Clinical and microbiologic features of Shigella and enteroinvasive Escherichia coli infections detected by DNA hybridization.

To determine the clinical and microbiologic features of Shigella and enteroinvasive Escherichia coli (EIEC) infections, we investigated 410 children with diarrhea and 410 control children without diarrhea who were seen at Children's Hospital, Bangkok, Thailand, from January to June 1985. Shigella spp. were isolated from 96 (23%) and EIEC were isolated from 17 (4%) of 410 children with diarrhea and from 12 (3%) and 6 (1%) of 410 control children, respectively. The isolation rates of both pathogens increased with age and peaked in children 3 to 5 years old from whom Shigella spp. were isolated from 38% and EIEC were isolated from 9%. Shigella spp. were isolated from 52% and EIEC were isolated from 7% of 91 children with bloody diarrhea and from 15 and 3% of 319 children with nonbloody diarrhea. Fifteen (65%) of 23 EIEC were lactose positive, and all belonged to recognized EIEC serotypes. Among children with diarrhea, the stool blots of 76% of 17 children infected with EIEC, 45% of 96 children infected with Shigella spp., and 1% of 297 culture-negative children hybridized with the 17-kilobase EcoRI digestion fragment of pRM17, a recombinant plasmid containing DNA derived from the 140-megadalton Shigella flexneri plasmid. Although EIEC colonies can be reliably detected by DNA hybridization, detection by stool blot is less sensitive. Shigella spp. and EIEC are important causes of endemic diarrhea among children greater than 1 year old in Thailand.

HeLa cell-adherent enteropathogenic Escherichia coli in children under 1 year of age in Thailand.

Enteropathogenic Escherichia coli (EPEC) was isolated from 11% of 148 Hmong children under 1 year old with diarrhea at a refugee camp in northern Thailand. Of 16 children with EPEC-associated diarrhea, 11 were infected with EPEC that adhered to HeLa cells in a diffuse pattern, 3 were infected with EPEC that adhered to HeLa cells in a localized adherence (LA) pattern, and 2 were infected with EPEC that were nonadherent. In Bangkok, EPEC was isolated from 6% of 64 children under 1 year old with diarrhea and 7% of 56 children of the same age without diarrhea. Of four children with diarrhea, two were infected with EPEC with an LA pattern, and two were infected with nonadherent EPEC. Of four children without diarrhea, one was infected with EPEC with an LA pattern, one was infected with EPEC that adhered in a diffuse pattern, and two were infected with nonadherent EPEC. The 21 EPEC isolates with an LA pattern hybridized with the EPEC adherence factor DNA probe. EPEC was the only enteric pathogen identified in 16 (80%) of 20 children with EPEC-associated diarrhea. EPEC was as frequently isolated from children under 1 year old as were other bacterial enteric pathogens. The problem of identifying EPEC with pools of polyvalent antisera are described, and the need to identify additional enteropathogenic determinants of EPEC is discussed.

Trimethoprim-resistant Shigella and enterotoxigenic Escherichia coli strains in children in Thailand.

The percentage of Shigella and enterotoxigenic Escherichia coli (ETEC) strains resistant to trimethoprim (TMP)-sulfamethoxazole isolated from children with diarrhea at the outpatient department of the Children's Hospital in Bangkok increased from 3 and 0%, respectively, in 1982 to 29% and 25% in 1986. One hundred thirty-nine Shigella and 22 ETEC strains resistant to greater than 1024 micrograms/ml of trimethoprim (TMPr) isolated from children with diarrhea in Bangkok in 1984 and 1985 were analyzed for the presence of type I, II and III plasmid-specific dihydrofolate reductase (DHFR) genes. Thirty-two percent (45 of 139) of TMPR Shigella had genes encoding type II and 9% (13 of 139) had genes encoding type I DHFR genes. Fifty percent (11 of 22) of TMPR ETEC had type II and 14% (3 of 22) had type I DHFR genes. Plasmids encoding DHFR were identified by the Southern technique in 24% (14 of 58) of Shigella and 1 of 14 ETEC that contained genes encoding DHFR. Plasmids coding for type II DHFR were transferred to E. coli K12 by conjugation from 13 of 14 Shigella and a plasmid coding for type I DHFR was transferred from the single ETEC containing a plasmid coding for type I DHFR. Genes coding for DHFR were presumably situated on the chromosome in 76% (44 of 58) of Shigella and 93% (13 of 14) of ETEC that contained genes encoding DHFR. Since 58% (81 of 139) of TMPR Shigella and 36% (8 of 22) of TMPR ETEC strains examined did not contain genes encoding type I, II or III DHFR, high level TMP resistance was presumably caused by other types of DHFR genes.(ABSTRACT TRUNCATED AT 250 WORDS)

Potential sources of enterotoxigenic Escherichia coli in homes of children with diarrhoea in Thailand.

PIP: A year-long study was performed to identify potential sources of enterotoxigenic Escherichia coli (ETEC) within the homes of children with diarrhea in Bangkok. ETEC was identified in 8% (10 of 130) of the inhabitants of 42 homes with children with ETEC diarrhea and 6% (8 of 137) of their neighbors, but in only 2% (49 of 3077) of those individuals living in 866 homes not associated with children with ETEC diarrhea. While 46% (13 of 28) of the children under age 2 infected with ETEC were identified on home visits as having had a recent history of diarrhea, only 13% (5 of 39) of those over age 2 presented such a history. ETEC was isolated from 14% of the mothers' hands, 13% of the children's hands, and 7% of jars containing bath water that was used for washing the children after defecation. Drinking water was identified as a probable source of infection in 1 of 42 cases. Further studies are needed to determine whether ETEC from water stored in the home can spread and cause secondary infections.^ieng

Identification of enteric pathogens in the small and large intestine of children with diarrhea.

Enteric pathogens were identified in children with diarrhea from duodenal specimens obtained with a string capsule and from fecal specimens. Rotavirus was identified in stools of 43 of 100 children, and was recovered from the small intestine from nine (21%) children who were excreting this virus. Shigella was isolated from stools from 22, Salmonella from 17, enterotoxigenic Escherichia coli from eight, and Aeromonas hydrophila from one of 100 children with diarrhea. In contrast to rotavirus, Salmonella, Shigella, enterotoxigenic Escherichia coli, and A. hydrophila were not isolated from the small intestine. Nonenterotoxigenic Aeromonas species were recovered from the small intestine, but not the stool of five children. These children were also infected with Shigella or with rotavirus; this suggests that Aeromonas was not the cause of their diarrhea. None of 51 Escherichia coli isolated with the string capsules, or 67 isolated from stool that agglutinated in commercial enteropathogenic Escherichia coli antisera were of classical enteropathogenic Escherichia coli serotypes. One hundred and five of these 118 Escherichia coli did not hybridize with a deoxyribonucleic acid probe for plasmid mediated factors conferring adherence to HeLa cells. Examination of specimens collected with a string capsule from children with diarrhea did not identify any more enteric pathogens than examining stools. Furthermore testing Escherichia coli for agglutination in commercial enteropathogenic Escherichia coli antisera did not identify Escherichia coli of enteropathogenic serotypes.

A non-radioactive DNA probe to identify Shigella and enteroinvasive Escherichia coli in stools of children with diarrhoea.

A non-radioactive biotinylated DNA probe was constructed to detect Shigella and enteroinvasive Escherichia coli (EIEC). Specimens were examined with the biotinylated probe after removing streptavidin-binding glycoproteins with proteinase K. Both biotinylated and radioactive probes detected 125 pg of target-cell DNA after hybridisation for 24 h and exposure to indicator dyes or X-ray film for 4 h. Both probes hybridised with 52 EIEC and none of 16 non-EIEC examined; they also hybridised with stool blots from 11 of 13 children with culture-proven shigellosis or EIEC diarrhoea and were negative with stool blots from 43 children who were culture negative for Shigella and EIEC. Biotinylated DNA probes can be as sensitive as radiolabelled probes, but have the advantage of a longer shelf-life and greater availability.

Colonization factors associated with enterotoxigenic Escherichia coli isolated in Thailand.

Eighty-six percent (72 of 84) of heat-labile and heat-stable, none of 141 heat-labile, and 24% (27 of 111) heat-stable enterotoxigenic Escherichia coli isolates from Thailand aggregated in less than 1 M (NH4)2SO4, hemagglutinated human group A and bovine erythrocytes in 1% D-mannose, and possessed either colonization factor I or colonization factor II. No other colonization factors were identified by these two methods.

Identification by DNA hybridisation of enterotoxigenic Escherichia coli in homes of children with diarrhoea.

The DNA hybridisation technique to detect genes coding for Escherichia coli enterotoxin was used to identify enterotoxigenic E coli (ETEC) in homes of children with diarrhoea in Thailand. ETEC was found in 30 (14%) of 221 children with diarrhoea and in 9% (8/88) of their household contacts, 8% (8/101) of their neighbours, and 2% (32/1379) of inhabitants of 382 homes not associated with ETEC infections. ETEC was found significantly more often in water and food and on mothers' hands in homes of children with ETEC-associated diarrhoea and of their neighbours than in homes of children without ETEC infections (8/360 vs 3/2290; p less than 0.001). ETEC was identified in 80% (71/89) of specimens that hybridised with the enterotoxin gene probes by testing E coli isolated from the same specimen in the Y-1 adrenal and suckling-mouse assays. The DNA hybridisation assay to detect genes coding for E coli enterotoxin is an effective method of identifying ETEC in a large number of human and environmental specimens and will be a valuable tool to define further the epidemiology of this enteric pathogen.

Viruses and bacteria in pediatric diarrhea in Thailand: a study of multiple antibiotic-resistant enteric pathogens.

Children with diarrhea admitted to a rehydration ward of a children's hospital in Bangkok were investigated to determine the prevalence of enteric pathogens, the extent of children's previous antibiotic therapy, and the frequency of plasmid-mediated antibiotic resistance among infecting bacteria. Rotavirus (36%), enterotoxigenic Escherichia coli (18%), Shigella (9%), Salmonella (6%), Campylobacter jejuni/coli (4%), and Vibrio cholerae (2%) infections were among 105 children with diarrhea. Antibiotics were detected in 29% of urines collected from children on admission. All Shigella, 83% of enterotoxigenic E. coli, and 40% of Salmonella were resistant to more than one antibiotic. Sixty-two percent of 24 antibiotic-resistant enteric pathogens transferred R factors to E. coli K12 by conjugation. Four of four multiresistant E. coli that produced heat-labile and heat-stable toxin transferred resistance and the ability to produce heat-labile toxin. An analysis of plasmids by agarose gel electrophoresis indicated enterotoxigenicity and antibiotic resistance were associated with separate plasmids in transconjugants from these six matings. Antibiotics are used frequently in the treatment of pediatric diarrhea in Bangkok, which has undoubtedly contributed to the high frequency of plasmid-mediated resistance among enteric pathogens.