Molecular analysis of subtilase cytotoxin genes of food-borne Shiga toxin-producing Escherichia coli reveals a new allelic subAB variant

Abstract

BackgroundThe open reading frames of subAB genes and their flanking regions of 18 food-borne Shiga toxin-producing E. coli (STEC) strains were analyzed.ResultsAll but one subAB open reading frames (ORF) were complete in all STEC strains. The subAB1 genes of nine STEC strains were located on large plasmids. The subAB2 allele (here designated subAB2-1), which was recently described by others to be present in the Subtilase-Encoding PAI (SE-PAI) was found in 6 STEC strains. A new chromosomal subAB2 variant, designated subAB2-2 was detected in 6 strains and was linked to a chromosomal gene hypothetically encoding an outer membrane efflux protein (OEP). Three STEC strains contained both subAB2 variants. DNA analysis indicated sequence conservation in the plasmid-located alleles and sequence heterogeneity among the chromosomal subAB2 genes.ConclusionsThe results of this study have shown that 18 subAB-PCR positive STEC strains contain complete subAB open reading frames. Furthermore, the new allelic variant subAB2-2 was described, which can occur in addition to subAB2-1 on a new chromosomal locus.

Cytotoxic and Apoptotic Effects of Recombinant Subtilase Cytotoxin Variants of Shiga Toxin-Producing Escherichia coli

“…The subAB-positive STEC strains TS30/08, LM27564, and LM14603/08 used in this study were described previously (12,25). Laboratory E. coli strain BL21(DE3) and the BL21 derivative C41(DE3) (26) were used for protein expression.…”

“…Based on previous results of variant-specific PCRs and sequencing analysis (25), STEC strains TS30/08, LM27564, and LM14603/08 were used as templates for cloning of subAB 1 , subAB 2-1 , and subAB 2-2 , respectively. Whereas subA 1 and subB 1 could be cloned directly into the expression vector, the subunit genes of both chromosomal variants were cloned after amplification of complete operons and surrounding genetic regions by a nested PCR approach as outlined in Fig.…”

“…The subAB 2-1 genes are about 90% homologous to the plasmid-carried subAB 1 with the catalytic triad being present in subA 2-1 , but the subB 2-1 open reading frame is one triplet shorter than that of subB 1 . The subAB 2-2 operon recently was described by our group (25), and it could be demonstrated that subAB was located next to a gene encoding an outer membrane efflux protein and associated genes of a type one secretion system. This variant also shows about 90% sequence identity to the plasmid-encoded subAB 1 operon and differs by about 2% from subAB 2-1 .…”

“…This variant also shows about 90% sequence identity to the plasmid-encoded subAB 1 operon and differs by about 2% from subAB 2-1 . Furthermore, it could be demonstrated that both chro-mosomal variants can be present as single operons or in parallel (25).…”

Estudo dinâmico da expressão gênica global durante a interação STEC-enterócito utilizando séries temporais

“…As cepas eae negativas possuem outros fatores de aderência e virulência associados, tais como Saa, STEC agglutinating adhesin (Paton et al, 2001) e SubAB, subtilase cytotoxin (Paton et al, 2004), ambos localizados no megaplasmídeo pO113 (Funk et al, 2013). Paton et al (2001) mutagenizaram o gene saa do megaplasmídeo da O113:H21, o que resultou na redução de aderência da bactéria.…”