Identification of novel endolysins from Streptococcus agalactiae prophages by bioinformatic analysis.

KOVACEC V; DI GREGORIO S; DE MEDTS M; DI DONATO M; GHIGLIONE B; POWER P; POKLEPOVICH T; CAMPOS J; MOLLERACH M; BONOFIGLIO L

Conferencia; FEMS Online Conference on Microbiology,; 2020

Federation of European Microbiological Societies

Streptococcus agalactiae (GBS) causes severe infections in neonates, pregnant women and adults with comorbidities. Phage lysins are promising agents to fight antimicrobial resistance.We aimed to characterize endolysins of 13 prophages from 10 representative local GBS strains recovered from a National Multicentre Study, clone and purify one lysin and test its activity against GBS. Lysin sequences were analyzed with ExPASy and NCBI tools. Phylogeny, including lysins from 50 publicly available GBS prophages, was performed with IQTree. Genomic DNA was used for PCR amplification of lysin S2P1. Primers with NdeI and XhoI restriction sites were designed with Oligo Explorer. Vectors pJET1.2/blunt and pET22b, E. coli Top10 and BL21(DE3) were used for cloning and expression, respectively. Anion exchange chromatography was used for purification. Lytic activity was tested against host GBS cultures. Lysins were found in all prophages (n=16), ranging from 6-53kDa. They were classified, according to their cleavage site, into three of the four major endolysin classes. Bifunctional lysins were found. Our lysins were present in the five phylogenetic clusters identified, 9/16 having 90-100% identity with already characterized GBS lysins, 7/16 being novel GBS endolysins. Lysin S2P1, identical to LambdaSa1 lysin (Pritchard, 2007), was purified and showed lytic activity against host GBS strain S2 on preliminary assays.We will keep on studying S2P1 lytic activity against our GBS collection, alongside novel lysins we are currently cloning. The variety of endolysin classes found in our prophages is encouraging as these lysins could potentially be used against the diverse GBS strains circulating locally and internationally.