Genetic diversity, virulence, antimicrobial resistance genes, and antimicrobial susceptibility of group B Streptococcus (GBS) linked to mass mortalities of cultured Nile tilapia in Brazil

Streptococcus agalactiae, group B Streptococcus (GBS), is the main bacterial pathogen of cultured Nile tilapia in many countries and causes extensive deaths in all stages of the farming cycle throughout the year. This study investigated the genetic diversity, virulence, presence of antibiotic resistance genes and antimicrobial susceptibility of 72 GBS linked to mass mortalities of cultured Nile tilapia in Brazil. Isolate identity was confirmed by morphological, biochemical and molecular analyses. Capsular serotype, multi-locus sequence typing (MLST) allelic profiles and putative pathogenic factors were identified by polymerase chain reaction (PCR), gel electrophoresis, sequencing and molecular analyses of different genes. The presence of antimicrobial resistance genes and antimicrobial susceptibility to florfenicol (FFC), oxytetracycline (OTC), thiamphenicol (TAP) and their combination were evaluated by PCR, followed by gel electrophoresis, and broth microdilution antimicrobial susceptibility testing, respectively. All clinical isolates studied were confirmed to be GBS, one from serotype III (IA2022) and 71 from serotype Ib, suggesting that serotype Ib was the most prevalent strain between 2011 and 2016 in the south and southern and southeastern regions of Brazil. Eight different allelic profiles were identified for the first time, being adhP- 52, pheS-2, atr-31, glnA-4, sdhA-2, tkt-19 the most predominant. Between one (glcK) and three (adhP and glnA) alleles were present at each locus. All strains, except IA2022, showed a partial gene deletion event on the glcK gene. The surface protein Rib and hypervirulent GBS adhesin BibA were detected in all strains, except for 18P, which was negative for rib. On the other hand, α and β antigens of the C protein were only detected in IA2022. All antimicrobials showed high minimum inhibitory concentration (MIC ≥ 16 µg/mL) values against several strains with negative results for resistance genes. Despite indifference and antagonism being the most predominant activities in all combinations evaluated, the record of synergism, including in a strain with a resistance gene and phenotypic resistance, suggests that combination therapy can have therapeutic efficacy when well planned. The combination involving OTC and TAP or FFC is a likely candidate for improving the treatment of streptococcosis using combination therapy, even for strains showing phenotypic and genotypic resistance to OTC. This study provides pertinent data on pathogenic GBS genetic diversity, virulence, the presence of antibiotic resistance genes and antimicrobial susceptibility, which may be useful in the development of effective vaccines and therapeutic strategies for the prevention and control of streptococcosis in aquaculture farms.