Molecular prediction of the O157: H-negative phenotype prevalent in Australian Shiga toxin-producing Escherichia coli cases improves concordance of in silico serotyping with phenotypic motility

Shiga toxin-producing Escherichia coli (STEC) is a foodborne pathogen, and serotype O157:H7 is typically associated with severe disease. Australia is unique in its STEC epidemiology, as severe cases are typically associated with non-O157 serogroups, and locally acquired O157 isolates are H-negative/nonmotile. The H-negative phenotype and reduced severity of disease compared to that associated with H7/motile strains are distinct features of Australian O157 strains, but the molecular mechanism behind this phenotype has not been reported. Accurate characterization of the H-negative phenotype is important in epidemiological surveillance of STEC. Serotyping is moving away from phenotype-based methods, as next generation sequencing allows rapid extrapolation of serotype through in silico detection of the O-antigen processing genes, wzx, wzy, wzm, and wzt, and the H-antigen gene, fliC. The detection and genotyping of fliC alone is unable to determine the motility of the strain. Typically, most Australian O157:H-negative strains carry an H7 genotype yet phenotypically are nonmotile; thus, many are mischaracterized as H7 strains by in silico serotyping tools. Comparative genomic analysis of flagellar genes between Australian and international isolates was performed and an insertion at nucleotide (nt) 125 in the flgF gene was identified in H-negative isolates. Chi-square results showed that this insertion was significantly associated with the H-negative phenotype (P 0.0001). Phylogenetic analysis was also completed and showed that the Australian H-negative isolates with the insertion in flgF represent a clade within the O157 serogroup, distinct from O157:H7 serotypes. This study provides a genetic target for inferring the nonmotile phenotype of Australian O157 STEC, which increases the predictive value of in silico serotyping.</p

Comparative genomics identifies distinct lineages of <i>S</i>. Enteritidis from Queensland, Australia

<div><p><i>Salmonella enterica</i> is a major cause of gastroenteritis and foodborne illness in Australia where notification rates in the state of Queensland are the highest in the country. <i>S</i>. Enteritidis is among the five most common serotypes reported in Queensland and it is a priority for epidemiological surveillance due to concerns regarding its emergence in Australia. Using whole genome sequencing, we have analysed the genomic epidemiology of 217 <i>S</i>. Enteritidis isolates from Queensland, and observed that they fall into three distinct clades, which we have differentiated as Clades A, B and C. Phage types and MLST sequence types differed between the clades and comparative genomic analysis has shown that each has a unique profile of prophage and genomic islands. Several of the phage regions present in the <i>S</i>. Enteritidis reference strain P125109 were absent in Clades A and C, and these clades also had difference in the presence of pathogenicity islands, containing complete SPI-6 and SPI-19 regions, while P125109 does not. Antimicrobial resistance markers were found in 39 isolates, all but one of which belonged to Clade B. Phylogenetic analysis of the Queensland isolates in the context of 170 international strains showed that Queensland Clade B isolates group together with the previously identified global clade, while the other two clades are distinct and appear largely restricted to Australia. Locally sourced environmental isolates included in this analysis all belonged to Clades A and C, which is consistent with the theory that these clades are a source of locally acquired infection, while Clade B isolates are mostly travel related.</p></div

Maximum-likelihood phylogeny of QLD <i>S</i>. Enteritidis isolates, <i>S</i>. Typhimurium LT2 and <i>S</i>. Gallinarum 287/91 rooted to <i>S</i>. Berta.

<p>Isolates cluster into three clades as indicated. The scale bar shows substitutions per site. Sequence type and Phage type are indicated by colour as shown in the legend. The presence and absence of regions of difference (RODs) and prophages are also indicated in the following order: φSE10, φSE12, φSE12A, φSE14, φSE20, RE-2010, ROD4, ROD9, ROD13, ROD17, ROD21, ROD25, ROD28, ROD34, ROD37, ROD42, with a white box indicating absence and a black box indicating presence. Blue stars, red triangles and blue boxes indicate partial presence of the prophage.</p

Heatmap showing the presence and absence of acquired antimicrobial resistance genes and point mutations in <i>gyrA</i>.

<p>Blue boxes indicate absence and red boxes indicate presence of genes or point mutations.</p

Culture-independent detection of chlorhexidine resistance genes qacA/B and smr in bacterial DNA recovered from body sites treated with chlorhexidine-containing dressings

Purpose. Dressings containing chlorhexidine gluconate (CHG) are increasingly used in clinical environments for prevention of infection at central venous catheter insertion sites. Increased tolerance to this biocide in staphylococci is primarily associated with the presence of qacA/B and smr genes. Methodology. We used a culture-independent method to assess the prevalence of these genes in 78 DNA specimens recovered from the skin of 43 patients at catheter insertion sites in the arm that were covered with CHG dressings. Results. Of the 78 DNA specimens analysed, 52 (67 %) possessed qacA/B and 14 (18 %) possessed smr; all samples positive for smr were also positive for qacA/B. These prevalence rates were not statistically greater than those observed in a subsample of specimens taken from non-CHG treated contralateral arms and non-CHG-dressing exposed arms. A statistically greater proportion of specimens with greater than 72 h exposure to CHG dressings were qac-positive (P=0.04), suggesting that the patients were contaminated with bacteria or DNA containing qacA/B during their hospital stay. The presence of qac genes was not positively associated with the presence of DNA specific for Staphylococcus epidermidis and Staphylococcus aureus in these specimens. Conclusion. Our results show that CHG genes are highly prevalent on hospital patients’ skin, even in the absence of viable bacteria