Teresa Estrada-Garcia

Single Multiplex Polymerase Chain Reaction To Detect Diverse Loci Associated with Diarrheagenic Escherichia coli

We developed and tested a single multiplex polymerase chain reaction (PCR) that detects enterotoxigenic, enteropathogenic, enteroinvasive, and Shiga-toxin–producing Escherichia coli. This PCR is specific, sensitive, and rapid in detecting target isolates in stool and food. Because of its simplicity, economy, and efficiency, this protocol warrants further evaluation in large, prospective studies of polymicrobial substances.

Multiplex PCR for Detection of Three Plasmid-Borne Genes of Enteroaggregative Escherichia coli Strains

ABSTRACT We developed a novel multiplex PCR assay for enteroaggregative Escherichia coli (EAEC) detection, by using three plasmid-borne genes (the aggregative adherence [AA] probe, aap, and aggR). One or more of the loci were detected in 24 (86%) of 28 patient isolates analyzed. The multiplex PCR assay is a fast, convenient, and sensitive molecular test to detect EAEC.

Association of diarrheagenic Escherichia coli pathotypes with infection and diarrhea among Mexican children and association of atypical enteropathogenic E. coli with acute diarrhea.

ABSTRACT Seventy-six children ≤2 years old were prospectively followed for 1 year in a peri-urban community of Mexico City to determine asymptomatic infection and acute diarrhea associated with diarrheagenic Escherichia coli pathotypes (DEPs). By use of a pathogen-specific multiplex PCR, DEPs were sought in 795 stool samples, of which 125 (16%) were positive for DEP; of these, 4 represented shedding episodes and 4 parasite coinfections. Most single-DEP infections (85/117) were asymptomatic (P < 0.001), and of the 32 DEP diarrhea episodes, 41% were associated with atypical enteropathogenic E. coli (aEPEC), 37.5% with enterotoxigenic E. coli, 9% with typical EPEC, 9% with enteroinvasive E. coli, and 3% with Shiga toxin-producing E. coli strains. Among the 76 children, 54 had at least one stool positive for DEP, of which 23 experienced a DEP-associated diarrhea episode. In the last group of children, DEP infection was significantly associated with a diarrhea episode (relative risk [RR] = 2.5; 95% confidence interval [CI], 1.79 to 3.57; P < 0.001), with ETEC (RR = 2.30; 95% CI, 1.49 to 3.54; P = 0.003) and aEPEC (RR = 1.92; 95% CI, 1.23 to 3.0; P = 0.019) being the pathotypes associated with diarrhea. aEPEC-associated diarrhea episodes were frequently in the <12-month age group (RR = 2.57; 95% CI, 1.05 to 6.27; P = 0.04). aEPEC infections were distributed all year round, but associated diarrheal episodes were identified from April to October, with a May-June peak (rainy season). Most ETEC infections and diarrhea episodes characteristically occurred during the summer (rainy season), with a diarrhea peak in August. Of all DEPs, only aEPEC was associated with acute diarrhea episodes lasting 7 to 12 days (P = 0.019). DEPs are important causes of community-acquired enteric infection and diarrhea in Mexican children.

Drug-resistant Diarrheogenic Escherichia coli, Mexico

Diarrheogenic Escherichia coli isolates from 45 (73%) of 62 hospitalized patients were resistant to common antimicrobial drugs. Sixty-two percent were multidrug resistant, and >70% were resistant to trimethoprim-sulfamethoxazole and ampicillin. Ciprofloxacin and cefotaxime were uniformly active. Effective and safe oral agents are needed to treat children with bacterial diarrhea.

Associations between Mucosal Innate and Adaptive Immune Responses and Resolution of Diarrheal Pathogen Infections

ABSTRACT The identification of immune response mechanisms that contribute to the control of diarrheal disease in developing countries remains an important priority. We addressed the role of fecal chemokines and cytokines in the resolution of diarrheal Escherichia coli and Giardia lamblia infections. Stools collected from 127 Mexican children 5 to 15 months of age enrolled in a randomized, double-blind, placebo-controlled, vitamin A supplementation trial were screened for enteropathogenic Escherichia coli (EPEC), enterotoxigenic E. coli (ETEC), and Giardia lamblia. Fecal concentrations of tumor necrosis factor alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1), interleukin-4 (IL-4), IL-5, IL-6, IL-8, IL-10, and interferon-γ (IFN-γ) were determined. Hazard models incorporating cytokine variables were fit to durations of asymptomatic and symptomatic pathogen infections, controlling for treatment group. Increased levels of TNF-α and IL-6 were associated with decreased durations of EPEC infection and increased ETEC durations. Increased IL-4 and IFN-γ levels were associated with decreased and increased durations, respectively, of both EPEC and ETEC infections. Increased IL-10 levels were associated with increased and decreased durations of asymptomatic and symptomatic EPEC infections, respectively, and increased durations of both asymptomatic and symptomatic ETEC infections. Increased levels of MCP-1, IFN-γ, IL-4, and IL-5 were associated with increased G. lamblia infection duration, while increased IL-8 levels were associated with decreased durations. Differences in proinflammatory and Treg cytokine levels are associated with differences in the resolution of inflammatory and noninflammatory pathogen infections.

Impact of Vitamin A on Selected Gastrointestinal Pathogen Infections and Associated Diarrheal Episodes among Children in Mexico City, Mexico

BACKGROUND The overall effect of vitamin A supplementation on diarrheal disease in community trials may result from its effect on specific diarrheal pathogens. METHODS We conducted a placebo-controlled, double-blind trial of the prophylactic effect of vitamin A on gastrointestinal pathogen infections and clinical symptoms among 188 children in Mexico City, Mexico, from January 1998 to May 1999. Children 6-15 months of age were randomly assigned to receive either a vitamin A supplement (for children <12 months of age, 20,000 international units [IU] of retinol; for children > or =12 months of age, 45,000 IU of retinol) every 2 months or a placebo and were followed for up to 15 months. Stool samples, collected semimonthly, were screened for enteropathogenic Escherichia coli (EPEC), enterotoxigenic E. coli (ETEC), Shiga toxin-producing E. coli (STEC), enteroinvasive E. coli (EIEC), and Giardia lamblia. RESULTS Vitamin A supplementation reduced the prevalence of EPEC infections (rate ratio [RR], 0.52 [95% confidence interval {CI}, 0.23-0.86]) and led to shorter durations of both EPEC and ETEC infections. Supplementation also reduced the prevalence of EPEC-associated diarrhea (RR, 0.41 [95% CI, 0.16-1.00]), EPEC-associated fever (RR, 0.15 [95% CI, 0.02-0.98]), and G. lamblia-associated fever (RR, 0.27 [95% CI, 0.13-0.80]). Finally, children who received vitamin A supplementation had shorter durations of EPEC-associated diarrhea than did children who did not receive supplementation but had longer durations of G. lamblia-associated diarrhea. CONCLUSIONS These results suggest that the effect of vitamin A supplementation on clinical outcomes may be pathogen dependent.

Faecal contamination and enterotoxigenic Escherichia coli in street-vended chili sauces in Mexico and its public health relevance.

The street-vended food industry provides employment and cheap ready-to-eat meals to a large proportion of the population in developing countries like Mexico, yet little is known about its role in the transmission of food borne diseases (FBD). Because of its wide consumption, street-vended chili sauces in Mexico are potential vehicles of FBD. An observational study was performed in Mexico City collecting 43 street-vended chili sauces. These sauces were prepared under poor hygienic conditions of handling and selling. Consumers add 4-8 ml of chili sauce per taco, ingest 2-5 tacos per meal and on average, 50 consumers frequent a stall per day. Seventeen (40%) samples were faecally contaminated and 2(5%) sauces harboured sufficient enterotoxigenic Escherichia coli to cause disease. Weestimate that the consumption of only one of these chili sauces could result in ETEC disease inat least 21,000 consumers per year, making them important potential vehicles of FBD.

Prevalence of Escherichia coli and Salmonella spp. in street-vended food of open markets ( tianguis ) and general hygienic and trading practices in Mexico City

Street-vendors in Mexico City provide ready-to-eat food to a high proportion of the inhabitants. Nevertheless, their microbiological status, general hygienic and trading practices are not well known. During spring and summer 2000, five tianguis (open markets) were visited and 48 vendors in 48 stalls interviewed. A total of 103 taco dressings were sampled for E. coli and Salmonella spp.: 44 (43%) contained E. coli and 5 (5%) Salmonella (2 S. Enteritidis phage type 8, 1 S. Agona, 2 S. B group). Both E. coli and salmonellas were isolated from three samples. Of Salmonella-positive stalls 80% (4/5) had three or more food-vendors and 80% of vendors were males, compared with 37.3% (16/43) and 46.4% (20/43) in the Salmonella-negative stalls respectively. Food-vendors kept water in buckets (reusing it all day), lacked toilet facilities, and prepared taco dressings the day before which remained at the tianguis without protection for 7.8 h on average. Consumption of street-vended food by local and tourist populations poses a health risk.

Vitamin A supplementation modifies the association between mucosal innate and adaptive immune responses and resolution of enteric pathogen infections

BACKGROUND The efficacy of vitamin A supplementation on diarrheal disease morbidity may reflect the divergent effects that supplementation has on pathogen-specific immune responses and pathogen-specific outcomes. OBJECTIVE We examined how vitamin A supplementation modified associations between gut-cytokine immune responses and the resolution of different diarrheal pathogen infections. DESIGN Stools collected from 127 Mexican children who were 5-15 mo old and enrolled in a randomized, placebo-controlled vitamin A supplementation trial were screened for enteropathogenic Escherichia coli (EPEC), enterotoxigenic E. coli (ETEC), and Giardia lamblia. Fecal concentrations of interleukin (IL)-6, IL-8, IL-4, IL-5, IL-10, monocyte chemoattractant protein 1 (MCP-1), tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ) were measured by using an enzyme-linked immunosorbent assay. Hazard models that incorporated categorized cytokine variables (ie, nondetectable, less than the median of detectable concentrations, and at least the median of detectable concentrations) were fit to the length of pathogen infections stratified by treatment group. RESULTS Vitamin A-supplemented children with fecal MCP-1 or IL-8 concentrations less than the median of detectable concentrations and IL-10 concentrations of at least median concentrations had longer durations of EPEC infection than did children in the placebo group. In supplemented children, detectable fecal TNF-α or IL-6 concentrations were associated with shorter ETEC infection durations, whereas MCP-1 concentrations of at least the median were associated with longer infection durations. Children in this group who had IL-4, IL-5, or IFN-γ concentrations of at least median detectable concentrations had shorter durations of G. lamblia infection. CONCLUSION The effect of supplementation on associations between fecal cytokine concentrations and pathogen infection resolution depends on the role of inflammatory immune responses in resolving specific pathogen infections.

Diarrheagenic Escherichia coli Carrying Supplementary Virulence Genes Are an Important Cause of Moderate to Severe Diarrhoeal Disease in Mexico

Diarrheagenic Escherichia coli (DEC) cause acute and persistent diarrhoea worldwide, but little is known about their epidemiology in Mexico. We determined the prevalence of bacterial enteropathogens in 831 children with acute diarrhoea over a four-year period in Yucatan, Mexico. Six DEC supplementary virulence genes (SVG), mainly associated with enteroaggregative E. coli (EAEC), were sought in 3100 E. coli isolates. DEC was the most common bacterial enteropathogen (28%), surpassing Salmonella (12%) and Shigella (9%). Predominant DEC groups were diffusely adherent E. coli (DAEC) (35%), EAEC (24%), and enteropathogenic E. coli (EPEC) (19%). Among children with DEC infections, 14% had severe illness mainly caused by EPEC (26%) and DAEC (18%); 30% had moderate diarrhoea mainly caused by DAEC (36%), mixed DEC infections (33%) and EAEC (32%). DAEC was most prevalent during spring, while ETEC, EAEC and EPEC predominated in summer. EAEC was more frequent in children 6–24 months old than in those younger than 6 months of age (P = 0.008, OR = 4.2, 95% CI, 1.3–13.9). The presence of SVG dispersin, (aatA), dispersin-translocator (aatA), enteroaggregative heat-stable toxin 1 (astA), plasmid encoded toxin (pet), cytolethal distending toxin (cdt) was higher in DEC than non-DEC strains, (36% vs 26%, P <0.0001, OR = 1.5, 95% CI, 1.3–1.8). 98% of EAEC-infected children harboured strains with SVG; 85% carried the aap-aatA gene combination, and 33% of these also carried astA. 28% of both EPEC and ETEC, and 6% of DAEC patients had strains with SVG. 54% of EPEC patients carried pet-positive strains alone or in combination with astA; only this DEC group harboured cdt-positive isolates. All ETEC patients carried astA- or astA-aap-positive strains. astA and aap were the most common SVG in DAEC (3% and 2%) and non-DEC strains (21% and 13%). DEC carrying SVG are an important cause of moderate to severe bacterial diarrhoea in Mexican children.