Caracterização fenotípica e genotípica de fatores de virulência e resistência à oxacilina em Staphylococcus spp. coagulase-positivos isolados de mastite bovina

Abstract

The present study was done to characterize phenotypic and genotypically the virulence factors and resistance profile to antibiotics, specifically to the oxacillin, of species of Staphylococcus spp. coagulase-positives isolated from milk samples of cows with mastitis. Of 150 isolated, the species S.aureus, S.intermedius and S.hyicus were identified. The antimicrobial susceptibility test yielded high resistance to penicillin and ampicillin and distinct antibiotyps had been identified, being of sensitivity to all the antibiotics prevalent. A total of 94,6% and 70,7% was positive for the production of betalactamases and presence of the mecA gene, respectively. The femA gene was detected in 80,9% of isolated mecA positive. The isolated presented distinct standards of growth in the tests of susceptibility to oxacillin. The tests of agar screen and the evaluation of the microdilution in plate, presented considerable values of sensitivity and specificity when the isolated ones were considered as an only group (Staphylococcus spp coagulase-positives) or when the species had been discriminated, respectively. The addition of 4% of NaCl in the agar plate can have effect in the increase of the sensitivity of the test of diffusion using the oxacillin and the test of diffusion presented the better capacity in the detention of the resistance isolates with cefoxitin disk. A total of 80% of the S.aureus was classified as MRSA and had presented resistance higher than 15% to the ampicillin and sulbactam association, vancomicin and gentamicin. About the virulence factors, the technique of the microplate disclosed 82.8% of isolated producers of slime and the intensity of the production was not associated with the presence of the icaA and icaD genes. In addition, the agar with the congo red presented effectiveness in the detention of isolated the not producing ones of slime . In all hemolitics isolates were detected the beta-hemolisin and 11.3% of the non-hemolitics had produced hemolisins in the presence of beta-hemolitic strain, indicating the presence of the delta hemolisin. spaA, coa and agr genes had been carried through the amplifications of the genes the S.aureus. The spaA gene, that it codifies protein A region X, was positive in all isolated ones, disclosed only amplicon for each isolated one and presented polimorfism being the prevalent size of 315pb. The amplification of the coa gene presented three distinct polimorfics types, with only amplicon for each isolated and the prevalent profile was of 900pb. The amplification of the agr gene resulted in amplicons with 350 and 550 pb in 74% of the isolated. The test of biotyping disclosed the bovine ecovar as the prevalent in the S.aureus. Seven S.aureus that had presented the same profiles of the coa, spaA, agr genes, antibiotyps and ecovars was detected being 4 pertaining the same properties and 3 isolated from distinct properties.