Genetic diversity of locus of enterocyte effacement genes of enteropathogenic Escherichia coli isolated from Peruvian children.

The aim of this study was to determine the frequency and allele associations of locus of enterocyte effacement encoded esp and tir genes among 181 enteropathogenic Escherichia coli (EPEC) strains (90 diarrhoea-associated and 91 controls) isolated from Peruvian children under 18 months of age. We analysed espA, espB, espD and tir alleles by PCR-RFLP. EPEC strains were isolated with higher frequency from healthy controls (91/424, 21.7 %) than from diarrhoeal samples (90/936, 9.6 %) (P,0.001); 28.9% of diarrhoeal and 17.6% of control samples were typical EPEC (tEPEC). The distribution of espA alleles (alpha, beta, beta2 and gamma) and espD alleles (alpha, beta, gamma and a new variant, espD-N1) between tEPEC and atypical EPEC (aEPEC) was significantly different (P,0.05). espD-alpha was more common among acute episodes (P,0.05). espB typing resulted in five alleles (alpha, beta, gamma and two new suballeles, espB-alpha2 and espB-alpha3), while tir-beta and tir-gamma2 were the most common intimin receptor subtypes. Seventy-two combinations of espA, espB, espD and tir alleles were found; the most prevalent combination was espA-beta, espB-beta, espD-beta, tir-beta (34/181 strains), which was more frequent among tEPEC strains (P,0.05). Our findings indicate that there is a high degree of heterogeneity among EPEC strains isolated from Peruvian children and that aEPEC and tEPEC variants cluster.
Authors: Contreras, C. A.; Ochoa, T. J.; Ruiz, J.; Lacher, D. W.; Durand, D.; DebRoy, C.; Lanata, C. F.; Cleary, T. G.
Source: J Med Microbiol

URL: http://hdl.handle.net/10757/314308

Genotypic and Phenotypic Characterization of Enterotoxigenic Escherichia coli Strains Isolated from Peruvian Children

Enterotoxigenic Escherichia coli (ETEC) is a major cause of childhood diarrhea. The present study sought to determine the prevalence and distribution of toxin types, colonization factors (CFs), and antimicrobial susceptibility of ETEC strains isolated from Peruvian children. We analyzed ETEC strains isolated from Peruvian children between 2 and 24 months of age in a passive surveillance study. Five E. coli colonies per patient were studied by multiplex real-time PCR to identify ETEC virulence factors. ETEC-associated toxins were confirmed using a GM1-based enzyme-linked immunosorbent assay. Confirmed strains were tested for CFs by dot blot assay using 21 monoclonal antibodies. We analyzed 1,129 samples from children with diarrhea and 744 control children and found ETEC in 5.3% and 4.3%, respectively. ETEC was more frequently isolated from children >12 months of age than from children <12 months of age (P < 0.001). Fifty-two percent of ETEC isolates from children with diarrhea and 72% of isolates from controls were heat-labile enterotoxin (LT) positive and heat-stable enterotoxin (ST) negative; 25% and 19%, respectively, were LT negative and ST positive; and 23% and 9%, respectively, were LT positive and ST positive. CFs were identified in 64% of diarrheal samples and 37% of control samples (P < 0.05). The most common CFs were CS6 (14% and 7%, respectively), CS12 (12% and 4%, respectively), and CS1 (9% and 4%, respectively). ST-producing ETEC strains caused more severe diarrhea than non-ST-producing ETEC strains. The strains were most frequently resistant to ampicillin (71%) and co-trimoxazole (61%). ETEC was thus found to be more prevalent in older infants. LT was the most common toxin type; 64% of strains had an identified CF. These data are relevant in estimating the burden of disease due to ETEC and the potential coverage of children in Peru by investigational vaccines.
Authors: Rivera, F. P.; Ochoa, T. J.; Maves, R. C.; Bernal, M.; Medina, A. M.; Meza, R.; Barletta, F.; Mercado, E.; Ecker, L.;Gil, A. I.; Hall, E. R.; Huicho, L.; Lanata, C. F.
Source: J. Clin. Microbiol

URL: http://hdl.handle.net/10757/314292

Global Causes of Diarrheal Disease Mortality in Children more 5 Years of Age: A Systematic Review.

Estimation of pathogen-specific causes of child diarrhea deaths is needed to guide vaccine development and other prevention strategies. We did a systematic review of articles published between 1990 and 2011 reporting at least one of 13 pathogens in children ,5 years of age hospitalized with diarrhea. We included 2011 rotavirus data from the Rotavirus Surveillance Network coordinated by WHO. We excluded studies conducted during diarrhea outbreaks that did not discriminate between inpatient and outpatient cases, reporting nosocomial infections, those conducted in special populations, not done with adequate methods, and rotavirus studies in countries where the rotavirus vaccine was used. Age-adjusted median proportions for each pathogen were calculated and applied to 712 000 deaths due to diarrhea in children under 5 years for 2011, assuming that those observed among children hospitalized for diarrhea represent those causing child diarrhea deaths. 163 articles and WHO studies done in 31 countries were selected representing 286 inpatient studies. Studies seeking only one pathogen found higher proportions for some pathogens than studies seeking multiple pathogens (e.g. 39% rotavirus in 180 single-pathogen studies vs. 20% in 24 studies with 5–13 pathogens, p,0?0001). The percentage of episodes for which no pathogen could be identified was estimated to be 34%; the total of all age-adjusted percentages for pathogens and no-pathogen cases was 138%. Adjusting all proportions, including unknowns, to add to 100%, we estimated that rotavirus caused 197 000 [Uncertainty range (UR) 110 000–295 000], enteropathogenic E. coli 79 000 (UR 31 000–146 000), calicivirus 71 000 (UR 39 000–113 000), and enterotoxigenic E. coli 42 000 (UR 20 000–76 000) deaths. Rotavirus, calicivirus, enteropathogenic and enterotoxigenic E. coli cause more than half of all diarrheal deaths in children ,5 years in the world.
Authors: Claudio F. Lanata; Christa L. Fischer-Walker; Ana C. Olascoaga; Carla X. Torres; Martin J. Aryee; Robert E. Black
Source: PLoS ONE

URL: http://hdl.handle.net/10757/314287

High frequency of antimicrobial drug resistance of diarrheagenic Escherichia coli in infants in Peru.

 In a prospective passive diarrhea surveillance cohort study of 1,034 infants of low socioeconomic communities in Lima, Peru, we determined the prevalence and antimicrobial drug susceptibility of the diarrheagenic Escherichia coli . The prevalence of diarrheagenic E. coli was 29% (161 of 557) in children with gastroenteritis and 30% (58 of 195) in the control group without diarrhea. The most common E. coli pathogens in diarrhea were enteroaggregative E. coli (EAEC) (14%), enteropathogenic E. coli (EPEC) (7%), diffusely adherent E. coli (DAEC) (4%), and enterotoxigenic E. coli (ETEC) (4%). Diarrheagenic E. coli as a group exhibited high levels of antimicrobial drug resistance in diarrheal cases to ampicillin (85%), cotrimoxazole (79%), tetracycline (65%), and nalidixic acid (28%). Among individual E. coli groups in patients with diarrhea, DAEC and EAEC exhibited significant higher frequencies of resistance to ampicillin, cotrimoxazole, tetracycline and nalidixic acid than EPEC and ETEC. Antimicrobial drug resistance to ampicillin and cotrimoxazole were more frequent in E. coli isolated from diarrheal samples than controls, which reflected greater antibiotic exposure in patients with gastroenteritis.
Authors: Ochoa, Theresa J.; Ruiz, Joaquím; Molina, Margarita; Del Valle, Luis J.; Vargas, Martha; Gil, Ana I.; Ecker, Lucie;Barletta, Francesca; Hall, Eric; Cleary, Thomas G.; Lanata, Claudio F.
Source: Am. J. Trop. Med. Hyg

URL: http://hdl.handle.net/10757/314286

Fecal Leukocytes in Children Infected with Diarrheagenic Escherichia col

The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response.
Authors: Mercado, Erik H.; Ochoa, Theresa J.; Ecker, Lucie; Cabello, Martin; Durand, David; Barletta, Francesca; Molina, Margarita; Gil, Ana I.; Huicho, Luis; Lanata, Claudio F.; Cleary, Thomas G.
Source: JOURNAL OF CLINICAL MICROBIOLOGY
URL: http://hdl.handle.net/10757/314005