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Vibrio cholerae, an estuarine bacterium, is the causative agent of cholera, a severe diarrheal disease that demonstrates seasonal incidence in Bangladesh. In an extensive study of V. cholerae occurrence in a natural aquatic environment, water and plankton samples were collected biweekly between December 2005 and November 2006 from Mathbaria, an estuarine village of Bangladesh near the mangrove forests of the Sundarbans. Toxigenic V. cholerae exhibited two seasonal growth peaks, one in spring (March to May) and another in autumn (September to November), corresponding to the two annual seasonal outbreaks of cholera in this region. The total numbers of bacteria determined by heterotrophic plate count (HPC), representing culturable bacteria, accounted for 1% to 2.7% of the total numbers obtained using acridine orange direct counting (AODC). The highest bacterial culture counts, including toxigenic V. cholerae, were recorded in the spring. The direct fluorescent antibody (DFA) assay was used to detect V. cholerae O1 cells throughout the year, as free-living cells, within clusters, or in association with plankton. V. cholerae O1 varied significantly in morphology, appearing as distinctly rod-shaped cells in the spring months, while small coccoid cells within thick clusters of biofilm were observed during interepidemic periods of the year, notably during the winter months. Toxigenic V. cholerae O1 was culturable in natural water during the spring when the temperature rose sharply. The results of this study confirmed biofilms to be a means of persistence for bacteria and an integral component of the annual life cycle of toxigenic V. cholerae in the estuarine environment of Bangladesh., IMPORTANCE Vibrio cholerae, the causative agent of cholera, is autochthonous in the estuarine aquatic environment. This study describes morphological changes in naturally occurring V. cholerae O1 in the estuarine environment of Mathbaria, where the bacterium is culturable when the water temperature rises and is observable predominantly as distinct rods and dividing cells. In the spring and fall, these morphological changes coincide with the two seasonal peaks of endemic cholera in Bangladesh. V. cholerae O1 cells are predominantly coccoid within biofilms but are rod shaped as free-living cells and when attached to plankton or to particulate matter in interepidemic periods of the year. It is concluded that biofilms represent a stage of the annual life cycle of V. cholerae O1, the causative agent of cholera in Bangladesh.

The treatment regimen for diarrhea depends greatly on correct diagnosis of its etiology. Recent diarrhea outbreaks in Bangladesh showed Vibrio cholerae to be the predominant cause, although more than 40% of the suspected cases failed to show cholera etiology by conventional culture methods (CMs). In the present study, suspected cholera stools collected from every 50th patient during an acute diarrheal outbreak were analyzed extensively using different microbiological and molecular tools to determine their etiology. Of 135 stools tested, 86 (64%) produced V. cholerae O1 by CMs, while 119 (88%) tested positive for V. cholerae O1 by rapid cholera dipstick (DS) assay; all but three samples positive for V. cholerae O1 by CMs were also positive for V. cholerae O1 by DS assay. Of 49 stools that lacked CM-based cholera etiology despite most being positive for V. cholerae O1 by DS assay, 25 (51%) had coccoid V. cholerae O1 cells as confirmed by direct fluorescent antibody (DFA) assay, 36 (73%) amplified primers for the genes wbe O1 and ctxA by multiplex-PCR (M-PCR), and 31 (63%) showed El Tor-specific lytic phage on plaque assay (PA). Each of these methods allowed the cholera etiology to be confirmed for 97% of the stool samples. The results suggest that suspected cholera stools that fail to show etiology by CMs during acute diarrhea outbreaks may be due to the inactivation of V. cholerae by in vivo vibriolytic action of the phage and/or nonculturability induced as a host response.

SUMMARYDuring epidemics of cholera in two rural sites (Bakerganj and Mathbaria), a much higher proportion of patients came for treatment with severe dehydration than was seen in previous years. V. cholerae O1 isolated from these patients was found to be El Tor in its phenotype, but its cholera toxin (CT) was determined to be that of classical biotype. Whether the observed higher proportion of severe dehydration produced by the El Tor biotype was due to a shift from El Tor to classical CT or due to other factors is not clear. However, if cholera due to strains with increased severity spread to other areas where treatment facilities are limited, there are likely to be many more cholera deaths.

UNLABELLED: A controlled, randomized, double-blind study in Bangladeshi children (ages 4-36 mo) with acute diarrhoea was undertaken to determine whether bismuth subsalicylate (BSS) would prevent the development of persistent diarrhoea (PD) in young children. The children were randomized to two groups: 226 were given liquid oral BSS, (as Pepto-Bismol), 100 mg/kg/d for 5 d; 225 were given placebo of identical appearance. On admission to the study, the two groups were comparable both clinically and microbiologically. Rotavirus was found in 56% of all the children, and enterotoxigenic E. coli in 31% of a subsample studied. Children treated with BSS had less severe and less prolonged illness than those treated with placebo (p = 0.057). There was, however, no difference in the development of PD between the two groups (8% and 11%). Unexpectedly, patients treated with BSS gained significantly more weight (2.3%) than those treated with placebo (0.5%; p < 0.001) during the course of the study. No toxicity of BSS was detected. CONCLUSION: Treatment with BSS had a modest therapeutic effect on acute diarrhoea, as has been previously demonstrated, but with no suggestion of a therapeutic effect on the prevention of persistent diarrhoea in this group of patients.

Enterotoxigenic Escherichia coli (ETEC) is a common cause of bacterial infection leading to acute watery diarrhea in infants and young children. Although the prevalence of ETEC is high in Bangladesh and infections can be spread through food and contaminated water, limited information is available about ETEC in the surface water. We carried out studies to isolate ETEC from surface water samples from ponds, rivers, and a lake from a site close to field areas known to have a high incidence of diarrhea in Dhaka, Bangladesh, and Matlab, Bangladesh. ETEC strains isolated from the water sources were compared with ETEC strains isolated from patients with diarrhea at two hospitals in these areas. ETEC were isolated from 30% (45 of 150) of the samples from the surface water sources and 19% (518 of 2700) of the clinical specimens. One hundred ETEC strains isolated from patients with similar phenotypes as the environmental strains were compared for phenotypic and genotypic properties. The most common O serogroups on ETEC were O6, O25, O78, O115, and O126 in both types of strains. Pulsed-field gel electrophoresis analyses of the ETEC strains showed that multiple clones of ETEC were present within each colonization factor type and that some clones detected in the environment were also isolated from the stools of patients. The strains showed multiple and similar antibiotic resistance patterns. This study shows that ETEC is prevalent in surface water sources in Bangladesh suggesting a possible reason for the endemicity of this pathogen in Bangladesh.Key words: enterotoxigenic Escherichia coli (ETEC), surface water samples, colonization factors, pulsed-field gel electrophoresis, toxin types.

The organization of the CTX prophage in environmental strains of Vibrio mimicus was investigated. Sixteen hundred non-sucrose fermenting vibrios were examined for ctx gene by hybridization. Out of 1,600 isolates, 6 V. mimicus isolates contained ctxA gene. The organization of CTX prophage was determined by RFLP using ctxA probe. The CTX prophage integrated at a single site in V. mimicus genome which was present as a single copy flanked by at least a single RS element. Ribotype pattern revealed that a particular clone of V. mimicus acquired the CTXPhi in the aquatic environment. This study demonstrated that V. mimicus could act as a reservoir of CTXPhi in the aquatic environment.

The diversity of toxigenic V. cholerae O1 in the aquatic environment of Bangladesh is not known. A total of 18 environmental and 18 clinical strains of toxigenic V. cholerae O1 were isolated simultaneously from four different geographical areas and tested for variation by the pulsed-field gel electrophoresis method. Environmental strains showed diversified profiles and one of the profiles was common to some environmental strains and most clinical strains. It appears that one clone has an advantage over others to cause disease. These findings suggest that the study of the molecular ecology of V. cholerae O1 in relation to its environmental reservoir is important in identifying virulent strains that cause disease.

A comparative study was carried out to see the differences in pathogenicity of rough and smooth strains. A total of 10 strains including 5 each of rough and smooth strains of Vibrio cholerae O1 were tested and found positive for toxin production by enzyme-linked immunosorbent assay (ELISA) in Richardson's and AKI media. All the smooth and rough strains, except one, showed a titre of 1: 10 and 1: 100 in Richardson's and AKI media, respectively. Both types of strains produced enterotoxin in rabbit ileal loop (RIL). The differences in multiplication abilities of smooth and rough strains in RIL were statistically significant (P0.05). However, these differences in multiplying abilities did not influence the adherence potential or enterotoxin production as there was no significant difference (P0.05) between these properties. This study demonstrated that the rough strains are equally pathogenic and as important as smooth strains.

Recently, a new strain of cholera, Vibrio cholerae O139, has emerged as an epidemic strain, but there is little information about its environmental reservoir. The present investigation was aimed to determine the role of cyanobacteria in the persistence of V. cholerae O139 in microcosms. An environmental isolate of V. cholerae O139 and three cyanobacteria (Anabaena sp., Nostoc sp., and Hapalosiphon sp.) were used in this study. Survival of culturable V. cholerae O139 in microcosms was monitored using taurocholate-tellurite gelatin agar medium. Viable but nonculturable V. cholerae O139 were detected using a fluorescent antibody technique. Vibrio cholerae O139 could be isolated for up to 12 days in a culturable form in association with cyanobacteria but could not be isolated in the culturable form after 2 days from control water without cyanobacteria. The viable but nonculturable V. cholerae O139 could be detected in association with cyanobacteria for up to 15 months. These results, therefore, suggest that cyanobacteria can act as a long-term reservoir of V. cholerae O139 in an aquatic environment.Key words: cyanobacteria, persistence, Vibrio cholerae, microcosm, reservoir.

Duckweed has been used for the treatment of wastewater and as fish feed. A comparative study was carried out to determine (i) the efficacy of duckweed in treating hospital-based wastewater and (ii) the level of the microbial contamination of fish fed on wastewater-grown duckweed. There were two groups of ponds where fish farming was done. In one group of ponds (control ponds), duckweed that was grown using artificial fertilizer was used as fish feed; in another group (study ponds), wastewater-grown duckweed was used as fish feed. The faecal contamination of water, duckweed, and fish from study and control ponds were monitored by faecal coliform estimation. The presence of enteric pathogens among handlers, water, duckweed, and fish samples was also examined. It was observed that the faecal coliform counts of raw wastewater were 4.7 Log10CFU/mL, which was reduced to 10CFU/mL after treating with duckweed. There was no significant difference (P > 0.05) in faecal coliform counts in water collected from duckweed ponds and fish ponds of study and control areas. The wastewater-grown duckweed did not pose any health hazard to the handlers. These results demonstrated that the wastewater-treated duckweed may be safely used as fish feed.Key words: wastewater, duckweed, fish feed, contamination, safety.

This paper defines high-risk areas of cholera based on environmental risk factors of the disease in an endemic area of Bangladesh. The risk factors include proximity to surface water, high population density, and low educational status, which were identified in an earlier study by the authors. Cholera data were analyzed by spatially referenced extended household units for two time periods, 1983-1987 and 1992-1996. These periods were chosen because they had different dominant cholera agents. From 1983-1987 classical cholera was dominant and from 1992-1996 El Tor was dominant. By defining high-risk areas based on risk factors, this study builds a spatial risk model for cholera. The model is then evaluated based on the locations of observed cholera cases. The study also identifies the determinants of death due to cholera for the two different time periods dominated by the different cholera agents. The modeled risk areas that were based on the risk factors were found to correspond with actual distributions of cholera morbidity and mortality. The high-risk areas of the dominant cholera agents are relatively stable over time. However, from 1983-1987 El Tor cholera, which was not the dominant agent during that period, was not associated with high-risk areas, suggesting that the El Tor habitat may have changed over time. The case fatality rate for cholera was related to proximity to a diarrhea treatment hospital in the study area.

Background: , Vibrio cholerae are known to be normal inhabitants pf surface water. However, the environmental niches of the different strains of cholera are not well known, and therefore, populations at risk for cholera outbreaks cannot be clearly identified. Methods: This study identifies environmental risk factors for cholera caused by V. cholerae 01 El Tor and 0139 and environmental niches of the two strains present in Matlab, a cholera endemic area of Bangladesh. The study year was 1993, the year that the O139 strain first appeared in the study area. Patients who had either strain of cholera identified in a laboratory were included in the study. A geographic information system was used to map the household locations of the patients, to describe the human sanitary environment and population density, and to address potential anthropogenic and environmental risk factors of the disease. Spatial point pattern and exploratory spatial data analysis techniques were used to define the environmental niches of the two cholera strains. Results: The study suggests the niches of O1 El Tor and O139 strains of V. cholerae appear to be similar, based on common environmental risk factors. Conclusions: The results of this study support a theory that O1 El Tor could possibly be replaced by the newer O139 strain in the future.

The need for further studies on relationships between deaths and environmental variables has been reported in the literature. This case-control study was, therefore, carried out to find out the associations between several social and environmental variables and deaths of children due to infectious diseases such as those leading to diarrhoea, acute respiratory infection, measles and other diseases. Six hundred and twenty-five deaths (cases) and an equal number of matched living children (controls) aged 1–59 months, were studied in rural Matlab. An analysis of crude and adjusted odds ratio showed differential associations. Sources of drinking water, amount of stored water, conditions of latrines, number of persons sleeping with the child and the type of cooking site were statistically significantly associated with deaths due to infectious diseases after controlling for breast feeding, immunization, and the family size. Significant associations were also observed between: (i) the sources of drinking water and deaths due to ARI, and (ii) conditions of latrines and deaths due to diarrhoeal diseases, after controlling for the confounding variables. Several other environmental factors also showed associations with these various death groups, but they were not statistically significant. The size of the samples in death groups (small) and the prevalence of more or less homogeneous environmental health conditions probably diminished the magnitude of the effects. The results of the study reconfirm the importance of environmental health intervention in child survival, irrespective of breast-feeding, immunization, and selected social variables.

Vibrio cholerae O139 has recently emerged as the second etiologic agent of cholera in Asia. A study was carried out to evaluate the induction of specific immune responses to the organism in V. cholerae O139-infected patients. The immune responses to V. cholerae O139 Bengal were studied in patients by measuring antibody-secreting cells (ASC), as well as vibriocidal and antitoxic antibodies in the circulation. These responses were compared with those in patients with V. cholerae O1 disease. Strong immunoglobulin A (IgA) and IgM ASC responses were seen against the homologous lipopolysaccharide or serogroup of V. cholerae. The magnitude and isotype of the responses were similar in O139- and O1-infected patients. Vibriocidal antibody responses were seen against bacteria of the homologous but not heterologous serogroup, and these responses reflect the lack of cross-protection between the infections caused by the two serogroups. The two groups of patients showed comparable cholera toxin-specific ASC responses, with the IgG isotype dominating over the IgA isotype, as well as comparable antitoxic immune responses in plasma. These results suggest that despite having a polysaccharide capsule, V. cholerae O139 induces systemic and intestine-derived ASC responses in peripheral blood comparable to those seen in patients with V. cholerae O1 disease.

During development of Peru-15, a new live oral vaccine for cholera, the role of buffer needed to be evaluated. Generally, oral bacterial vaccines are acid labile and need to be administered by using a formulation which protects them from gastric acid. We compared three different buffers for use with Peru-15, including a standard bicarbonate-ascorbic acid buffer, Alka-Seltzer, and a new electrolyte-rice buffer, CeraVacx. Saline served as the control. Thirty-nine healthy adult volunteers received Peru-15 (10(8) CFU) with one of the three buffers or saline in a double-masked study. The volunteers were monitored for symptoms for 7 days after the dose, serum was tested for antibody responses by vibriocidal antibody and immunoglobulin G antitoxin enzyme-linked immunosorbent assays, and stool samples were tested for excretion of the vaccine strain. Side effects were minimal in all groups. All 30 volunteers who took Peru-15 with a buffer showed significant rises in vibriocidal antibody titer. The magnitude of the rises was higher in the CeraVacx group than in the other two buffer groups. Four of nine volunteers who took the vaccine with saline also showed increased titers, but they were lower than those in any of the three buffer groups. Excretion of the vaccine strain was similar in the buffer groups, but excretion was not associated with the magnitude of the vibriocidal responses. Excretion of Peru-15 was not detected in the saline group. We conclude that buffer does amplify the serological response to Peru-15 and that CeraVacx may provide benefits not provided by other buffers.

The emergence of the new strain Vibrio cholerae O 139 and its rapid spread in Bangladesh and India together with its detection in several other countries, have raised the question whether this constitutes the beginning of the eighth pandemic of cholera, and if so, how large a threat it poses. In an attempt to answer this question, epidemic spread patterns of Vibrio cholerae O 139 strain in Bangladesh were studied. Initially the epidemic moved quickly and affected the entire coastal and estuarine tidal plains of southern Bangladesh. In the flood plains of the northern regions it affected mostly the north-eastern and north-central areas, at a slower pace than in the southern areas. In the beginning the new strain totally displaced both biotypes (classic and El Tor) of Vibrio cholerae O 1 . Nearly 2 years after its initial detection, striking differences in the distribution of V. cholerae O 139 and O 1 were observed. In most northern areas, the new strain was replaced by V. cholerae O 1 , whereas in the southern coastal regions, the O 139 strain continues to dominate epidemics. The study suggests that the O 139 strain may become endemic in the coastal ecosystem. The threat of a pandemic, therefore, may not be as large as it first seemed.

Objectives. To report on the burden and outcome of episodes of acute otitis media (AOM) based on awareness of AOM and compliance to referral by community health workers (CHWs) in rural children aged under 2 in Bangladesh. Study design. Retrospective population-based cohort study. Method. Secondary data analysis on episodes of AOM, care-seeking pattern, compliance with referral, antibiotic use and duration of episodes from morbidity data collected by CHWs on 252 children aged under 2 who completed biweekly household surveillance starting from birth. Results. A total of 375 episodes of AOM were observed in 45.6% (115/252) of the study subjects; 19.4% (49/252) experienced single and 26.2% (66/252) recurrent episodes. Just over 34% (129/375) of episodes were recognised by caregivers and reported to CHWs, and the remaining 246 episodes, unrecognised by caregivers, were detected by CHWs during home visits. CHWs referred all subjects with AOM to the study doctor. The caregivers complied with referral in all episodes they had recognised, but in only 21.5% (53/246) of episodes they had not recognised (p

We undertook a controlled study of children younger than 5 years in Bangladesh to determine whether enterotoxigenic Bacteroides fragilis (ETBF) was associated with diarrhea in this population. ETBF was isolated from 22 (6.1%) of 358 patients and 5 (1.2%) of 425 controls (P = 0.0001). In children younger than 1 year, however, low isolation rates (2 to 3%) were found in both patients and controls. In children older than 1 year, the rates were significantly higher in children with diarrhea (16 [9%] of 177) than in controls (2 [1%] of 264; P = 0.00001). When children with mixed infections with other known diarrheal pathogens were removed, the differences in children older than 1 year were still significant (7 [4%] of 177 versus 2 [1%] of 264; P = 0.033). The syndrome associated with ETBF was secretory in nature, with watery diarrhea, and of mild severity. These epidemiological and clinical findings are similar to those from a previous study of White Mountain Apaches in the United States and are the first to suggest that ETBF may also be an important diarrheal pathogen in other geographic areas and in the developing world where diarrhea is highly endemic.

Mouse monoclonal antibodies (MAbs) were derived against acetone-treated whole cells of the newly recognized Vibrio cholerae O139 serogroup which is causing epidemics of cholera-like disease in India and Bangladesh. Four MAbs specifically recognized the lipopolysaccharide antigens of V. cholerae O139. MAbs ICL9 and ICL13 were of the immunoglobulin M (IgM) isotype, ICL11 was of the IgG3 isotype, and ICL12 was of the Ig2b isotype. A fifth MAb, ICL10, of the IgG2b isotype cross-reacted with V. cholerae O91. All five MAbs recognized V. cholerae O139 in an enzyme-linked immunosorbent assay, slide agglutination test, motility inhibition test, and indirect immunofluorescence test. During a 1-month evaluation of these MAbs in our clinical laboratory, all 86 cases diagnosed as V. cholerae O139 by a rabbit polyclonal antiserum were also detected by these MAbs, establishing their utility as highly sensitive and specific diagnostic reagents. With these MAbs, it should now be possible to screen for the V. cholerae O139 serogroup in epidemic and endemic diarrhea cases and in environmental and food samples.

Despite the known presence of rotavirus-associated diarrhoea in Bangladesh, its prevalence, including records of hospitalization in rural health facilities, is largely unknown. In a systematic surveillance undertaken in two government-run rural health facilities, 457 children, aged less than five years, having acute watery diarrhoea, were studied between August 2005 and July 2007 to determine the prevalence of rotavirus. Due to limited financial support, the surveillance of rotavirus was included as an addendum to an ongoing study for cholera in the same area. Rotavirus infection was detected in 114 (25%) and Vibrio cholerae in 63 (14%) children. Neither rotavirus nor V. cholerae was detected in 280 (61%) samples; these were termed ‘non-rotavirus and non-cholera’ diarrhoea. Both rotavirus and cholera were detected in all groups of patients (

A community-based longitudinal study was carried out at Matlab, Bangladesh, to investigate the magnitude of the problem of persistent diarrhea; 705 children aged < 5 y were followed, yielding 7300 child-months of observation in 1 y. Morbidity data were collected every fourth day by home visit, anthropometric status was determined monthly, and cell-mediated immune status was assessed every 3 mo. The incidence of persistent diarrhea was 34 episodes per 100 child-years; rates were highest in infancy and declined through the remainder of childhood. In a logistic-regression model, weight-for-height status and immune status were significant predictors of persistent diarrhea. Compared with those at zero Z score, children with weight-for-height at -2 would have a 3.5 times increased risk of persistent diarrhea. Compared with immunocompetent children, immunodeficient children had about twice the risk of developing persistent diarrhea. Thus, nutritional status and cell-mediated immune status were important independent risk factors for persistent diarrhea.

Epidemics of cholera caused by Vibrio cholerae 01 occur regularly in Bangladesh, but until lately V cholerae non-01 has been associated only with sporadic cases of diarrhoeal disease in many parts of the world, including Bangladesh. We describe a large epidemic of cholera-like disease in Bangladesh that is due to a V cholerae non-01. The epidemic began in December, 1992, in southern Bangladesh and spread throughout the country. By the end of March 107 297 cases of diarrhoea and 1473 deaths had been reported. The disease is indistinguishable from cholera in clinical features and response to treatment, but most of the cases are in adults, which suggests that the population has no previous immunological experience of the organism. At two centres 375 (40%) of 938 and 236 (48%) of 492 rectal swabs were positive for V cholerae non-01, as were 5 of 54 surface water samples. 55 isolates of V cholerae non-01 were studied in detail. They resembled El Tor vibrios in being resistant to polymyxin B and positive for agglutination of chicken erythrocytes. The strain did not belong to any of the 138 known V cholerae serogroups; so a new serogroup 0139, with the suggested name Bengal, is proposed. All the isolates studied produced large amounts of an enterotoxin apparently identical to cholera toxin. This strain seems to have pandemic potential. It is important that other countries in southeast Asia are aware of the strain's potential to cause severe morbidity and mortality.

Morbidity and mortality data are important for planning and implementing healthcare strategies of a country. To understand the major causes for hospitalizations in rural Bangladesh, demographic and clinical data were collected from the hospital-records of five government-run rural health facilities (upazila health complexes) situated at different geographical regions of the country from January 1997 to December 2001. During this period, 75,598 hospital admissions in total were recorded, of which 54% were for male, and 46% were for female. Of all the admissions, diarrhoeal disease was the leading cause for hospitalization (25.1%), followed by injuries (17.7%), respiratory tract diseases (12.6%), diseases of the gastrointestinal tract (10.5%), obstetric and gynaecological causes (8.5%), and febrile illnesses (6.7%). A considerable proportion (8.3%) of the hospitalized patients remained undiagnosed. Despite the limitations of hospital-based data, this paper gives a reasonable insight of the important causes for hospitalizations in upazila health complexes that may guide the policy-makers in strengthening and prioritizing the healthcare needs at the upazila level in Bangladesh.

To improve the understanding of the relative importance of serotypes of rotavirus in dehydrating diarrhea, we examined the correlation of clinical characteristics and disease severity with serotype in 2,441 diarrheal episodes among children younger than 2 years of age in rural Bangladesh. Of 764 rotavirus-associated episodes, a single G type (serotype 1, 2, 3, or 4) was determined by oligonucleotide probe in 485 (63%), while 233 episodes were nontypeable. Episodes with G types 2 and 3 were associated with more-severe dehydration than episodes associated with G type 1 or 4 or with nontypeable rotavirus. Episodes did not differ by G type in prevalence of vomiting, copious diarrhea, fever, abdominal pain, or length of treatment center stay. Rotavirus reinfections were detected in seven children, with homologous reinfection (G type 2) in one. Twelve children with diarrhea who died had rotavirus detected in stool specimens within 30 days of death. Children who died were more likely to be malnourished than were surviving children with rotavirus diarrhea. Of 40 specimens tested by polymerase chain reaction, 29 (72.5%) were P type 1, 9 (22.5%) were P type 2, 1 (2.5%) was P type 3, and 1 (2.5%) was nontypeable. One severely symptomatic diarrheal episode was associated with P type 3 rotavirus, a serotype usually found in asymptomatic nursery infections. Although G types 2 and 3 were associated with more-severe dehydration than other serotypes, the differences do not appear to be of major clinical importance. Effective vaccines should protect against all four major G types.

We studied the restriction endonuclease cleavage patterns of rRNA genes (ribotypes) of 72 clinical isolates of Shigella flexneri representing eight serotypes to determine whether ribotyping could be used to distinguish S. flexneri strains and to compare the discriminating ability of the method with that of serotyping. By using a cloned Escherichia coli rRNA operon as the probe, Southern blot hybridization of restriction endonuclease-digested total DNA was carried out. Ribotyping of the isolates with each of the five restriction endonucleases BamHI, EcoRI, HindIII, PstI, and SalI generated reproducible restriction patterns. However, HindIII produced the optimum digestion pattern of the rRNA genes and was more useful than the other enzymes used in differentiating strains. Analysis of the 72 isolates showed 11 different HindIII cleavage patterns of their rRNA genes. Four of these HindIII-generated ribotypes could be further differentiated into two to four subribotypes by using PstI. The results indicate that ribotyping has an application for differentiation of S. flexneri strains and can complement serotyping. Definition of strains in terms of both serotype and ribotype may be of greater use in epidemiological studies.

EnterotoxigenicEscherichia coli(ETEC) is the most common bacterial cause of childhood diarrhoea in Bangladesh. Among the virulence factors of ETEC, toxins and colonization factors (CFs) play a major role in pathogenesis. UnlikeVibrio cholerae, the relationship between ETEC and ETEC-specific phages is poorly understood and the possible role of ETEC phages in the evolution of ETEC strains in the environment is yet to be established. This study was designed specifically to isolate phages that are specific for ETEC virulence factors. Among the 49 phages isolated from 12 different surface water samples, 13 were tested against 211 ETEC strains collected from clinical and environmental sources. One phage, designated IMM-001, showed a significant specificity towards CS7 CF as it attacked all the CS7-expressing ETEC. Electron microscopic analyses showed that the isolated phage possessed an isomeric hexagonal head and a long filamentous tail. An antibody blocking method and phage neutralization assay confirmed that CS7 pilus is required for the phage infection process, indicating the role of CS7 fimbrial protein as a potential receptor for IMM-001. In summary, this study showed the presence of a lytic phage in environmental water that is specific for the CS7 CF of ETEC.

Rectal swabs from 391 infants less than 18 months of age who were hospitalized with acute diarrhea and from 138 similarly aged healthy infants were examined for the etiologic agents of diarrhea. Aeromonas spp. were recovered from 205 of 391 (52.4%) diarrheic patients, whereas they were recovered from 12 of 138 (8.7%) controls (P less than 10(-11). Among the 205 Aeromonas-positive diarrheic patients, 118 (57.6%) were found to be coinfected with other common enteropathogens. Of the 164 Aeromonas-positive initial diarrheic specimens, 82 (50.0%) had one or more other enteropathogens present; 30 patients were coinfected with rotavirus, 20 with enterotoxigenic Escherichia coli, 16 with Campylobacter spp., 14 with Shigella spp., 13 with enteropathogenic E. coli, 4 with Vibrio spp., 1 with Salmonella spp., and 1 with Plesiomonas spp. of Aeromonas strains from cases compared with that from controls supports an etiologic role for this organism. However, frequent concomitant infections with other well-recognized enteropathogens and a lack of disease correlation with common Aeromonas phenotypes suggest that only a subset of Aeromonas strains may be diarrhea causing and that such strains may be common to several of the existing species.

The introduction of a soy-based, lactose-free formula during the acute phase of diarrheal illness in infants has been shown to reduce stool output and duration of diarrhea in hospitalized patients. In the United States, most infants with acute diarrhea are treated as outpatients. In the present study, infants with diarrhea who were treated as outpatients were randomly assigned to receive either a soy-based, lactose-free formula alternating with oral rehydration solution from the beginning of therapy ("early feeding") or oral rehydration solution alone for the first 24 hours of therapy, followed by a soy-based, lactose-free formula alternating with oral rehydration solution (control group). Twenty-nine infants were randomly assigned to the early-feeding group and 27 to the control group. Twenty-one (72%) of 29 in the early-feeding group resolved their diarrhea at the end of 48 hours of therapy compared with 12 (44%) of 27 in the group fed oral rehydration solution only (P = .02). It is concluded that the introduction of a soy-based, lactose-free formula from the beginning of therapy for acute diarrhea in children treated as outpatients is safe and may shorten duration of diarrhea while maintaining adequate caloric intake.

The intestinal colonization rate of Aeromonas spp. was determined for 52 cesarean-born Peruvian neonates. Rectal swabs were obtained daily from newborns during their postdelivery hospitalization (mean = 5.5 days), and the gross appearances of their feces (blind determinations) were recorded. Aeromonas spp. were recovered from rectal swabs of 12 of 52 (23.1%) infants during their first week of life; the isolates were obtained from 5 of 9 (55.6%) infants with at least one stool with a watery consistency and from 7 of 43 (16.3%) neonates with no watery stools (P = 0.022). None of the infected infants became clinically ill. No other commonly recognized enteropathogens were detected in watery stools. An environmental survey indicated that hospital water was the probable source of infection. These and other data indicated that Aeromonas colonization occurs transiently at a very early age in Peruvian neonates and that in some instances, initial infection may be followed several days later by one or more watery stools of normal volume.

Limited information is available on the etiology of acute lower respiratory infection (ALRI) particularly pneumonia in the rural community of developing countries since most etiological studies are carried out in the hospital settings. This study examined the etiology of pneumonia among young children in a rural community of Bangladesh. A cohort of 252 newborns was followed till 24 months of age during 1993-1996. Community health workers (CHWs) identified cases of ALRI during household surveillance and recommended hospitalization. On admission, nasopharyngeal aspirates (NPA) and blood were collected for bacterial and viral identification, and chest x-rays were done. Multiple regression analysis identified factors associated with a viral etiology. Physicians diagnosed 67 pneumonia; 45% of NPA were positive for viral agents of pneumonia, and respiratory syncytial virus (RSV) was predominant (81%); 6 of 48 blood cultures were positive. X-ray was done for 58 cases; 52% had pneumonic consolidation. Of the RSV cases, 33% were found in children without pneumonic consolidation. Children living in a one-room house were 3 times more likely to develop viral pneumonia (odds ratio (OR) = 3.67, CI 1.05-12.83) than children living in a larger house. Counseling on avoiding crowding where a newborn is accommodated might reduce pneumonia incidence.

In the present study we compared the ELISPOT and antibody in lymphocyte supernatants (ALS) assays as surrogate measures of mucosal immunity. In separate studies, 20 inpatient volunteers received oral doses of 6 x 10(8) or 4 x 10(9)cfu of ETEC strain E24377A (LT+, ST+, CS1+, CS3+) and 20 subjects received 1 (n = 9) or 2 (n = 11) oral doses of the attenuated ETEC vaccine, PTL-003 expressing CFA/II (CS1+ and CS3+) (2 x 10(9)cfu/dose). Peripheral blood mononuclear cells (PBMCs) from all subjects were assayed for anti-colonization factor or toxin-specific IgA antibody responses using the ALS and ELISPOT procedures. ALS responses were measured using a standard ELISA, as well as by time-resolved fluorescence (TRF). Following challenge with E24377A, significant anti-CS3, CS1 and LT ALS responses were detected in the lymphocyte supernatants of 75-95% of the subjects. A similar proportion (75%) of subjects mounted an ALS response to CFA/II antigen after vaccination with the PTL-003 vaccine. Inter-assay comparisons between ALS and ELISPOT methods also revealed a high degree of correlation in both immunization groups. ALS sensitivity versus the ELISPOT assay for LT, CS3 and CS1-specific responses following challenge were 95%, 94% and 78%, respectively and 83% for the ALS response to CFA/II antigen after vaccination with PTL-003. Correlation coefficients for the LT and CS3 antigens were 0.94 (p

A monoclonal antibody, ICT11, specific for the toxin of enterotoxigenic Bacteroides fragilis (ETBF) neutralized the cytotoxic effect of the toxin on human colonic cell line HT-29/C1. In an evaluation using 115 diarrheal stool specimens and culture as the "gold standard," the assay showed a sensitivity of 85% and a specificity of 100%. An ICT11-based sandwich enzyme-linked immunosorbent assay showed a sensitivity of 100% and a specificity of 98% for direct detection of toxin from stool samples compared with those of culture. Thus, ICT11-based assays will be useful for screening for ETBF.

A year-long community-based study of diarrhoeal diseases was conducted in Canto Grande, a periurban community in Lima, Peru. In 109 (34%) houses out of 323 that were visited, at least one individual was detected with shigellosis. The frequency of the 161 shigella isolates obtained was as follows: 117 S. flexneri (73%), 21 S. boydii (13%), 15 S. dysenteriae (9%), and 8 S. sonnei (5%). Using a non-radioactive ipaH gene probe as a molecular epidemiological tool, a total of 41 S. flexneri strains were shown to be distributed in 25 intra-family comparisons by pairs (icp). Further subdivision, based on a comparison of the serotype, plasmid profile, antibiotic resistances and ipaH hybridization patterns indicated that Group I, with 11 icp (44%), had strains that were identical, Group II with 8 icp (32%), had strains that were different and Group III with 6 icp (24%), had strains with the same serotype and identical ipaH profiles but with differences in other markers. This data indicates that a diversity of shigella clones circulated in this community resulting from both clonal spread and horizontal transfer of genetic elements. Furthermore, ipaH profiling of isolates can be used not only to differentiate between closely related shigella strains but also with other parameters, help to understand the dynamics of the generation of new clones of pathogenic bacteria.

A monoclonal antibody-based test, Bengal SMART, was developed for rapid detection of Vibrio cholerae O139 synonym Bengal directly from stool specimens. The test, which takes about 15 min to complete, was used to screen 189 diarrheal stool specimens. The results were compared with those of a monoclonal antibody-based coagglutination test (COAT) and the conventional culture methods used as the "gold standard" for detection of V. cholerae O139. The Bengal SMART test showed a sensitivity of 100% and a specificity of 97% in comparison with the gold standard. It also fared better than COAT, which had a sensitivity of 96% for rapid detection of V. cholerae O139 synonym Bengal. These results show that Bengal SMART is suitable for use in field settings for rapid diagnosis of cholera caused by V. cholerae O139.

Mucinase is a soluble haemagglutinin protease, which may be important for the survival of Vibrio cholerae in association with mucilaginous blue-green algae (cyanobacteria). A comparative survival study was carried out with an Anabaena sp. and a wild-type V. cholerae O1 strain hap+ gene (haemagglutinin-protease), together with its isogenic mutant hap (hap-deleted gene). A simple spread plate technique was followed to count culturable V. cholerae O1 on taurocholate tellurite gelatin agar plate. The fluorescent antibody technique of Kogure et al. (1979) was used for the microscopical viable count of V. cholerae O1. Polymerase chain reaction (PCR) and Southern blot hybridization were carried out to detect a lower number of viable but nonculturable (VBNC) V. cholerae O1 from the laboratory-based experiments. The wild and mutant V. cholerae O1 strains survived in culturable form for 22 and 10 days, respectively, in association with the Anabaena sp., with the difference being statistically significant (P < 0.01). The fluorescent antibody technique, PCR, and hybridization results also showed that the wild strain survived better in the VBNC state than did the mutant VBNC strain in association with an Anabaena sp. These results indicate that the enzyme mucinase may play an important role in the association and long-term survival of V. cholerae O1 with a mucilaginous blue-green alga, Anabaena sp.Key words: Vibrio cholerae, Anabaena, mucinase, reservoir.

The bacteria that cause cholera are known to be normal inhabitants of surface water, however, the environmental risk factors for different biotypes of cholera are not well understood. This study identifies environmental risk factors for cholera in an endemic area of Bangladesh using a geographic information systems (GIS) approach. The study data were collected from a longitudinal health and demographic surveillance system and the data were integrated within a geographic information system database of the research area. Two study periods were chosen because they had different dominant biotypes of the disease. From 1992 to 1996 El Tor cholera was dominant and from 1983 to 1987 classical cholera was dominant. The study found the same three risk factors for the two biotypes of cholera including proximity to surface water, high population density, and poor educational level. The GIS database was used to measure the risk factors and spatial filtering techniques were employed. These robust spatial methods are offered as an example for future epidemiological research efforts that define environmental risk factors for infectious diseases.

Vibrio cholerae are known to be normal inhabitants of surface water. However, the environmental niches of the different strains of cholera are not well known, and therefore, populations at risk for cholera outbreaks cannot be clearly identified.This study identifies environmental risk factors for cholera caused by V. cholerae O1 El Tor and O139 and environmental niches of the two strains present in Matlab, a cholera endemic area of Bangladesh. The study year was 1993, the year that the O139 strain first appeared in the study area. Patients who had either strain of cholera identified in a laboratory were included in the study. A geographic information system was used to map the household locations of the patients, to describe the human sanitary environment and population density, and to address potential anthropogenic and environmental risk factors of the disease. Spatial point pattern and exploratory spatial data analysis techniques were used to define the environmental niches of the two cholera strains.The study suggests the niches of O1 El Tor and O139 strains of V. cholerae appear to be similar, based on common environmental risk factors.The results of this study support a theory that O1 El Tor could possibly be replaced by the newer O139 strain in the future.

Infections due to non-typhoid Salmonella, resistant to antibiotics, have recently emerged as an important health problem worldwide. Antibiotic resistance was studied by the disc-diffusion method among 3,876 (2.78%) non-typhoid Salmonella isolates cultured from 139,279 faecal samples in a diarrhoea treatment centre in Dhaka, Bangladesh, during 1989-1996. Of 499 salmonellae isolated in 1989, serogroup C (1.12%) was the most common, followed by Salmonella Typhi (0.72%) and serogroup B (0.71%). Isolation rate of serogroup B increased significantly to 2.18% (p0.01) in 1992 compared to 0.56% in 1991, 2.86% in 1995, and 2.48% in 1996. Serotyping of 194 serogroup B isolates revealed Salmonella Typhimurium (52%) and Salmonella Gloucester (45%) as predominant serotypes. Resistance to ampicillin (A), chloramphenicol (C), and trimethoprim-sulphamethoxazole (Sxt) (R type-ACSxt) increased to 89-100% during 1992-1996 from 20-28% during 1989-1991 (p0.01) among S. Typhimurium and S. Gloucester isolates. In 1993, 8-10% of the strains of both the serotypes, resistant to ampicillin, chloramphenicol, and trimethoprim-sulphamethoxazole, acquired resistance to ceftriaxone (Cr) (R type-ACSxtCr), which increased to 85-92% in 1996 (p0.01). All were susceptible to ciprofloxacin. A 157-kb conjugative plasmid transferred R type-ACSxt from both the serotypes to Escherichia coli K-12. The findings of the study suggest the emergence of multidrug-resistant S. Gloucester and S. Typhimurium for the first time as a significant health problem in Bangladesh, and surveillance is essential to monitor the resistant non-typhoid Salmonella and identify its sources and modes of transmission.

Studies have shown that various objects, such as utensils, toys, and clothes, can serve as vehicles for transmission of Shigella spp. Shigellae can become viable but non-culturable (VBNC) when exposed to various environmental conditions as shown in earlier studies. The present study was carried out to detect VBNC Shigella dysenteriae type 1 on various fomites by direct viable counting, polymerase chain reaction (PCR), and fluorescent antibody methods. S. dysenteriae type 1 was inoculated onto cloth, wood, plastic, aluminum, and glass objects. Results showed that 1.5-4.0 hours after inoculation, S. dysenteriae type 1 became non-culturable, and after five days, non-culturable but viable S. dysenteriae type 1 could be detected by both PCR and fluorescent antibody techniques. Fomites can be considered an important potential route of transmission of VBNC S. dysenteriae type 1 and a significant factor in the epidemiology of shigellosis.

A controlled, randomized, double-blind study in Bangladeshi children (ages 4-36 mo) with acute diarrhoea was undertaken to determine whether bismuth subsalicylate (BSS) would prevent the development of persistent diarrhoea (PD) in young children. The children were randomized to two groups: 226 were given liquid oral BSS, (as Pepto-Bismol), 100 mg/kg/d for 5 d; 225 were given placebo of identical appearance. On admission to the study, the two groups were comparable both clinically and microbiologically. Rotavirus was found in 56% of all the children, and enterotoxigenic E. coli in 31% of a subsample studied. Children treated with BSS had less severe and less prolonged illness than those treated with placebo (p = 0.057). There was, however, no difference in the development of PD between the two groups (8% and 11%). Unexpectedly, patients treated with BSS gained significantly more weight (2.3%) than those treated with placebo (0.5%; p0.001) during the course of the study. No toxicity of BSS was detected.Treatment with BSS had a modest therapeutic effect on acute diarrhoea, as has been previously demonstrated, but with no suggestion of a therapeutic effect on the prevention of persistent diarrhoea in this group of patients.

Aeromonads are causative agents of a number of human infections. Even though aeromonads have been isolated from patients suffering from diarrhea, their etiological role in gastroenteritis is unclear. In spite of a number of virulence factors produced by Aeromonas species, their association with diarrhea has not been clearly linked. Recently, we have characterized a heat-labile cytotonic enterotoxin (Alt), a heat-stable cytotonic enterotoxin (Ast), and a cytotoxic enterotoxin (Act) from a diarrheal isolate of Aeromonas hydrophila . Alt and Ast are novel enterotoxins which are not related to cholera toxin; Act is aerolysin related and has hemolytic, cytotoxic, and enterotoxic activities. We studied the distribution of the alt , ast , and act enterotoxin genes in 115 of 125 aeromonads isolated from 1,735 children with diarrhea, in all 27 aeromonads isolated from 830 control children ( P = 7 × 10 −4 for comparison of rates of isolation of aeromonads from cases versus those from controls), and in 120 randomly selected aeromonads from different components of surface water in Bangladesh. Aeromonas isolates which were positive only for the presence of the alt gene had similar distributions in the three sources; the number of isolates positive only for the presence of the ast gene was significantly higher for the environmental samples than for samples from diarrheal children; and isolates positive only for the presence of the act gene were not found in any of the three sources. Importantly, the number of isolates positive for both the alt and ast genes was significantly higher for diarrheal children than for control children and the environment. Thus, this is the first study to indicate that the products of both the alt and ast genes may synergistically act to induce severe diarrhea. In 26 patients, Aeromonas spp. were isolated as the sole enteropathogen. Analysis of clinical data from 11 of these patients suggested that isolates positive for both the alt and ast genes were associated with watery diarrhea but that isolates positive only for the alt gene were associated with loose stools. Most of the isolates from the three sources could be classified into seven phenospecies and eight hybridization groups. For the first time, Aeromonas eucrenophila was isolated from two children, one with diarrhea and another without diarrhea.

We have compared the B cell responses evoked in Bangladeshi, adults (n=11, median age 25 years) and children (n=21, median age 4.5 years), 7 days after intake of each of two doses of an oral, inactivated enterotoxigenic Escherichia coli (ETEC) vaccine composed of formalin-killed ETEC strains expressing the colonization factors, CFA/I, CFA/II and CFA/IV together with 1 mg of recombinant cholera toxin B-subunit (rCTB). The vaccine was well tolerated and only gave rise to negligible side effects. Peak antibody-secreting cell (ASC) response of the IgA isotype were seen 7 days after the first dose of the vaccine. The ASC responses to the different colonization factors (CFs) increased from a 29- to 46-fold (responder frequency 90-100%) in the adults and 13- to 24-fold (responder frequency 67-90%) in the children. The IgA-ASC response to rCTB also peaked after the first dose in the adults (426-fold, responder frequency 100%) and the children (46-fold, responder frequency 95%). Increased IgA antibody levels against CFA/I as well as IgA and IgG antibody levels to rCTB were seen in plasma after immunisation. About 86% of the children and 80% of the adults responded with faecal antibodies to rCTB, whereas about 67% of both groups responded to CFA/I. These results show that a single dose of the ETEC vaccine may elicit significant mucosal immune responses in both children and adults residing in an ETEC-endemic country such as Bangladesh.

It has been hypothesized that Vibrio cholerae is an autochthonous flora of the estuarine and brackish water environment. Zooplankton and phytoplankton have been considered as possible reservoirs. The present study was carried out in microcosms to confirm the role of a cyanobacterium, Anabaena sp., as a reservoir of V. cholerae O1 using culture, polymerase chain reaction (PCR) and immunoelectron microscopy. Survival of culturable V. cholerae in microcosms was monitored by using tellurite taurocholate gelatin agar. Culturable V. cholerae were detected for up to 1 h in association with Anabaena sp. from a microcosm. However, viable but nonculturable (VBNC) V. cholerae O1 were detected for up to 25 months using PCR and immunoelectron microscopy. Results also showed that VBNC V. cholerae can multiply and maintain their progeny in the mucilaginous sheath of Anabaena sp. This is the first time that PCR and immunoelectron microscopy have been used to detect nonculturable V. cholerae in association with Anabaena sp. This study further clarifies the role of Anabaena sp. as a possible reservoir of cholera.

The need for further studies on relationships between deaths and environmental variables has been reported in the literature. This case-control study was, therefore, carried out to find out the associations between several social and environmental variables and deaths of children due to infectious diseases such as those leading to diarrhoea, acute respiratory infection, measles and other diseases. Six hundred and twenty-five deaths (cases) and an equal number of matched living children (controls) aged 1-59 months, were studied in rural Matlab. An analysis of crude and adjusted odds ratio showed differential associations. Sources of drinking water, amount of stored water, conditions of latrines, number of persons sleeping with the child and the type of cooking site were statistically significantly associated with deaths due to infectious diseases after controlling for breast feeding, immunization, and the family size. Significant associations were also observed between: (i) the sources of drinking water and deaths due to ARI, and (ii) conditions of latrines and deaths due to diarrhoeal diseases, after controlling for the confounding variables. Several other environmental factors also showed associations with these various death groups, but they were not statistically significant. The size of the samples in death groups (small) and the prevalence of more or less homogeneous environmental health conditions probably diminished the magnitude of the effects. The results of the study reconfirm the importance of environmental health intervention in child survival, irrespective of breast-feeding, immunization, and selected social variables.Findings are reported from a case-control study conducted to assess the associations between several social and environmental variables and deaths of children due to infectious diseases, such as those leading to diarrhea, acute respiratory infection (ARI), measles, and other diseases. 625 deaths (cases) and an equal number of matched living children (controls) aged 1-59 months were studied in rural Matlab. Sources of drinking water, the amount of stored water, latrine conditions, the number of persons sleeping with the child, and the type of cooking site were statistically significantly associated with deaths due to infectious diseases after controlling for breast-feeding, immunization, and family size. Significant associations were also observed between sources of drinking water and deaths due to ARI, and conditions of latrines and deaths due to diarrheal diseases, after controlling for confounding variables. No other statistically significant environmental factors were associated with these various death groups. Study results confirm the importance of environmental health interventions in child survival, irrespective of breast-feeding, immunization, and selected social variables.

Three hundred sixty-two Streptococcus pneumoniae strains were isolated from children under 5 years of age at Dhaka Shishu (Children) Hospital from 1993 to 1997. The strains were isolated from blood ( n = 105), CSF ( n = 164), ear swab ( n = 61), eye swab ( n = 20), and pus ( n = 12). Of the 362 isolates, 42 (11.6%) showed intermediate resistance (MIC, 2.0 μg/ml) to penicillin. Penicillin resistance exhibited a strong relationship with serotype 14; 47.8% of the penicillin-resistant strains belonged to this type. A remarkably high (64.1%) resistance to co-trimoxazole was observed, along with a significant increase during the time period studied; there was no relationship to capsular type. By way of contrast, penicillin resistance did not show any significant change during the study period. Resistance to chloramphenicol (2.2%) and erythromycin (1.1%) was rare. The high resistance to co-trimoxazole and its increasing trend demand elucidation of the clinical impact of pneumonia treatment by this antimicrobial and reconsideration of the World Health Organization recommendation for co-trimoxazole administration to children with community-acquired pneumonia at the health care worker level in Bangladesh.

A total of 14915 shigella isolates obtained in 1991–1996 from patients attending the Dhaka (urban) and Matlab (rural) treatment centres of the International Centre for Diarrhoeal Disease Research Bangladesh were examined for susceptibility to ampicillin co-trimoxazole nalidixic acid mecillinam and ciprofloxacin by a disc diffusion method. There were no ciprofloxacinresistant shigella isolates. The prevalence of resistance to ampicillin co-trimoxazole and nalidixic acid varied between isolates. It increased to similar degrees in isolates from both Matlab and Dhaka. However resistance to mecillinam was more prevalent among isolates from Matlab than from Dhaka. The increase in mecillinam-resistant shigellae in the community may have grave implications for the empirical treatment of shigellosis in Bangladesh and other developing countries. (authors)

Considerable numbers of measles cases occur below the target age for vaccination in the Indian sub-continent. The immunogenicity of measles vaccine in infancy is dependent on the rate of decay in maternal antibody since this antibody interferes with vaccine induced seroconversion. This study investigated maternal antibody decay in a rural population in Bangladesh and evaluated possible risk factors for early decay. Measles antibodies were assessed using both ELISA and Plaque Reduction Neutralization (PRN) test in 330 infant-mother pairs in a cross-sectional survey. PRN was more sensitive method than ELISA for determining antibody levels. Antibody levels decreased rapidly in infants with increasing age. By the age of 5 months, 67% (2842) infants had practically no protective antibody left (30 mIU ml−1 or below). Only 12% infants at 5 months of age, and 5% at 8 months, had levels greater than 120 mIU ml−1—stated to ‘protect’ children. Multiple regression showed that maternal age was the only variable associated with the level of antibody (maternal weight, height and MUAC were not associated), decreasing by 1.06 mIU ml−1 for each year of age (P = 0.002). Infant's antibody concentration decreased with age by an average 2 mIU ml−1 for every month of life (P

A community-based longitudinal study conducted in rural Bangladesh investigated the association between nutritional status, cell-mediated immune status and acute upper respiratory infections (URI). A total of 696 children aged 0-59 months was followed prospectively for 1 y yielding 183,865 child-days' observation. Trained field workers visited each child every 4th d and collected morbidity data on symptoms suggesting URI (cough, fever, nasal discharge) for the preceding 3 d by recall. On the day of visit they examined each child reporting cough and/or fever to record the temperature, presence of nasal discharge, rate of respiration and presence of chest indrawing. Anthropometry for all children was conducted monthly. Cell-mediated immune competence was assessed by a multiple antigen skin test at baseline and thereafter every 3 months. The incidence of URI was 5.3 episodes per child-year observed. Approximately three-quarters of the study children were below -2 Z-score weight for age and height for age, and a quarter below -2 Z-score weight for height. During different test periods 9-21% of the study children did not respond to any of the test antigens. In a regression model children < -2 Z-score for weight for height had 16% [odds ratio (OR) 1.16, 95% confidence interval (CI) 1.03-1.31, p = 0.01] higher risk of developing URI. Anergic children had 20% higher risk (OR 1.20, CI 1.05-1.38, p = 0.009) of URI than immunocompetent children. The study demonstrated that wasting and depressed cell-mediated immunity (CMI), but not stunting, were associated with the incidence of URI among rural Bangladeshi children.