Molecular characterization and clonal diversity of Staphylococcus aureus isolated from milk of cows with subclinical mastitis in Sao Paulo State

Author(s)

Erika Carolina Romão Bonsaglia

• Document type: Doctoral Thesis
• Press: Botucatu. 2017-08-29.
• Institution: Universidade Estadual Paulista (Unesp). Instituto de Biociências. Botucatu
• Defense date: 2017-07-31
• Advisor: Vera Lúcia Mores Rall

Abstract

Staphylococcus aureus is a common agent of bovine mastitis, causing economic losses in Brazilian livestock. This microorganism has well known virulence factors such as the production of hemolysins, leukotoxins and superantigens such as toxic shock syndrome toxin and enterotoxins. The objective of the study was to characterize molecularly isolates of S. aureus from milk of cows with subclinical mastitis, of the several regions of São Paulo State, through Multilocus sequence typing (MLST), spatyping, Pulse Field Electrophoresis Gel (PFGE) and agr. Phenotypic and genotypic tests of antimicrobial resistance, as well as the search for genes of some virulence factors such as pvl (Panton Valentine toxin), tst (Toxic Shock Syndrome toxin). In the present study, all isolates of S. aureus were sensitive to methicillin (MSSA), as the mecA and mecC genes were not found. About 12 antibiotics tested, intermediate resistance to erythromicyn was observed and none resistance to oxacillin, vancomycin and gentamicin. The tetracycline resistant isolates showed the tetK gene. Molecular characterization showed 23 different spa types with prevalence of t605 and t127. The most of the strains (48.1%) presented as belonging to the agr II group, followed by 20.1% of the agr III group and 8.1% of the agrI group, and the agr IV group was not found. About the virulence factors studied, the pvl gene was not observed. In relation to super antigens, the tst gene was observed in 105 of the 285 strains (37.1%) and the distribution of genes coding for enterotoxin production was very variable. Among the 285 isolates, 126 (44.2%) were positive for at least one of the genes. The most frequent was seg, occurring alone or in different combinations in 275 (96.5%) isolates, followed by seh (250/90%), sec (227/ 79,6%), sei (44/15,4%), sea (31/10,8) and sed (10/3.5%). The seb and see genes were not observed as well as pvl gene.There were observed 23 different spa types (t605, t127, t458, t521, t342, t318, t693, t177, t11659, t021, t2164, t1192, t7335, t114, t6811, t6980, t002, t3324, t559, t138, t321, t456, T2066). The most frequent was t605, occurring 168 strains (58.9%), followed by t127, in 27 (9.5%) and the presence of t321 in cows causing mastitis had not been previously reported. The most frequent sequence types (ST) were ST126, corresponding to 59% of the isolates, followed by ST1 (9%), belonging to the clonal complex (CC) 126 and 1, respectively. The results showed notable differences between profiles in PFGE compared to other techniques, strains of the same spa type with different profiles of PFGE suggesting that spatyping is not a good molecular marker. (AU)

• FAPESP's process: 13/18338-8 - Clonal diversity and molecular characterization of Staphylococcus aureus isolated from milk of cows with subclinical mastitis in the state of Sao Paulo.
• Grantee: Erika Carolina Romão Bonsaglia
• Support type: Scholarships in Brazil - Doctorate