Identification and genotyping of the blaTEM and blaSHV genes of extended-spectrum beta-lactamase (ESBL)-producing bacteria isolated from Cabalyorisa Cave, Mabini, Pangasinan through a polymerase chain reaction (PCR)

Abstract

Extended-spectrum beta-lactamase (ESBL) producing bacteria although have originally emerged in clinical isolates have recently been found to increasingly spread among various environments worldwide. This study identified and detected ESBLproduction among bacteria isolated from bat guano in Cabalyorisa Cave, Mabini, Pangasinan. A total of 28 strains were initially screened for antimicrobial susceptibility test against four 3rd generation cephalosporin and aztreonam. Eighteen (64.3%) out of 28 samples were found to be resistant to more than one of the antibiotics tested based on CLSI guidelines thus, were considered as putative ESBL-producers. The production of betalactamases by these strains was then confirmed through Phenotypic Confirmatory Combination Disc Diffusion Test (PCCDDT) and results showed that 15 (83.3%) of the strains were ESBL-positive. The identity of the phenotypically confirmed ESBL-producers were then confirmed by the amplification and sequencing of the 16S rRNA gene. Results showed that the strains were found to be members of the family Enterobacteriaceae under the genus Salmonella, Enterobacter, and Escherichia. The type of beta-lactamase genes present from the different ESBL-producers was determined by the amplification and sequencing of the blaSHV, and blaTEM genes through Polymerase Chain Reaction (PCR). Among the ESBL- producers, 13 (86.6%) were found to possessed the blaSHV while 12 (80%) harbored the blaTEM gene. The high prevalence of these genes is not surprising because the blaSHV, and blaTEM genes are the most common plasmid-mediated betalactamases and most ESBL variants are derivatives of these genes. Interestingly, two strains which were found to be positive in PCCDDT did not possessed any of the ESBL genes tested indicating that other ESBL genes might be responsible for the expression the beta-lactamase enzyme. This study therefore, is the first to report the presence of ESBLproducers with multiple genes from bacteria isolated in bat guano from Philippine caves. Further studies on the detection and sequencing of the specific gene variants harbored by the different strains should be conducted along with transconjugation studies to determine the mode of transfer and spread of these genes.