Shiga toxin-producing Escherichia coli Terry Arthur Research Microbiologist Meat Safety and Quality Research Unit U.S. Meat Animal Research Center Use of product names by USDA implies no An equal opportunity approval to the provider and employer exclusion of others that may also be suitable
Outline -Background -Non-O157 STEC prevalence pre- and post-harvest -Non-O157 STEC response to interventions -Detection difficulties -Knowledge gaps
Nomenclature - Shiga toxin initially discovered as a product of Shigella dysenteriae - Antibodies against Shiga toxin shown to inhibit cytotoxicity of an E. coli strain - E. coli strain discovered to be toxic to vero cells Shiga-like toxin (SLTEC) Vero toxin (VTEC) Shiga toxin (STEC) Cause disease in humans Enterohemorrhagic E. coli (EHEC)
E. coli serotyping O157:H7 O111:H8 O26:H11 Lipopolysaccharide (LPS) = O antigen Flagella = H antigen Y O1-O185 Y H1-H56
Non-O157 STEC – emerging? • Initial STEC isolation from clinical cases were O111 and O26 strains, not O157. (Konowalchuk et al. 1977) • Retrospective analysis found STEC O20 isolates dating from the early 1970’s. (Bettelheim et al. 1982) • A 1954 disease outbreak in New England thought to be caused by STEC O111, but no isolates were screened for Shiga toxin carriage. (Johnson et al. 1996) • O157:H7 not associated with human disease until 1983. (Riley et al. 1983) • Retrospective search identified O157:H7 isolates in CDC repository from 1975. (Wachsmuth et al. 1997)
Non-O157 STEC Estimated to cause one-half of the clinical EHEC cases. Over 200 STEC serotypes have been isolated from cattle. KG = The proportion of non-O157 STEC able to cause disease in humans is unknown.
STEC emphasis progression • Initially O157:H7 • Then O157:H7 and non-O157 • Now O157, Top 6 non-O157, & the rest of the non-O157 (non-Top 6 non-O157) – (O26, O45, O103, O111, O121, and O145) • Breadth of STEC strains viewed as a continuum.
To assess the clinical and public health risks associated with non-O157 STEC a seropathotype classification defined. A O157:H7 and O157:NM, which are common causes of outbreaks and HUS in most countries. B Associated with outbreaks and HUS, but less frequently than seropathotype A (CDC Top-6). C Associated with sporadic HUS but not epidemics. D Associated with diarrhea but not with outbreaks or HUS. E Multiple STEC serotypes that have never been associated with human disease and appear to be linked exclusively to animal infections in an agricultural setting.
CDC Top Six non-O157 STEC Serotypes 22% O26 :H11 or NM 7% O45 :H2 or NM 12% O103 :H2, H11, H25 or NM 16% O111 :H8 or NM 9% O121 :H19 or H7 5% O145 :NM 71% The cause of 71% of non-O157 diseases in the US Brooks et al, 2005, JID 192:1422.
STEC virulence factors Shiga toxins – two types: stx 1 and stx 2 - ribosome inactivating proteins Intimin ( eae) – attachment to epithelial cells tir/espE - translocated intimin receptor espA and espB required for intimate attachment/attaching and effacing (AE) lesions characteristic of STEC infection hlyA - pO157 enterohemolysin releasing hemoglobin from red blood cells katP - pO157 catalase peroxidase that defends the cell against oxidative damage etpD - pO157, encodes part of a type II secretory pathway transporting proteins across the outer membrane lpf - chromosomal long polar fimbriae espP - pO157 extracellular serine protease autotransporter saa - pO113 STEC agglutinating adhesion subA – subtilase cytotoxin – serine protease KG = What are essential virulence components?
Determinants of infection Virulence of E. coli O157:H7 organism E. coli + stx “Interestingly, different virulence gene profiles were detected within strains from the same serotype, for example, O26:H11 (10 isolates) displayed 3 different virulence profiles: stx1/stx2/stx2d/eaeA/hlyA/tir/lpfAO113/espP/toxB/iha (7 isolates), stx1/eaeA/hlyA/tir/lpfAO113/espP/toxB/iha (2 isolates) and stx1/eaeA/hlyA/lpfAO113/espP/toxB/iha (1 isolate).” Monaghan et al., 2011
Determinants of infection Virulence of E. coli O157:H7 organism E. coli + stx Dose Immune status of individual
http://bites.ksu.edu/nonO157outbreaks
Outbreaks Since 1994, there have been approximately 31 non- O157 STEC outbreaks in the U.S. From 2000 to 2006 the CDC reported an average of 40 E. coli O157:H7 outbreaks per year.
Outline -Background - Non-O157 STEC prevalence pre- and post-harvest
Prevalence of non-O157 STEC in Cattle Country Prevalence Method Reference United States 5.9% colony hybridization Cray et al. 1996 19% vero cell assay Wells et al. 1991 Argentina 37% vero cell assay Blanco et al. 1997 Canada 38.2% vero cell assay Van Donkersgoed et al. 1999 France 70% PCR Pradel et al. 2000 Japan 78.9% PCR Shinagawa et al. 2000 100% nested PCR Kobayashi et al. 2001 “Of the 1800 samples analysed, 40% (480/1200) of faecal and 27% (162/600) of soil samples were stx1 and/or stx2 positive. STEC were cultured from 1.9% (23/1200) of faecal and 0.7% (4/600) of soil samples …” Monaghan et al., 2011
Prevalence of STEC in Cattle Calves appear to be more susceptible to STEC colonization than older cows. STEC strains harboring stx 1 are more commonly isolated from cattle than those harboring stx 2. Bovine-related STEC isolates lack accessory virulence factors intimin and hemolysin EHEC serotypes (O157:H7, O111:H8, and O26:H11) are infrequently isolated form cattle when using unbiased methods.
bovine human - Majority of bovine STEC strains lack accessory virulence factors and are potentially less virulent. - Cannot distinguish between virulent and nonvirulent STEC.
Colonization • Tissue tropism difference reported for O157 and non- O157 STEC (van Dieman et al. I&I 2005) – 4-day old calves inoculated with 10 10 CFU O26 = spiral colon O157 = distal colon/RAJ • Later work with tissue explants on 6 wk old calves determines similar binding patterns. (Girard et al. AEM 2007) – O26 and O111 can bind at RAJ – O157 can adhere and induce A/E lesions at intestinal sites other than the terminal rectum
Colonization Naturally colonized dairy cattle shown to excrete non-O157 STEC (O113, O22) for >6 months (Monrath et al. 2011) Persistent shedder – yes, Super shedder - ? KG = Will swab sampling of RAJ be useful for detecting non-O157 STEC?
Prevalence of STEC by Sample Type Barkocy-Gallagher et al., 2003.
What do we know from the processing plant studies regarding the Top6 STEC Hides Pre-evis Final Trim % stx positive 91.7 96.5 16.2 30.0 % STEC isolate 56.6 58.0 8.9 5.7 ND 0.0 0.0 % Top 6 11.1 Arthur et al., 2002; Barkocy-Gallagher et al., 2003; Bosilevac et al., 2007.
Prevalence of Non-O157 contamination of post-intervention beef cattle carcasses U.S. 1 France 2 Hong Kong 3 Total Samples 326 851 986 Total non-O157 STEC positive 27 (8.3) 16 (1.9) 17 (1.7) PCR positive for stx genes 43 (13.4) 3 91 (10.7) 112 (11.4) 1 Arthur et al. 2002. 2 Rogerie et al. 2001. 3 Leung et al. 2001.
STEC virulence # of factors Isolates Preevis Post stx 1 152 135 17 stx 2 93 78 15 stx 1 , stx 2 15 15 0 stx 1 , eae 2 2 0 stx 1 , hlyA 8 3 5 stx 2 , hlyA 19 17 2 stx 1 , stx 2 , hlyA 31 23 8 stx 1 , stx 2 , eae 1 1 0 stx 1 , eae , hlyA 8 6 2 stx 2 , eae, hlyA 20 20 0 stx 1 , stx 2, eae, hlyA 12 10 2 Total 361 310 51
Prevalence of STEC in beef Number (%) of ground beef samples STEC Potential Total stx positive confirmation EHEC 4,133 (100) 1,006 (24.3) 300 (7.3) 10 (0.24) Bosilevac and Koohmaraie, 2011. New Zealand Australia Uruguay USA n 220 223 256 487 % stx gene 30 9.7 28 30 % STEC isolated 3.6 1.8 15.6 5.7 % Potential EHEC 0 0.9 2.3 1.0 isolated Bosilevac et al., 2007.
Prevalence of STEC in beef 16.8% retail meat samples positive for stx genes (n= 296, Samadpour et al. 2002) However, it is important to note that mere possession of the Stx toxin gene is not sufficient to convert a nonpathogenic E. coli strain into a pathogen. Although the prevalence of STEC in retail ground beef is much higher than that of E. coli O157:H7, which is a member of the group, it does not follow that the public health impact of the group is higher by the same magnitude. (Samadpour et al. 2002)
Outline -Background -Non-O157 STEC prevalence pre- and post-harvest - Non-O157 STEC response to interventions
Genome comparison EHEC strains carry large virulence plasmid (pEHEC) Ogura et al. PNAS 2009
Genome comparison EHEC Commensal EAEC ExPEC EPEC Shigella Ogura et al. PNAS 2009
Genome comparison These results indicate that the whole-gene repertoires of the EHECs are more similar to each other than to any of the other strains. Ogura et al. PNAS 2009 KG = What are the selective pressures and mechanisms driving the development of EHEC? • Does parallel evolution have implications for non-virulence-related attributes? • O104 outbreak – horizontal transmission
Recommend
More recommend
