Colonization factors of enterotoxigenic Escherichia coli isolated from children with diarrhea in Argentina.

A prospective study was performed to evaluate the presence of colonization factor antigens (CFAs) in enterotoxigenic Escherichia coli (ETEC) strains isolated from 1,211 children with diarrhea in Argentina. One hundred nine ETEC strains that were isolated from seven different laboratories in various regions of the country were tested for CFAs by using monoclonal antibodies against CFA/I and E. coli surface antigens CS1, CS2, and CS3 of CFA/II and CS4 and CS5 of CFA/IV; a polyclonal antiserum against CS6 was used. The CFAs searched for were found in 52% of the ETEC strains: 23% of the strains carried CFA/I, 17% carried CFA/IV, and 12% carried CFA/II. All of the CFA/I strains produced heat-stable enterotoxin, and several of them were of the prevalent serotypes O153:H45 and O78:H12. Among the 19 strains expressing CFA/IV, 16 expressed CS5 and CS6 and produced the heat-stable enterotoxin and most were of serotype O128:H21; the remaining 3 strains produced CS6 only. No ETEC strains expressing CS4 were found. Most (11 of 13) of the CFA/II-carrying ETEC strains expressed CS1 and CS3, and 10 of them were of the O6:K15:H16 serotype and produced both heat-labile and heat-stable toxins. As many as 24 of the 109 CFA-negative ETEC strains gave mannose-resistant hemagglutination with erythrocytes from different species; 4 strains had high surface hydrophobicity, suggesting the presence of additional, as yet undefined, colonization factors in up to 25% of the ETEC isolates.     

Plasmids coding for colonization factor antigen I and heat-stable enterotoxin production isolated from enterotoxigenic Escherichia coli: comparison of their properties.

We examined seven enterotoxigenic Escherichia coli strains which produced colonization factor antigen I (CFA/I). Four of these strains were from South Africa (three serotype O78:H12 and one serotype O63:H-), one was from Ethiopia (O78:H12), and two were from Bangladesh (O78:H11 and O78:H12). Plasmids coding for CFA/I were mobilized from six of these strains by using resistance or enterotoxin factors. No plasmid was mobilized from the serotype O78:H12 Bangladesh strain. The transconjugants obtained from crosses with the O78 strains also produced heat-stable enterotoxin (ST), and additional investigations showed that CFA/I and ST were coded for by a single non-autotransferring plasmid. These plasmids were fertility inhibition negative, did not restrict any of the coliphages with which they were tested, and were incompatible with each other. Four had molecular weights of approximately 60 X 10(6), and one had a molecular weight of 52 X 10(6). Like the other CFA/I plasmids, the CFA/I plasmid transferred from the O63:H- strain coded for ST, but this plasmid also coded for heat-labile enterotoxin. In most other respects the properties of this plasmid were similar to those of the CFA/I-ST plasmids previously described. The molecular weight of this plasmid was 65 X 10(6). The IncT R-factor Rtsl was marked with a transposon for tetracycline resistance and then transferred into the two Bangladesh wild-type strains. Plasmids which coded for tetracycline resistance, CFA/I, and ST were transferred from these strains. These plasmids were incompatible with Rtsl and with the CFA/I-ST plasmids described above and were recombinants between Rtsl and a CFA/I-ST plasmid. Their properties are also described.     

Relationship between enterotoxin production and serotype in enterotoxigenic Escherichia coli.

We examined the relationship between serotype and enterotoxin production in 109 enterotoxigenic Escherichia coli strains isolated from 109 patients with severe cholera-like diarrhea in Dacca, Bangladesh. Of 69 strains producing both heat-labile and heat-stable toxins, 59 (86%) belonged to the one of four O serogroups, and 56 (81%) of these strains belonged to one of six O:K:H serotypes. In contrast, 34 strains producing only heat-stable toxin were distributed among 15 O serogroups, and six strains producing only heat-labile toxin were distributed among six O serogroups. Twelve strains producing heat-labile and heat-stable toxins and five strains producing heat-stable toxin were found which had the same serotype (O78:K-:H12) and biotype. It appears that at least in one geographic setting E. coli strains producing both heat-labile and heat-stable toxins are more restricted in their O groups and O:K:H serotypes than E. coli that produce only heat-stable toxin and that certain serobiotypes may commonly include strains which produce both toxin types.     

Enterotoxigenic Escherichia coli in a population of infants with diarrhea in Chile.

The incidence of enterotoxigenic Escherichia coli (ETEC) was investigated in 95 E. coli strains isolated from 48 infants with diarrhea in Santiago, Chile. By using standard biological assays and DNA-DNA hybridization procedures, ETEC was found in 31.2% of the cases: 14 strains produced heat-stable enterotoxin (ST) only, three strains produced heat-labile enterotoxin (LT) and ST, and two strains produced LT only. DNA probes detected all enterotoxin producers except one ST-producing strain. The ST strains hybridized with one or both of the human ST probes (ST Ib and ST A2). Two of the LT-ST strains hybridized with the ST Ia and ST Ib probes, and the third strain did not hybridize with any of the ST probes. Only the ST group expressed multiple resistance (85.7%) and colonization factor antigen I (CFA I) (92.8%); CFA II was found in two of three LT-ST strains. The O153:H45 serotype was found in 10 of 14 ST strains, and O6:K15:H16 was found in one LT strain and in two LT-ST strains. These findings suggest that ETEC, especially strains that produce ST, may be an important cause of diarrhea among Chilean infants.     

Characterization of monoclonal antibodies against putative colonization factors of enterotoxigenic Escherichia coli and their use in an epidemiological study.

Monoclonal antibodies (MAbs) against five putative colonization factors (PCFs), i.e., colonization factor antigen (CFA)/III, coli surface antigen (CS)7 and CS17, PCFO159, and PCFO166 of enterotoxigenic Escherichia coli (ETEC), were produced. Hybridomas (one each) producing specific antibodies against the respective PCFs were selected. All the MAbs reacted with the corresponding fimbriae but not with CFA/I, CFA/II, or CFA/IV or the heterologous PCFs in bacterial agglutination and enzyme-linked immunosorbent assays (ELISAs). In immunoelectron microscopy these MAbs bound along the fimbriae, and they also reacted with the corresponding subunits in immunoblots. The five MAbs were used to evaluate the prevalence of CFA/III, CS7, CS17, PCFO159, and PCFO166 in ETEC strains isolated from children with diarrhea in Argentina. One hundred five ETEC isolates negative for CFA/I, CFA/II, and CFA/IV were tested in slide agglutination or in a dot blot test for spontaneously agglutinating strains; positive results were confirmed by inhibition ELISAs. It was found that 27% of the CFA-negative ETEC strains carried one of the PCFs. The sensitivity of slide agglutination with these MAbs was similar to that with specific polyclonal antisera; however, the specificity was higher. PCFO166 was found in 9.5% of the strains tested, mainly in ETEC of serogroup O78 producing heat-stable toxin alone. CS17 and CS7 were identified in 6.7 and 5.7%, respectively, of strains producing heat-labile toxin only, most of which belonged to serogroup O114. PCFO159 was found in 3.8% of the isolates tested, whereas CFA/III was detected in only one ETEC strain.     

Genetic and molecular studies of plasmids coding for colonization factor antigen I and heat-stable enterotoxin in several escherichia coli serotypes.

Plasmids coding for colonization factor antigen I (CFA/I) and heat-stable enterotoxin (ST) were identified in 10 strains of human enterotoxigenic Escherichia coli. The strains, which belonged to serogroups O63, O114, O128, and O153, were isolated in Bangladesh, Latin America, Spain, and South Africa. Two strains produced heat-labile enterotoxin in addition to ST. CFA/I-ST plasmids were mobilized from two O128 strains into E. coli K-12 with the R factor R1-19K-. Like the prototype CFA/I-ST plasmid NTP113, mobilized previously from an E. coli O78 strain into K-12, these two plasmids were non-autotransferring. All 10 CFA/I-ST plasmids were incompatible with NTP113 and had molecular weights ranging from 59 X 10(6) to 72 X 10(6). The molecular properties of seven of these plasmids were compared with those of six CFA/I-ST plasmids previously mobilized from O78 strains from Ethiopia, South Africa, and Bangladesh and with those of one plasmid coding for CFA/I, ST and heat-labile enterotoxin from a South African strain of serogroup O63. Digestion with the restriction endonuclease HindIII showed that several plasmids had very similar fragment patterns and two were identical. Generally, a larger proportion of HindIII fragments were of common size in digests of plasmids identified in strains from related geographical areas, regardless of serogroup. However, all except one plasmid shared five or six HindIII fragments of the same size, one of which had been shown previously to be involved in CFA/I production. There was at least 90% DNA homology between CFA/I-ST plasmids with a molecular weight of about 58 X 10(6) from O78 strains from different sources. Most of the DNA sequences of these plasmids were present in a larger CFA/I-ST plasmid (72 X 10(6) from an O128 strain. The results of genetic and molecular studies suggest that CFA/I and ST production is determined by very similar plasmids in different serogroups of human enterotoxigenic E. coli from several sources.     

Immunity to Enterotoxigenic Escherichia coli

Enterotoxigenic Escherichia coli strains represent the most frequent etiological agent of travelers diarrhea. Challenge studies with several of these strains were undertaken in volunteers to evaluate the mechanisms of disease-induced immunity. Seventeen students and other community volunteers were given 10(6) or 10(8) organisms of E. coli B7A (O148:H28), which produces heat-labile and heat-stable enterotoxins. Ten individuals developed diarrheal illness closely resembling natural travelers diarrhea; of these ten, rises in titer of serum antitoxin and anti-O antibody occurred in eight (80%). Eight of the volunteers who developed diarrhea in the first test agreed to undergo rechallenge 9 weeks later with 10(8) B7A organisms. Only one of these eight "veterans" developed diarrhea versus seven of twelve controls given the same challenge (P = 0.05). Despite clinical protection, all "veterans" excreted B7A after rechallenge. Four controls who developed diarrhea during the homologous B7A rechallenge test were rechallenged 9 weeks later with 10(9) organisms of E. coli strain E2528-C1 (O25:H-), which produces only heat-labile enterotoxin and possesses a different O, H, and pili antigen composition than B7A. Three of four "veterans" and two of six controls developed comparable diarrhea. These studies demonstrate that prior disease due to enterotoxigenic E. coli confers homologous immunity against subsequent challenge, and the operative mechanism apparently is not bactericidal and is not mediated by serum anti-O antibodies. Heterologous protection was not conferred where the only common antigen was heat-labile enterotoxin, indicating that serum infection-derived antitoxin to heat-labile enterotoxin also is not protective.     

Relationship among enterotoxigenic phenotypes, serotypes, and sources of strains in enterotoxigenic Escherichia coli.

The relationship among O groups, O:H serotypes and enterotoxigenic phenotypes was examined in 76 Escherichia coli strains isolated in Brazil from different sources. Of the 17 heat-labile and -stable enterotoxin (LT/ST)-producing strains whose O antigens were identified, 15 belonged to serotypes O6:H16 (7 strains), O63:H- (5 strains), and O139:H28 (3 strains). All 11 ST strains were in group OO128PAC, which was represented by four O:H serotypes. The 23 LT strains with the O antigen identified were distributed among serotypes of 14 O groups. Colonization factor CFA/I was not found in any of the LT strains, but it was found in six LT/ST and three ST strains. On the whole, each E. coli O:H serotype had a particular fermentation pattern. LT/ST as well as ST strains were all isolated from patients with diarrhea, whereas LT strains were isolated from patients with diarrhea, normal children, food, and river water.     

Evaluation of antisera used for detecting enterotoxigenic Escherichia coli in Sao Paulo.

The usefulness of antisera in detecting enterotoxigenic Escherichia coli (ETEC) strains in Sao Paulo was evaluated. Polyvalent antisera detected 49% of ETEC isolates and were more effective in identifying E. coli that produced heat-labile and heat-stable enterotoxins and in strains that produced only heat-stable enterotoxin. ETEC strains not detected by the antisera belonged to different serogroups not isolated in Sao Paulo before; 34% of these strains had undetermined O antigens, and most of them produced only heat-labile toxin. A variation of serogroups over time was especially observed among strains that produced heat-stable toxin. The importance of H-antigen determinations in the effectiveness of ETEC diagnosis by serological methods became evident, as non-ETEC strains were also detected by polyvalent antisera, but their serotypes were different from those of ETEC strains. Although antisera can be used to identify O:H types of ETEC strains with accuracy, serotyping cannot be recommended for routine diagnosis. However, such a procedure may be useful for studying outbreaks of ETEC diarrhea if the involved serotypes are already known.     

Survey in Vanuatu on enterotoxigenic Escherichia coli in children and infants with and without acute diarrhea.

We have studied the incidence of enterotoxigenic Escherichia coli (ETEC) strains isolated from infants with and without diarrheal diseases in Vanuatu, South Pacific. Over a period of 5 months we have isolated enterotoxigenic E. coli strains from 29 (26.6%) of 109 children with acute diarrhea and from 13 (21.6%) of 60 children of the control group. In the group with diarrhea, 7 (6.4%) strains released heat-labile toxin, 7 (6.4%) released heat-stable toxin, and 15 (13.7%) produced both heat-labile and heat-stable toxin. In the control group, only one strain (1.6%) produced heat-stable toxin, 12 (20%) produced heat-labile toxin, and none produced both. Association of strains releasing heat-stable toxin or both heat-labile and heat-stable toxin with diarrhea was highly significant as shown by statistical analysis. The O serogroups and colonization factors CFA/I and CFA/II are presented.     

Enterotoxigenic Escherichia coli associated with infant diarrhoea in Galicia, north-western Spain

To assess the role of enterotoxigenic Escherichia coli (ETEC) in infantile diarrhoea, 482 children with diarrhoea and 103 healthy controls, from three localities of Galicia, north-western Spain, were investigated between 1985 and 1988. Rotavirus (37.3%) and Salmonella spp. (12.8%) were the most common causal agents, followed by ETEC (3.9%), Campylobacter jejuni (2.3%), Shigella spp. (0.9%) and Yersinia enterocolitica (0.5%). ETEC were significantly more frequently isolated from children with diarrhoea who were under 1 month of age (26.5%) than from older diarrhoeic children (2.2%) (p less than 0.001) or from healthy children who were under 1 month of age (0%) (p less than 0.05). Among children who harboured ETEC, five of the nine children under 1 month of age developed diarrhoea in hospital, whereas none of the 10 children over 1 month of age did so. Seventeen ETEC isolates produced heat-stable enterotoxin (STa) only, four produced only heat-labile enterotoxin (LT), and two produced both toxins. Colonisation factor antigens CFA/I and CFA/II were detected in 11 (55.0%) of the 20 ETEC isolates that remained enterotoxigenic after maintenance in the laboratory. Most ETEC isolates belonged to serotypes O153:K-:H45 (nine STa+ CFA/I+ isolates), O27:K-:H7 (three STa+ isolates) or O6:K15:H16 (two LT+ STa+ CFA/II+ isolates). Our results suggest that ETEC constitute an important cause of neonatal diarrhoea in this part of Spain.     

Characterization of enterotoxigenic Escherichia coli isolated from U.S. troops deployed to the Middle East.

Enterotoxigenic Escherichia coli (ETEC) was a common cause of traveler's diarrhea in U.S. soldiers in the Middle East in 1989 and 1990. To determine which bacterial components would be useful in a vaccine, potential protective antigens (toxin, colonization factor antigen [CFA], and serotype) from 189 ETEC isolates were examined. Nearly half of the isolates expressed both ETEC toxins, 39% had only heat-stable enterotoxin (ST), and 17% had heat-labile enterotoxin (LT). CFA/I was the least common colonization factor antigen (11%), CFA/II was common (34%), as was CFA/IV (31%), and 24% expressed none of these CFAs. Fifty-seven O:H serotypes were found. Serotype O6:H16 was the most common, occurring in 29% of the ETEC isolates, usually with LT-ST and CFA/II. Generally, CFA/II was associated with expression of both toxins, CFA/IV was associated with expression of ST, and none of the CFAs was routinely found with LT. We conclude that ETEC from soldiers in the Middle East expressed a variety of antigens and that an effective vaccine will require multiple protective antigens.     

Heat-labile enterotoxin produced by Escherichia coli serogroup O149 isolated from diarrheic calves.

Nine strains of Escherichia coli serogroup O149 isolated from diarrheic calves were found to produce a heat-labile enterotoxin. The heat-labile enterotoxin activity was destroyed by heat (85 degrees C/30 min) and was neutralized by rabbit anti-heat-labile enterotoxin. One of the strains also produced a heat-stable enterotoxin.     

Outbreak of infantile enteritis caused by enterotoxigenic Escherichia coli O6.H16.

Ten out of 18 babies at risk developed enteritis in an outbreak in a special-care baby unit. Salmonella, Shigella, and Escherichia coli belonging to the traditional infantile enteropathogenic serogroups were not found in faeces from the babies and the staff, and no virus particles were found by electron microscopy. Detailed serotyping of E. coli showed that five of the 10 babies with diarrhoea and one of the eight without diarrhoea were excreting E. coli O6.H16. All six isolates of this serotype produced both heat-labile and heat-stable enterotoxin. Enterotoxigenic strains of E. coli O6.H16 have caused outbreaks of enteritis in adults in the USA and Japan.     

Phenotypic Diversity of Enterotoxigenic Escherichia coli Strains from a Community-Based Study of Pediatric Diarrhea in Periurban Egypt

No past studies of diarrhea in children of the Middle East have examined in detail the phenotypes of enterotoxigenic Escherichia coli (ETEC) strains, which are important pathogens in this setting. During a prospective study conducted from November 1993 to September 1995 with 242 children under 3 years of age with diarrhea living near Alexandria, Egypt, 125 episodes of diarrhea were positive for ETEC. ETEC strains were available for 98 of these episodes, from which 100 ETEC strains were selected and characterized on the basis of enterotoxins, colonization factors (CFs), and O:H serotypes. Of these representative isolates, 57 produced heat-stable toxin (ST) only, 34 produced heat-labile toxin (LT) only, and 9 produced both LT and ST. Twenty-three ETEC strains expressed a CF, with the specific factors being CF antigen IV (CFA/IV; 10 of 23; 43%), CFA/II (5 of 23; 22%), CFA/I (3 of 23; 13%), PCFO166 (3 of 23; 13%), and CS7 (2 of 23; 9%). No ETEC strains appeared to express CFA/III, CS17, or PCFO159. Among the 100 ETEC strains, 47 O groups and 20 H groups were represented, with 59 O:H serotypes. The most common O serogroups were O159 (13 strains) and O43 (10 strains). O148 and O21 were each detected in five individual strains, O7 and O56 were each detected in four individual strains, O73, O20, O86, and O114 were each detected in three individual strains, and O23, O78, O91, O103, O128, and O132 were each detected in two individual strains. The most common H serogroups were H4 (16 strains), 12 of which were of serogroup O159; H2 (9 strains), all of which were O43; H18 (6 strains); H30 (6 strains); and H28 (5 strains); strains of the last three H serogroups were all O148. Cumulatively, our results suggest a high degree of clonal diversity of disease-associated ETEC strains in this region. As a low percentage of these strains expressed a CF, it remains possible that other adhesins for which we either did not assay or that are as yet undiscovered are prevalent in this region. Our findings point out some potential barriers to effective immunization against ETEC diarrhea in this population and emphasize the need to identify additional protective antigens commonly expressed by ETEC for inclusion in future vaccine candidates.     

Plasmids coding for colonization factor antigens I and II, heat-labile enterotoxin, and heat-stable enterotoxin A2 in Escherichia coli.

Colonization factor antigens I and II (CFA/I and CFA/II) are important in the pathogenesis of diarrhea in humans caused by some enterotoxigenic Escherichia coli (ETEC). Plasmid DNA from 16 CFA/I+ and five CFA/II+ ETEC were examined by Southern blot analysis with enterotoxin gene probes and were compared with plasmid DNA from derivatives of the same ETEC that had lost the ability to produce these colonization factors. Among the 16 CFA/I+ ETEC strains, the loss of CFA/I was accompanied by the loss of a plasmid of between 34 and 68 megadaltons (MDa) coding for heat-stable enterotoxin A2 (ST-A2) in 12 strains, by the loss of a 60-MDa plasmid coding for heat-labile enterotoxin (LT) and ST-A2 in one strain, or by deletions of a segment of DNA encoding for ST-A2 in three strains. Among five CFA/II+ ETEC strains, the loss of CFA/II was associated with the loss of a plasmid of 75 MDa coding for LT and ST-A2 in three strains, with the loss of genes coding for LT and ST-A2 from a 68-MDa plasmid in one strain, or with no discernible loss of a plasmid or DNA sequences coding for enterotoxins in the remaining strain. The loss of CFA/I and CFA/II production was associated with the loss of DNA sequences encoding for ST-A2 in 20 of 21 ETEC examined.     

Phenotypic profiles of enterotoxigenic Escherichia coli associated with early childhood diarrhea in rural Egypt

Enterotoxigenic Escherichia coli (ETEC) causes substantial diarrheal morbidity and mortality in young children in countries with limited resources. We determined the phenotypic profiles of 915 ETEC diarrheal isolates derived from Egyptian children under 3 years of age who participated in a 3-year population-based study. For each strain, we ascertained enterotoxin and colonization factor (CF) expression, the O:H serotype, and antimicrobial susceptibility. Sixty-one percent of the strains expressed heat-stable enterotoxin (ST) only, 26% expressed heat-labile enterotoxin (LT) alone, and 12% expressed both toxins. The most common CF phenotypes were colonization factor antigen I (CFA/I) (10%), coli surface antigen 6 (CS6) (9%), CS14 (6%), and CS1 plus CS3 (4%). Fifty-nine percent of the strains did not express any of the 12 CFs included in our test panel. Resistance of ETEC strains to ampicillin (63%), trimethoprim-sulfamethoxazole (52%), and tetracycline (43%) was common, while resistance to quinolone antibiotics was rarely detected. As for the distribution of observed serotypes, there was an unusually wide diversity of O antigens and H types represented among the 915 ETEC strains. The most commonly recognized composite ETEC phenotypes were ST CS14 O78:H18 (4%), ST (or LTST) CFA/I O128:H12 (3%), ST CS1+CS3 O6:H16 (2%), and ST CFA/I O153:H45 (1.5%). Temporal plots of diarrheal episodes associated with ETEC strains bearing common composite phenotypes were consistent with discrete community outbreaks either within a single or over successive warm seasons. These data suggest that a proportion of the disease that is endemic to young children in rural Egypt represents the confluence of small epidemics by clonally related ETEC strains that are transiently introduced or that persist in a community reservoir.     

Does enteropathogenic Escherichia coli produce heat-labile enterotoxin, heat-stable enterotoxins a or b, or cholera toxin A subunits?

Although most enteropathogenic Escherichia coli strains do not produce recognized enterotoxins, we wished to examine whether they produce any factors like heat-stable enterotoxin b or cholera toxin active subunits that might be missed by conventional assay methods. E. coli strains E851 (O142) and E2348 (O127) that had caused diarrhea in volunteers were negative for heat-labile enterotoxin and heat-stable enterotoxin a in Chinese hamster ovary cell and suckling mouse assays, failed to cause secretion in ligated small bowel loops from 6- to 8-week-old pigs after 4 to 5 h (used to show heat-stable enterotoxin b), and did not activate adenylate cyclase in pigeon erythrocyte lysates (used to demonstrate cholera toxin A subunit). We conclude that crude, unconcentrated culture filtrates and sonicates do not mimic heat-labile or heat-stable enterotoxins or cholera toxin or its A subunit and that enteropathogenic strains of E. coli probably have yet another mechanism or group of mechanisms by which they cause diarrhea.     

Prevalence and association of the longus pilus structural gene (lngA) with colonization factor antigens, enterotoxin types, and serotypes of enterotoxigenic Escherichia coli.

Human enterotoxigenic Escherichia coli (ETEC) produces a plasmid-encoded type IV pilus termed longus (for long pilus). Regardless of the geographic origins of ETEC strains, the longus structural gene lngA was found to have the highest level of association with ETEC producing colonization factor antigen (CFA) CFA/II, followed by ETEC producing CFA/I and CFA/IV. ETEC bearing the less prevalent CFA/III and putative colonization factors and ETEC negative for CFA and putative colonization factor also contained lngA-related sequences. lngA was found in a considerable number of ETEC serotypes and was more often associated with ETEC producing heat-stable enterotoxins than with ETEC producing both heat-labile and heat-stable enterotoxins or heat-labile enterotoxin alone. lngA was found more often in strains isolated from children with diarrhea than in strains from healthy children, suggesting an association with intestinal disease. We conclude that longus is a widely distributed antigenic determinant in ETEC that is highly associated with known plasmid-encoded virulence factors, namely, CFAs and enterotoxins. A longus-specific probe may be a helpful epidemiological tool to assist in the identification of ETEC.     

New Surface-Associated Heat-Labile Colonization Factor Antigen (CFA/II) Produced by Enterotoxigenic Escherichia coli of Serogroups O6 and O8

Enterotoxigenic Escherichia coli (ETEC) belonging to serogroups O6 and O8 do not possess the H-10407-type colonization factor antigen (CFA/I). However, these frequently isolated ETEC were found to possess a second and distinct heat-labile surface-associated colonization factor antigen, termed CFA/II. Whereas CFA/I mediates mannose-resistant hemagglutination of human group A erythrocytes, CFA/II does not. CFA/II mediates mannose-resistant hemagglutination of bovine erythrocytes, and mannose-resistant hemagglutination is rapid only at reduced temperature (4 degrees C). Because CFA/II, like CFA/I, is spontaneously lost by many ETEC isolates in the laboratory, it was possible to produce specific anti-CFA/II serum by preparing antiserum against living cells of a prototype strain (PB-176) and adsorbing this serum with living and heat-treated cells of its CFA/II-negative derivative strain PB-176-P. This serum, which neutralized the colonization factor activity of CFA/II-positive strains in infant rabbits, was employed to confirm the presence of CFA/II on ETEC which exhibited mannose-resistant hemagglutination of bovine but not human erythrocytes. CFA/II, like CFA/I, mediates adherence of the bacteria to the mucosal surface of the small intestine, as demonstrated by indirect immunofluorescence. CFA/II appears to be an important virulence factor for humans since CFA/II-positive ETEC are frequently isolated from diarrhea cases, particularly travelers' diarrhea, in Mexico; these ETEC were not uncommon in a collection of isolates from Bangladesh. The O6:H16 strain of ETEC responsible for an outbreak of diarrhea in the United States was also shown to be CFA/II positive. CFA/I and CFA/II were never found on the same serotypes of ETEC, but 98% of the heat-stable and heat-labile enterotoxin-producing ETEC belonging to the frequently isolated serogroups O6, O8, O15, O25, O63, and O78 were positive for either CFA/I or CFA/II.