Pseudintermedius Strains Research Articles

Genomic Analyses of Methicillin-Susceptible and Methicillin-Resistant Staphylococcus pseudintermedius Strains Involved in Canine Infections: A Comprehensive Genotypic Characterization.

Staphylococcus pseudintermedius is frequently associated with several bacterial infections in dogs, highlighting a One Health concern due to the zoonotic potential. Given the clinical significance of this pathogen, we performed comprehensive genomic analyses of 28 S. pseudintermedius strains isolated from canine infections throughout whole-genome sequencing using Illumina HiSeq, and compared the genetic features between S. pseudintermedius methicillin-resistant (MRSP) and methicillin-susceptible (MSSP) strains. Our analyses determined that MRSP genomes are larger than MSSP strains, with significant changes in antimicrobial resistance genes and virulent markers, suggesting differences in the pathogenicity of MRSP and MSSP strains. In addition, the pangenome analysis of S. pseudintermedius from canine and human origins identified core and accessory genomes with 1847 and 3037 genes, respectively, which indicates that most of the S. pseudintermedius genome is highly variable. Furthermore, phylogenomic analysis clearly separated MRSP from MSSP strains, despite their infection sites, showing phylogenetic differences according to methicillin susceptibility. Altogether our findings underscore the importance of studying the evolutionary dynamics of S. pseudintermedius, which is crucial for the development of effective prevention and control strategies of resistant S. pseudintermedius infections.

Virulence and host specificity of staphylococci from Staphylococcus intermedius group of pigeon origin with an emphasis on Staphylococcus intermedius

The Staphylococcus intermedius group (SIG) includes coagulase-positive staphylococci commonly found in animals. The taxonomic classification within the SIG has evolved with molecular techniques distinguishing five species. Despite their similarities, these species exhibit varied host affinities, with unclear implications for virulence and host interaction.This study aimed to investigate the presence of coagulase-positive staphylococci in pigeons and to detect genes encoding for selected virulence factors in isolated strains. Another goal was to determine the adhesion capabilities of randomly selected pigeon S. intermedius, S. delphini, and canine S. pseudintermedius strains to canine and pigeon corneocytes and their adhesion and invasion abilities to canine keratinocytes in vitro.In total, 121 coagulase-positive strains were isolated from domestic and feral pigeons. The most prevalent species were S. delphini B and S. intermedius in domestic and feral pigeons, respectively. We proved that pigeon strains carried genes encoding for exfoliative toxin SIET and leukotoxin Luk-I. Moreover, we found that S. intermedius showed higher adherence to pigeon than to canine corneocytes, aligning with its presumed natural host. No difference in adherence abilities of S. pseudintermedius to canine and pigeon corneocytes was observed. In this study, we also observed that S. pseudintermedius could successfully invade the canine keratinocytes, in contrary to S. delphini and S. intermedius. Moreover, only S. intermedius was not able to invade canine keratinocytes at all.These findings highlight the complex interplay between SIG bacteria, and their hosts, underscoring the need for further research to understand the mechanisms of host adaptation and pathogenicity within this group.

Brief communication: Carrier rate, antimicrobial resistance and molecular typing of Staphylococcus aureus and Staphylococcus pseudintermedius in healthy dogs from Morogoro, Tanzania.

Staphylococcus pseudintermedius and S. aureus are important pathogens in dogs. This study established carrier rates, strain diversity and antimicrobial resistance of these bacteria among healthy dogs in Tanzania. Based on cultures of mouth and perineal swabs, 11.3% and 50.3% of 151 healthy dogs were carriers of S. aureus and S. pseudintermedius, respectively. Only four dogs (3%) carried meticillin-resistant S. aureus (MRSA), while none of the S. pseudintermedius strains were meticillin-resistant. 12 of 19 S. aureus strains tested were resistant to penicillin G, and resistance to enrofloxacin and tetracycline was also commonly detected. The most common resistances in 103 S. pseudintermedius strains tested were to penicillin G (28.2%) and tetracycline (22.3%). S. pseudintermedius strains showed 65 different random amplified polymorphic DNA (RAPD) fingerprints, and S. aureus strains belonged to eight different spa types, including two novel types (t18988 and t18989). MRSA strains carried SCCmec type V. Healthy dogs in Tanzania were carriers of MRSA at low frequency, and half of the dogs carried S. pseudintermedius with high strain diversity.

Antibacterial Potential of Essential Oils and Silver Nanoparticles against Multidrug-Resistant Staphylococcus pseudintermedius Isolates.

Staphylococcus pseudintermedius is an emergent zoonotic agent associated with multidrug resistance (MDR). This work aimed to describe the antibacterial activity of four essential oils (EOs) and silver nanoparticles (AgNPs) against 15 S. pseudintermedius strains isolated from pyoderma. The four EOs, namely Rosmarinus officinalis (RO), Juniperus communis (GI), Citrus sinensis (AR), and Abies alba (AB), and AgNPs were used alone and in combination to determine the Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC). All strains were MDR and methicillin-resistant. Among the antibiotic cohort, only rifampicin, doxycycline, and amikacin were effective. EOs' chemical analysis revealed 124 compounds belonging to various chemical classes. Of them, 35 were found in AR, 75 in AB, 77 in GI, and 57 in RO. The monoterpenic fraction prevailed over the sesquiterpenic in all EOs. When EOs were tested alone, AB showed the lowest MIC followed by GI, AR, and RO (with values ranging from 1:128 to 1:2048). MBC increased in the following order: AB, AR, GI, and RO (with values ranging from 1:512 to 1:2048). MIC and MBC values for AgNPs were 10.74 mg/L ± 4.23 and 261.05 mg/L ± 172.74. In conclusion, EOs and AgNPs could limit the use of antibiotics or improve the efficacy of conventional therapies.

In Vitro Activity of Allium cepa Organosulfur Derivatives against Canine Multidrug-Resistant Strains of Staphylococcus spp. and Enterobacteriaceae.

The increase of multi-resistant bacteria, especially Staphylococcus spp. and Enterobacteriaceae, constitutes a challenge in veterinary medicine. The rapid growth of resistance is outpacing antibiotic discovery. Innovative strategies are needed, including the use of natural products like Allium species (Allium sativum L. and Allium cepa L.), which have been used empirically for centuries to treat infectious diseases in humans and farm and aquaculture animals due to their antibacterial properties. This study aimed to evaluate the in vitro activity of two Allium-derived compounds, propyl propane thiosulfinate (PTS) and propyl propane thiosulfonate (PTSO), against multi-resistant Staphylococcus spp. (n = 30) and Enterobacteriaceae (n = 26) isolated from dogs referred to a veterinary teaching hospital in Madrid. The results indicated the in vitro efficacy of PTSO/PTS against the tested bacterial strains, and 56.7% of Staphylococcus pseudintermedius and 53.8% of Enterobacteriaceae showed sensitivity to PTS and PTSO compared with classic antibiotics. In addition, 50% of S. pseudintermedius strains resistant to erythromycin, ibofloxacin, difloxacin and orbifloxacin and 50% of Enterobacteriaceae strains resistant to tetracycline and doxycycline were sensitive to PTS and PTSO. Although studies are needed to verify their efficacy in vivo, the combined use of PTS and PTSO exhibits promise in enhancing bacterial sensitivity against S. pseudintermedius and Enterobacteriaceae infections, providing a first insight into the potential of both compounds in veterinary practice.

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.

Biofilm forming ability of coagulase-positive staphylococci isolated from animals in Ukraine

Staphylococcal biofilms are an important virulence factor that allows for effective infectious effects and colonization of the animal body. This study was devoted to the evaluation of the biofilm forming ability of different strains of Staphylococcus pseudintermedius and Staphylococcus aureus isolated from animals in Ukraine. In addition, the presence of extracellular adhesin genes icaA and icaD in S. pseudintermedius strains was determined. The density of the biofilms was determined by culturing microorganisms in 96-well plates and staining the resulting structures with crystal violet. The genes responsible for biofilm formation were identified by classical polymerase chain reaction (PCR) using primers selected from the literature. The data obtained from this study showed a direct correlation between the density of the biofilm and the number of viable cells involved in its formation. Thus, 23.1% of S. pseudintermedius strains and 25% of S. aureus strains isolated from dogs demonstrated the ability to form a dense biofilm, while 46.2% of S. pseudintermedius strains and 50% of S. aureus strains formed a weak biofilm. The origin of the isolates had no significant effect on the biofilm characteristics. Coagulase-positive staphylococci obtained from cats did not form dense biofilms. 42.9% of methicillin-resistant Staphylococcus aureus (MRSA) isolates from cows with mastitis had moderate to strong biofilm forming properties. Isolates that showed resistance to three or more antibiotic groups tended to form denser biofilms. In addition, 73.3% of the studied S. pseudintermedius strains were found to contain the icaA gene, and 90% – the icaD gene. A genotypic profile combining both icaA and icaD genes was present in 66.7% of the bacteria, while one strain lacked both genes. Understanding the biofilm forming properties of staphylococcal isolates is important in the context of developing optimal treatment strategies and effective antibiotic use, which will contribute to better control of infections caused by these microorganisms.

Microbiological and molecular genetic characterization of Staphylococcus aureus and Staphylococcus pseudintermedius

Coagulase-positive staphylococci are an important infectious agentcausing numerous infections in animals. Staphylococcus aureus andStaphylococcus pseudintermedius share a number of similar cultural andbiochemical characteristics, which makes their differentiation difficult.Since these species have different zoonotic potential, it is advisable todevelop rapid and specific schemes for species differentiation of pathogens.We have studied the cultural and biochemical characteristics ofStaphylococcus spp. isolated from dogs, cats and cows. In total, 103halophilic coccal cultures were isolated from companion animals and45 from cows, of which 55 and 30 cultures were coagulase-positivestaphylococci, respectively. The reactions that can be used to differentiateS. pseudintermedius and S. aureus were studied. Growth inhibition zonesaround the disk with polymyxin B antibiotic for S. pseudintermediuswere statistically higher (p<0.001) than for S. aureus. The determinationof acetone production to differentiate between these pathogens hasless specificity, as 30% of S. pseudintermedius showed a false-positivereaction. The belonging of two isolates to the species Staphylococcuspseudintermedius was confirmed by MALDI-TOF.The virulence of staphylococci is due to the presence of genesthat regulate the synthesis of various pathogenicity factors and causeantibiotic resistance. Molecular genetic methods can detect the presenceof gene specificity and help to assess the risk of a particular strain causinginfection. Using classical and real-time PCR, the mecA gene was detectedin 8 S. aureus strains and 1 S. pseudintermedius strain that showedphenotypic resistance to methicillin. The pathogenicity genes lukF andsiet were present in 100%, and the lukS gene in 90% of the studiedStaphylococcus pseudintermedius.The study highlights a number of aspects of the diagnosis anddifferentiation of coagulase-positive staphylococci. The possibility ofusing the Neonatal FAST well D-ONE microculture system for use inveterinary laboratories was also studied. The data obtained can be usedto develop methodological approaches to the identification of pathogenicstaphylococci using a combination of different methods. Key words: S. pseudintermedius, resistance to methicillin,pathogenicity genes, MALDI-TOF MS.

Detection of Antimicrobial Resistance and Biofilm Production Among Staphylococcus pseudintermedius from Canine Skin Lesions.

Aims: Staphylococcus pseudintermedius is an opportunistic pathogen also indicated as one of the major causes of skin infections in dogs. This study aimed to identify S. pseudintermedius isolated from canine skin lesions, evaluate their antibiotic resistance profile and biofilm production ability. Methodology: Lesions from 50 rural dogs with different skin lesions were sampled after pyoderma diagnosis by private practices. Bacterial species determination was investigated and susceptibility to nine antimicrobials were determined by means of Kirby-Bauer assay. Then seven antibiotic resistance genes, including mecA, blaZ, tetK, tetM, blaSHV, blaOXA-1, and blaTEM were screened by PCR. Moreover, biofilm formation ability of the strains was determined using the microtiter plate assay along with the presence of icaADBC genes. Results: A total of 37 (74%) isolates were identified as S. pseudintermedius. All S. pseudintermedius isolates were resistant to multiple drugs. Resistance to penicillin, amoxicillin-clavulanic acid, and cefazolin were higher than other antimicrobials. All the beta-lactam-resistant isolates carried blaZ, whereas mecA was found in 6 (16.21%) of them. Among tetracycline-resistant strains, the frequency of tetK and tetM determinants were 19 (90.47%) and 21 (100%), respectively. Finally, most cefazolin-resistant strains 31 (91.89%) were positive for blaTEM gene. The rate of biofilm production ability among S. pseudintermedius isolates was 35 (94.59%). Furthermore, the presence of icaA/D genes was detected in 35 (100%) and 29 (82.85%) of S. pseudintermedius strains, respectively. Conclusion: The high rates of antimicrobial resistance and biofilm production ability among S. pseudintermedius isolates, emphasize the urgent need to use more effective infection control policies and guidelines for antimicrobial use.

An investigation of household dogs as the source in a case of human bacteraemia caused by Staphylococcus pseudintermedius

ABSTRACT Staphylococcus pseudintermedius is a commensal and an opportunistic pathogen in dogs, and is also an opportunistic pathogen in humans. Here we report about a case of bacteraemia with a fatal outcome in a 77-year-old co-morbid male likely caused by a S. pseudintermedius and the investigation into the possible transmission from the two dogs in the patient’s household. The two dogs carried the same S. pseudintermedius strain, but this dog strain was unrelated to the strain from the patient. In contrast to the patient strain, the dog strain showed reduced susceptibility to several antibiotics and both dogs had received antibiotic treatment prior to sampling. So, it is conceivable that these treatments can have eliminated the patient’s strain between the transmission event and the dog sampling. It is also worth noting that the patient strain was positive for the expA gene, which encodes an exfoliative toxin closely related to the S. aureus exfoliative toxin B. This toxin has been linked to canine pyoderma, but its effect on humans remains unknown. Transmission of S. pseudintermedius was confirmed in the household between the dogs. However, we could not verify that the dogs were the source for the S. pseudintermedius in the patient.

Potential natural antimicrobial and antibiofilm properties of Piper betle L. against Staphylococcus pseudintermedius and methicillin-resistant strains

Ethnopharmacological relevancePiper 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. Aim of the studyThe antimicrobial and antibiofilm activities of P. betle extract were assessed against S. pseudintermedius and MRSP strains. Materials and methodsEthanol 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. ResultsThe 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. ConclusionsP. 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.

Prevalence and clonal diversity of methicillin-resistant Staphylococcus aureus andmethicillin-resistant Staphylococcus pseudintermedius isolated from dogs andcatswith eye discharge.

Methicillin-resistant Staphylococcus (MRS) has originated, spread extensively, and become a prominent source of bacterial infections in both human and animal. 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. 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. Inaddition, the lineages ST4155 and ST7217 of two methicillin-resistant S. aureus strains of cats were connected epidemiologically to previously reported cases. 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 veterinarymedicine under the One Health concept.

Cheilitis in an atopic dermatitis patient associated with co-infection of Staphylococcus pseudintermedius and Staphylococcus aureus

BackgroundAtopic dermatitis (AD) is an inflammatory skin condition distinguished by an activated Th2 immune response. The local skin microbial dysbiosis is a contributing factor to the development of AD. The pathogenic coagulase-positive Staphylococcus aureus is the primary species responsible for the progression of AD. Even though Staphylococcus pseudintermedius is an animal-origin pathogen, it is increasingly becoming a source of concern in human diseases. As another coagulase-positive Staphylococci, it is crucial to pay more attention to S. pseudintermedius isolated from the lesion site.ResultsIn our investigation, we presented a case of cheilitis in a patient with atopic dermatitis (AD). We utilized culture and next-generation genomic sequencing (NGS) to identify the bacteria present on the skin swabs taken from the lip sites both prior to and following treatment. Our findings indicated that the predominant bacteria colonizing the lesion site of AD were S. pseudintermedius and S. aureus, both of which were eradicated after treatment. The Multi-locus sequence typing (MLST) of S. pseudintermedius and S. aureus demonstrated coordinated antibiotic susceptibility, with ST2384 and ST22 being the respective types. Although the skin abscess area resulting from S. pseudintermedius infection was significantly smaller than that caused by S. aureus in mice, the expression of cytokines interleukin-4 (IL-4) and interleukin-5 (IL-5) were significantly higher in the S. pseudintermedius-infected mice.ConclusionsThe S. pseudintermedius strain isolated from the lesion site of the AD patient exhibited a higher expression of IL-4 and IL-5 when colonized on mouse skin, as compared to S. aureus. This observation confirms that S. pseudintermedius can effectively induce the Th2 response in vivo. Our findings suggest that animal-origin S. pseudintermedius may play a role in the development of AD when colonized on the skin, emphasizing the importance of taking preventive measures when in contact with animals.

Exploring Efflux as a Mechanism of Reduced Susceptibility towards Biocides and Fluoroquinolones in Staphylococcus pseudintermedius.

Staphylococcus pseudintermedius is the main bacterial cause of skin and soft tissue infections (SSTIs) in companion animals, particularly dogs. The emergence of methicillin-resistant S. pseudintermedius (MRSP) strains, frequently with multidrug resistance phenotypes is a public health concern. This study aimed to evaluate efflux, a resistance mechanism still poorly characterized in S. pseudintermedius, as a contributor to biocide and fluoroquinolone resistance. Susceptibility to the efflux pump substrates ethidium bromide (EtBr), tetraphenylphosphonium bromide (TPP) and ciprofloxacin (CIP) was evaluated by minimum inhibitory concentration (MIC) determination for 155 SSTIs-related S. pseudintermedius in companion animals. EtBr and TPP MIC distributions were analyzed to estimate cut-off (COWT) values. The effect of the efflux inhibitors (EIs) thioridazine and verapamil was assessed upon MICs and fluorometric EtBr accumulation assays, performed with/without glucose and/or EIs. This approach detected a non-wild type population towards TPP with increased efflux, showed to be strain-specific and glucose-dependent. Resistance to fluoroquinolones was mainly linked to target gene mutations, yet a contribution of efflux on CIP resistance levels could not be ruled out. In sum, this study highlights the relevance of efflux-mediated resistance in clinical S. pseudintermedius, particularly to biocides, and provides a methodological basis for further studies on the efflux activity on this important pathogen of companion animals.

Virulent potential of methicillin-resistant and methicillin-susceptible Staphylococcus pseudintermedius in dogs

Staphylococcus pseudintermedius is a zoonotic pathogen responsible for several infectious diseases in pet animals, yet its pathogenic potential is not fully understood. Thus, this study aims to unravel the virulence profile of S. pseudintermedius from canine origin. Methicillin-resistant (MRSP) and methicillin-susceptible (MSSP) strains were isolated from different infection sites and their genotypic and phenotypic features were compared to determine the clinical implications of MRSP and MSSP strains. Bacterial identification was performed using MALDI-TOF and 16S-rDNA sequencing. In addition, we used multilocus sequence typing (MLST) for strains’ sequence type (ST) determination and phylogenetic relationship. The strains were screened for toxin genes, including cytotoxins (lukS, lukF), exfoliative toxin (siet), enterotoxins (sea, seb, sec, secCanine, sel, sem, and seq) and toxic shock syndrome toxin (tst-1). In vitro phenotypic analyses assessing antimicrobial susceptibility profile, biofilm formation ability, and expression of extracellular matrix components were performed. The investigated S. pseudintermedius strains belong to 17 unique ST, most of which were classified as ST71. MSSP and MRSP strains shared siet, lukS, and lukF virulence markers. Our findings showed that some MSSP strains also harbored sel, seq, and sem enterotoxin genes, suggesting a more diverse virulence profile. All MRSP strains and 77% of MSSP strains were classified as multidrug resistant (MDR). Moreover, all investigated S. pseudintermedius strains showed strong biofilm formation ability. In summary, our findings highlight the wide spread of highly virulent and drug-resistant zoonotic S. pseudintermedius strains, being a potential concern for One Health issues.

In Vitro Antimicrobial Activity of Piper betle Leaf Extract and Some Topical Agents against Methicillin-Resistant and Methicillin-Susceptible Staphylococcus Strains from Canine Pyoderma.

As multidrug-resistant methicillin-resistant staphylococci (MRS) is becoming more prevalent in canine pyoderma, the discovery of new therapeutic options is required. This study aimed to test the antimicrobial activity of crude Piper betle leaf extract and some topical antimicrobial agents against canine Staphylococcus clinical strains by determining the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC). The results showed that the mean MICs of chlorhexidine, miconazole, crude P. betle leaf extract, azelaic acid, and benzoyl peroxide against Staphylococcus strains were 1.41, 1.62, 252.78, 963.49, and 1342.70 mg/L, respectively. Therefore, betel leaf extract demonstrated a superior efficacy to azelaic acid and benzoyl peroxide. Furthermore, the ratio of MBC/MIC of betel leaf extract was 1.75, indicating its bactericidal action. When applied to methicillin-resistant S. pseudintermedius (MRSP) and methicillin-susceptible S. pseudintermedius (MSSP), betel leaf extract was equally efficient towards both groups. S. pseudintermedius strains were more susceptible to betel leaf extract than S. schleiferi subsp. coagulans. In gas chromatography-mass spectrometry analysis, eugenol and hydroxychavicol appeared to be the major components of betel leaf extract. Given its efficacy, dogs with pyoderma could benefit from the use of betel leaf extract as a topical antimicrobial alternative.

1H NMR metabolic profiling of Staphylococcus pseudintermedius isolated from canine uroliths.

Staphylococcus pseudintermedius is a urease-producing bacteria which is a major cause of magnesium ammonium phosphate (MAP) urolithiasis in canine. A positive urolith culture is an important risk factor for MAP urolithiasis in canine. The mechanism underlying the metabolic changes of S. pseudintermedius after crystallization in artificial urine (AU) needs more defined baseline metabolic information. Therefore, we extensively investigated the metabolic changes of S. pseudintermedius extensively after crystallization in AU. A high urease activity and positive biofilm formation strain, entitled the S. pseudintermedius (SPMAP09) strain, was isolated from canine MAP uroliths, and analyzed using nuclear magnetic resonance (NMR) spectroscopy-based metabolomics. The molecular mechanism-specific metabolic phenotypes were clearly observed after crystallization in AU at day 3. The crystals induced by SPMAP09 were also confirmed and the major chemical composition identified as struvite. Interestingly, our findings demonstrated that a total of 11 identified metabolites were significantly changed. The levels of formate, homocarnosine, tyrosine, cis-aconitate, glycolate, ethyl malonate, valine and acetate level were significantly higher, accompanied with decreased levels of inosine, glucose, and threonine at day 3 compared with the initial time-point (day 0). In addition, our results exhibited that the glyoxylate and dicarboxylate metabolism was significantly related to the SPMAP09 strain at day 3 in AU. Thus, metabolic changes of the SPMAP09 strain after crystallization in AU potentially helps to explain the preliminary molecular mechanism for the crystals induced by S. pseudintermedius.

Pathogenic potential and antimicrobial resistance of Staphylococcus pseudintermedius isolated from human and animals

Crossing of interspecies barriers by microorganisms is observed. In recent years, Staphylococcus pseudintermedius—a species formerly thought to be animal—has also been isolated from human clinical materials. Many virulence factors are responsible for the colonization, which is the first step an infection, of the new host organism. We analyzed the factors influencing this colonization as well as susceptibility to antibiotics in fourteen S. pseudintermedius strains isolated from clinical cases from humans and animals. The occurrence of genes responsible for binding elastin, fibronectin, and fibrinogen and some phenotypic features, although different between strains, is comparable in both groups. However, the animal isolates had more genes coding for virulence factors. All isolates tested had the exfoliating toxin gene and the leukotoxin determining genes, but only the human strains had enterotoxin genes. The assessment of antibiotic resistance of strains of both groups indicates their broad resistance to antibiotics commonly used in veterinary medicine. Antibiotic resistance was more common among animal isolates. The multilocus sequence typing analysis of the studied strains was performed. The results indicated a large diversity of the S. pseudintermedius population in both studied groups of strains. Equipped with important virulence factors, they showed the ability to infect animals and humans. The clonal differentiation of the methicillin-susceptible strains and the multidrug resistance of the strains of both studied groups should be emphasized. The considerable genetic diversity of strains from a limited geographical area indicates the processes of change taking place within this species. Thus, careful observation of the ongoing process of variation is necessary, as they may lead to the selection of S. pseudintermedius, which will pose a significant threat to humans.

Risk Factors for Antimicrobial Resistance of Staphylococcus Species Isolated from Dogs with Superficial Pyoderma and Their Owners

The microbial communities on the skin of dogs include several species of bacteria, which contribute to skin health and disease. Staphylococcus pseudintermedius, cultured at high frequency from the skin of dogs, is an opportunistic pathogen causing superficial pyoderma. Effective treatment against S. pseudintermedius infections is an important issue in veterinary medicine. However, multiple antibiotic-resistant mechanisms gradually developed by bacteria make treatment more challenging nowadays. Drug-resistant genes may have the chance to be transferred from infected dogs to other staphylococci in humans. The objective of this survey is to investigate the bacterial species that cause canine superficial pyoderma and characterize the antibiotic-resistant profiles and drug-resistant genes of isolated S. pseudintermedius. In addition, the possible risk factors causing S. pseudintermedius colonizing owners were also evaluated by a questionnaire survey. Sixty-five bacteria were isolated from dogs with superficial pyoderma, which included 47 S. pseudintermedius (72.3%), 12 other staphylococci (18.5%), 4 other Gram-positive bacteria (6.2%) and 2 Gram-negative bacteria (3.1%). Strains containing mecA and blaZ genes showed multiple-drug resistance characteristics. Dogs that received antimicrobial treatment within a recent month were at significantly higher risk of MRSP infections. Only five S. pseudintermedius strains (8.33%) were isolated from 60 samples of owners. Risk factor analysis indicated there was no significant association between S. pseudintermedius isolated from dogs and owners, but the “Keeping three or more dogs” and “Dogs can lick the owner’s face” have high odds ratios of 3.503 and 5.712, respectively. MRSP isolates belonged to three different dru types, including dt11y (29.41%), dt11a (47.06%) and dt10cp (23.53%). In conclusion, the major pathogen of canine superficial pyoderma is found to be S. pseudintermedius in Taiwan, and isolates which are mecA- or blaZ-positive are generally more resistant to commonly used antibiotics. Although S. pseudintermedius isolated from the owners might be transferred from their dogs, definite risk factors should be examined in the future study.

Interspecies transfer of plasmid-borne gentamicin resistance between Staphylococcus isolated from domestic dogs to Staphylococcus aureus.

As preconized by the One Health concept, the intimate relationship between pets and owners is a common source for the trade of microorganisms with zoonotic potential, and with them, antimicrobial resistance genes. In this work, we evaluated the presence of antimicrobial resistance genes, that are usually within mobile genetic elements, in a laboratory collection of 79 canine Staphylococcus strains, mostly Staphylococcus pseudintermedius and Staphylococcus coagulans. Resistance to tetracycline was observed in 34% of the strains, followed by resistance to erythromycin (21%) and gentamicin (19%). These phenotypes were partially correlated with the presence of the tetracycline resistance genes tet(M) and tet(K) in 64% and 44% of all strains, respectively; erythromycin resistance genes erm(A) and erm(C) in 53% and 23%; and gentamicin resistance gene aac(6')-aph(2″) in 26% of the strains. At least 45% of the strains harbored high- and/or low-molecular weight plasmids, whose transfer may be facilitated by their widespread biofilm-forming capacity, and absence of restrictive CRISPR systems. We selected eight plasmid-bearing and multidrug resistant strains, which were submitted to plasmid curing by stress with SDS. No strain lost resistance during stressing cultivation but, by conjugation experiments, the S. pseudintermedius strain 27 transferred its plasmid-borne resistance to gentamicin, conferred by the aac(6')-aph(2″) gene, to Staphylococcus aureus. The frequent empirical use of gentamicin to treat skin and ear infections in domestic dogs is likely to select resistant strains. Also, as demonstrated by our study, these strains can serve as gene reservoirs for human pathogens, such as S. aureus.