blaBioF, a novel B2 metallo- B-lacatamase from Pseudomonas sp isolated from an on-farm biopurification system

CAFIERO JH; MARTINI M. C; DEL PAPA M. F.; VACCA C.; LAGARES A.; LOZANO M. J; TOMATIS PE

Virtual

Congreso; SAIB-SAMIGE 2020; 2020

Antibiotic resistant (AR) bacteria represent a major global health concern and environmental bacteria are considered as a source of AR genes. We have previously performed a high-throughput sequencing of plasmids present in a bacterial collection isolated from a biopurification system (BPS), used on-farm to treat pesticide-polluted water. Among the predicted genes identified, a novel metallo-β-lactamase (MBL) gene was found. In this study, we designated this gene as blaBioF and show by sequence comparison that it belongs to the B2 family of MBLs and is closely related to the recently characterized PFM enzymes. BioF retains conserved amino acids present in B2 MBLs crucial for their catalytic activity. The BPS isolate carrying the blaBioF gene was identified as a Pseudomonas sp. strain BF61 according to 16S DNA sequencing. Susceptibility testing performed for β-lactams by disk diffusion showed that strain BF61 was resistant to meropenem and imipenem, which confirms the presence of an active β-lactamase in this isolate. Next, we performed a high-throughput sequencing of the whole genome of strain BF61 to accurately identify this isolate species and the genomic location of the blaBioF gene. Whole-genome ANIb comparisons with Pseudomonas spp. type strains showed that isolate BF61 belongs to the Pseudomonas fluorescens complex and the closest type strain was P. gessardii DSM17152. However, the ANIb and dDDH values between BF61 and DSM17152 were below the cut-off boundary required to assign both strains as the same species. These genomic comparison studies revealed that strain BF61 belongs to a novel species of the genus Pseudomonas. Regarding the genomic location of blaBioF, the transfer of this gene into E. coli by electroporation or conjugation was unsuccessful. Thus, we explored current in silico tools to assemble or detect plasmids in bacterial whole genome sequences or reads. Nevertheless, the results were inconclusive, with the blaBioF containing contig sometimes predicted as plasmidic and sometimes as chromosomal. Interestingly, IslandViewer4 predicted that the blaBioF gene is coded in a small genomic island. This tool also predicted that genes coding MBLs homologous to BioF (carried by P. synxantha MCP106 and P. libanensis BS2975) were also present in genomic islands with conserved gene synteny and high nucleotide identity with the blaBioF containing region of Pseudomonas sp. BF61. These results suggest that the blaBioF gene might have been horizontally acquired. The discovery of blaBioF in a BPS isolate emphasizes the importance of the exploration of AR in the environmental microbiota, as a first step towards the prediction of novel resistance mechanisms before they emerge clinically.