Colonization of the oropharynx with Gram-negative bacilli in children with severe protein-calorie malnutrition.

Oral pharyngeal isolation of Gram-negative bacteria was compared in four groups of Bengali children; acutely ill, severely malnourished outpatients swabbed on hospital admission; ill but less severely malnourished outpatients from the same area as the malnourished children; orphans also less severely malnourished but not acutely ill; and well controls drawn from a priviledged socioeconomic group. The expected weight for height percentage (National Center Health Statistics/Center for Disease Control median) of the four groups was respectively 67, 91, 97, and 97%. Isolation of Gram-negative bacteria from 74 of 87 (85%) severely malnourished children was significantly greater (p less than 0.01) compared to 43 of 113 (38%) outpatients, to 20 of 93 (22%) orphans, and to five of 51 (10%) controls. A total of 71 malnourished children under 5 yr of age (90%) had higher rates of Gram-negative throat colonization than did 16 older children (63%) (p less than 0.01). Thus there was an increased rate of Gram-negative colonization in severely malnourished children especially among the younger age group. In the subset of ill children, Gram-negative pharyngeal colonization was significantly associated inversely with nutritional indices and age. The high rate of such carriage may be partly responsible for the increased susceptibility of Gram-negative infection demonstrated in these children.

Simultaneous infection with multiple serotypes of Shigellae in a patient.

We isolated three different serotypes of Shigella on admission from a patient with dysentery as well as a Shigella-like organism and Campylobacter jejuni upon follow-up. The patient produced serum antibodies to all three serotypes of shigellae.

Moraxella septicemia in children with diarrheal disease.

We report 10 cases of Moraxella septicemia associated with diarrheal disease. Their clinical presentations and outcomes are discussed. Recognition of the pathogenicity of these microorganisms in appropriate clinical setting should result in prompt and specific therapy.

Characterisation of Plesiomonas shigelloides strains that share type-specific antigen with Shigella flexneri 6 and common group 1 antigen with Shigella flexneri spp. and Shigella dysenteriae 1.

Three strains of Plesiomonas shigelloides isolated from patients with diarrhoea were agglutinated with Shigella flexneri 6 antiserum in slide and tube tests. All the strains were also agglutinated with a monoclonal antibody to the common group 1 antigen shared between S. flexneri serotypes and S. dysenteriae type 1. Further studies with one strain also showed sharing of antigenicity in an enzyme-linked immunosorbent assay. The results suggest that the strains share type-specific antigen with S. flexneri 6 and the common group 1 antigen with S. flexneri serotypes and S. dysenteriae 1. The sharing of antigens may have implications for cross-protection. One strain adhered to HEp-2 cell monolayers. None of the strains contained high mol. wt plasmids and there was no sequence homology with the invasiveness plasmid of Shigella spp. in DNA probe hybridisation. They were susceptible to the commonly used antibiotics. However, they possessed four other virulence-associated properties of Shigella spp. that included Congo-red binding, hydrophobicity, toxicity to HeLa cells and HEp-2 cell invasiveness (although they gave negative results in the Sereny test for invasiveness). These data suggest that the three unique strains might be considered pathogenic. Studies in animal models and human volunteers would be necessary to establish their pathogenic potential.

A simple assay to detect Escherichia coli producing heat labile enterotoxin: results of a field study of the Biken tests in Bangladesh.

In the Biken test a precipitin line forms in the agar between an Escherichia coli colony producing heat-labile enterotoxin (LT) and anti-cholera-toxin or anti-LT sera placed in an adjacent well. E. Coli isolates form 273 unselected patients with diarrhoea and 516 stock cultures of known toxin type were screened blindly for LT production by means of the Biken tests and the standard Chinese hamster ovary (CHO) cell assay. The Biken tests was 98% sensitive and 99% specific. All results of the Biken test which did not accord with the CHO cell assay were attributable to mixed stocks of LT-producing and non-LT-producing E. coli strains. The Biken test performed on agar plates with commercially available reagents offers clinical laboratories in developed and developing countries a simple, reliable, and inexpensive method of diagnosing diarrhoea caused by LT-producing strains of E. coli.

Isolation of Shigella dysenteriae serotypes 11, 12, and 13 from patients with diarrhea in Bangladesh.

Nine isolates of bacteria biochemically resembling Shigella dysenteriae but not belonging to the 10 recognized serotypes were isolated from patients with diarrhea in Bangladesh. Further studies suggested that two, one, and six isolates belonged to the recently recognized S. dysenteriae serotypes 11, 12, and 13, respectively.

First isolation of Yersinia intermedia from stool during an epidemiological investigation in Bangladesh.

Following the first isolation of Yersinia intermedia (Y. intermedia) in the Microbiology Branch of ICDDR, B from two postmortem cases, one of whom suffered from pneumonia and septicaemia and the other from enteric fever, an epidemiological investigation was carried out in the communities from where these postmortem cases had come. The stools from both postmortem cases were negative for Yersinia. One hundred and twenty five contacts of the first postmortem case and 135 contacts of the second postmortem case along with 175 animals of Dhaka Zoo were examined. Y. intermedia was isolated from only a five-year-old healthy contact of the first postmortem case. This was the first isolation of Y. intermedia from stool of a healthy child in Bangladesh.

Isolation of sucrose late-fermenting and nonfermenting variants of Vibrio cholerae O139 Bengal: implications for diagnosis of cholera.

The sucrose-containing selective medium thiosulfate-citrate-bile salt-sucrose agar missed a sucrose nonfermenting and four sucrose late-fermenting variant strains of Vibrio cholerae O139 Bengal from diarrheal stools. These strains were, however, correctly identified as V. cholerae O139 on a sucrose-deficient selective medium, taurocholate-tellurite-gelatin agar.

Detection of provisional serovars of Shigella dysenteriae and designation as S. dysenteriae serotypes 14 and 15.

In initial studies of slide and tube agglutination tests of shigella-like isolates from diarrheal stools from Bangladesh, 10 and 8 strains of Shigella dysenteriae were identified as provisional serovars (serovars that have been defined but not given a number in the antigenic schema) E22383 and E23507, respectively. Further screening of diarrheal stool isolates in a slide agglutination test with antisera (made with the international reference strains) to the two provisional serovars of S. dysenteriae identified an additional 36 isolates as belonging to E22383 and 8 isolates as belonging to E23507. All strains had properties typical of invasive Shigella strains in that they were positive in the Sereny test and had the 120- to 140-MDa plasmid associated with invasiveness. On the basis of these markers of pathogenicity and isolation from various geographical locations, now we recommend that provisional serovars E22383 and E23507 be designated S. dysenteriae serotypes 14 and 15, respectively.

An outbreak of dysentery caused by Shigella dysenteriae type 1 on a coral island in the Bay of Bengal.

An epidemic of severe dysentery occurred on St. Martin Island in the Bay of Bengal at a time when the island, with a population of 1,318, was isolated from the mainland because of the monsoon season. There were 434 cases and 28 deaths within a three-month period. Mortality (2%) was limited to the very young and the elderly. Shigella dysenteriae type 1 (Shiga bacillus) was isolated from 12 of 65 rectal swab specimens. All strains gave similar biochemical reactions and were resistant to tetracycline, chloramphenicol, streptomycin, and the sulfonamides but were sensitive to ampicillin, kanamycin, and gentamicin. No other organism was implicated in the outbreak. The disease disappeared from the island after institution of chlorination of the sources of drinking water and effective treatment of patients.

Use of colony pools for diagnosis of enterotoxigenic Escherichia coli diarrhea.

Diagnosis of enterotoxigenic Escherichia coli diarrhea was made in 109 adult males with an acute dehydrating cholera-like syndrome in Dacca, Bangladesh, by testing 10 colonies isolated from admission stool specimens for production of heat-labile and heat-stable toxins. Toxin testing of one colony yielded a diagnosis in 92% of the cases, testing of two colonies yielded a diagnosis in 95% of the cases, testing of a pool of 5 colonies yielded a diagnosis in 95% of the cases, and testing of a pool of 10 colonies yielded a diagnosis in 96% of the cases. From stool cultures obtained on subsequent days, toxin testing of individual colonies and pools revealed diminished efficacy of pooling with decreasing numbers of enterotoxin-positive isolates in the pool. To detect the presence of enterotoxigenic E. coli in stools, toxin testing of 5 individual isolates and a pool of 10 colonies was found to be almost as effective as the testing of 10 individual isolates.

Campylobacter jejuni bacteraemia in children with diarrhoea in Bangladesh: report of six cases.

Campylobacter jejuni was isolated from blood cultures from 6 of 6,275 diarrhoeal children seeking treatment at the Clinical Research Centre of the International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR,B) between April 1989 and December 1990. The clinical records of these 6 children were reviewed. All children were male; 5 were less than 1-year old and were severely malnourished. Five patients presented with watery diarrhoea and one with bloody diarrhoea. Two children died in the hospital. All strains of Campylobacter isolated from the 6 children were negative for cell invasive properties as tested by the Sereny test. Of the two strains tested for serum bactericidal activity, one strain was serum sensitive (growth inhibition), and the other resistant. The ability of C. jejuni to cause bacteraemia suggests that the organisms may be responsible for diarrhoeal diseases especially in young and malnourished children. An early attempt to detect Campylobacter and start effective antimicrobial therapy is indicated.

Epidemiologic and clinical features of endemic Campylobacter jejuni infection in Bangladesh.

Epidemiologic and clinical features of infection with Campylobacter jejuni in Bangladesh were examined in (1) diarrheal patients infected with C jejuni, (2) healthy control subjects, and (3) village children who were cultured monthly and at each diarrheal episode during a 10-month period. C jejuni was isolated from 437 (14%) of 3,038 outpatients with diarrhea. These patients had no distinct clinical presentation and were more likely to have a mixed infection than were patients infected with other pathogens (59% vs 42%, P less than 0.01). Age-specific infection rates were greatest in infants and did not differ significantly from those in control subjects. C jejuni was isolated less frequently from village children with diarrhea than from those cultured routinely (5% vs 9%, P less than 0.05). Forty percent of 47 patients with C jejuni vs 23% of 48 control subjects (P less than 0.01) had an elevated convalescent-phase antibody titer as determined by complement fixation test. In Bangladesh, enteric infection with C jejuni is common but often asymptomatic, although pathogenicity is suggested by serologic response in some patients.

Phage specific for Vibrio cholerae O139 Bengal.

From the stool of a Vibrio cholerae O139 Bengal-infected patient, a phage that specifically lysed capsulated V. cholerae O139 strains only was isolated. The phage is useful for the confirmatory diagnosis of V. cholerae O139 infection and for the differentiation of variants that lack the capsule.

Outbreak of keratoconjunctivitis due to Salmonella weltevreden in a guinea pig colony.

The purpose of this report is to demonstrate that the ability to produce keratoconjunctivitis (KC) is a property found in Salmonella weltevreden. This observation is contrary to previous reports that Salmonella spp. do not produce KC. An outbreak of KC due to S. weltevreden occurred in a guinea pig colony, and the animals carried the organism in the intestinal tract. The same Salmonella serotype that caused an epidemic of diarrhea in humans and a routine laboratory isolate also possessed the ability to induce KC. Unlike Shigella spp. (the prototype organisms positive for KC), S. weltevreden induced KC and bound Congo red dye even when grown at 30 degrees C. It invaded HeLa cells in culture but did not hybridize with a DNA probe for invasiveness of Shigella spp. and enteroinvasive Escherichia coli even though it harbored plasmids. It was susceptible to all the antibiotics tested, was hydrophobic, and showed mannose-sensitive hemagglutination. It did not have enterotoxic or cytotoxic activities.

Hafnia alvei, a probable cause of diarrhea in humans.

Hafnia alvei, a member of the family Enterobacteriaceae, was the only species of bacteria cultured from the stool of a 9-month-old child who was admitted with a 3-day history of watery diarrhea. The isolated strain of H. alvei failed to produce heat-labile or heat-stable enterotoxins or Shiga-like toxin I or II and did not invade HeLa cells, nor did it cause keratoconjunctivitis (determined by the Sereny test) in a guinea pig's eye. The strain, however, induced diarrhea in 8 of 12 adult rabbits with removable intestinal ties (removable intestinal tie-adult rabbit diarrhea [RITARD] assay) and in 1 of 2 orally fed animals. No diarrhea could be induced with Escherichia coli K-12 in eight RITARD assay rabbits and three orally fed rabbits, respectively. Microscopic examination of affected animals revealed moderate inflammatory cellular infiltration of the intestinal mucosa, in which bacterial attachment to the surface epithelium and loss of the microvillus border were evident in the ileum and colon. Electron microscopy demonstrated cellular modifications of the apical surface, with cupping or pedestal formation and increased terminal web density at sites of bacterial "attachment-effacement," a well-known characteristic and mechanism of diarrhea of enteropathogenic E. coli. Identical lesions were also induced by H. alvei in rabbit ileal loops, which ruled out naturally occurring rabbit enteropathogenic E. coli strains, which are known to produce similar lesions. It is concluded that at least some strains of H. alvei have the potential to cause diarrhea and that attachment-effacement is a virulence characteristic shared by bacteria other than E. coli.

Characterization of Aeromonas caviae antigens which cross-react with Shigella boydii 5.

Live and boiled cells of 16 strains of Aeromonas caviae, isolated from patients with diarrhea, agglutinated with Shigella boydii 5 antiserum in a slide test. Further studies with seven selected strains showed agglutination with boiled cells in a tube test. Lipopolysaccharide antigen extracted from one of these strains cross-reacted with S. boydii 5 in enzyme-linked immunosorbent assay and immunoblot studies. Either all or the majority of the seven strains possessed properties deemed to be diarrheagenic.