Characterization of linezolid- and methicillin-resistant coagulase-negative staphylococci in a tertiary hospital in China.

BACKGROUND: Recently, linezolid-resistant staphylococci have become an emerging problem worldwide. Understanding the mechanisms of resistance, molecular epidemiology and transmission of linezolid-resistant CoNS in hospitals is very important. METHODS: The antimicrobial susceptibilities of all isolates were determined by the microdilution method. The resistance mechanisms and molecular characteristics of the strains were determined using whole-genome sequencing and PCR. RESULTS: All the strains were resistant to oxacillin and carried the mecA gene; 13 patients (36.1%) had prior linezolid exposure. Most S. epidermidis and S. hominis isolates were ST22 and ST1, respectively. MLST typing and evolutionary analysis indicated most linezolid-resistant CoNS strains were genetically related. In this study, we revealed that distinct CoNS strains have different mechanisms of linezolid resistance. Among ST22-type S. epidermidis, acquisition of the T2504A and C2534T mutations in the V domain of the 23 S rRNA gene, as well as mutations in the ribosomal proteins L3 (L101V, G152D, and D159Y) and L4 (N158S), were linked to the development of linezolid resistance. In S. cohnii isolates, cfr, S158Y and D159Y mutations in the ribosomal protein L3 were detected. Additionally, emergence of the G2576T mutation and the cfr gene were major causes of linezolid resistance in S. hominis isolates. The cfr gene, G2576T and C2104T mutations, M156T change in L3 protein, and I188S change in L4 protein were found in S. capitis isolates. CONCLUSION: The emergence of linezolid-resistant CoNS in the environment is concerning because it involves clonal dissemination and frequently coexists with various drug resistance mechanisms.

Allosteric Probe-Based Colorimetric Assay for Direct Identification and Sensitive Analysis of Methicillin Resistance of Staphylococcus aureus.

The accurate and rapid detection of methicillin-resistance of Staphylococcus aureus (SA) holds significant clinical importance. However, the methicillin-resistance detection strategies commonly require complicated cell lysis and gene extraction. Herein, we devised a novel colorimetric approach for the sensitive and accurate identification of methicillin-resistance of SA by combining allosteric probe-based target recognition with self-primer elongation-based target recycling. The PBP2a aptamer in the allosteric probe successfully identified the target MRSA, leading to the initiation of self-primer elongation based-cascade signal amplification. The peroxidase-like hemin/G-quadruplex undergo an isothermal autonomous process that effectively catalyzes the oxidation of ABTS2- and produces a distinct blue color, enabling the visual identification of MRSA at low concentrations. The method offers a shorter duration for bacteria cultivation compared to traditional susceptibility testing methods, as well as simplified manual procedures for gene analysis. The overall amplification time for this test is 60 min, and it has a detection limit of 3 CFU/ml. In addition, the approach has exceptional selectivity and reproducibility, demonstrating commendable performance when tested with real samples. Due to its advantages, this colorimetric assay exhibits considerable potential for integration into a sensor kit, thereby offering a viable and convenient alternative for the prompt and on-site detection of MRSA in patients with skin and soft tissue infections.

Methicillin-resistant staphylococci in clinical bovine mastitis: occurrence, molecular analysis, and biofilm production.

Staphylococcus aureus is an important pathogen that causes mastitis in cattle, and the emergence of methicillin-resistant S. aureus (MRSA) poses a threat to veterinary and human medicine. The aims of the study were to investigate the prevalence of MRSA and methicillin-resistant coagulase-negative staphylococci (MR-CoNS) isolated from clinical mastitis, their ability to form biofilms, and the antimicrobial susceptibility of S. aureus strains. In addition, the Staphylococcal Cassette Chromosome mec (SCCmec) type, spa type and the presence of Panton-Valentine Leucocidin in MRSA were evaluated. A total of 326 staphylococcal strains were screened by multiplex-PCR for S. aureus and Staphylococcus intermedius group (SIG) identification. The S. aureus strains (n = 163) were subjected to phenotypic testing for antimicrobial susceptibility and biofilm formation. Molecular analysis was performed on MRSA mecA-positive strains. Of 163 S. aureus isolates, 142 strains (87.1%) were resistant to at least one antibiotic, and all 19 MRSA strains were resistant to at least four out of five antibiotics tested. All S. aureus strains harboured the icaA gene and were biofilm producers. Nineteen MR-CoNS strains were also isolated. The most prevalent spa types among MRSA were t001 (57.9%) and t037 (31.6%), while one MRSA was type t008 and one was type t041. Most MRSA were SCCmec type I (63.2%) and III (31.6%) and only one strain was type IV. None of the MRSA isolates had the PVL gene. The prevalence of multidrug-resistant S. aureus in bovine mastitis is a serious concern. The finding of MRSA with spa types predominant in humans and infrequent in Italian cows and with SCCmec infrequently found in bovine milk or cheese suggest a human origin of these strains. The ability of MRSA and MR-CoNS involved in bovine mastitis to be transferred to humans and vice versa poses a public health concern.

Recombination-mediated dissemination of Methicillin-resistant S. aureus clonal complex 1 in the Egyptian health care settings.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) is a rapidly evolving pathogen that is frequently associated with outbreaks and sustained epidemics. This study investigated the population structure, resistome, virulome, and the correlation between antimicrobial resistance determinants with phenotypic resistance profiles of 36 representative hospital-acquired MRSA isolates recovered from hospital settings in Egypt. RESULTS: The community-acquired MRSA lineage, clonal complex 1 (CC1) was the most frequently detected clone, followed by three other globally disseminated clones, CC121, CC8, and CC22. Most isolates carried SCCmec type V and more than half of isolates demonstrated multi-drug resistant phenotypes. Resistance to linezolid, a last resort antibiotic for treating multidrug resistant MRSA, was observed in 11.11% of the isolates belonging to different genetic backgrounds. Virulome analysis indicated that most isolates harboured a large pool of virulence factors and toxins. Genes encoding aureolysin, gamma hemolysins, and serine proteases were the most frequently detected virulence encoding genes. CC1 was observed to have a high pool of AMR resistance determinants including cfr, qacA, and qacB genes, which are involved in linezolid and quaternary ammonium compounds resistance, as well as high content of virulence-related genes, including both of the PVL toxin genes. Molecular clock analysis revealed that CC1 had the greatest frequency of recombination (compared to mutation) among the four major clones, supporting the role of horizontal gene transfer in modulating AMR and hypervirulence in this clone. CONCLUSIONS: This pilot study provided evidence on the dissemination success of CA-MRSA clone CC1 among Egyptian hospitals. Co-detection of multiple AMR and virulence genes in this lineage pose a broad public health risk, with implications for successful treatment. The results of this study, together with other surveillance studies in Egypt, should be used to develop strategies for controlling MRSA infections in Egyptian health-care settings.

Molecular and Phenotypic Characterization of Ocular Methicillin-Resistant Staphylococcus epidermidis Isolates in Taiwan.

Purpose: Staphylococcus epidermidis, a commensal, has emerged as an important opportunistic pathogen, particularly methicillin-resistant S. epidermidis (MRSE). The mechanism behind this transformation remains unclear. This study aimed to investigate the molecular and phenotypic characteristics of MRSE isolated from healthy conjunctiva and ocular infections. Methods: We collected MRSE isolates from two groups: healthy conjunctiva from patients undergoing cataract surgeries and ocular infections at our hospital. Genotypic analysis included pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), staphylococcal cassette chromosome mec (SCCmec), and biofilm-related genes (icaA, aap, and bhp). Additionally, phenotypic data on biofilm production and antibiotic susceptibility were recorded. Results: A total of 86 isolates, including 42 from healthy conjunctiva and 44 from ocular infections, were analyzed. MLST identified 21 sequence types (STs), with ST59 being the most frequent (n = 33, 39.5%), followed by ST130 (n = 10, 11.6%), ST57 (n = 6, 7.0%), and ST2 (n = 6, 7.0%). All isolates were categorized in 23 PFGE types, and SCCmec IV was the most prevalent SCCmec type (n = 52, 60.5%). The two sources of isolates exhibited overlapping molecular types and phenotypic traits, although the ocular infection isolates exhibited significantly higher multidrug resistance compared to healthy conjunctiva isolates (P = 0.032). When contrasting ST59 with non-ST59, ST59 displayed a significantly higher presence of aap (100%) and bhp (69.7%) while lacking icaA (0%). ST59 also showed lower susceptibility to fluoroquinolones compared to non-ST59 (42.4%-54.5% vs. 75.5%-83.0%; P < 0.01). Conclusions: MRSE isolates from healthy conjunctiva and ocular infections demonstrated a degree of resemblance. Specific strains, notably ST59, exhibited distinctive characterizations.

A signal-off Cas14a1-based platform for highly specific detection of methicillin-resistant Staphylococcus aureus.

Antibiotic usage has become very widespread in aquaculture, and the abuse or overuse of antibiotics has led to the evolution of antibiotic-resistance bacteria, which has adverse effects on aquatic products and ecosystems. Moreover, this evolution can potentially cause harm to human health. Thus, there is an urgent need for diagnostic tools for antibiotic-resistant microorganisms. Herein, we proposed a signal-off Cas14a1-based platform (SOCP) for the detection of methicillin-resistant Staphylococcus aureus (MRSA). In this SOCP, we have designed single-stranded DNA (ssDNA) that not only can activate the trans-cleavage ability of dual Cas14a1-sgRNA complex but also can be used as the primers for the amplified methicilin-resistant gene (mecA). When MRSA is present, the primers can be transformed into products with amplification, leading to the signal decrease of trans-cleavage activity of Cas14a1. The SOCP showed high specificity and fair sensitivity for mecA gene and MRSA. In the detection of real samples, this platform also showed consistent results compared with qPCR. The SOCP could provide an alternative tool for the diagnosis of antibiotic-resistant bacteria in aquaculture, food industry and other fields.

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.

Evaluation of the performance of GeneSoC®, a novel rapid real-time PCR system, to detect Staphylococcus aureus and methicillin resistance in blood cultures.

Staphylococcus aureus bacteremia results in substantial mortality. Rapid identification and the determination of methicillin susceptibility are crucial for immediate treatment with appropriate antibiotics. In the present study, we aimed to evaluate the basic assay performance of GeneSoC®, a novel rapid quantitative polymerase chain reaction (qPCR) method, for the detection of methicillin-susceptible (MS) or -resistant (MR) S. aureus in blood culture (BC) bottles. qPCR pimers and probes were desinged for femA and mecA genes to diagnose S. aureus and its methicilline-resistance status. GeneSoC® system can detect target genes within 12 min per sample using microfludic thermal cycling. A total of 100 BC-positive samples, showing clusters of gram-positive cocci using microscopy, were tested. The analytical sensitivity was demonstrated for the target sequence of femA and mecA genes at 10 copies/µL, respectively. The detection limit of the MRSA bacterial burden using this system was 104 and 103 CFU/mL for femA and mecA, respectively. Compared with culture-based identification and susceptibility testing, the sensitivity and specificity for the detection of femA (+)/mecA (+) MRSA using GeneSoC® were 90.9 and 98.9%, respectively, whereas the sensitivity and specificity for detection of femA (+)/mecA (-) MSSA were 96.2% and 97.3%, respectively. In conclusion, although this was a small sample and pilot study, the GeneSoC® system is beneficial for rapid, reliable, and highly sensitive real-time testing of MRSA and MSSA in BC bottles.

Genomic analysis of a multidrug methicillin-resistant staphylococcus epidermidis recovered from the urine of a guinea pig (Cavia porcellus) with suspected pyelonephritis.

Staphylococcus epidermidis is an emerging pathogen causing infant pyelonephritis. There is a lack of genomic data on Staphylococcus epidermidis as the etiology of pyelonephritis and its resistant determinants. In this study, we have conducted a genomic and microbiologic investigation of an S. epidermidis recovered from the urine of a guinea pig with suspected pyelonephritis in Brazil. The genome was sequenced using the Illumina MiSeq platform and de novo assembled using SPades. Resistome, virulome, and plasmidome were in silico predicted using bioinformatics tools. Data analysis revealed that S. epidermidis USP-LZB-G06 belonged to sequence type ST332. Strikingly, a broad resistome (antibiotics, hazardous heavy metals, and biocides) was predicted, including the presence of the clinically relevant mecA, blaZ, and qacA efflux pump genes. SNP-based analysis revealed that strain USP-LZB-G06 was clustered along mecA positive S. epidermidis strains of ST332 isolated between 2008 and 2016 from humans in Australia and the United States of America. Our results indicate that the detection of this microorganism should be considered as a urinary tract infection agent in exotic pets, particularly guinea pigs. In addition, there is an urgent need to update veterinarians regarding the detection and therapeutic management of these microorganisms.

Lactoferrin modulates the biofilm formation and bap gene expression of methicillin-resistant Staphylococcus epidermidis.

Lactoferrin is an innate glycoprotein with broad antibacterial and antibiofilm properties. The autonomous antibiofilm activity of lactoferrin against Gram-positive bacteria is postulated to involve the cell wall and biofilm components. Thus, the prevention of biomass formation and eradication of preformed biofilms by lactoferrin was investigated using a methicillin-resistant Staphylococcus epidermidis (MRSE) strain. Additionally, the ability of lactoferrin to modulate the expression of the biofilm-associated protein gene (bap) was studied. The bap gene regulates the production of biofilm-associated proteins responsible for bacterial adhesion and aggregation. In the in vitro biofilm assays, lactoferrin prevented biofilm formation and eradicated established biofilms for up to 24 and 72 h, respectively. Extensive eradication of MRSE biofilm biomass was accompanied by the significant upregulation of bap gene expression. These data suggest the interaction of lactoferrin with the biofilm components and cell wall of MRSE, including the biofilm-associated protein.

Novel cassette chromosome recombinases CcrA8B9 catalyse the excision and integration of the staphylococcal cassette chromosome mec element.

OBJECTIVES: A defining feature of MRSA is the SCCmec element. The excision and integration of SCCmec elements are catalysed by Ccr recombinases. Currently, seven ccrA, eight ccrB and two ccrC allotypes have been described. However, there have been no recent reports of a novel Ccr recombinase and thus this area should be explored. METHODS: According to the proposed criteria of the International Working Group on the Classification of Staphylococcal Cassette Chromosome Elements (IWG-SCC) committee, novel ccr genes were explored by searching the genome of our laboratory staphylococcal strains, which were isolated from bovine mastitis in Northwest China. The biological activity of the novel Ccr recombinases to excise and integrate SCCmec elements was determined. The distribution of the novel ccr genes in staphylococci was conducted by querying the NCBI nr/nt database. RESULTS: We report a set of novel Ccr recombinases CcrA8B9, which share nucleotide identities of 46.6%-50.2% and 47.4%-52.8% with the ccrA and ccrB alleles, respectively. We used PCR to show that CcrA8B9 can excise and integrate the SCCmec element. Furthermore, using NCBI BLAST we showed that the ccrA8B9 genes exist in other staphylococcal strains. Unlike the common ccr genes, ccrA8B9 is located outside the SCCmec/SCC element. CONCLUSIONS: The novel Ccr recombinases CcrA8B9 can help excise and integrate SCCmec/SCC from the genome and provide a new way to facilitate the transmission of SCCmec/SCC elements among staphylococci.

Characteristics of population structure, antimicrobial resistance, virulence factors, and morphology of methicillin-resistant Macrococcus caseolyticus in global clades.

Macrococcus caseolyticus is an opportunistic pathogen that is frequently isolated from dairy products and veterinary infections. Recent studies have reported the possibility of methicillin resistance that be transferred among staphylococcal species in foods. The present study examined the population structure, antimicrobial resistance, virulence factors, and morphology of methicillin-resistant M. caseolyticus by investigation of 94 genomes derived from both isolates in beef (n = 7) and pork (n = 2) at Shanghai and those deposited in public domain (n = 85). Phylogenetically, M. caseolyticus were divided into four clades, which each consisted of genomes isolated from continent of European countries (82.4%, n = 78), Asian countries (11.3%, n = 10), United States (4.1%, n = 4), Australia (1%, n = 1), and Sudan (1%, n = 1). The M. caseolyticus isolated from present study formed a genetically distinguished clade, which was characterized by novel alleles in the traditional 7-gene MLST scheme. Furthermore, we identified 24 AMR genes that were associated with 10 classes of antimicrobial agents in M. caseolyticus. Most AMR genes were carried by dominant plasmids such as rep7a, rep22 and repUS56. The genomes in the global clades carried significantly less AMR genes (p < 0.05) and more virulence factors (p < 0.001) than present clade. Virulence factors were detected in methicillin resistant M. caseolyticus including genes coding hemolysin, adherence, biofilm formation, exotoxin, and capsule that associated to human health and infection. Finally, as the close relative of the genus Staphylococcus, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were performed for morphological comparison that M. caseolyticus has a larger diameter and thicker cell wall compared with S. aureus ATCC 25,923. Taken together, our study suggested that M. caseolyticus mediating divergent antimicrobial resistance and virulence factors could serve as the vector for methicillin resistance habitats in foodborne microorganisms.

Frequent dissemination and carriage of an SCCmec-mecC hybrid in methicillin-resistant Mammaliicoccus sciuri in farm animals from Tunisia.

OBJECTIVES: In this study, we aimed to assess the extent of dissemination of methicillin-resistant Mammaliicoccus sciuri in animal farms in Tunisia and evaluate the distribution of virulence and methicillin resistance genes in the M. sciuri population. METHODS: Staphylococci and mammaliicocci isolated from unhealthy animals and healthy humans from adjacent farms in Tunisia were characterized for antimicrobial susceptibility, biofilm formation, agglutination, and hemolysis abilities. Mammaliicoccus sciuri relatedness and content in antibiotic resistance and virulence genes were analyzed by whole-genome sequencing (WGS). RESULTS: Mammaliicoccus sciuri was the most prevalent species (46.2%), showing the highest resistance rates to fusidic acid (94.6%), oxacillin (73%), penicillin (40.5%), clindamycin (37%), ciprofloxacin (27%), and cefoxitin (24.3%). Some isolates carried genes encoding resistance to nine different antibiotic classes. mecA was found in 35% of M. sciuri and mecC in 16.2%. All isolates carrying mecC were of S. sciuri subspecies carnaticus and carried the hybrid element SCCmec-mecC. Mammaliicoccus sciuri were able to produce strong biofilm (27%) and have clumping ability (16%). Additionally, they carried genes for capsule production (cap8, 100%), iron-regulated surface determinants (isdE, 24%; isdG, 3%), and virulence regulation (clpC and clpP, 100%). Single nucleotide polymorphisms (SNPs) analysis showed that 17 M. sciuri cross-transmission events probably occurred between different animal species and farms. Moreover, SCCmec was estimated to have been acquired five times by S. sciuri subsp. carnaticus. CONCLUSION: Multidrug resistant and pathogenic M. sciuri were frequently disseminated between different animal species within the farm environment. mecA and mecC can be disseminated by both frequent acquisition of the SCCmec element and clonal dissemination.

Nasal Staphylococcus aureus and S. pseudintermedius carriage in healthy dogs and cats: a systematic review of their antibiotic resistance, virulence and genetic lineages of zoonotic relevance.

The molecular ecology of Staphylococcus aureus, Staphylococcus pseudintermedius and their methicillin-resistant strains in healthy dogs and cats could serve as good models to understand the concept of bacterial zoonosis due to animal companionship. This study aims to provide insights into pooled prevalence, genetic lineages, virulence and antimicrobial resistance (AMR) among healthy dogs and cats. Original research and brief communication articles published from 2001 to 2021 that reported the nasal detection of S. aureus and S. pseudintermedius in healthy dogs and cats in the community, homes and outside veterinary clinics were examined and analysed. Forty-nine studies were eligible and included in this systematic review. The pooled prevalence of nasal carriage of S. aureus/methicillin-resistant S. aureus (MRSA) in healthy dogs and cats were 10.9% (95% CI: 10.1-11.9)/2.8% (95% CI: 2.4-3.2) and 3.2% (95% CI: 1.9-4.8)/0.5% (95% CI: 0.0-1.1), respectively. Conversely, the pooled prevalence of S. pseudintermedius/methicillin-resistant S. pseudintermedius (MRSP) in healthy dogs and cats were 18.3% (95% CI: 17.1-19.7)/3.1% (95% CI: 2.5-3.7) and 1.3% (95% CI: 0.6-2.4)/1.2% (95% CI: 0.6-2.3), respectively. Although highly diverse genetic lineages of S. aureus were detected in healthy dogs and cats, MSSA-CC1/CC5/CC22/CC45/CC121/CC398 and MRSA-CC5/CC93/CC22/CC30 were mostly reported in dogs; and MSSA-CC5/CC8/CC15/CC48 and MRSA-CC22/CC30/CC80 in cats. Of note, MSSA-CC398 isolates (spa-types t034 and t5883) were detected in dogs. Genetic lineages often associated with MSSP/MRSP were ST20/ST71, highlighting the frequent detection of the epidemic European MRSP-ST71 clone in dogs. S. aureus isolates carrying the luk-S/F-PV, tst, eta, etb and etd genes were seldomly detected in dogs, and luk-S/F-PV was the unique virulence factor reported in isolates of cats. S. pseudintermedius isolates harbouring the luk-S/F-I, seint and expA genes were frequently found, especially in dogs. High and diverse rates of AMR were noted, especially among MRSA/MRSP isolates. There is a need for additional studies on the molecular characterization of isolates from countries with under-studied nasal staphylococci isolates.

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.

Genetic Characteristics of Methicillin-Resistant Staphylococcus argenteus Isolates Collected in the Dutch National MRSA Surveillance from 2008 to 2021.

Staphylococcus argenteus is a recently described member of the Staphylococcus aureus complex (SAC) and is associated with human disease. The frequency and intensity of infections caused by S. argenteus are similar to those of Staphylococcus aureus. S. argenteus can harbor antibiotic resistance genes and a variety of virulence factors analogous to methicillin-resistant S. aureus (MRSA). The aim of our study was to analyze a collection of isolates in the Dutch national MRSA surveillance from January 2008 until March 2021 that were nontypeable by multilocus variable-number tandem-repeat analysis (MLVA). Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-ToF MS) was used for identifying the S. argenteus isolates, and whole-genome sequencing and SeqSphere were used to generate an in-house whole-genome multilocus sequence typing (wgMLST) scheme for typing the isolates. Furthermore, the presence of antibiotic resistance genes, replicons, and virulence genes was determined. Of 52,467 isolates submitted as MRSA from January 2008 until March 2021, 64 isolates (0.12%) were nontypeable with MLVA, and 54 of them were identified with mass spectrometry (MALDI-ToF MS) as S. argenteus. It appeared in retrospect that the first methicillin-resistant S. argenteus (MRSArg) was already submitted in 2008. An in-house-developed S. argenteus wgMLST scheme revealed that S. argenteus isolates clustered in 5 genomic groups which were characterized by distinct MLST types, resistomes, plasmid replicon families, and virulence factors. All but one isolate carried the staphylococcal chromosomal cassette mec (SCCmec) type IV harboring the methicillin resistance gene mecA and represent MRSArg. Most of the isolates with SCCmec subtype IVc(2B) had a trimethoprim resistance gene, dfrG, and harbored a blaZ-carrying plasmid, and most MRSArg isolates have the immune-modulating genes scn and sak. Nine of the 47 isolates carried enterotoxin-encoding genes seg, sei, sem, seo, and seu, which might be able to cause food poisoning. In some persons there was long-term persistence of MRSArg, and there were several genetically related MRSArg isolates in people living in close proximity, suggesting direct human-human transmission. IMPORTANCE We show that MRSArg has been circulating in the Netherlands since at least 2008. Although MRSArg is distinct from MRSA, it has a comparable population structure and carries similar resistance and virulence genes. The Dutch national MRSA surveillance has been expanded to include other methicillin-resistant members of the S. aureus complex, such as S. argenteus and Staphylococcus schweitzeri.

Alveolar osteitis associated with methicillin-resistant Staphylococcus epidermidis.

A critical point in dentistry is the empiric prescription of broad-spectrum antibiotics that could increase the levels of antimicrobial resistance. Alveolar osteitis is one of the most common post-op- erative complications in which antibiotic use is controversial. A 35-year-old female, with pain in the right mandibular region and treated with cefixime, was diagnosed with cracked tooth syndrome and pulpitis. The tooth was extracted and a massive purulent bleeding drainage was observed. Irrigation of the socket and a new therapy with azithromycin were done. Bacteriological analysis, a specific mecA gene PCR for the methicillin resistance, and the antimicrobial susceptibility test were per- formed on the bacterial isolate. A Staphylococcus epidermidis isolate was methicillin-resistant and showed resistance to erythromycin, azithromycin, clarithromycin, and sulfamethoxazole + trimeth- oprim. After 7 days, intraoral examination showed a complete resolution. The aim of this report is to suggest that systemic antibiotics may provide insufficient efficacy during alveolar osteitis, especially when caused by a multidrug-resistant organism.

Methicillin Resistant Staphylococci Isolated from Goats and Their Farm Environments in Saudi Arabia Genotypically Linked to Known Human Clinical Isolates: a Pilot Study.

We conducted a pilot whole genome sequencing (WGS) study to characterize the genotypes of nine methicillin resistant staphylococci (MRS) isolates recovered from goats and their farm environments in Eastern Province, Saudi Arabia, between November 2019 to August 2020. Seven out of nine isolates were methicillin resistant Staphylococcus aureus (MRSA), and two were methicillin resistant Staphylococcus epidermidis (MRSE). All MRSA isolates possessed genotypes previously identified to infect humans, including isolates harboring ST6-SCCmec IV-t304 (n = 4), ST5-SCCmec VI- t688 (n = 2) and ST5-SCCmec V-t311 (n = 1). 2 MRSA isolates possessed plasmids that were genetically similar to those identified in S. aureus isolates recovered from humans and poultry. In contrast, plasmids found in three MRSA isolates and one MRSE isolate were genetically similar to those recovered from humans. All MRSA isolates harbored the host innate modulate genes sak and scn previously associated with human infections. The genotypes of MRSE isolates were determined as ST35, a well-known zoonotic sequence type and ST153, which has been associated with humans. However, the MRSE isolates were untypeable due to extra ccr complexes identified in their SCCmec elements. Moreover, we identified in ST153 isolate SCCmec element also harbored the Arginine Catabolic Mobile Element (ACME) IV. All MRS isolates were phenotypically resistant to trimethoprim-sulfamethoxazole, an antibiotic for the decolonization of MRS. Three isolates carried antibiotic resistance genes in their SCCmec elements that were not previously described, including those encoding fusidic acid resistance (fusC) and trimethoprim resistance (dfrC) incorporated in the MRSA SCCmec VI. IMPORTANCE Our findings demonstrate a possible cross-transmission of methicillin resistant staphylococci between goats and their local environments and between goats and humans. Due to ever increasing resistance to multiple antibiotics, the burden of MRS has a significant impact on livestock farming, public health, and the economy worldwide. This study highlights that implementing a holistic approach to whole genome sequencing surveillance in livestock and farm environments would aid our understanding of the transmission of methicillin resistant staphylococci and, most importantly, allow us to implement appropriate infection control and hygiene practices.