Prospective Genomic Characterization of the German Enterohemorrhagic Escherichia coli O104:H4 Outbreak by Rapid Next Generation Sequencing Technology

Abstract

An ongoing outbreak of exceptionally virulent Shiga toxin (Stx)-producing Escherichia coli O104:H4 centered in Germany, has caused over 830 cases of hemolytic uremic syndrome (HUS) and 46 deaths since May 2011. Serotype O104:H4, which has not been detected in animals, has rarely been associated with HUS in the past. To prospectively elucidate the unique characteristics of this strain in the early stages of this outbreak, we applied whole genome sequencing on the Life Technologies Ion Torrent PGM™ sequencer and Optical Mapping to characterize one outbreak isolate (LB226692) and a historic O104:H4 HUS isolate from 2001 (01-09591). Reference guided draft assemblies of both strains were completed with the newly introduced PGM™ within 62 hours. The HUS-associated strains both carried genes typically found in two types of pathogenic E. coli, enteroaggregative E. coli (EAEC) and enterohemorrhagic E. coli (EHEC). Phylogenetic analyses of 1,144 core E. coli genes indicate that the HUS-causing O104:H4 strains and the previously published sequence of the EAEC strain 55989 show a close relationship but are only distantly related to common EHEC serotypes. Though closely related, the outbreak strain differs from the 2001 strain in plasmid content and fimbrial genes. We propose a model in which EAEC 55989 and EHEC O104:H4 strains evolved from a common EHEC O104:H4 progenitor, and suggest that by stepwise gain and loss of chromosomal and plasmid-encoded virulence factors, a highly pathogenic hybrid of EAEC and EHEC emerged as the current outbreak clone. In conclusion, rapid next-generation technologies facilitated prospective whole genome characterization in the early stages of an outbreak.

Defects in RNA polyadenylation impair both lysogenization by and lytic development of Shiga toxin-converting bacteriophages

“…The severity of medical problems caused by STEC strains can be exemplified by the outbreak that occurred in Germany in 2011. Its fatal results, summarized as over 4000 symptomatic infections and over 50 cases of death, besides confirming the importance of the problem, also underlined our incomplete knowledge about regulatory mechanisms operating during development of the Stx phages (Mellmann et al, 2011;Beutin & Martin, 2012;Bloch et al, 2012;Karch et al, 2012;Werber et al, 2012).…”

Shiga toxins and stx phages: highly diverse entities

“…However, many questions remain unanswered and some old statements need to be revised, as demonstrated in the HUS outbreak in Germany in 2011. This outbreak was caused by an unusual E. coli strain, occurred in a high proportion of adult patients and had no evidence of zoonotic origin (Frank et al, 2011;Mellmann et al, 2011;Piérard et al, 2012). We learnt that strains lacking in typical virulence factors or belonging to infrequent serotypes may be highly virulent.…”

Shiga toxin-producing Escherichia coli O104:H4: An emerging important pathogen in food safety

“…Concurrent with the epidemiological investigation, scientists around the world rushed to sequence the whole genome of the outbreak strains to understand their phenotypic and virulent features, as well as acquire the strain-specific sequences using in designation of PCR diagnostic primers [37,38,[40][41][42]. We sequenced one outbreak strain, TY2482, isolated by the University Medical Center Hamburg-Eppendorf during the early phase of the outbreak from a 16-yearold female patient suffering from hemorrhagic enterocolitis.…”

“…These findings imply that the 01-09591 is closer to the current outbreak strain. Although the genome sequence of the strain 01-09591 indicated that it was different from the 2011 outbreak strain in terms of plasmid content and fimbrial genes [41], the fact that the German-origin O104 : H4 isolate is closely related to the Germany outbreak strain is still of scientific interest.…”