Abstract
The Long Polar Fimbriae (Lpf) is one of few adhesive factors of Shiga toxin-producing Escherichia coli (STEC) and it is associated with colonization of the intestine. Studies have demonstrated the presence of lpf genes in several pathogenic E. coli strains and classification of variants based on polymorphisms in the lpfA1 and lpfA2 genes has been adopted. Using a collection of Argentinean Locus of Enterocyte Effacement (LEE)-negative STEC strains, we determined that the different lpfA types were present in a wide variety of serotypes and no apparent association was observed between the types of lpfA1 or lpfA2 genes and the severity of human disease. The lpfA2-1 was the most prevalent variant identified, which was present in 95.8% of the isolates, and the lpfA1-3 and lpfA2-2, proposed as specific biomarkers of E. coli O157:H7, were not found in any of the serotypes studied. The prevalence of lpf genes in a large number of strains is useful to understand the genetic diversity of LEE-negative STEC and to define the association of some of these isolates carrying specific lpf-variants with disease.
Keywords: LEE-negative, Shiga toxin-producing Escherichia coli, lpf variant, virulence profile, human strain, cattle strain
Introduction
Shiga toxin-producing Escherichia coli (STEC) strains are foodborne enteric pathogens associated with different clinical manifestations such as non-bloody diarrhea, hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS) (reviewed in Nataro & Kaper, 1998). Although E. coli O157:H7 is the most prevalent serotype associated with sporadic cases and large outbreaks of HC and HUS, there is growing concern over the emergence of highly virulent STEC non-O157 serotypes that become globally distributed and associated with outbreaks and/or severe human illness (Coombes et al., 2008). Ruminants, particularly cattle, are recognized as the main natural reservoir for non-O157 STEC and cattle-derived food products have been implicated in many outbreaks (Caprioli et al., 2005).
Shiga toxins (Stx) are considered to be the major virulence markers of STEC, but adherence to the intestinal epithelium and colonization of the gut is an essential component of pathogenic process (reviewed in Torres et al., 2005). Some STEC serotypes harbor a large pathogenicity island, termed the Locus of Enterocyte Effacement (LEE), required for the formation of Attaching and Effacing (A/E) lesions (McDaniel & Kaper, 1997). The LEE carries the eae gene encoding the adhesin intimin, responsible for the intimate adherence of the bacteria to the enterocyte and linked to cytoskeleton rearrangements. However, the presence of the LEE does not seem to be essential for full virulence, as a wide number of LEE-negative STEC strains have been associated with sporadic cases and small outbreaks of HC and HUS (reviewed in Bettelheim, 2007). Of these LEE-negative organisms, O113:H21 is one of the most commonly isolated STEC serotypes in many regions. Clinical isolates of LEE-negative STEC typically express Stx2 and also harbor a ca. 90-kb plasmid encoding several virulence factors, including EHEC hemolysin (Newton et al., 2009). Some studies have elucidated different mechanisms by which these strains interact with the host intestinal mucosa and induce disease; i.e. it has been demonstrated that STEC O113:H21 can invade tissue cultured cells (Luck et al., 2006). Besides the knowledge that some STEC strains do not carry the LEE, very little is known about other strategies used by these pathogens to adhere and to colonize the host intestine.
Analysis of the genome sequence of E. coli O157:H7 showed that several O157-specific islands contain putative fimbrial biosynthesis operons, including two regions encoding the Long Polar Fimbriae (Lpf). The Lpf loci are related at the genetic and protein level to Lpf of Salmonella enterica serovar Typhimurium and the latter have been shown to facilitate attachment of the bacteria to intestinal Peyer´s patches (Bäumler et al., 1996). In E. coli O157:H7, expression of the lpf operon 1 (lpf1) in an E. coli K-12 strain was linked to increased adherence to tissue-cultured cells and has been associated with the appearance of long fimbriae (Torres et al., 2002, 2004). The lpf2 operon has also been associated to adherence to epithelial cells and its expression in some pathogenic E. coli strains is believed important for the development of severe diarrhea (Doughty et al., 2002; Osek et al., 2003). E. coli O157:H7 strains harboring mutations in one or both of the lpf loci (named lpf1 and lpf2 operons) have diminished intestinal colonization abilities and persistence in several animal models of infection (Jordan et al., 2004; Torres et al., 2007; Newton et al. 2004). Further, these lpf mutant strains also displayed an altered human intestinal tissue tropism (Fitzhenry et al., 2006).
Cumulative evidence indicates that homologues to the lpf genes are found in other pathogenic E. coli, Salmonella, Shigella and even in some commensal E. coli isolates (Doughty et al., 2002; Szalo et al., 2002; Toma et al., 2004, 2006; Cergole-Novella et al., 2006; Galli et al., 2009). A recent study identified several polymorphisms within lpfA (encoding the major fimbrial Lpf subunit) genes, and this finding was used to classify the lpfA genes in distinct variants. The lpfA1 gene was classified in 5 different types (named as alleles 1, 2, 3, 4 and 5) and the lpfA2 gene in 3 distinct types (alleles 1, 2 and 3) (Torres et al., 2009). In the current study, we investigated the presence of these lpf variants in a collection of 120 LEE-negative STEC strains, 70 isolated from human infections and 50 from cattle; and explore the relationship between the presence of determined combination of lpf variants with other virulence determinants and severity of disease.
Materials and methods
Bacterial strains
A total of 120 randomly selected LEE-negative STEC strains belonging to different non-O157 serotypes were included in this study. Seventy human strains isolated during surveillance of HUS and diarrheal diseases, in a period from 2001 through 2009, and submitted to the Argentinean National Reference Laboratory, were included. The human strains were isolated from diarrheal cases (n=26), HUS (n=28) and asymptomatic household contacts (n=16). For comparison purposes, 50 strains isolated from fecal samples and carcasses from healthy Argentinean beef cattle, obtained during surveys and research programs carried out from 2005 to 2007 were also included. All the strains were previously serotypified and the presence of virulence genes (stx, eae, ehxA, saa, iha, fimA, efa1, astA, subAB, cdt-V) also determined (Galli et al., 2009).
Detection of lpfA variants
The primers and conditions employed in the PCR assays for identification of lpfA gene variants were identical to those reported by Torres et al. (2009). The DNA template was prepared boiling isolated single colonies of the strains in 150 µl of 1% Triton X-100 in TE buffer by 15 min. All amplifications began with a five-minute hot start at 94°C followed by 35 cycles of denaturing at 94°C for 30s, annealing for 30s in a range of temperatures ranging from 52°C–72°C (depending of the lpfA variant amplified), and extension at 72°C for 30s. E. coli strain EDL933 was used as positive control for lpfA1-3 and lpfA2-2; E. coli EH41 (O113:H21) for lpfA2-1 (kindly provided by Elizabeth Hartland); enteropathogenic E. coli (EPEC) 2348/69 (O127:H6) for lpfA1-1, and EPEC H30 (O26:H11) for lpfA1-2.
Statistical analysis
ANOVA and Pearson`s Chi square test were used to test associations between clinical courses (diarrhea, HUS and asymptomatic carriers) and the presence of the lpfA variant genes.
Results
Presence of lpf1 and lpf2 gene variants in LEE-negative STEC strains
Using the experimental classification of lpfA gene variants described by Torres et al. (2009), we found that the lpfA2-1 was the most commonly found variant in our isolates. As shown in Table 1, 95.8% of the strains carried the lpfA2-1 variant, while the lpfA2-3 variant was present in only one strain and 3.3% of the strains were negative for both lpfA1 and lpfA2 genes. The frequency of lpfA1-2 was 56.6% and it seems that this gene is frequently linked to lpfA2-1. We did not find lpfA1 variants 1, 3, 4 and 5, or lpfA2 variant 2 in any of the strains studied.
Table 1.
Characteristics of STEC strains regarding lpfA variants, origin, associated disease, stx genotype and serotype.
| Antigen |
No. of strains |
Origin/associated disease (No. of strains) |
stx genotype | lpfA1 | lpfA2 | |
|---|---|---|---|---|---|---|
| O | H | |||||
| 2 | 6 | 1 | A | 2 | 2 | 1 |
| 2 | 25 | 1 | C | 2 | 1 | |
| 5 | NM | 1 | A | 1 | 1 | |
| 7 | NM | 2 | C | 1 | 1 | |
| 7 | 21 | 1 | C | 2 | 1 | |
| 7 | 21 | 1 | C | 1+2 | 1 | |
| 8 | 16 | 1 | HUS | 1+2 | 2 | 1 |
| 8 | 16 | 1 | HUS | 2 | - | - |
| 8 | 16 | 1 | C | 1+2 | 2 | 1 |
| 8 | 19 | 1 | HUS | 2 | 2 | 1 |
| 8 | 19 | 2 | HUS | 2 | 1 | |
| 8 | 19 | 1 | D | 2 | 2 | 1 |
| 15 | 27 | 2 | HUS(1), C(1) | 2 | 1 | |
| 15 | 27 | 1 | D | 1+2 | 1 | |
| 20 | 19 | 1 | HUS | 2 | 1 | |
| 22 | 8 | 2 | C | 2 | 2 | 1 |
| 22 | 16 | 2 | C | 2 | 1 | |
| 22 | NT | 1 | D | 2 | 2 | 1 |
| 39 | 49 | 1 | C | 2 | 1 | |
| 46 | 38 | 2 | C | 1+2 | 1 | |
| 58 | 40 | 1 | D | 1 | 1 | |
| 59 | 19 | 4 | HUS | 2 | 2 | 1 |
| 74 | 12 | 1 | C | 1 | 1 | |
| 74 | 28 | 1 | C | 2 | 2 | 1 |
| 74 | 42 | 1 | C | 1+2 | 2 | 1 |
| 79 | 19 | 2 | C | 1+2 | 1 | |
| 82 | 8 | 1 | C | 1+2 | 2 | 1 |
| 91 | 16 | 1 | C | 2 | 2 | 1 |
| 91 | 21 | 5 | D(3), HUS(2) | 2 | 2 | 1 |
| 91 | 21 | 1 | C | 1+2 | 2 | 1 |
| 91 | NT | 1 | A | 1 | 2 | 1 |
| 91 | NM | 1 | HUS | 1+2 | 2 | 1 |
| 113 | 21 | 4 | HUS(2)*, D(1), C(1) | 2 | 1 | |
| 113 | NM | 1 | D | 2 | 2 | 1 |
| 116 | 21 | 2 | C | 1+2 | 1 | |
| 116 | 21 | 1 | C | 2 | 1 | |
| 117 | 7 | 1 | A | 1 | - | - |
| 130 | 11 | 2 | D(1), C(1) | 1+2 | 2 | 1 |
| 130 | 11 | 1 | C | 1 | 2 | 1 |
| 136 | 12 | 2 | C | 1 | 1 | |
| 141 | 49 | 2 | C | 2 | 2 | 1 |
| 143 | NM | 1 | HUS* | 2 | - | 3 |
| 163 | 19 | 2 | D(1), C(1) | 2 | 2 | 1 |
| 171 | 2 | 2 | D | 2 | 1 | |
| 174 | 8 | 1 | A | 2 | 1 | |
| 174 | 21 | 11 | HUS(1), D(5), A(4), C(1) | 2 | 2 | 1 |
| 174 | 28 | 2 | D(1), HUS(1)* | 2 | 1 | |
| 174 | 28 | 1 | C | 1+2 | 1 | |
| 174 | NM | 1 | D | 1+2 | 1 | |
| 174 | NM | 1 | HUS | 2 | 2 | 1 |
| 178 | 19 | 1 | C | 2 | 1 | |
| 178 | 19 | 1 | C | 2 | 2 | 1 |
| 178 | 19 | 2 | HUS(1), C(1) | 1+2 | 2 | 1 |
| 179 | 8 | 2 | C | 2 | 2 | 1 |
| NT | 2 | 2 | C | 2 | 1 | |
| NT | 4 | 1 | HUS | 2 | - | - |
| NT | 4 | 1 | HUS | 2 | 2 | 1 |
| NT | 7 | 2 | D(1), C(1) | 2 | 2 | 1 |
| OR | 11 | 1 | A | 2 | - | 1 |
| NT | 19 | 2 | A(1), C(1) | 2 | 1 | |
| NT | 19 | 1 | HUS | 2 | 2 | 1 |
| NT | 21 | 1 | C | 2 | 1 | |
| NT | 21 | 1 | C | 2 | 2 | 1 |
| NT | 28 | 2 | C | 2 | 2 | 1 |
| NT | 46 | 2 | A(1), C(1) | 2 | 2 | 1 |
| NT | 49 | 1 | D | 1+2 | 1 | |
| NT | NM | 5 | D(2), HUS(3) | 2 | 2 | 1 |
| NT | NM | 1 | D | 1 | - | - |
| NT | NM | 1 | A | 1+2 | - | 1 |
| NT | NM | 1 | C | 1 | 1 | |
| OR | NM | 1 | A | 2 | 2 | 1 |
| NT | NT | 1 | A | 1 | 2 | 1 |
| NT | NT | 2 | D(1), HUS(1) | 2 | 2 | 1 |
| NT | NT | 1 | HUS | 2 | 2 | 1 |
| NT | NT | 1 | A | 1 | 2 | 1 |
| Total | 120 | |||||
HUS patient died; C, cattle; A, asymptomatic patient; D, diarrheal patient; NM, non motile strains; NT, non-typeable.
The four lpfA-negative STEC strains identified in our study were of human origin (serotypes O8:H16, O117:H7, ONT:H4 and ONT:HNM). Two of them, serotypes O8:H16 and ONT:H4, were isolated from HUS cases and the only putative virulence factor currently identified in these strains is encoded by the iha gene. The ONT:HNM strain was isolated from a patient suffering from diarrhea and the only virulence factor found in this serotype is encoded by the stx1 toxin gene. Finally, the O117:H7 strain was isolated from an asymptomatic carrier with prolonged shedding and had the particularity to be non-sorbitol fermenting and carrying the putative virulence factors iha and astA.
In the current study, we could not find an statistical association between the presence of a particular lpfA variant and the severity of the disease. However, we observed that most serotypes maintained the same combination of lpf variants independent from the source of isolation. Therefore, we observed a good association between the lpfA variant and the serotype of the strain, i.e. we identified two strains from serotype O22:H8 that carried lpfA1-2 and lpfA2-1, and two strains from serotype O22:H16 which carried only lpfA2-1. Interestingly, we found that these strains belonging to the same serogroup and which were isolated from cattle, shared the same virulence profiles (Table 1). One more isolate from serotype O22:HNT and which was isolated from a human diarrheal case, carried the lpfA1-2 and lpfA2-1 genes. Similar results occurred in the O174 serogroup, where all the O174:H21 isolates carried the lpfA1-2 and lpfA2-1 gene variants, while the other O174 serotypes (O174:H8, O174:H28, O174:NM) carried the lpfA2-1 gene and not a common theme with respect to the virulence profile or the source of isolation was observed.
The most variable serogroup with respect to lpfA gene variant content was O8 from which we identified three O8:H16 and four O8:H19 isolates. In the case of the O8:H16 isolates, two were lpfA1-2 and lpfA2-1-positive and carried the same virulence profile, while a third isolate was lpfA-negative and iha was the only putative virulence marker. Another difference in these strains, apart from the source of isolation, was the stx genotype, while the lpfA-negative strain was stx2-positive, the others were stx1/stx2-positive. In the case of the O8:H19 isolates, two carried the lpfA1-2 and lpfA2-1 genes, and two strains carried only the lpfA2-1 gene. Further, all the strains of this sertotype carried different virulence gene profiles.
Another serotype identified in our study was O178:H19, which included two strains sharing the same origin, and carrying the same stx gene and a common virulence profile, but differing in the type of lpfA variant present. While one strain was lpfA1-2-positive, the other was lpfA1-2 and lpfA2-1-positive.
The virulence profiles found in the LEE-negative STEC strains are presented in Table 2, and differences were observed between human and cattle isolates. Sixteen different virulence profiles were identified among the 70 human strains, being the combination of iha fimA genes the most prevalent (19 of 70 strains, 27.1%). Within this group, the presence of lpfA1-2 and lpfA2-1(12 of 19 strains, 63%), only lpfA2-1 (6 of 19 strains, 32%) and only lpfA2-3 (1 of 19 strains, 5%) was detected. Among the 50 bovine strains, 9 different virulence profiles were observed, being the combination of iha fimA saa ehxA subAB the most prevalent (14 of 50 strains, 28%). From these, 8 of 14 strains (57%) carried the lpfA1-2 and lpfA2-1 variants, while 6 of the 14 strains (43%) contained the lpfA2-1 variant.
Table 2.
Comparison of virulence profiles and lpfA variants identified in human and cattle LEE-negative STEC strains
| Virulence profile | No. of strains | ||||||
|---|---|---|---|---|---|---|---|
| Human | Cattle | Total | |||||
|
lpfA1-2 lpfA2-1 |
lpfA2-1 | lpfA2-3 |
lpfA- negative |
lpfA1-2 lpfA2-1 |
lpfA2-1 | ||
| iha fimA | 20 | 6 | 1 | 3 | 6 | 36 | |
| iha fimA saa ehxA subAB | 3 | 4 | 8 | 6 | 21 | ||
| iha fimA saa ehxA subAB | 6 | 3 | 2 | 5 | 16 | ||
| cdt-V | |||||||
| iha fimA saa ehxA | 2 | 2 | 6 | 2 | 12 | ||
| iha fimA astA | 5 | 1 | 6 | ||||
| iha astA | 2 | 1 | 1 | 4 | |||
| fimA ehxA | 1 | 2 | 1 | 4 | |||
| fimA | 2 | 2 | |||||
| fimA astA | 1 | 1 | |||||
| fimA saa ehxA | 1 | 1 | |||||
| fimA saa ehxA subAB | 1 | 1 | |||||
| iha fimA saa | 1 | 1 | |||||
| iha fimA ehxA | 1 | 1 | |||||
| iha fimA saa ehxA cdt-V | 3 | 3 | |||||
| iha saa ehxA | 1 | 1 | |||||
| Iha | 2 | 2 | |||||
| astA | 6 | 6 | |||||
| fimA ehxA astA | 1 | 1 | |||||
| All negative | 1 | 1 | |||||
| Total | 45 | 20 | 1 | 4 | 24 | 26 | 120 |
Discussion
The virulence factors of LEE-negative STEC strains can not only be limited to the production of Stx toxin variants, but also to the presence of adhesins that mediate binding to the intestinal epithelium and eventually, contribute to the colonization of the gut. Some studies have suggested that LEE-negative STEC are invasive and that a particular flagellin type may contribute to cell invasion and gut colonization (Luck et al., 2005, 2006). Besides those observations, still little is known about other adhesins associated with colonization of the intestine and other mechanisms of pathogenesis. Recently, Torres et al. (2009) identified several polymorphisms within the lpfA genes, which were used to classify the major fimbrial subunit genes in distinct variants. The expression of Lpf in LEE-negative STEC strains is believed to be important for development of severe diarrhea and hence its identification is potentially clinically relevant (Dougthy et al., 2002; Osek et al., 2003).
In an attempt to characterize some fimbrial adhesins in these pathogens, we investigated the distribution of lpfA gene variants in a wide range of LEE-negative STEC strains isolated in Argentina from human infections and healthy cattle. We found that the lpfA1 and lpfA2 genes are present in 56.6% and 96.6% of the STEC strains studied, respectively, and only 3.3% of the human strains were lpfA-negative. These data confirmed that the presence of lpf genes in LEE-negative STEC strains seems to be a common characteristic, particularly the presence of the lpfA2-1 variant. It is plausible to speculate that the four lpfA-negative strains identified in this study either contain novel and unidentified adherence factors required for colonization or perhaps the strains possess another lpf operon that we could not identified with our detection approach.
The majority of the strains possessed the lpfA2-1 allele (95.8%). Indeed, 39.1% of the strains were only lpfA2-1-positive and 56.6% were positive for both lpfA1-2 and lpfA2-1 genes. It is interesting to mention that the most common variant in bovine isolates was that encoded by the lpfA2-1 gene, while the combination lpfA1-2 and lpfA2-1 was the common genotype in human isolates. This finding suggests that, perhaps the presence of the lpfA2-1 variant might be linked to those isolates that are cattle colonizers, while the strains containing the lpfA1-2 and lpfA2-1 genes had an advantage to colonize the human intestine and eventually to cause disease. It is evident that additional experiments are required to confirm Lpf expression in these strains and to establish the association of those strains expressing specific variants of Lpf with human disease. Interestingly, some of our prior studies evaluating adherence of the strains to HEp-2 cell showed different adhesive profiles between those strains possessing different lpfA variants, i.e. a lpfA2-3 positive strain adhere to the HEp-2 cell surface in a localized adherence like pattern, while three cattle lpfA2-1-positive strains adhere but also invade these tissue cultured cells (Galli et al., 2009).
Although a great diversity of serotypes and virulence profiles were observed among human and bovine LEE-negative STEC strains, seven of the eighteen profiles were common in both groups. This observation reinforces the idea that cattle are the main natural reservoir of LEE-negative STEC strains, and potentially, the principal source of infection to humans. We also confirmed that LEE-negative STEC strains are not a clonal group of pathogens, as we observed differences in their virulence profiles, including strains from the same serotype. Some of these determinants are not considered essential factors for human infection, although their presence could facilitate survival and persistence of the strains in different environments.
In agreement with previous data described by Torres et al. (2009), none of the strains analyzed in this study carried the lpfA1-3 or the lpfA2-2 gene variants, either alone or in combination. Therefore, our study strongly supports their observation that those two gene variants are specific for the O157:H7 lineage and are not present in any other STEC isolates, regardless of the source or their association with disease. Interestingly, the only virulence factor that has been associated with the presence of specific lpf genes is the adhesin intimin (Torres et al., 2009). That study indicated that different intimin alleles are associated to specific lpfA gene variants, and the presence of both lpfA1 and lpfA2 alleles is also linked to specific pathogenic E. coli strains, particularly with those belonging to the STEC pathotype group (Torres et al., 2009). However, that study did not include those strains that are LEE-negative (intimin-negative), which adds a significant value to our current study, because in addition to confirm some of their findings, we now provided cumulative evidence regarding the distribution of lpfA gene variants in other STEC strains that are significant human pathogens. Finally, the presence of the lpfA genes in a large number of strains supports the idea that Lpf might play an important but not fully characterized role in adherence and/or colonization capabilities of LEE-negative STEC strains.
Acknowledgements
We want to thank Patricio Valenzuela for technical assistance, and Kinue Irino for previously serotype the strains. This work was partially supported by grants from Agencia Nacional de Promoción Científica y Tecnológica, PICT 26093/2004. LG was supported by a fellowship from Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET). Research in the AGT laboratory is supported in part by NIH AI079154 -01A2 grant.
References
- Bäumler AJ, Tsolis RM, Heffron F. The lpf fimbrial operon mediates adhesion of Salmonella typhimurium to murine Peyer´s patches. Proc Natl Acad Sci USA. 1996;93:279–283. doi: 10.1073/pnas.93.1.279. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bettelheim KA. The Non-O157 Shiga-Toxigenic (Verocytotoxigenic) Escherichia coli; under-rated pathogens. Crit Rev Microbiol. 2007;33:67–87. doi: 10.1080/10408410601172172. [DOI] [PubMed] [Google Scholar]
- Caprioli A, Morabito S, Brugreb H, Oswald E. Enterohaemorrhagic Escherichia coli: emerging issues on virulence and modes of transmission. Vet Res. 2005;36:289–311. doi: 10.1051/vetres:2005002. [DOI] [PubMed] [Google Scholar]
- Cergole-Novella MC, Nishimura LS, Dos Santos LF, Irino K, Vaz TM, Bergamini AM, Guth BEC. Distribution of virulence profiles related to new toxins and putative adhesins in Shiga toxin-producing Escherichia coli isolated from diverse sources in Brazil. FEMS Microbiol Lett. 2007;274:329–334. doi: 10.1111/j.1574-6968.2007.00856.x. [DOI] [PubMed] [Google Scholar]
- Coombes BK, Wickham ME, Mascarenhas M, Gruenheid S, Brett Finlay B, Karmali MA. Molecular analysis as an aid to assess the Public Health risk of Non-O157 Shiga toxin-producing Escherichia coli strains. Appl Eviron Microbiol. 2008;74:2153–2160. doi: 10.1128/AEM.02566-07. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Doughty S, Sloan J, Bennet-Wood V, Robertson M, Robins-Browne RM, Hartland EL. Identification of a novel fimbrial gene cluster related to long polar fimbriae in locus of enterocyte effacement-negative strains of enterohemorrhagic Escherichia coli. Infect Immun. 2002;70:6761–6769. doi: 10.1128/IAI.70.12.6761-6769.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fitzhenry R, Dahan S, Torres AG, Chong Y, Heuschkel R, Murch S, Thomson M, Kaper JB, Frankel G, Phillips AD. Long polar fimbriae and tissue tropism in Escherichia coli O157:H7. Int J Med Microbiol. 2006;296:547–552. doi: 10.1016/j.micinf.2006.02.012. [DOI] [PubMed] [Google Scholar]
- Galli L, Leotta GA, Irino K, Rivas M. Virulence profile comparison between LEE-negative Shiga toxin-producing Escherichia coli strains isolated from cattle and humans. Vet Microbiol. 2009 doi: 10.1016/j.vetmic.2009.11.028. Epub ahead of print. [DOI] [PubMed] [Google Scholar]
- Galli L, Larzábal M, Leotta GA, Mercado EC, Rivas M. Differential adherence of locus of enterocyte effacement-negative Shiga-toxin producing Escherichia coli strains harboring lpfAO113 and/or iha genes to HEp-2 cells, p. 80; Abstr. 7th International Symposium on Shiga Toxin (Verocytotoxin) Producing E. coli Infections; Buenos Aires, Argentina: Asociación Argentina de Microbiología Publishing; 2009. [Google Scholar]
- Jordan DM, Cornick N, Torres AG, Dean-Nystrom EA, Kaper JB, Moon HW. Long Polar Fimbriae Contribute to Colonization by Escherichia coli O157:H7 in vivo. Infect Immun. 2004;72:6168–6171. doi: 10.1128/IAI.72.10.6168-6171.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Luck SN, Bennet-Wood V, Poon R, Robins-Browne RM, Hartland EL. Invasion of epithelial cells by locus of enterocyte effacement-negative enterohemorragic Escherichia coli. Infect Immun. 2005;73:3063–3071. doi: 10.1128/IAI.73.5.3063-3071.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Luck SN, Badea L, Bennet-Wood V, Robins-Browne RM, Hartland EL. Contribution of FliC to epithelial cell invasion and by enterohemorrhagic Escherichia coli O113:H21. Infect Immun. 2006;74:6999–7004. doi: 10.1128/IAI.00435-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
- McDaniel TK, Kaper JB. A cloned pathogenicity island from enteropathogenic Escherichia coli confers the attaching and effacing phenotype on E. coli K12. Mol Microbiol. 1997;2:399–407. doi: 10.1046/j.1365-2958.1997.2311591.x. [DOI] [PubMed] [Google Scholar]
- Nataro JP, Kaper JB. Diarrheagenic Escherichia coli. Clin Microbiol Rev. 1998;11:142–201. doi: 10.1128/cmr.11.1.142. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Newton HJ, Sloan J, Bennet-Wood V, Adams LM, Robins-Browne RM, Hartland EL. Contribution of long polar fimbriae to the virulence of rabbit-specific enteropathogenic Escherichia coli. Infect Immun. 2004;72:1230–1239. doi: 10.1128/IAI.72.3.1230-1239.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Newton HJ, Sloan J, Bulach DM, Seemann T, Allison CC, Tauschek M, Robins-Browne RM, Paton JC, Whittam TS, Paton AW, Hartland EL. Shiga toxin-producing Escherichia coli strains negative for Locus of Enterocyte Effacement. Emerg Infect Dis. 2009;15:372–378. doi: 10.3201/eid1502.080631. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Osek J, Weiner M, Hartland EL. Prevalence of the lpfO113 gene cluster among Escherichia coli O157 isolates from different sources. Vet Microbiol. 2003;96:259–266. doi: 10.1016/j.vetmic.2003.07.002. [DOI] [PubMed] [Google Scholar]
- Toma C, Martinez EE, Song T, Miliwebsky E, Chinen I, Iyoda S, Iwanaga M, Rivas M. Distribution of putative adhesins in different seropathotypes of Shiga toxin-producing Escherichia coli. J Clin Microbiol. 2004;42:4937–4946. doi: 10.1128/JCM.42.11.4937-4946.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Toma C, Nakasone N, Miliwebsky E, Higa N, Rivas M, Suzuki T. Differential adherence of Shiga toxin-producing Escherichia coli harboring saa to epithelial cells. Int. J Med Microbiol. 2007;298:571–578. doi: 10.1016/j.ijmm.2007.12.003. [DOI] [PubMed] [Google Scholar]
- Torres AG, Giron JA, Perna NT, Burland V, Blattner FR, Avelino-Flores F, Kaper JB. Identification and characterization of lpfABCC'DE, a fimbrial operon of enterohemorrhagic Escherichia coli O157:H7. Infect Immun. 2002;70:5416–5427. doi: 10.1128/IAI.70.10.5416-5427.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Torres AG, Kanack KJ, Tutt CB, Popov V, Kaper JB. Characterization of the second long polar (LP) fimbriae of Escherichia coli O157:H7 and distribution of LP fimbriae in other pathogenic E. coli strains. FEMS Microbiol Lett. 2004;238:333–344. doi: 10.1016/j.femsle.2004.07.053. [DOI] [PubMed] [Google Scholar]
- Torres AG, Zhou X, Kaper JB. Adherence of diarrheagenic Escherichia coli strains to epithelial cells. Infect Immun. 2005;73:18–29. doi: 10.1128/IAI.73.1.18-29.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Torres AG, Milflores-Flores L, Garcia-Gallegos JG, Patel SD, Best A, La Ragione RM, Martinez-Laguna Y, Woodward MJ. Environmental regulation and colonization attributes of the Long Polar Fimbriae of Escherichia coli O157:H7. Int J Med Microbiol. 2007;297:177–185. doi: 10.1016/j.ijmm.2007.01.005. [DOI] [PubMed] [Google Scholar]
- Torres AG, Blanco M, Valenzuela P, Slater TM, Patel1 SD, Dahbi G, López C, Fernández Barriga X, Blanco JE, Gomes TAT, Vidal R, Blanco J. The Long Polar Fimbriae genes of pathogenic Escherichia coli strains as reliable markers to identify virulent isolates. J Clin Microbiol. 2009;47:2442–2451. doi: 10.1128/JCM.00566-09. [DOI] [PMC free article] [PubMed] [Google Scholar]