzoyl peroxide, chlorine solution, ethyl lactate, and miconazole can also be used for treatment. For local lesions, local antibiotics are sometimes prescribed – gentamicin, fusidine, silver sulfadiazine.

Screening canine sources for novel antimicrobials reveals the circular broad-spectrum bacteriocin, caledonicin, produced by Staphylococcus caledonicus.

Antimicrobial-resistant pathogens present an ongoing threat to human and animal health, with deaths linked to antimicrobial resistance (AMR) predicted to increase annually. While the misuse and overuse of antibiotics in humans undoubtedly contribute to this escalation, antibiotic use in the veterinary field, including companion animals, also plays a contributing role. Pet owners' desire to improve the quality of life of their pets is likely to support antibiotic use in this field. Consequently, there is a need for antibiotic alternatives to treat bacterial infections. This study set out to screen for antimicrobial peptides known as bacteriocins from bacterial isolates of aerobic/microaerophilic environments of canine sources and determine their potential as antibiotic alternatives against clinically relevant pathogens. Following a laboratory-based protocol, 22 bacterial isolates were subjected to whole-genome sequencing (WGS), and a total of 14 putative novel bacteriocins were identified from both class I and II bacteriocin classes. One particular bacteriocin, herein named caledonicin, was identified via in silico analysis from a Staphylococcus caledonicus strain and partially purified for further in vitro evaluation. Caledonicin is a 64-amino acid (IAANLGVSSGTAYS MANALNNISNVATA LTIIGTFTGVGTIGSGIA ATILAILKKKGVAAAAAF) novel circular bacteriocin most closely related to enterocin_NKR-5-3B based on core peptide alignment (39.1%), with a molecular weight of 6077.1 Da. Caledonicin exhibits a broad-spectrum of activity against a range of pathogenic bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant Staphylococcus pseudintermedius (MRSP), and Listeria monocytogenes; and the gut-related bacterium associated with Crohn's disease, Mediterraneibacter gnavus ATCC 29149 (previously Ruminococcus gnavus ATCC 29149). This represents the first bacteriocin screening study involving bacteria from canine sources and confirms this is a rich environment for bacteriocin-producing strains. This study also identifies and characterises the first novel bacteriocin from the staphylococcal species, Staphylococcus caledonicus.

Pharmacokinetic study of local and systemic gentamicin concentrations after subcutaneous implantation of a gentamicin-impregnated collagen sponge in dogs

This study evaluated the pharmacokinetics of commercial gentamicin-impregnated collagen sponges (GICS) applied subcutaneously in dogs. In six healthy beagles, an 11 ×6 cm subcutaneous pocket was created, a folded 10×10 cm GICS was inserted, and saline was injected to mimic a seroma. Wound fluid samples were aspirated, and the gentamicin concentration was determined. Simultaneously, blood samples were collected to evaluate the corresponding systemic gentamicin concentration. All samples were collected before and 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, 18, 24, 36, 48, 72, 96, 120, and 168 hours after GICS placement. The local Cmax of gentamicin was reached after 0.5 hours (range, 0.5–1.0 hours) post-implantation in 5/6 dogs at a median concentration of 2053.3 µg/mL (range, 918.0–2791.9 µg/mL). Whitin 24 hours, the local concentration dropped below the MIC for Staphylococcus sp. (4 µg/mL) in 5/6 dogs. Plasma Cmax was achieved at a median of 1.2 hours post-implantation (range, 1.0–2.0 hours) and reached a median concentration of 10.3 µg/mL (range, 8.8–18.03 µg/mL). After 6 hours, the gentamicin concentration in the plasma was below 4 µg/mL in all dogs. The GICS provided a high local concentration of gentamicin in a short time with a local Cmax:MIC ratio of 513:1, largely sufficient to eliminate susceptible bacteria, including methicillin-resistant Staphylococcus pseudintermedius (MRSP) and Pseudomonas sp., in a clinical setting. The repeated administration of saline in the present study seemed to have induced a quicker gentamicin release from the GICS than described in previous studies that typically dealt with "drier" wounds.

Comparative Analysis of Methicillin-Resistant Staphylococcus pseudintermedius Prevalence and Resistance Patterns in Canine and Feline Clinical Samples: Insights from a Three-Year Study in Germany.

The emergence of methicillin-resistant Staphylococcus pseudintermedius (MRSP) presents a significant public health concern globally, particularly within veterinary medicine. MRSP's resistance to multiple antibiotics is limiting treatment options and potentially leading to severe infections in companion animals. This study aimed to understand antimicrobial resistance in dogs and cats, focusing on MRSP resistance patterns and its prevalence in Germany. We analyzed results of bacterial diagnostic samples from canines and felines, sourced from a German veterinary diagnostic microbiology laboratory between 2019 and 2021. This dataset included samples from 3491 veterinary practices, covering 33.1% of veterinary practices and clinics in Germany. MRSP rates were detailed by host species, sample types and co-resistance patterns. Analysis of 175,171 bacterial examination results revealed S. pseudintermedius in 44,880 samples, yielding a 25.6% isolation rate. S. pseudintermedius was more prevalent in dogs (35.0%) than cats (3.6%). Methicillin resistance was found in 7.5% of all S. pseudintermedius isolates. MRSP prevalence was higher in feline samples (16.1%, 95% CI 14.4-17.8) compared to canine samples (7.1%, 95% CI 6.8-7.0). S. pseudintermedius showed high resistance rates to ampicillin (cats: 48.6%, dogs: 67.6%) and clindamycin (cats: 37.2%, dogs: 32.7%), while MRSP exhibited high co-resistance to clindamycin (cats: 82.8%, dogs: 85.4%) and sulfamethoxazole + trimethoprim (cats: 66.4%, dogs: 66.2%). Our study revealed distinct resistance patterns of MRSP in cats compared to dogs, highlighting the need for tailored treatment approaches and the importance of antimicrobial resistance surveillance.

Diagnostic ultrasonography and antimicrobial resistance of different pathogens associated with canine and feline lower urinary tract disorders

There is a significant issue concerning the dissemination of antimicrobial-resistant bacteria within companion animals. Urinary tract infections (UTIs) are a common problem in veterinary medicine for which empirical antibiotics are utilized. This study aimed to investigate the antimicrobial resistance of different uropathogens associated with UTIs in canine and feline cases. A total of 146 dogs and 162 cats suffered from lower urinary tract disorders were subjected to ultrasonographic and microbiological examination. Cystitis, urinary sediment, and cystic calculi are the most common ultrasonographic abnormalities associated with bacterial UTIs. Bacterial UTIs were obtained in 36.98 % and 25.92 % of cases in dogs and cats, respectively. A low rate of mixed infection was detected in canine cases (3.7 %). E. coli was the most prevalent pathogen isolated from 46.4 % and 66.7 % of canine and feline isolates, respectively followed by Proteus spp. in canine isolates (16.1 %) and Klebsiella spp. in feline isolates (14.3 %). Staphylococcus spp. was isolated from canine cases only with the detection of methicillin-resistant Staphylococcus pseudintermedius (MRSP) strains at 3.6 %. The majority of the isolated strains were resistant to various antibiotic classes, particularly β-lactams. All gram-negative bacteria were susceptible to amikacin, whereas gram-positive strains exhibited 100 % sensitivity to nitrofurantoin and linezolid. Different bacterial species displayed low resistance to carbapenems and fluoroquinolones. Multi-drug resistance was reported in canine and feline strains at 64.3 % and 54.8 %, respectively. These findings prove the crucial necessity to restrict antibiotic consumption depending on urine culture and antibiotic sensitivity tests.

Chromosomal and plasmid localization of ileS2 in high-level mupirocin-resistant Staphylococcus pseudintermedius and Staphylococcus aureus isolated from canine and feline origins.

To characterize the mobile genetic elements and genetic localization of ileS2 in high-level mupirocin-resistant (Hi-MupR) methicillin-resistant Staphylococcus pseudintermedius (MRSP) and MRSA isolates recovered from canine and feline clinical samples. The identification of bacterial species and presence of mecA and ileS2 genes in MRSP and MRSA isolates were performed using MALDI-TOF MS and PCR, respectively. Antimicrobial resistance (AMR) phenotypes were determined by broth microdilution assays. The genome characteristics, ileS2-containing elements and staphylococcal cassette chromosome mec (SCCmec) were illustrated using complete circular genomes obtained from hybrid assembly of Illumina short-reads and Oxford Nanopore Technologies long-reads. These were analysed through phylogenetic and bioinformatics approaches. A total of 18 MRSP clinical isolates and four MRSA clinical isolates exhibited the Hi-MupR phenotype and carried multiple AMR genes, including mecA and ileS2 genes. MRSP ST182-SCCmec V (n = 6) and ST282-ΨSCCmec57395-t10 (n = 4) contained the ileS2 transposable unit associated with IS257 on the chromosome. Three MRSA ST398-SCCmec V-t034/t4652 isolates carried ∼42 kb pSK41-like ileS2 plasmids, whereas similar ileS2 plasmids lacking tra genes were found in MRSP ST282-ΨSCCmec57395-t72/t21 isolates. Furthermore, a new group of ileS2 plasmids, carried by MRSP ST45-ΨSCCmec57395, ST433-ΨSCCmecKW21-t05 and ST2165-SCCmec IV-t06, and by one MRSA ST398-SCCmec V-t034 strain, shared the plasmid backbone with the cfr/fexA-carrying plasmid pM084526_1 in MRSA ST398. This study provides the first evidence of ileS2 integration into the S. pseudintermedius chromosome, which is a rare occurrence in staphylococcal species, and plasmids played a pivotal role in dissemination of ileS2 in both staphylococcal species.

In vitro activity of selected antimicrobials against methicillin-resistant Staphylococcus pseudintermedius of canine origin in Poland.

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) is an important veterinary pathogen. In general, only a few antimicrobials show in vitro activity against MRSP isolates. The objective of this study was to determine the in vitro activity of selected antimicrobials, including last-choice drugs, against clinical MRSP isolates of canine origin. The activity of 10 selected agents was evaluated against 41 clinical MRSP isolates. The disk diffusion method and minimal inhibitory concentration values were used for antimicrobial susceptibility testing (AST). The guidelines for staphylococci of canine or human origin were employed for the interpretation of the results. Among the examined MRSP isolates, resistance to enrofloxacin and clindamycin was the most prevalent (n=40; 97.6%). Resistance to doxycycline and gentamicin was observed in 83.0% (n=34) and 68.3% (n=28) of the isolates, respectively. Single isolates were resistant to chloramphenicol (n=5; 12.2%) and rifampicin (n=3; 7.3%), whereas all showed susceptibility to amikacin, vancomycin, mupirocin and linezolid. Predominantly, the results of AST obtained by both methods were consistent. Some discrepancies were observed for gentamicin; however, clinical breakpoints for staphylococci of human origin were used. Amikacin and chloramphenicol constitute potential treatment options in infections caused by MRSP and may be included in extended susceptibility testing in our geographical region. The determination of clinical breakpoints for some antimicrobials not incorporated in the recommendations should be a high priority in the veterinary diagnostics.

Emergence of novel methicillin resistant Staphylococcus pseudintermedius lineages revealed by whole genome sequencing of isolates from companion animals and humans in Scotland.

Staphylococcus pseudintermedius is an opportunistic pathogen in dogs, and infection in humans is increasingly found, often linked to contact with dogs. We conducted a retrospective genotyping and antimicrobial susceptibility testing study of 406 S. pseudintermedius isolates cultured from animals (dogs, cats and an otter) and humans across Scotland, from 2007 to 2020. Seventy-five sequence types (STs) were identified, among the 130 isolates genotyped, with 59 seen only once. We observed the emergence of two methicillin resistant Staphylococcus pseudintermedius (MRSP) clones in Scotland: ST726, a novel locally-evolving clone, and ST551, first reported in 2015 in Poland, possibly linked to animal importation to Scotland from Central Europe. While ST71 was the most frequent S. pseudintermedius strain detected, other lineages that have been replacing ST71 in other countries, in addition to ST551, were detected. Multidrug resistance (MDR) was detected in 96.4% of MRSP and 8.4% of MSSP. A single MRSP isolate was resistant to mupirocin. Continuous surveillance for the emergence and dissemination of novel MDR MRSP in animals and humans and changes in antimicrobial susceptibility in S. pseudintermedius is warranted to minimise the threat to animal and human health.

Intervention with impact: Reduced isolation of methicillin-resistant Staphylococcus pseudintermedius from dogs following the introduction of antimicrobial prescribing legislation in Germany.

Legislation was introduced in Germany in 2018, requiring bacterial culture and antimicrobial susceptibility testing before the prescription of fluoroquinolones and third-generation cephalosporins to dogs. We hypothesised that, following this intervention, the number of clinical samples testing positive for methicillin-resistant Staphylococcus pseudintermedius (MRSP) would reduce. Reports of S. pseudintermedius isolated from canine clinical samples by three German veterinary diagnostic microbiology laboratories during the 38 months before the introduction of the legislation and the 46 months after were compared. Bacterial identification was performed by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry, and antimicrobial susceptibility testing followed recognised recommendationsbut with changes during the study period. Among a total of 120,571 S. pseudintermedius isolates, MRSP accounted for 7.1% overall. Following the legislative intervention, monthly submissions yielding S. pseudintermedius increased at all three laboratories. The MRSP percentage was lower in the period after the intervention in two of the three laboratories (p < 0.001); in the third laboratory, there was no change between periods, but a year-on-year reduction in MRSP percentages occurred after the intervention (p = 0.0004). Changing susceptibility testing methods limited the direct comparison of resistance patterns among laboratories. The reduction in MRSP in canine clinical samples following the introduction of this legislation suggests a positive impact of compulsory laboratory testing on reducing antimicrobial resistance.

Epidemiologic case investigation on the zoonotic transmission of Methicillin-resistant Staphylococcus pseudintermedius among dogs and their owners

Dogs often carry methicillin-resistant Staphylococci asymptomatically. These bacteria are frequently linked to conditions such as canine pyoderma and otitis. Close interaction between dogs and humans can facilitate the exchange of resistant strains, particularly Methicillin-resistant Staphylococcus pseudintermedius (MRSP). This represents a public health issue, since these strains, in addition to occasionally causing infections in humans, can also serve as a source of resistance and virulence genes for strains of greater importance in human medicine, such as Staphylococcus aureus. Furthermore, MRSP strains are often multidrug resistant, which ends up compromising the treatment of infections. This study aimed to assess the potential transmission of Staphylococcus pseudintermedius among dogs and their owners. We examined a total of one hundred canine samples collected from cases of pyoderma and otitis to detect the presence of staphylococci. Simultaneously, we conducted evaluations on all dog owners. Staphylococci strains were identified using MALDI-TOF MS and PCR targeting the nuc gene. Methicillin resistance screening was also performed by detecting the mecA gene using PCR. Among the sampled dogs, 64 carried S. pseudintermedius. Nine were identified as MRSP. In six instances, dogs and their owners exhibited S. pseudintermedius. These samples underwent genome sequencing and were screened for antimicrobial resistance genes, SCCmec typing, MLST characterization, and Single Nucleotide Polymorphisms (SNP) analyses. The results of the phylogenetic analysis revealed that in three cases, dogs and owners had closely related isolates, suggesting interspecies transmission. Two of these cases involved MRSP and one MSSP. Moreover, in the two MRSP cases, the same SCCmec type (type V) was detected. Additionally, the sequence type was consistent across all three cases involving dogs and owners (MSSP ST2277, MRSP ST2282, and ST2286). These findings strongly indicate a transmission event. Since Staphylococcus pseudintermedius is primarily isolated from canine samples, it is plausible that dogs may have acted as a potential source. In the remaining three cases, despite identifying the same species in both samples, they had notable phylogenetic differences.

The molecular epidemiology and antimicrobial resistance of Staphylococcus pseudintermedius canine clinical isolates submitted to a veterinary diagnostic laboratory in South Africa.

Staphylococcus pseudintermedius is an important cause of clinical infections in small-animal-veterinary medicine. Evolutionary changes of strains using multilocus sequence typing (MLST) have been observed among S. pseudintermedius in European countries and the United States. However, there are limited or no studies on the detection of methicillin resistant Staphylococcus pseudintermedius (MRSP) and predominating MLST strains in South Africa. Therefore, this study aimed to determine the molecular epidemiology of S. pseudintermedius in South Africa. Twenty-six, non-duplicate, clinical isolates from dogs were obtained as convenience samples from four provinces in South Africa. The Kirby Bauer disk diffusion test was used to determine antimicrobial susceptibility. We used Resfinder and the Comprehensive Antibiotic Resistance Database (CARD) to detect antimicrobial resistance genes. Virulence genes were identified using the virulence factor database and Basic Local Alignment Search Tool (BLASTN) on Geneious prime. geoBURST analysis was used to study relationships between MLST. Finally, the maximum likelihood phylogeny was determined using Randomized Axelerated Maximum Likelihood (RAxML). Twenty-three isolates were confirmed as S. pseudintermedius of which 14 were MRSP. In addition to β-lactam antimicrobials, MRSP isolates were resistant to tetracycline (85.7%), doxycycline (92.8%), kanamycin (92.8%), and gentamicin (85.7%). The isolates harbored antimicrobial resistance genes (tetM, ermB, drfG, cat, aac(6')-Ie-aph(2")-Ia, ant(6)-Ia, and aph(3')-III) and virulence genes (AdsA, geh, icaA, and lip). MLST analysis showed that ST2228, ST2229, ST2230, ST2231, ST2232, ST2318, ST2326 and ST2327 are unique sequence types in South Africa. Whereas, previously reported major STs including ST45, ST71, ST181, ST551 and ST496 were also detected. The geoBURST and phylogenetic analysis suggests that the isolates in South Africa are likely genetically related to isolates identified in other countries. Highly resistant MRSP strains (ST496, ST71, and ST45) were reported that could present challenges in the treatment of canine infections in South Africa. Hence, we have gained a better understanding of the epidemiology of MRSP in the African continent, the genes involved in resistance and virulence factors associated with these organisms.

Temporal changes in antibiotic resistance and population structure of methicillin-resistant Staphylococcus pseudintermedius between 2010 and 2021 in the United States

The aim of this study was to perform a phenotypic and molecular epidemiological survey to determine temporal changes in the antimicrobial resistance and population structure of methicillin-resistant Staphylococcus pseudintermedius (MRSP) in the United States. Samples from 200 S. pseudintermedius isolates were obtained from veterinary diagnostic facilities located in geographic regions sampled approximately ten years ago and compared to samples obtained in 2021. Kirby-Bauer disk diffusion was used to determine antimicrobial susceptibility. geoBURST analysis and MrBayes were used to infer relationships of isolates using MLST data. Almost all MRSP isolates (98%) in 2021 were multidrug-resistant with 21% of these isolates resistant to more than 16 antimicrobials. In 2010, 190 S. pseudintermedius isolates were collected and 141 of them were MRSP. From 2010–2021 there was a significant increase in resistance observed to all antibiotics tested except cephalothin and sulfonamides. Whereas ten years previously multilocus sequence types (ST) ST68 (35.7%), ST71 (10%), and ST84 (17.4%) predominated, these strains have been supplanted by other STs, notably ST45 (n = 14), ST155 (n = 9), ST181 (n = 13), ST496 (n = 9) and ST551 (n = 9). The newly prevalent STs are distantly related to ST68, ST71 and ST84 and most likely do not share any recent common ancestors. The population structure of MRSP is far more elastic than expected with new, highly resistant strains replacing the ones that predominated in the United States a decade ago. Antibiotic use may play a role in selection; however, the strains that were replaced were also multidrug-resistant and other factors are likely involved.

In vitro and in vivo susceptibility to cefalexin and amoxicillin/clavulanate in canine low-level methicillin-resistant Staphylococcus pseudintermedius.

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) lineages harbouring staphylococcal cassette chromosome (SCC) mec types IV, V and ΨSCCmec57395 usually display low oxacillin MICs (0.5-2 mg/L). To evaluate how oxacillin MICs correlate with PBP mutations and susceptibility to β-lactams approved for veterinary use. Associations between MICs and PBP mutations were investigated by broth microdilution, time-kill and genome sequence analyses in 117 canine MRSP strains harbouring these SCCmec types. Clinical outcome was retrospectively evaluated in 11 MRSP-infected dogs treated with β-lactams. Low-level MRSP was defined by an oxacillin MIC <4 mg/L. Regardless of strain genotype, all low-level MRSP isolates (n = 89) were cefalexin susceptible, whereas no strains were amoxicillin/clavulanate susceptible according to clinical breakpoints. Exposure to 2× MIC of cefalexin resulted in complete killing within 8 h. High (≥4 mg/L) oxacillin MICs were associated with substitutions in native PBP2, PBP3, PBP4 and acquired PBP2a, one of which (V390M in PBP3) was statistically significant by multivariable modelling. Eight of 11 dogs responded to systemic therapy with first-generation cephalosporins (n = 4) or amoxicillin/clavulanate (n = 4) alone or with concurrent topical treatment, including 6 of 7 dogs infected with low-level MRSP. Oxacillin MIC variability in MRSP is influenced by mutations in multiple PBPs and correlates with cefalexin susceptibility. The expert rule recommending that strains with oxacillin MIC ≥0.5 mg/L are reported as resistant to all β-lactams should be reassessed based on these results, which are highly clinically relevant in light of the shortage of effective antimicrobials for systemic treatment of MRSP infections in veterinary medicine.

Comparative Susceptibility of Pathogenic Methicillin-Resistant and Methicillin-Susceptible Staphylococcus pseudintermedius to Empirical Cotrimazole for Canine Pyoderma.

The prevalence of methicillin-resistant Staphylococcus pseudintermedius (MRSP) that causes pyoderma has been gradually shifting, according to many surveillance studies, with annual changes. The empirical cotrimazole regimen remains interesting, but research on cotrimazole susceptibility to MRSP is limited. The objective of this study was to evaluate the susceptibility of cotrimazole to canine pyoderma MRSP isolates. Sixty isolates of S. pseudintermedius were identified as 16 MRSP and 44 methicillin-susceptible S. pseudintermedius (MSSP) using an oxacillin disk diffusion test and VITEK 2 system with VITEK GP card. Using the VITEK 2 system with a VITEK AST-GP81 card, the susceptibility rates of MRSP (15.00%) and MSSP (35.00%) to cotrimazole was observed. The median MIC of cotrimazole on MSSP (median, ≤10; IQR, 10-320) was lower than that of MRSP (median, ≥320; IQR, 10-320) (p = 0.5889, Mann-Whitney test). Percent attainment of PK/PD targets in MRSP (q 12 h, 43.75; q 8 h, 43.75) were lower than that of MSSP (q 12 h, 52.27; q 8 h, 52.27) (p = 0.7710). These findings show the moderately phenotypic cotrimazole susceptibilities of both MRSP and MSSP. Further study is required to develop clinical trials examining the use of cotrimazole in dogs with pyoderma.

Effects of Piper betle Extracts against Biofilm Formation by Methicillin-Resistant Staphylococcus pseudintermedius Isolated from Dogs.

Emergence of methicillin-resistant Staphylococcus pseudintermedius (MRSP) isolated from dogs with cutaneous and wound infections has significantly impacted veterinary medicine. This study aimed to isolate S. pseudintermedius from canine pyoderma and investigate the effects of ethanolic extracts of Piper betle (PB), P. sarmentosum (PS), and P. nigrum (PN) on the bacterial growth and biofilm formation of S. pseudintermedius and MRSP. Of the isolated 152 isolates, 53 were identified as S. pseudintermedius using polymerase chain reaction, and 10 isolates (6.58%) were identified as MRSP based on the presence of mecA. Based on phenotype, 90% of MRSPs were multidrug-resistant. All MRSP had moderate (10%, 1/10) and strong (90%, 9/10) biofilm production ability. PB extracts were the most effective in inhibiting planktonic cells, and the minimum inhibitory concentration at which ≥50% of the isolates were inhibited (MIC50) was 256 µg/mL (256-1024 µg/mL) for S. pseudintermedius isolates and 512 µg/mL (256-1024 µg/mL) for MRSP isolates. The MIC90 for S. pseudintermedius and MRSP was 512 µg/mL. In XTT assay, PB at 4× MIC showed an inhibition rate of 39.66-68.90% and 45.58-59.13% for S. pseudintermedius and MRSP, respectively, in inhibiting biofilm formation. For PB at 8× MIC, the inhibition rates for S. pseudintermedius and MRSP were 50.74-81.66% and 59.57-78.33%, respectively. Further, 18 compounds were identified in PB using gas chromatography-mass spectrometry, and hydroxychavicol (36.02%) was the major constituent. These results indicated that PB could inhibit bacteria growth of and biofilm formation by S. pseudintermedius and MRSP isolated from canine pyoderma in a concentration-dependent manner. Therefore, PB is a potential candidate for the treatment of MRSP infection and biofilm formation in veterinary medicine.

Risk Factors, Genetic Diversity, and Antimicrobial Resistance of Staphylococcus spp. Isolates in Dogs Admitted to an Intensive Care Unit of a Veterinary Hospital.

Intensive Care Units (ICU) usually provide an excellent environment for the selection of pathogens associated with hospital-acquired infections (HAI), leading to increased mortality and hospitalization costs. Methicillin-resistant Staphylococcus pseudintermedius (MRSP) is a major cause of HAI in dogs worldwide, but the risk factors and dynamics of colonization by MRSP are largely unknown. This study aimed to evaluate the risk factors associated with the acquisition of MRSP in dogs admitted to an ICU, and to report the antimicrobial resistance profiles and genetic relatedness of MRSP isolates. Sterile swabs from the nostril, axilla, and rectum were collected daily during the hospitalization of 54 dogs. Samples were subjected to Mannitol Salt Agar, and colonies were identified by MALDI-ToF, polymerase chain reaction (PCR), and sequencing of the rpoB gene. Antimicrobial susceptibility testing and PCR detection of mecA were performed. Staphylococcus spp. was isolated from 94% of the dogs, and the most frequently isolated species was S. pseudintermedius (88.2%). Carriage of multidrug resistant (MDR) staphylococci was observed in 64.4% of the dogs, and approximately 39% had methicillin-resistant Staphylococcus sp. (MRS), of which 21.6% had MRSP and 1.9% had methicillin-resistant S. aureus (MRSA). The acquisition of MRSP during ICU hospitalization was associated with sex (female), age (>7 years), and dogs that had previously been treated with antimicrobials. Animals colonized by MRSP resistant to ≥9 antimicrobial classes had longer hospital stays than those colonized by other MRS strains. Among the 13 MRSP isolates that were subjected to whole-genome sequencing, ten were classified as ST71. A single nucleotide polymorphism (SNP) analysis revealed three clones, including one that was detected in infected dogs outside the ICU. This study indicates novel risk factors associated with colonization by MRSP. The detection of the same MRSP clone causing HAI outside the ICU reinforces the need for improved infection prevention and control practices at veterinary hospitals in general and at the ICU in particular.

Bacteriocins Revitalize Non-Effective Penicillin G to Overcome Methicillin-Resistant Staphylococcus pseudintermedius.

The rise of antibiotic-resistant bacteria is among the biggest challenges in human and veterinary medicine. One of the major factors that contributes to resistance is use of frontline clinical antibiotics in veterinary practices. To avoid this problem, searching for antimicrobials aimed at veterinary applications is becoming especially important. Thiopeptide micrococcin P1 and leaderless peptide EntEJ97s are two different bacteriocins that are very active against many gram-positive bacteria; however, sensitive bacteria can rapidly develop resistance towards those bacteriocins. To overcome this problem, we searched for synergy between those bacteriocins and conventional antibiotics against methicillin-resistant Staphylococcus&amp;nbsp;pseudintermedius (MRSP): a common pathogen in animal skin infections. The two bacteriocins acted synergistically with each other and with penicillin G against MRSP clinical isolates in both planktonic and biofilm assays; they also prevented resistance development. The therapeutic potential was further validated in a murine skin infection model that showed that a combination of micrococcin P1, EntEJ97s and penicillin G reduced cell-forming units of MRSP by 2-log10 CFU/g. Taken together, our data show that a combination of bacteriocins with conventional antibiotics can not only prevent resistance development but also pave the way to revitalize some old, less useful antibiotics, such as penicillin, which by itself has no effect on methicillin-resistant pathogens.

Detection of the international lineage ST71 of methicillin-resistant Staphylococcus pseudintermedius in two cities in Rio de Janeiro State.

Staphylococcus pseudintermedius is the main coagulase-positive staphylococci associated with canine skin/soft tissue infections (SSTI), otitis externa, and surgical site infections. The international spread of an epidemic and multiresistant lineage of methicillin-resistant Staphylococcus pseudintermedius (MRSP), the so-called European clone-displaying sequence type (ST) 71-requires attention. The first isolation of an MRSP ST71 isolate in South America was reported in Rio de Janeiro city, in 2010; however, a limited number of canine isolates were analyzed. Thus, to have a better panel of the MRSP spread in this city, we were stimulated to continue this study and search for the presence of MRSP in 282 colonized or infected dogs in the city of Rio de Janeiro. Among the MRSP isolates collected (N = 17; 6.1%), the pulsed-field gel electrophoresis (PFGE) patterns were similar to those of European clone. All 17 isolates were classified as ST71 by multilocus sequence typing (MLST). In order to assess whether isolates of MRSP ST71 may have also spread to the Rio de Janeiro state countryside, we collected samples from 124 infected dogs in the city of Campos dos Goytacazes (232km away from Rio de Janeiro city). Our data showed the presence of ST71 lineage in one isolate among three MRSP detected. S. pseudintermedius was isolated from 40.6% of the clinical samples (N = 165/406). A relatively high incidence of methicillin resistance, detected by a PCR-based method, was found in 12.1% of the S. pseudintermedius recovered from animals (N = 20/165). The resistance profile of these isolates was similar to that described for the international ST71 strains whose genomes are publicly available in the GenBank. The prospect of ST71 isolates being resistant to virtually all antimicrobials used in veterinary medicine is alarming and should be considered a central issue considering that MRSP ST71 spreads over large geographic distances and its transmission from animals to humans.

Antibiotypes and high frequency of toxin genes in methicillin-resistant Staphylococcus pseudintermedius from nares of dogs and dog guardians in Nigeria

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.

Prevalence of methicillin resistance in Staphylococcus pseudintermedius isolates from dogs with skin and ear infections in South Africa

Staphylococcus pseudintermedius (SP) is an important opportunistic pathogen, frequently associated with pyoderma and otitis in dogs. The emergence and rapid expansion of methicillin-resistant Staphylococcus pseudintermedius (MRSP) is problematic due to multidrug resistance and reduced treatment options. The aim of this study was to determine i) the prevalence of MRSP in dogs with pyoderma or otitis externa, ii) the antimicrobial resistance patterns of MRSP from South African isolates, and iii) the risk factors for MRSP-associated pyoderma or otitis externa in dogs in South Africa (RSA). Sixty-eight presumptive clinical SP isolates (collected from 65 dogs) from five geographically dispersed laboratories in RSA were collected over 2 years. Possible MRSP isolates were flagged when resistance to oxacillin was observed. Thereafter, all isolates were confirmed as SP by polymerase chain reaction (PCR) and further genotyped for the mecA gene. Fifty-seven of 68 isolates were confirmed to be SP (83.8%), while 49/57 (85.9%) carried mecA. Our findings showed that preliminary phenotypic methods supplemented by genotypic methods increased the accuracy of correctly identifying SP. All isolates were resistant to at least one antimicrobial drug. There was a high incidence of amoxicillin (70.1%) and enrofloxacin (65%) resistance. Important risk factors for mecA positive carriage were previous hospital admission, pruritus, and previous antibacterial failure. This study demonstrates a high prevalence of mecA positive carriage (85.9% of samples) in MRSP pyoderma and otitis in dogs in RSA. There is an urgent need for better laboratory diagnosis of MRSP and surveillance of dogs presenting with pyoderma and otitis in South Africa.