Epidemiology, genetics, and ecology of toxigenic Vibrio cholerae.

Cholera caused by toxigenic Vibrio cholerae is a major public health problem confronting developing countries, where outbreaks occur in a regular seasonal pattern and are particularly associated with poverty and poor sanitation. The disease is characterized by a devastating watery diarrhea which leads to rapid dehydration, and death occurs in 50 to 70% of untreated patients. Cholera is a waterborne disease, and the importance of water ecology is suggested by the close association of V. cholerae with surface water and the population interacting with the water. Cholera toxin (CT), which is responsible for the profuse diarrhea, is encoded by a lysogenic bacteriophage designated CTXPhi. Although the mechanism by which CT causes diarrhea is known, it is not clear why V. cholerae should infect and elaborate the lethal toxin in the host. Molecular epidemiological surveillance has revealed clonal diversity among toxigenic V. cholerae strains and a continual emergence of new epidemic clones. In view of lysogenic conversion by CTXPhi as a possible mechanism of origination of new toxigenic clones of V. cholerae, it appears that the continual emergence of new toxigenic strains and their selective enrichment during cholera outbreaks constitute an essential component of the natural ecosystem for the evolution of epidemic V. cholerae strains and genetic elements that mediate the transfer of virulence genes. The ecosystem comprising V. cholerae, CTXPhi, the aquatic environment, and the mammalian host offers an understanding of the complex relationship between pathogenesis and the natural selection of a pathogen.

Female reproductive output in a Mediterranean shrub: effects from inflorescence to population.

Inflorescence effects have been poorly studied, in spite of the functional relevance of the inflorescence in fruit and seed ecology. The present study focused on the effects of inflorescence size and phenology, and flower position within the inflorescence, in relation to fruit and seed production of the Mediterranean shrub Ononis fruticosa. Variability in fruit and seed production, seed weight and germination were estimated and modelled. Results confirmed that the most important predictors in seed production were inflorescence flowering time and flower position within the inflorescence. Thus, the number of mature seeds per fruit was higher in earlier inflorescences and in basal positions. On the other hand, predation was higher in fruits in basal positions. In fact, seed predation seemed to be the most important factor controlling final seed production. Models at the plant level suggested a negative incidence of geitonogamous pollination and resource limitation, which were also observed at the fruit level. This study confirmed the relevance of inflorescence effects on the reproductive output of O. fruticosa. Although the underlying processes could not be identified, our results provide several hypotheses for future experimental studies.

Effects of season and population size on pollination and reproductive output in a Mediterranean shrub.

Pollinator guilds may change throughout extended flowering periods, affecting plant reproductive output, especially in seasonal climates. We hypothesised a seasonal shift in pollinator guild and an autumn reduction in pollinator abundance, especially in small and sparse populations. We recorded pollinator identity, abundance and behaviour in relation to flower density from plant to population throughout the extended flowering of Ononis tridentata. We evaluated female reproductive output by recording pollination success and pre-dispersal seed predation in eight populations of contrasting size and density. Offspring quality was also characterised through seed weight and germination. A diverse guild of insects visited O. tridentata in spring, while only Apis mellifera was observed in autumn. Visitation frequency did not vary seasonally, but the number of flowers per foraging bout was lower, and seeds were heavier and had a higher germination rate in autumn. Plant and neighbourhood flowering display were not related to pollinator visitation frequency or behaviour. However, the rate of fertilised ovules, seed set and autumn flowering display size were positively related to population density. The maintenance of pollination in autumn enhances the reproductive performance of O. tridentata due to higher quality of autumn seed, and to a large reduction in seed predator pressure. We also suggest that observed changes in pollinator behaviour could be one of the processes behind seasonal variation in seed performance, since geitonogamous crosses were less likely to occur in autumn.

Treatment of Helicobacter pylori infection in infants in rural Bangladesh with oral immunoglobulins from hyperimmune bovine colostrum.

BACKGROUND: Antibodies from hyperimmune bovine colostrum have been shown to be effective in treatment against a variety of microorganisms, including Helicobacter pylori in adults. AIM: To test this form of treatment in a small group of H. pylori infected children in a periurban community in Bangladesh. METHODS: Twenty-four infants, 4-29 months old (mean age 16.5+/-7.7 months) and infected with H. pylori, were treated with purified immunoglobulins from hyperimmune bovine colostrum for 1 month, in a placebo-controlled, double-blind pilot study. Diagnosis was established with 13C-urea breath test (UBT) before and after the treatment period and at a 1-month follow-up. RESULTS: None of the hyperimmune bovine colostrum-treated children became UBT negative. Five children initially positive in the UBT screening spontaneously became negative by the start of the study with hyperimmune bovine colostrum/placebo. At the end of the 1-month study period, three had became positive again. CONCLUSION: Hyperimmune bovine colostrum does not eradicate H. pylori infection in infants. Transient H. pylori infection is common among infants in high endemic areas, as is reinfection after clearance. This presents obstacles to evaluation of therapeutic investigations in young children in areas where H. pylori is prevalent.

Sociodemographic, hygienic and nutritional correlates of Helicobacter pylori infection of young Bangladeshi children.

BACKGROUND: By the age of 10 years most children in developing countries have been infected by Helicobacter pylori Identification of clues to modes of transmission of this organism to children, as well as evaluation of the sequelae of childhood infections, constitute important research priorities for developing countries. OBJECTIVES: To evaluate demographic, socioeconomic and hygienic factors associated with acquisition of infection by H. pylori early in childhood among Bangladeshi children ages 2 to 5 years and to assess whether infection by H. pylori was associated with poor nutritional status in these children and in an older group ages 6 to 9 years. METHODS: A random population-based survey of 257 rural Bangladeshi children ages 2 to 5 years and 312 children ages 6 to 9 years. Seropositivity for H. pylori, as manifested by the presence of serum IgG anti-H. pylori antibodies, was correlated with nutritional status of the sampled children and with sociodemographic features and access to clean water and latrine facilities among families of the children. RESULTS: Among children ages 2 to 5 years, the 123 (48%) who were infected by H. pylori were similar to the 134 noninfected children with respect to socioeconomic level, family access to tube well water and family ownership of a latrine. However, families of infected children had more persons per sleeping room in the home (3.8 vs. 3.2, P < 0.05) and were more likely to be Hindu (20% vs. 10%, P < 0.05). Infected children did not differ significantly from noninfected children in Z scores for weight-for-age (-2.66 vs. -2.78), weight-for-height (-1.17 vs. -1.28) or height-for-age (-3.58 vs. -3.56). Analysis of survey children ages 6 to 9 years also revealed similar nutritional indexes among infected vs. noninfected children. CONCLUSIONS: Household crowding and behaviors that differ between Hindus and Muslims, but not lack of access to clean water and latrines, may enhance the transmission of H. pylori to rural Bangladeshi children. Although confirming the high frequency of infections in young Bangladeshi children, our findings do not support the notion that H. pylori is responsible for the high prevalence of malnutrition in this setting.

Neonatal rotavirus infection in Bangladesh: strain characterization and risk factors for nosocomial infection.

BACKGROUND: Rotavirus (RV) diarrhea is an important cause of childhood morbidity and mortality in Bangladesh and is responsible for 24% of hospital admissions for diarrhea in children from 3 months to 2 years of age. However, the prevalence of neonatal RV infections and characteristics of RV strains infecting neonates have not been explored in Bangladesh. METHODS: We investigated neonates at six hospitals in Bangladesh to determine the prevalence of neonatal RV infection, to identify risk factors for infection and to characterize neonatal RV strains by reverse transcription-polymerase chain reaction. RESULTS: Of 381 neonates screened at 6 hospitals 61 of 146 infants (42%) at 2 hospitals in Dhaka were RV-positive. Of these 62% were detected within the first 5 days of life. We found an increased risk for neonatal RV infection among infants whose mothers reported no handwashing during care of the neonate (P = 0.03). Analysis of RV strains in enzyme-linked immunosorbent assay-positive specimens identified P[6]G4 and P[6]G1 genotypes to be most common; 7% (2 of 27) of strains were nontypable. A concurrent analysis of RV strains circulating in Bangladesh suggested that RV genotypes infecting neonates had a distinct P genotype, because most community strains were P-nontypable compared with neonatal strains, which carried the P[6] genotype. CONCLUSIONS: Hospitalized neonates in Dhaka have increased risk for infection with RV as early as the first week of life with strains having the unusual P[6] genotype. Our findings confirm studies in India showing that neonatal RV infection can be common and may occur with strains distinct from those circulating in the community. Neonatal RV infections could alter a child's response to the RV vaccine as well as the calculation of RV vaccine efficacy in these populations.

Anticipating rotavirus vaccines: hospital-based surveillance for rotavirus diarrhea and estimates of disease burden in Bangladesh.

OBJECTIVES: Rotavirus is the most common cause of severe diarrhea in children worldwide, and a vaccine may soon be licensed and available for use in immunization programs. To assess the need for a rotavirus vaccine in Bangladesh, we estimated the disease burden of rotavirus diarrhea from national vital statistics for births and diarrheal deaths, together with hospital surveillance data on the proportion of severe childhood diarrhea attributed to rotavirus. METHODS: From 1990 through 1993, hospital surveillance was conducted of a systematic, random 4% sample of >80,000 patients with diarrhea who sought care each year at the International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR,B). RESULTS: Rotavirus was detected in 20% (1561 of 7709) of fecal specimens from children with diarrhea <5 years of age; 92% of all cases (1436) occurred in children <2 years of age, but only 3% (50) of cases occurred in infants <3 months of age. Children infected with rotavirus were more likely to have watery stools (P < 0.001), severe vomiting (P < 0.001) but less severe dehydration (P = 0.007) than children infected with other enteropathogens. CONCLUSIONS: We estimate that in this setting, where 18% of children die by age 5 and about 25% of these succumb to diarrhea, between 14,850 and 27,000 of the 3 million Bangladeshi children born in 1994 will die of rotavirus by the age of 5 years, equivalent to 1 rotavirus death per 111 to 203 children. The estimated burden of rotavirus diarrhea in Bangladesh is sufficiently great to warrant field testing of rotavirus vaccines for possible inclusion in the current immunization program.

Successful treatment of rotavirus diarrhea in children with immunoglobulin from immunized bovine colostrum.

BACKGROUND: Oral ingestion of immunoglobulins in humans has been shown to be effective as prophylaxis against enteric infections. However, its therapeutic effect in children with infectious diarrhea has hitherto not been proven. We treated children with rotavirus diarrhea with immunoglobulins extracted from immunized bovine colostrum (IIBC) containing high titers of antibodies against four rotavirus serotypes. METHODS: In this double blind placebo-controlled trial, 80 children with rotavirus diarrhea were randomly assigned to receive orally either 10 g of IIBC (containing 3.6 g of antirotavirus antibodies) daily for 4 days or the same amount of a placebo preparation. The daily stool output (grams/kg/day), intake of oral rehydration solution (ml/kg/day), stool frequency (number of stools/day) and presence of rotavirus in stool were monitored for the 4 days during treatment. RESULTS: Children who received IIBC had significantly less daily and total stool output and stool frequency and required a smaller amount of oral rehydration solution than did children who received placebo (P < 0.05). Clearance of rotavirus from the stool was also earlier in the IIBC group compared with the placebo group (mean day, 1.5 vs. 2.9, P < 0.001). No adverse reactions from the colostrum treatment were observed. CONCLUSIONS: Treatment with antirotavirus immunoglobulin of bovine colostral origin is effective in the management of children with acute rotavirus diarrhea.

Astrovirus infection in association with acute, persistent and nosocomial diarrhea in Bangladesh.

BACKGROUND: Diarrhea is an important public health concern in developing countries such as Bangladesh. Diarrhea in children that persists for 14 days or more occurs in 7% of patients in Bangladesh and frequently results in death. Astrovirus has been demonstrated as a cause of acute and nosocomial diarrhea and can be excreted for prolonged periods, yet its importance as a cause of diarrhea among children in a developing country like Bangladesh has not been investigated. METHODS: We tested 629 stool specimens from patients with acute diarrhea, 153 from patients with persistent diarrhea, 175 specimens from 76 patients hospitalized for diarrhea who were sampled repeatedly to detect nosocomial infection and 428 from nonhospitalized healthy children (controls). All children enrolled in the study were <5 years of age. Astrovirus was detected by enzyme immunoassay and other enteropathogens were detected by standard techniques. RESULTS: The detection of astrovirus increased significantly with the duration of diarrhea. Astrovirus was found in 23 (15%) specimens from patients with persistent diarrhea, 26 (4%) patients with acute diarrhea, but only 8 (2%) healthy controls. This trend remained when we limited our analysis to infants <12 months of age and to episodes in which astrovirus was the sole pathogen. Among patients with nosocomial diarrhea, 16% of postadmission specimens were positive for astrovirus when the admission specimen was negative. CONCLUSION: The observation that astrovirus is detected more frequently with diarrhea of increasing duration suggests the need for further studies to determine whether astrovirus plays a causative role in persistent diarrhea or is a secondary agent.

Neonatal diarrhea caused by Vibrio cholerae 0139 Bengal.

Cholera rarely occurs in children under 2 years of age. We describe diarrhea due to Vibrio cholerae 0139 Bengal, the newly described etiologic agent of cholera in a 4-day-old breast-fed baby. However, the diarrhea was mild and was successfully treated with rehydration therapy and erythromycin.

PIP: Cholera due to Vibrio cholerae 01 has been endemic in Bangladesh. In endemic cholera, the most susceptible age group is children aged 3-5 years, while those under age 2 years are rarely affected. Breast-fed children are protected from cholera due to the lack of extraneous contamination of breast milk and the protective effect of breast milk antibodies. The authors describe the case of diarrhea due to Vibrio cholerae 0139 Bengal in a 4-day old infant admitted to the Clinical Research and Service Center of the International Center for Diarrheal Disease Research, Bangladesh, for treatment. Vibrio cholerae 0139 Bengal caused large epidemics of clinical cholera in India during 1992-93, and is now established as the second etiologic agent of cholera. The male baby from a poor, rural family of Bangladesh was born at term, weighed 2.1 kg, and was breast fed. He had a history of watery diarrhea for the preceding 2 days and evidence of jaundice, but no indication of abnormality based upon systemic examination. After isolating V. cholerae 0139 in a stool specimen, the baby was treated with a total of 300 ml IV cholera saline and oral erythromycin, after which he recovered from diarrhea and was discharged on the third day after admission.

Monoclonal antibodies specific for Shigella dysenteriae serotype 13. Production, characterization, and diagnostic application.

Three mouse monoclonal antibodies (mAbs) (ICL3, ICL4, and ICL5) were produced that specifically recognized the lipopolysaccharide antigen of the newly recognized Shigella dysenteriae serotype-13 strain. All three mAbs reacted with all nine reference isolates of S. dysenteriae 13 in different tests. The mAbs also detected colonies of S. dysenteriae-13 isolates by direct slide agglutination test. The mAbs also reacted with the reference Escherichia coli 0150 strain and showed its close antigenic relationship with S. dysenteriae 13. Use of these mAbs in our clinical laboratory during an 8-month period detected three S. dysenteriae-13 isolates that were also detected by a polyclonal rabbit antiserum. It should now be possible to define the epidemiologic importance of S. dysenteriae serotype 13 in diarrhea by using these mAbs.

Fluorescent antibody staining test for rapid diagnosis of Shigella dysenteriae 1 infection.

An indirect fluorescent antibody test for rapid detection of Shigella dysenteriae 1 in diarrheal stools was developed. A diagnosis could be made within 90 min of submission of specimens to the laboratory. On comparison with culture results, the test had a sensitivity of 92%, a specificity of 93%, and positive and negative predictive values of 94% and 92%, respectively.

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.

Multiple-drug-resistant Salmonella gloucester infections in Bangladesh.

We describe three different cases of extraintestinal infections caused by multidrug-resistant Salmonella gloucester from Bangladesh that were associated with bad prognoses. These clinical manifestations were not previously reported for S. gloucester.

Diarrhea associated with Cyclospora Sp. in Bangladesh.

A spherical acid-fast organism measuring approximately 10 microns in diameter (Cyclospora sp.) has recently been implicated in diarrheal diseases in many parts of the world. We detected this organism in the stools of six Bangladeshi patients with diarrhea. Four patients had chronic diarrhea and two had acute diarrhea at the time of presentation. This is the first report of infection with this organism in the indigenous population from this region.

Septicemia due to Vibrio cholerae 0139 Bengal.

We describe septicemia due to Vibrio cholerae 0139 Bengal, the newly described causative agent of cholera, in a child who also had simultaneous intestinal infection with Shigella boydii. Because V. cholerae 0139 is capsulated, its propensity to cause extraintestinal infection is stressed.

Adherence to human small intestines of capsulated Vibrio cholerae O139.

Capsulated cells of V. cholerae O139 adhered to formalin-fixed or native mucosa of the small intestines from an adult and a child. The primary adherence target was mucus. Capsulated O139 cells adhered better to the antigen sampling cells (M cells) of ileal Peyer's patch than to the absorptive cells. O139 cells on the mucosa appeared as small aggregates. Similar organisms were found on the mucosa of duodenal biopsy samples from patients infected with V. cholerae O139. The findings indicated that capsulated cells of V. cholerae O139 tend to autoagglutinate and contribute to the effective adherence to the intestinal mucosa.

Purification and characterization of vibrio cholerae O139 fimbriae.

A Vibrio cholerae O139 (strain Al-1841) isolated from a patient with a cholera-like disease in Bangladesh predominantly produced new curved, wavy fimbriae (Al-1841 fimbriae) and small numbers of previously reported V. cholerae non-O1 S7-like pili. The former was purified and characterized. The molecular mass of the Al-1841 fimbrial subunit was less than 2.5 kDa, and it was immunologically different from that of V. cholerae non-O1 S7 pili. This novel fimbrial antigen was detected in all 182 Gram-negative strains from five genera tested but was absent from the Gram-positive bacteria tested. The purified Al-1841 fimbriae did not agglutinate human or rabbit erythrocytes.

A monoclonal antibody to Shigella dysenteriae serotype 13 cross-reacting with Shiga toxin.

A monoclonal antibody (mAb ICT6) was produced against the newly described Shigella dysenteriae serotype type 13. The mAb was of IgM isotype and recognized purified Shiga toxin in ELISA and immunoblot. It also recognized periplasmic extract S. dysenteriae type 13 in immunoblot as did an affinity-purified polyclonal rabbit antiserum and a previously described monoclonal antibody to the B subunit of Shiga toxin. The mAb ICT6 did not neutralize the cytotoxic effects or S. dysenteriae type 13, Shiga toxin or periplasmic extracts of S. dysenteriae type 1 for HeLa cells.