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A multi-virulence-locus sequence typing (MVLST) scheme was developed for subtyping Listeria monocytogenes, and the results obtained using this scheme were compared to those of pulsed-field gel electrophoresis (PFGE) and the published results of other typing methods, including ribotyping (RT) and multilocus sequence typing (MLST). A set of 28 strains (eight different serotypes and three known genetic lineages) of L. monocytogenes was selected from a strain collection (n > 1,000 strains) to represent the genetic diversity of this species. Internal fragments (ca. 418 to 469 bp) of three virulence genes (prfA, inlB, and inlC) and three virulenceassociated genes (dal, lisR, and clpP) were sequenced and analyzed. Multiple DNA sequence alignment identified 10 (prfA), 19 (inlB), 13 (dal), 10 (lisR), 17 (inlC), and 16 (clpP) allelic types and a total of 28 unique sequence types. Comparison of MVLST with automated EcoRI-RT and PFGE with ApaI enzymatic digestion showed that MVLST was able to differentiate strains that were indistinguishable by RT (13 ribotypes; discrimination index = 0.921) or PFGE (22 profiles; discrimination index = 0.970). Comparison of MVLST with housekeeping-gene-based MLST analysis showed that MVLST provided higher discriminatory power for serotype ½a and 4b strains than MLST. Cluster analysis based on the intragenic sequences of the selected virulence genes indicated a strain phylogeny closely related to serotypes and genetic lineages. In conclusion, MVLST may improve the discriminatory power of MLST and provide a convenient tool for studying the local epidemiology of L. monocytogenes. [ABSTRACT FROM AUTHOR]

Salmonella enterica subsp. enterica serovar Enteritidis is a major cause of food-borne salmonellosis in the United States. Two major food vehicles for S. Enteritidis are contaminated eggs and chicken meat. Improved subtyping methods are needed to accurately track specific strains of S. Enteritidis related to human salmonellosis throughout the chicken and egg food system. A sequence typing scheme based on virulence genes (fimH and sseL) and clustered regularly interspaced short palindromic repeats (CRISPRs)—CRISPR-including multi-virulence-locus sequence typing (designated CRISPR-MVLST)—was used to characterize 35 human clinical isolates, 46 chicken isolates, 24 egg isolates, and 63 hen house environment isolates of S. Enteritidis. A total of 27 sequence types (STs) were identified among the 167 isolates. CRISPR-MVLST identified three persistent and predominate STs circulating among U.S. human clinical isolates and chicken, egg, and hen house environmental isolates in Pennsylvania, and an ST that was found only in eggs and humans. It also identified a potential environment-specific sequence type. Moreover, cluster analysis based on fimH and sseL identified a number of clusters, of which several were found in more than one outbreak, as well as 11 singletons. Further research is needed to determine if CRISPR-MVLST might help identify the ecological origins of S. Enteritidis strains that contaminate chickens and eggs. [ABSTRACT FROM AUTHOR]

Abstract: Extraintestinal pathogenic Escherichia coli (ExPEC) are known to cause important diseases of humans and animals, and they have been shown to carry a variety of plasmids associated with increased virulence and decreased antimicrobial susceptibility. Here, the completed DNA sequence of a human uropathogenic E. coli (UPEC; O6:H31 isolate) plasmid, pEC14_114, was determined. The plasmid was 114,222bp in length and was highly similar to plasmid sequences or draft contiguous sequences from three other human cystitis-associated UPEC isolates. pEC14_114 contained 141 coding regions, including a number of genes associated with mobile genetic elements, F-type transfer, plasmid maintenance and stability, colicin immunity, and plasmid replication. This plasmid also possessed a “genetic load” region containing genes with predicted similarity to iron acquisition systems and virulence factors. The prevalence of pEC14-associated genes was determined for a collection of 1456 E. coli isolates, including those from food products, humans, dogs, cats, pigs, chickens, and turkeys. pEC14_114-associated genes were found significantly more often (16–35%) among human UPEC and neonatal meningitis-associated isolates than among food- and animal-source isolates (0–8%). Overall, this plasmid represents a novel IncFIB/FIIA plasmid type associated with human ExPEC belonging to the B2 phylogenetic group. The overall role of this plasmid, if any, in human ExPEC infections remains to be determined. [Copyright &y& Elsevier]