Dataset Information

Genotypic and phenotypic ?-lactam resistance and presence of PVL gene in Staphylococci from dry bovine udder.

ABSTRACT: Dairy cows affected with subclinical mastitis can be sources of virulent, antimicrobial-resistant Staphylococci to humans because of the excretion of the bacteria through their milk. This study focussed on the phenotypic and genotypic antibiotic resistance patterns of Staphylococci isolated from dairy cows in early dry period. Among 96 isolates of Gram positive cocci from 157 cows, 76 were identified as Coagulase Negative Staphylococci and the remaining 20 were Staphylococcus aureus. Typical amplicons of coagulase gene were obtained for all 20 samples of S. aureus with three major coagulase types being identified as giving 627 bp (40%), 910 bp (35%) and 710 bp (25%) long PCR products. The groEL gene was amplified in PCR of all 76 isolates of Coagulase Negative Staphylococci, and incubation of PCR products with restriction enzyme PvuII yielded three distinct PCR-RFLP fragment patterns bearing resemblance to S. chromogenes and S. hyicus. Highest sensitivity of Coagulase Negative Staphylococci was noted for Azithromycin (92.5%) and the least to Tetracyclines (76.3%), whereas for S. aureus, it was Cefoperazone (95%) and Azithromycin (72.2%) respectively. Phenotypic resistance to Oxacillin (25 isolates), and Cefoxitin (11 isolates) was detected by dilution method with a commercial strip (Ezy MICTM). Genotypic resistance to ?-Lactam antibiotics was found in 65 (34 with mecA gene and 31 with blaZ gene) isolates. Eighteen isolates possessed both the genes, with the PVL gene for virulence being detected in five of them. Nine isolates which had mecA gene were phenotypically susceptible to oxacillin while phenotypic resistance to oxacillin was observed in seven isolates that did not have either mecA or blaZ gene. This is the first report of persistent Staphylococcal infections possessing PVL gene and high level of genotypic resistance to ?-Lactam antibiotics in small- holder dairy cattle from India.

SUBMITTER: Kulangara V

PROVIDER: S-EPMC5665534 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Genotypic and phenotypic β-lactam resistance and presence of PVL gene in Staphylococci from dry bovine udder.

Kulangara Vinodkumar V Nair Neetha N Sivasailam Asok A Sasidharan Suchithra S Kollannur Justin Davis JD Syam Radhika R

PloS one 20171101 11

Dairy cows affected with subclinical mastitis can be sources of virulent, antimicrobial-resistant Staphylococci to humans because of the excretion of the bacteria through their milk. This study focussed on the phenotypic and genotypic antibiotic resistance patterns of Staphylococci isolated from dairy cows in early dry period. Among 96 isolates of Gram positive cocci from 157 cows, 76 were identified as Coagulase Negative Staphylococci and the remaining 20 were Staphylococcus aureus. Typical amp ...[more]

PMID: 29091956

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Project description:This study investigated 960 Slovak and Czech spotted cattle from four different conventional (non-organic) dairy herds located in Eastern Slovakia and Czechia during early lactation (14-100 days after calving). Dairy cows were examined clinically; milk from fore-stripping of each udder quarter was subjected to sensory examination and assessed by the California mastitis test (CMT), and laboratory analyses of bacterial pathogens in milk, including virulence factors, were conducted. Positive CMT scores (1-3) for one or more quarters were detected in 271 (28.2%) of the examined animals. Out of 230 infected milk samples, representing 24.0% of all dairy cows, staphylococci (59.1% of positive findings) were the most commonly isolated organisms, followed by E. coli (11.3%), streptococci Str. uberis (9.1%) and Str. agalactiae (3.4%), and enterococci (6.1%). From 136 isolates of S. aureus (38 isolates) and non-aureus staphylococci (NAS; 98 isolates), virulence factors and their resistance to 14 antimicrobials were detected using the disk diffusion method, with PCR detection of the methicillin resistance gene, mecA. An increased incidence of clinical and chronic forms of mastitis has been reported in mastitic cows in which staphylococci, especially S. aureus and NAS (S. chromogenes, S. warneri, and S. xylosus), have been detected and compared to other isolated udder pathogens. From those species, S. aureus and isolates of NAS mentioned above showed multiple virulence factors that are more likely to hydrolyze DNA, hemolysis, produce gelatinase and biofilm, and have multi-drug resistance as compared to other less virulent staphylococci. Generally, the isolated staphylococci showed 77.2% resistance to one or more antimicrobials, in particular to aminoglycosides, β-lactams, macrolides, or cephalosporins. Isolates that showed the ability to form a biofilm were more resistant to more than one antimicrobial than isolates without biofilm production. Multi-drug resistance to three or more antimicrobial classes was recorded in 16 isolates (11.7%), and the presence of the mecA gene was also confirmed in two isolates of S. aureus and two species of NAS.

Project description:The epidemiologically most important mechanism of antibiotic resistance in Staphylococcus aureus is associated with mecA-an acquired gene encoding an extra penicillin-binding protein (PBP2a) with low affinity to virtually all β-lactams. The introduction of mecA into the S. aureus chromosome has led to the emergence of methicillin-resistant S. aureus (MRSA) pandemics, responsible for high rates of mortality worldwide. Nonetheless, little is known regarding the origin and evolution of mecA. Different mecA homologues have been identified in species belonging to the Staphylococcus sciuri group representing the most primitive staphylococci. In this study we aimed to identify evolutionary steps linking these mecA precursors to the β-lactam resistance gene mecA and the resistance phenotype. We sequenced genomes of 106 S. sciuri, S. vitulinus and S. fleurettii strains and determined their oxacillin susceptibility profiles. Single-nucleotide polymorphism (SNP) analysis of the core genome was performed to assess the genetic relatedness of the isolates. Phylogenetic analysis of the mecA gene homologues and promoters was achieved through nucleotide/amino acid sequence alignments and mutation rates were estimated using a Bayesian analysis. Furthermore, the predicted structure of mecA homologue-encoded PBPs of oxacillin-susceptible and -resistant strains were compared. We showed for the first time that oxacillin resistance in the S. sciuri group has emerged multiple times and by a variety of different mechanisms. Development of resistance occurred through several steps including structural diversification of the non-binding domain of native PBPs; changes in the promoters of mecA homologues; acquisition of SCCmec and adaptation of the bacterial genetic background. Moreover, our results suggest that it was exposure to β-lactams in human-created environments that has driven evolution of native PBPs towards a resistance determinant. The evolution of β-lactam resistance in staphylococci highlights the numerous resources available to bacteria to adapt to the selective pressure of antibiotics.

Dataset Information

Genotypic and phenotypic ?-lactam resistance and presence of PVL gene in Staphylococci from dry bovine udder.

Publications

Genotypic and phenotypic β-lactam resistance and presence of PVL gene in Staphylococci from dry bovine udder.

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