MecA-related structure in methicillin-resistant coagulase-negative staphylococci from street food in Taiwan.
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ABSTRACT:
SUBMITTER: Yang TY
PROVIDER: S-EPMC5299846 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:mecA, the gene that mediates methicillin resistance, and its accompanying mec locus DNA, insert near the gyrA gene in Staphylococcus aureus. To investigate whether there is a similar relationship between mecA and gyrA in coagulase-negative staphylococci (CNS), mecA- and gyrA-specific DNA fragments were used to probe methicillin-resistant isolates of Staphylococcus epidermidis (MRSE) (n = 11) and Staphylococcus haemolyticus (MRSH) (n = 11). The gyrA probe hybridized to the same SmaI DNA fragment as the mecA probe in all strains tested. However, since the size of the SmaI fragments containing mecA and gyrA varied from 73 to 600 kb, the distance between the two genes was determined more precisely. Cloned mecA or gyrA fragments plus vector sequences each containing a SmaI site were introduced into the chromosome of three isolates each of MRSE and methicillin-resistant S. aureus (MRSA), and the sizes of the generated SmaI fragments were determined by pulsed-field gel electrophoresis. The distance between gyrA and mecA was found to be between 38 and 42 kb in both MRSE and MRSA, and the two genes were in the same relative orientation in all strains. Restriction fragment length polymorphism (RFLP) patterns around the gyrA gene in CNS were identical, but species specific, for all 10 MRSE and 10 MRSH isolates examined. In contrast, 8 of 11 methicillin-susceptible S. epidermidis isolates and 7 of 7 methicillin-susceptible S. haemolyticus isolates had different gyrA RFLP patterns. These data show that mecA is site and orientation specific, relative to gyrA, in both MRSE and MRSA. In addition, the local environment around gyrA in methicillin-resistant CNS, in contrast to methicillin-susceptible isolates, is similar, suggesting clonality or the requirement for specific DNA sequences with which the mec complex must interact for chromosomal integration to occur.
| S-EPMC105405 | biostudies-literature
Project description:Coagulase-negative staphylococci (CoNS) are an important group of opportunistic pathogenic microorganisms that cause infections in hospital settings and are generally resistant to many antimicrobial agents. We report on phenotypic and genotypic virulence characteristics of a select group of clinical, mecA-positive (encoding penicillin-binding protein 2a) CoNS isolates. All CoNS were resistant to two or more antimicrobials with S. epidermidis strain 214EP, showing resistance to fifteen of the sixteen antimicrobial agents tested. Aminoglycoside-resistance genes were the ones most commonly detected. The presence of megaplasmids containing both horizontal gene transfer and antimicrobial resistance genetic determinants indicates that CoNS may disseminate antibiotic resistance to other bacteria. Staphylococcus sciuri species produced six virulence enzymes, including a DNase, gelatinase, lipase, phosphatase, and protease that are suspected to degrade tissues into nutrients for bacterial growth and contribute to the pathogenicity of CoNS. The PCR assay for the detection of biofilm-associated genes found the eno (encoding laminin-binding protein) gene in all isolates. Measurement of their biofilm-forming ability and Spearman's rank correlation coefficient analyses revealed that the results of crystal violet (CV) and extracellular polymeric substances (EPS) assays were significantly correlated (ρ = 0.9153, P = 3.612e-12). The presence of virulence factors, biofilm-formation capability, extracellular enzymes, multidrug resistance, and gene transfer markers in mecA-positive CoNS clinical strains used in this study makes them powerful opportunistic pathogens. The study also warrants a careful evaluation of nosocomial infections caused by CoNS and may be useful in studying the mechanism of virulence and factors associated with their pathogenicity in vivo and developing effective strategies for mitigation.
| S-EPMC7284987 | biostudies-literature
Project description:Methicillin-resistant coagulase-negative staphylococci were isolated from the nares and skin of 1- to 8-week-old healthy chickens in three flocks from a farm. Isolation of methicillin-resistant coagulase-negative staphylococci was positive for 72 (25.7%) of the 280 chickens tested, with the frequency varying from 2.2 to 100% according to flock. A total of 45 appropriate isolates were selected and subjected to identification. Of the 45 methicillin-resistant coagulase-negative staphylococcal isolates selected, 37 were identified as Staphylococcus sciuri, 5 were identified as Staphylococcus epidermidis, and 3 were identified as Staphylococcus saprophyticus. The distribution of the species was different among the flocks. Comparative analysis of the SmaI-digested chromosomal DNA by pulsed-field gel electrophoresis revealed that the isolates could have originated from a single clone of each of S. sciuri and S. saprophyticus and three clones of S. epidermidis. By two methods based on the PCR technique, the mecA gene was detected in all five representative isolates of each methicillin-resistant coagulase-negative staphylococcal clone. The nucleotide sequence of a PCR fragment obtained from an isolate of S. sciuri was completely identical to the corresponding region of mecA genes reported in human methicillin-resistant Staphylococcus aureus isolates and Staphylococcus epidermidis isolates. The representative methicillin-resistant coagulase-negative staphylococcal isolates were resistant to many beta-lactam antibiotics, and some isolates were also resistant to macrolide and aminoglycoside antibiotics. This is the first evidence of the existence of methicillin-resistant coagulase-negative staphylococci from animals possessing the mecA gene.
| S-EPMC229192 | biostudies-other
Project description:BACKGROUND: Methicillin-resistant coagulase-negative staphylococci (MR-CoNS) are opportunistic pathogens and serve as a large reservoir of staphylococcal cassette chromosome mec (SCCmec). Characterization of SCCmec in MR-CoNS can generate useful information on the mobilization and evolution of this element. METHODOLOGY/PRINCIPAL FINDINGS: Non-repetitive MR-CoNS clinical isolates (n?=?84; 39 S. epidermidis, 19 S. haemolyticus, 9 S. hominis, 6 S. capitis, 4 S. warneri, 2 S. cohnii, 2 S. saprophyticus, 1 S. kloosii, 1 S. simulans and 1 S. massiliensis) were collected. All isolates could grow on plates with 4 mg/L cefoxitin and all had mecA as detected by PCR. Strain typing using RAPD and ERIC-PCR revealed that almost all isolates were of different strains. SCCmec typing was performed using multiplex PCR published previously. For isolates in which SCCmec could not be typed, the mec complex classes were determined by additional PCR and the ccr genes were amplified with published or newly-designed primers and then sequenced. SCCmec types were assigned for 63 isolates by multiplex PCR and were assigned for 14 other isolates by PCR targeting mec and ccr. Among 77 isolates with determined SCCmec types, 54 had a single type, including type III (n?=?19), IV (n?=?14), V (n?=?10), II (n?=?2), I (n?=?1), VIII (n?=?1) and five unnamed types (n?=?7), while 23 isolates had two types, III+V (n?=?12), II+V (n?=?8), II+IV (n?=?2) or IV+V (n?=?1). The five unnamed types were assigned UT1 (class A mec, ccrA1/ccrB4), UT2 (class C1 mec, ccrA4/ccrB4), UT3 (class A mec, ccrA5/ccrB3), UT4 (class C2 mec, ccrA2/ccrB2 plus ccrC1) and UT5 (class A mec, ccrA1/ccrB1 plus ccrC1). CONCLUSIONS/SIGNIFICANCE: SCCmec types III, IV and V were prevalent in MR-CoNS and many isolates could harbor more than one type. Several new types of SCCmec were identified, highlighting the great genetic diversity and the need of developing classification schemes for SCCmec in MR-CoNS.
| S-EPMC3102682 | biostudies-literature
Project description:The accurate performance of the Vitek 2 GP67 card for detecting methicillin-resistant coagulase-negative staphylococci (CoNS) is not known. We prospectively determined the ability of the Vitek 2 GP67 card to accurately detect methicillin-resistant CoNS, with mecA PCR results used as the gold standard for a 4-month period in 2012. Included in the study were 240 consecutively collected nonduplicate CoNS isolates. Cefoxitin susceptibility by disk diffusion testing was determined for all isolates. We found that the three tested systems, Vitek 2 oxacillin and cefoxitin testing and cefoxitin disk susceptibility testing, lacked specificity and, in some cases, sensitivity for detecting methicillin resistance. The Vitek 2 oxacillin and cefoxitin tests had very major error rates of 4% and 8%, respectively, and major error rates of 38% and 26%, respectively. Disk cefoxitin testing gave the best performance, with very major and major error rates of 2% and 24%, respectively. The test performances were species dependent, with the greatest errors found for Staphylococcus saprophyticus. While the 2014 CLSI guidelines recommend reporting isolates that test resistant by the oxacillin MIC or cefoxitin disk test as oxacillin resistant, following such guidelines produces erroneous results, depending on the test method and bacterial species tested. Vitek 2 cefoxitin testing is not an adequate substitute for cefoxitin disk testing. For critical-source isolates, mecA PCR, rather than Vitek 2 or cefoxitin disk testing, is required for optimal antimicrobial therapy.
| S-EPMC4313137 | biostudies-literature
Project description:Nasal carriage of methicillin-resistant coagulase-negative staphylococci (MR-CoNS) is highly prevalent in community subjects, but its dynamic has been little investigated. Nasal swabbing was performed in 2006 and 2008 in 154 Amerindians living isolated in French Guiana. MR-CoNS strains were identified and characterized by non-?-lactam susceptibility testing and staphylococcal cassette chromosome mec element (SCCmec) typing, characterizing the associations of ccr and mec gene complex allotypes, and for MR Staphylococcus epidermidis (MRSE), multilocus variable number of tandem repeats analysis (MLVA) was used. The impact of sociodemographic and medical characteristics on the persistence of MR-CoNS carriage was assessed by bivariate analysis. Prevalence of MR-CoNS carriage was 50.6% in 2006 and 46.8% in 2008. The 274 MR-CoNS isolates, including S. epidermidis (n = 89, 62 MLVA patterns), Staphylococcus haemolyticus (n = 78), and Staphylococcus hominis (n = 72), exhibited 41 distinct ccr and mec gene complex associations. Persistent carriage (in 2006 and 2008), intermittent carriage (either in 2006 or 2008), and noncarriage were documented in 25.3, 47.4, and 27.3% of the participants, respectively. Persistent carriage of a given MRSE isolate was rarely observed (n = 8 isolates). Furthermore, no epidemiological factor, including antibiotic exposure, was associated with persistent carriage. The high diversity of MRSE clones and their ccr and mec gene complex associations contrasted with the high carriage rates in this isolated community, which might reflect the occurrence of SCCmec rearrangement and the generation of new MR-CoNS strains.
| S-EPMC3256065 | biostudies-literature
Project description:Four methicillin-resistant coagulase-negative staphylococci (MRCoNS), one Staphylococcus haemolyticus and three Staphylococcus cohnii, from infections of humans collected via the Ministry of Health National Antimicrobial Resistance Surveillance Net (Mohnarin) program in China were identified as linezolid-resistant. These four isolates were negative for the 23S rRNA mutations, but positive for the gene cfr. Mutations in the gene for the ribosomal protein L3, which resulted in the amino acid exchanges Gly152Asp and Tyr158Phe, were identified in S. haemolyticus 09D279 and S. cohnii NDM113, respectively. In each isolate, the cfr gene was located on a plasmid of ca. 35.4 kb, as shown by S1 nuclease pulsed-field gel electrophoresis and Southern blotting experiments. This plasmid was indistinguishable from the previously described plasmid pSS-02 by its size, restriction pattern, and a sequenced 14-kb cfr-carrying segment. Plasmid pSS-02 was originally identified in staphylococci isolated from pigs. This is the first time that a cfr-carrying plasmid has been detected in MRCoNS obtained from intensive care patients in China. Based on the similarities to the cfr-carrying plasmid pSS-02 from porcine coagulase-negative staphylococci, a transmission of this cfr-carrying plasmid between staphylococci from pigs and humans appears to be likely.
| S-EPMC3577776 | biostudies-literature
Project description:Coagulase-negative staphylococci (CoNS) for a long time were considered avirulent constituents of the human and warm-blooded animal microbiota. However, at present, S. epidermidis, S. haemolyticus, and S. hominis are recognized as opportunistic pathogens. Although linezolid is not registered for the treatment of CoNS infections, it is widely used off-label, promoting emergence of resistance. Bioinformatic analysis based on maximum-likelihood phylogeny and Bayesian clustering of the CoNS genomes obtained in the current study and downloaded from public databases revealed the existence of international linezolid-resistant lineages, each of which probably had a common predecessor. Linezolid-resistant S. epidermidis sequence-type (ST) 2 from Russia, France, and Germany formed a compact group of closely related genomes with a median pairwise single nucleotide polymorphism (SNP) difference of fewer than 53 SNPs, and a common ancestor of this lineage appeared in 1998 (1986-2006) before introduction of linezolid in practice. Another compact group of linezolid-resistant S. epidermidis was represented by ST22 isolates from France and Russia with a median pairwise SNP difference of 40; a common ancestor of this lineage appeared in 2011 (2008-2013). Linezolid-resistant S. hominis ST2 from Russia, Germany, and Brazil also formed a group with a high-level genome identity with median 25.5 core-SNP differences; the appearance of the common progenitor dates to 2003 (1996-2012). Linezolid-resistant S. hominis isolates from Russia demonstrated associated resistance to teicoplanin. Analysis of a midpoint-rooted phylogenetic tree of the group confirmed the genetic proximity of Russian and German isolates; Brazilian isolates were phylogenetically distant. repUS5-like plasmids harboring cfr were detected in S. hominis and S. haemolyticus.
| S-EPMC8473885 | biostudies-literature
Project description:Methicillin-resistant coagulase negative staphylococci (MR-CoNS) are the major cause of infectious diseases because of their potential ability to form biofilm and colonize the community or hospital environments. This study was designed to investigate the biofilm producing ability, and the presence of mecA, icaAD, bap and fnbA genes in MR-CoNS isolates. The MR-CoNS used in this study were isolated from various samples of community environment and five wards of hospital environments, using mannitol salt agar (MSA) supplemented with 4 μg/ml of oxacillin. The specie level of Staphylococcus haemolyticus, Staphylococcus epidermidis, Staphylococcus hominis and Staphylococcus warneri was identified by specific primers of groESL (S. haemolyticus), rdr (S. epidermidis) and nuc (S. hominis and S. warneri). The remainder isolates were identified by tuf gene sequencing. Biofilm production was determined using Congo red agar (CRA) and Microtiter plate (MTP) assay. The mecA and biofilm associated genes (icaAD, fnbA and bap) were detected using PCR method. From the 558 samples from community and hospital environments, 292 MR-CoNS were isolated (41 from community environments, and 251 from hospital environments). S. haemolyticus (41.1%) and S. epidermidis (30.1%) were the predominant species in this study. Biofilm production was detected in 265 (90.7%) isolates by CRA, and 260 (88.6%) isolates were detected by MTP assay. The staphylococci isolates derived from hospital environments were more associated with biofilm production than the community-derived isolates. Overall, the icaAD and bap genes were detected in 74 (29.5%) and 14 (5.6%) of all isolates from hospital environments. When tested by MTP, the icaAD gene from hospital environment isolates was associated with biofilm biomass. No association was found between bap gene and biofilm formation. The MR-CoNS isolates obtained from community environments did not harbor the icaAD and bap genes. Conversely, fnbA gene presented in MR-CoNS isolated from both community and hospital environments. The high prevalence of biofilm producing MR-CoNS strains demonstrated in this study indicates the persisting ability in environments, and is useful in developing prevention strategies countering the spread of MR-CoNS.
| S-EPMC5578677 | biostudies-literature
Project description:Swine farming as a source of methicillin-resistant Staphylococcus aureus (MRSA) has been well documented. Methicillin-resistant coagulase-negative staphylococci (MRCoNS) have been less studied, but their importance as pathogens is increasing. MRCoNS are indeed considered relevant nosocomial pathogens; identifying putative sources of MRCoNS is thus gaining importance to prevent human health hazards. In the present study, we investigated MRSA and MRCoNS in animals and environment in five pigsties in a high farm-density area of northwestern Italy. Farms were three intensive, one intensive with antibiotic-free finishing, and one organic. We tested nasal swabs from 195 animals and 26 environmental samples from three production phases: post-weaning, finishing and female breeders. Phenotypic tests, including MALDI-TOF MS, were used for the identification of Staphylococcus species; PCR and nucleotide sequencing confirmed resistance and bacterial species. MRCoNS were recovered in 64.5% of nasal swabs, in all farms and animal categories, while MRSA was detected only in one post-weaning sample in one farm. The lowest prevalence of MRCoNS was detected in pigs from the organic farm and in the finishing of the antibiotic-free farm. MRCoNS were mainly Staphylococcus sciuri, but we also recovered S. pasteuri, S. haemolyticus, S. cohnii, S. equorum and S. xylosus. Fifteen environmental samples were positive for MRCoNS, which were mainly S. sciuri; no MRSA was found in the farms' environment. The analyses of the mecA gene and the PBP2-a protein highlighted the same mecA fragment in strains of S. aureus, S. sciuri and S. haemolyticus. Our results show the emergence of MRCoNS carrying the mecA gene in swine farms. Moreover, they suggest that this gene might be horizontally transferred from MRCoNS to bacterial species more relevant for human health, such as S. aureus.
| S-EPMC8227741 | biostudies-literature