RESUMEN
A 10 kb O-antigen gene cluster was sequenced from a Salmonella enterica subsp. enterica Dakar O28 reference strain and from two S. Pomona serogroup O28 isolates. The two S. Pomona O antigen gene clusters showed only moderate identity with the S. Dakar O28 gene cluster, suggesting that the O antigen oligosaccharides may contain one or more sugars conferring the O28 epitope but may otherwise be different. These novel findings are absolutely critical for the correct interpretation of molecular serotyping assays targeting genes within the O antigen gene clusters of these Salmonella serotypes and suggest the possibility that the O antigen gene clusters of other Salmonella serovars may also be heterogenous.
RESUMEN
BACKGROUND: A large, multi-province outbreak of listeriosis associated with ready-to-eat meat products contaminated with Listeria monocytogenes serotype 1/2a occurred in Canada in 2008. Subtyping of outbreak-associated isolates using pulsed-field gel electrophoresis (PFGE) revealed two similar but distinct AscI PFGE patterns. High-throughput pyrosequencing of two L. monocytogenes isolates was used to rapidly provide the genome sequence of the primary outbreak strain and to investigate the extent of genetic diversity associated with a change of a single restriction enzyme fragment during PFGE. RESULTS: The chromosomes were collinear, but differences included 28 single nucleotide polymorphisms (SNPs) and three indels, including a 33 kbp prophage that accounted for the observed difference in AscI PFGE patterns. The distribution of these traits was assessed within further clinical, environmental and food isolates associated with the outbreak, and this comparison indicated that three distinct, but highly related strains may have been involved in this nationwide outbreak. Notably, these two isolates were found to harbor a 50 kbp putative mobile genomic island encoding translocation and efflux functions that has not been observed in other Listeria genomes. CONCLUSIONS: High-throughput genome sequencing provided a more detailed real-time assessment of genetic traits characteristic of the outbreak strains than could be achieved with routine subtyping methods. This study confirms that the latest generation of DNA sequencing technologies can be applied during high priority public health events, and laboratories need to prepare for this inevitability and assess how to properly analyze and interpret whole genome sequences in the context of molecular epidemiology.
Asunto(s)
Contaminación de Alimentos , Microbiología de Alimentos , Genoma Bacteriano , Listeria monocytogenes/genética , Análisis de Secuencia de ADN/métodos , Canadá/epidemiología , Biología Computacional , ADN Bacteriano/genética , Brotes de Enfermedades , Enfermedades Transmitidas por los Alimentos/microbiología , Islas Genómicas , Ensayos Analíticos de Alto Rendimiento , Mutación INDEL , Listeria monocytogenes/clasificación , Listeria monocytogenes/aislamiento & purificación , Listeria monocytogenes/patogenicidad , Listeriosis/epidemiología , Productos de la Carne/microbiología , Filogenia , Plásmidos , Polimorfismo de Nucleótido Simple , Profagos , Alineación de Secuencia , Serotipificación , VirulenciaRESUMEN
The serotyping of O and H antigens is an important first step in the characterization of Salmonella enterica. However, serotyping has become increasingly technically demanding and expensive to perform. We have therefore sequenced additional S. enterica O antigen gene clusters to provide information for the development of DNA-based serotyping methods. Three S. enterica isolates had O antigen gene clusters with homology to the Escherichia coli O123 O antigen region. O antigen clusters from two serogroup O58 S. enterica strains had approximately 85 % identity with the E. coli O123 O antigen region over their entire length, suggesting that these Salmonella and E. coli O antigen regions evolved from a common ancestor. The O antigen cluster of a Salmonella serogroup O41 isolate had a lower level of identity with E. coli O123 over only part of its O antigen DNA cluster sequence, suggesting a different and more complex evolution of this gene cluster than those in the O58 strains. A large part of the Salmonella O41 O antigen DNA cluster had very close identity with the O antigen cluster of an O62 strain. This region of DNA homology included the wzx and wzy genes. Therefore, molecular serotyping tests using only the O41 or O62 wzx and wzy genes would not differentiate between the two serogroups. The E. coli O123 O-antigenic polysaccharide and its repeating unit were characterized, and the chemical structure for E. coli O123 was entirely consistent with the O antigen gene cluster sequences of E. coli O123 and the Salmonella O58 isolates. An understanding of both the genetic and structural composition of Salmonella and E. coli O antigens is necessary for the development of novel molecular methods for serotyping these organisms.