Evaluation of the impact of Shigella virulence genes on the basis of clinical features observed in patients with shigellosis.

INTRODUCTION: Shigella continues to cause significant morbidity and mortality each year, mostly in under-five children living in developing countries. We investigated the association between Shigella virulence genes and shigellosis. METHODOLOGY: We randomly selected 61 S. flexneri strains isolated from patients in Bangladesh between 2009 and 2013, and evaluated the presence of 140 MDa large-virulence-plasmid (p140), and 22 virulence genes including ipaH, ial, toxin, and T3SS-related genes. RESULTS: We found p140 in 79% (n = 48) and ipaBCD in 90% (n = 55) strains, while seven strains were missing the p140. The prevalence of ial was 89%, ipgC and ipgE was 85%, and the prevalence for the remaining genes was < 85%. During the multivariate analysis, we found that instead of sen, the Shigella enterotoxin gene set along with several other virulence genes such as ipgA, icsB, ipgB1, spa15, and mxiC, were significantly influencing multiple clinical features relevant to shigellosis, including bloody stool, mucoid stool, and rectal straining. CONCLUSIONS: We believe our model will help to determine the actual disease burden by directly looking for the genetic material in clinically suggestive patients, especially when detecting the causative organisms by traditional means is difficult.

Characterization of a serologically atypical Shigella flexneri Z isolated from diarrheal patients in Bangladesh and a proposed serological scheme for Shigella flexneri.

BACKGROUND: Atypical Shigella flexneri Z variant, that agglutinate with E1037 group factor specific monoclonal antisera against Shigella flexneri IV-I but not with other group or type specific antisera, has continuously being isolated in Bangladesh since 1997. Later this serotype has been reported in Indonesia, China and Argentina. Despite being a provisional serotype, continuous isolation of these strains in diverse geographical regions implicated a great necessity to study the overall characteristics of these strains. Therefore, we extensively characterized S. flexneri Z strains using various phenotypic and molecular tools. METHOD: Of 3569 S. flexneri isolated between 1997 and 2015, 95 strains were identified as S. flexneri Z using a panel of polyvalent absorbed antisera and monoclonal antisera of S. flexneri (MASF). Of them, randomly selected 65 strains were molecular O-serotyped using multiplex PCR and characterized using different phenotypic and molecular techniques (i.e.biotyping, plasmid profile, virulence marker and PFGE) to determine relationship with other subserotypes of S. flexneri. RESULTS: All these atypical S. flexneri Z strains were agglutinated with MASF B and IV-I antisera. Concordantly, these strains were positive to opt-gene, responsible for MASF IV-I sero-positive phenotype. However, molecular O-serotyping of all 65 strains could not differentiate between Z and Yb giving similar amplification products (wzx1-5 and opt). Contrarily, MASF based serotypic scheme distinguished among Z and Yb as well as Ya. All these S. flexneri Z showed typical biochemical reaction of S. flexneri, harboured a 140 MDa virulence plasmid and virulence markers namely ipaH, ial, sen, sigA and sepA genes. Along with the virulence plasmid, small plasmids (2.6, 1.8 and 1.6 MDa) were present as core plasmid. Moreover, a middle ranged plasmid and a 4.0 MDa sized plasmid were observed in 65% and 20% strains, respectively. Analysis of PFGE on XbaI-digested chromosomal DNA of Bangladeshi strains showed that S. flexneri Z had a close relatedness with Ya and Yb but completely different than the strains of Xa, Xb, 2a and 2b. This observation was found to be unequivocal while the overall result of biotyping, plasmid profile, and virulence factors was compared. Therefore, we conclude that these atypical serotype Z isolated in Bangladesh had a clonal relationship with Ya and Yb of Bangladesh and the opt gene played an important role in serotypic switching among them. Current serotyping scheme of S. flexneri strains fails to place many such atypical strains (1c, 1c+6, 1d, type 4, and 4c) including S. flexneri Z isolated from different parts of the world. Therefore, an updated serotyping scheme for identification of subserotypes of S. flexneri has been proposed to avoid multiple naming of the same subserotype having similar agglutination pattern.

Clonality, virulence and antimicrobial resistance of enteroaggregative Escherichia coli from Mirzapur, Bangladesh.

PURPOSE: This study investigates the virulence and antimicrobial resistance in association with common clonal complexes (CCs) of enteroaggregative Escherichia coli (EAEC) isolated from Bangladesh. The aim was to determine whether specific CCs were more likely to be associated with putative virulence genes and/or antimicrobial resistance. METHODOLOGY: The presence of 15 virulence genes (by PCR) and susceptibility to 18 antibiotics were determined for 151 EAEC isolated from cases and controls during an intestinal infectious disease study carried out between 2007-2011 in the rural setting of Mirzapur, Bangladesh (Kotloff KL, Blackwelder WC, Nasrin D, Nataro JP, Farag TH et al.Clin Infect Dis 2012;55:S232-S245). These data were then analysed in the context of previously determined serotypes and clonal complexes defined by multi-locus sequence typing. RESULTS: Overall there was no association between the presence of virulence or antimicrobial resistance genes in isolates of EAEC from cases versus controls. However, when stratified by clonal complex (CC) one CC associated with cases harboured more virulence factors (CC40) and one CC harboured more resistance genes (CC38) than the average. There was no direct link between the virulence gene content and antibiotic resistance. Strains within a single CC had variable virulence and resistance gene content indicating independent and multiple gene acquisitions over time. CONCLUSION: In Bangladesh, there are multiple clonal complexes of EAEC harbouring a variety of virulence and resistance genes. The emergence of two of the most successful clones appeared to be linked to either increased virulence (CC40) or antimicrobial resistance (CC38), but increased resistance and virulence were not found in the same clonal complexes.

MICROBIAL CONTAMINATION OF STREET VENDED FOODS FROM A UNIVERSITY CAMPUS IN BANGLADESH.

The microbiological quality of street vended food samples from Dhaka, Bangladesh was evaluated. The objective of the study was to identify the presence of common pathogens (Escherichia coli, Shigella spp, Salmonella and Vibrio spp) and to describe the molecular characterization of E coli, a commonly found pathogen in various street foods. Fifty food samples were collected from fixed and mobile vendors from two sampling locations (Mohakhali and Aftabnagar) in Dhaka city, Bangladesh. The tested samples included deep fried and fried snacks; quick lunch items; pickles; fruit chutney; baked items; spicy, sour and hot snacks etc: Juices, tamarind water and plain drinking water were also tested. Sterile polythene bags were used for collecting 200 g of each category of samples. They were tested for the presence of microorganisms following conventional microbiological processes. Biochemical tests followed by serology were done for the confirmation of Shigella and Salmonella. Serological reaction was carried out for confirmation of Vibrio spp. DNA was isolated for the molecular characterization to detect the pathogenic E. coli by polymerase chain reaction (PCR). Out of 50 food samples, six (12%) were confirmed to contain different species of E. coli and Shigella. Molecular characterization of E. coli revealed that three samples were contaminated with enteroaggregative E. coli (EAEC) and one was contaminated with enterotoxigenic E. coli (ETEC). Shigellaflexneri X variant was detected in one food item and Shigella flexneri 2a was found in drinking water. All these enteric pathogens could be the potential cause for foodborne illnesses.

Species-wide whole genome sequencing reveals historical global spread and recent local persistence in Shigella flexneri.

Shigella flexneri is the most common cause of bacterial dysentery in low-income countries. Despite this, S. flexneri remains largely unexplored from a genomic standpoint and is still described using a vocabulary based on serotyping reactions developed over half-a-century ago. Here we combine whole genome sequencing with geographical and temporal data to examine the natural history of the species. Our analysis subdivides S. flexneri into seven phylogenetic groups (PGs); each containing two-or-more serotypes and characterised by distinct virulence gene complement and geographic range. Within the S. flexneri PGs we identify geographically restricted sub-lineages that appear to have persistently colonised regions for many decades to over 100 years. Although we found abundant evidence of antimicrobial resistance (AMR) determinant acquisition, our dataset shows no evidence of subsequent intercontinental spread of antimicrobial resistant strains. The pattern of colonisation and AMR gene acquisition suggest that S. flexneri has a distinct life-cycle involving local persistence.

Enteroaggregative Escherichia coli have evolved independently as distinct complexes within the E. coli population with varying ability to cause disease.

Enteroaggregative E. coli (EAEC) is an established diarrhoeagenic pathotype. The association with virulence gene content and ability to cause disease has been studied but little is known about the population structure of EAEC and how this pathotype evolved. Analysis by Multi Locus Sequence Typing of 564 EAEC isolates from cases and controls in Bangladesh, Nigeria and the UK spanning the past 29 years, revealed multiple successful lineages of EAEC. The population structure of EAEC indicates some clusters are statistically associated with disease or carriage, further highlighting the heterogeneous nature of this group of organisms. Different clusters have evolved independently as a result of both mutational and recombination events; the EAEC phenotype is distributed throughout the population of E. coli.

Genomic analysis of the emergence of 20th century epidemic dysentery.

BACKGROUND: Shigella dysenteriae type 1 (Sd1) causes recurrent epidemics of dysentery associated with high mortality in many regions of the world. Sd1 infects humans at very low infectious doses (10 CFU), and treatment is complicated by the rapid emergence of antibiotic resistant Sd1 strains. Sd1 is only detected in the context of human infections, and the circumstances under which epidemics emerge and regress remain unknown. RESULTS: Phylogenomic analyses of 56 isolates collected worldwide over the past 60 years indicate that the Sd1 clone responsible for the recent pandemics emerged at the turn of the 20th century, and that the two world wars likely played a pivotal role for its dissemination. Several lineages remain ubiquitous and their phylogeny indicates several recent intercontinental transfers. Our comparative genomics analysis reveals that isolates responsible for separate outbreaks, though closely related to one another, have independently accumulated antibiotic resistance genes, suggesting that there is little or no selection to retain these genes in-between outbreaks. The genomes appear to be subjected to genetic drift that affects a number of functions currently used by diagnostic tools to identify Sd1, which could lead to the potential failure of such tools. CONCLUSIONS: Taken together, the Sd1 population structure and pattern of evolution suggest a recent emergence and a possible human carrier state that could play an important role in the epidemic pattern of infections of this human-specific pathogen. This analysis highlights the important role of whole-genome sequencing in studying pathogens for which epidemiological or laboratory investigations are particularly challenging.

Plasmid-mediated sulfamethoxazole resistance encoded by the sul2 gene in the multidrug-resistant Shigella flexneri 2a isolated from patients with acute diarrhea in Dhaka, Bangladesh.

In this study, mechanisms of plasmid-mediated sulfamethoxazole resistances in the clinical strains of multi-drug resistant (MDR) Shigella flexneri 2a were elucidated for the first time in Bangladesh. From 2006 to 2011, a total of 200 S. flexneri 2a strains were randomly selected from the stock of the Enteric and Food Microbiology Laboratory of icddr,b. Antimicrobial susceptibility of the strains showed 73%, 98%, 93%, 58%, 98%, 64% and 4% resistance to trimethoprim-sulfamethoxazole, nalidixic acid, ampicillin, erythromycin, tetracycline, ciprofloxacin and ceftriaxone respectively. Plasmid profiling revealed heterogeneous patterns and interestingly, all the trimethoprim-sulfamethoxazole resistant (SXT(R)) strains yielded a distinct 4.3 MDa plasmid compared to that of the trimethoprim-sulfamethoxazole susceptible (SXT(S)) strains. Curing of this 4.3 MDa plasmid resulted in the susceptibility to sulfamethoxazole alone suggesting the involvement of this plasmid in the resistance of sulfamethoxazole. Moreover, PCR analysis showed the presence of sul2 gene in SXT(R) strains which is absent in SXT(S) strains as well as in the 4.3 MDa plasmid-cured derivatives, confirming the involvement of sul2 in the resistance of sulfamethoxazole. Furthermore, pulsed-field gel electrophoresis (PFGE) analysis revealed that both the SXT(R) and SXT(S) strains were clonal. This study will significantly contributes to the knowledge on acquired drug resistance of the mostly prevalent S. flexneri 2a and further warrants continuous monitoring of the prevalence and correlation of this resistance determinants amongst the clinical isolates of Shigella and other enteric pathogens around the world to provide effective clinical management of the disease.

Poultry as a possible source of non-typhoidal Salmonella enterica serovars in humans in Bangladesh.

We investigated Salmonella enterica isolates from human clinical cases of gastroenteritis to determine the distribution of non-typhoidal Salmonella serovars in the human population, and compared them to isolates originating from poultry by serotyping, phage typing, plasmid profiling, pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) to evaluate the potential role of poultry in human non-typhoidal salmonellosis in Bangladesh. Nine different serovars were identified among the human isolates of which Salmonella Paratyphi B var Java (S. Java), S. Kentucky, S. Enteritidis, S. Virchow and S. Weltevreden also were commonly isolated from poultry. The poultry isolates belonging to S. Java, S. Kentucky and S. Enteritidis were indistinguishable from human isolates or genetically closely related, based on PFGE profiles and MLST. S. Kentucky clone ST198 and S. Java clone ST43 both well-known cause of human infections were also isolated from poultry.

Etiological diversity of diarrhoeal disease in Bangladesh.

BACKGROUND: This study compared the diversity of common diarrhoeal pathogens and antimicrobial susceptibility in four hospitals in Bangladesh. METHODOLOGY: A total of 13,959 diarrhoea patients, comprising rural Mirzapur (2,820), rural Matlab (2,865), urban Dhaka (5,287) and urban Mirpur (2,987) were included under the diarrhoeal disease surveillance system of icddr,b during 2010-2011; stool specimens were tested for Shigella spp., Vibrio cholerae, enterotoxigenic Escherichia coli and rotavirus. RESULTS: Rotavirus was highest in Mirzapur (28%) followed by Dhaka (24%), Matlab (19%) and Mirpur (18%). Overall, Shigella was significantly more prevalent in rural sites (Mirzapur 13% and Matlab 7%), than in urban sites (Dhaka 3% and Mirpur 3%). Vibrio cholerae was more common in the urban sites of Dhaka (14%) and Mirpur (12%). 72% of Shigella isolates were susceptible to ciprofloxacin in Mirzapur, and 88% to mecillinam. In Dhaka, the figures for Shigella were 65% and 50%, in Matlab 65% and 85%, and in Mirpur 59% and 92% respectively. Susceptibility of Shigella to azithromycin and ceftriaxone in Dhaka was 74% and 95%, and in Mirpur 88% and 92% respectively.  Vibrio cholerae showed the highest resistance to trimethoprim-sulfamethoxazole (100% in Mirpur) and lowest resistance to ciprofloxacin (0% in Dhaka, Matlab and Mirpur) and azithromycin (30% in Dhaka to 7% in Mirzapur). Multidrug resistance (≥3 antibiotics) for Shigella were: Mirzapur (50%); Dhaka (36%); Matlab (23%) and Mirpur (37%); and for V. cholerae it was 26%, 37%, 49% and 23% respectively. CONCLUSION: The isolation rates and antimicrobial susceptibility of Shigella spp. and V. cholerae along with rotavirus differed significantly in certain geographical sites.

Treatment with phenylbutyrate in a pre-clinical trial reduces diarrhea due to enteropathogenic Escherichia coli: link to cathelicidin induction.

Treatment of shigellosis in rabbits with phenylbutyrate reduces clinical severity and counteracts down-regulation of cathelicidin (CAP-18) in the large intestinal epithelia. We aimed to further evaluate whether in a rabbit model of enteropathogenic Escherichia coli (EPEC) diarrhea, CAP-18 is down-regulated in the small intestine and if oral phenylbutyrate treatment affects CAP-18 expression, clinical recovery, shedding of EPEC in stool and virulence properties of the isolated colonies. EPEC-induced diarrhea down-regulated CAP-18 in the small intestinal epithelia as revealed by immunohistochemistry. Phenylbutyrate treatment reduced clinical illness, improved histological features of inflammation and up-regulated CAP-18 in the epithelia. Active CAP-18 peptide was also released in the stool as noted in Western blot analysis. Multiplex PCR analysis of total bacterial DNA in the stool showed absence of EPEC specific genes eae and bfpA. Treated rabbits shed rough strains still harboring eae and bfpA genes, which were less potent in binding to HeLa cells and induced delayed onset of diarrhea in new rabbits. In conclusion, EPEC-mediated down-regulation of CAP-18 in the small intestinal epithelia was restored by phenylbutyrate treatment. Upregulation of CAP-18 in the epithelia was accompanied by healing of the epithelial lining, reduced shedding and virulence of EPEC and recovery from diarrhea.

Changing emergence of Shigella sero-groups in Bangladesh: observation from four different diarrheal disease hospitals.

BACKGROUND: Shigellosis continues to be a public health challenge for developing countries, including Bangladesh. The aim of the study is to demonstrate recent changes in Shigella sero-groups and their geographical diversity. METHODS: Data were extracted from data archive of four diarrheal disease surveillance systems. A 2% sub sample from urban Dhaka Hospital (2008-2011; n = 10,650), and 10% from urban Mirpur Treatment Centre (2009-2011; n = 3,585), were enrolled systematically; whereas, all patients coming from the Health and Demographic Surveillance System area in rural Matlab (2008-2011; n = 6,399) and rural Mirzapur (2010-2011; n = 2,812) were included irrespective of age, sex, and disease severity. A fresh stool specimen was collected for identification of Shigella spp. Of them, 315 (3%) were positive for Shigella in Dhaka, 490 (8%) from Matlab, 109 (3%) from Mirpur and 369 (13%) from Mirzapur and considered as analyzable sample size. RESULTS: Among all Shigella isolates regardless of age, significant decreases in percentage of S. flexneri over time was observed in Mirpur (55→29%; p value of χ(2)-for trend = 0.019) and Mirzapur (59→47%; p = 0.025). A non-significant decrease was also seen in Dhaka (58→48%), while in Matlab there was a non-significant increase (73→81%). Similar patterns were observed among under-5 children at all sites. Emergence of S. sonnei was found in Dhaka (8→25%; p<0.001) and Mirpur (10→33%; p = 0.015), whereas it decreased in Mirzapur (32→23%; p = 0.056). The emergence of S. boydii was seen in all ages in Mirzapur [(3→28%; p<0.001); (3→27%; p<0.001)]. On the other hand, we saw non-significant percent reductions in S. boydii in Dhaka [overall (25→16%); under-5 (16→9%)]. Decreasing rates of Shigella dysenteriae were observed in Matlab, Mirpur and Mirzapur; whereas, in Dhaka it remained unchanged. CONCLUSION AND SIGNIFICANCE: Emergence of S. sonnei and S. boydii as important infectious diarrhea etiologies and variations in geographical diversity underscore the need for monitoring, with possible implications for vaccine development.

Occurrence and characterization of multidrug-resistant New Delhi metallo-ß-lactamase-1-producing bacteria isolated between 2003 and 2010 in Bangladesh.

The purpose of this study was to screen for reduced susceptibility against imipenem and the presence of the New Delhi metallo-ß-lactamase-1 (NDM-1) gene in a collection of Enterobacteriaceae (Escherichia coli, Shigella spp. and Klebsiella pneumoniae) from different surveillance studies between 2003 and 2010 at the International Centre for Diarrhoeal Disease Research, Bangladesh. None of the E. coli (n = 1789) and Shigella spp. (n = 90) isolated between 2009 and 2010 from stool samples was resistant or had intermediate susceptibility to imipenem. Among 127 extended-spectrum ß-lactamase-producing strains isolated during 2003-2009, three Klebsiella pneumoniae isolates (2.4 %) were resistant to imipenem and were positive for bla(NDM-1). All these NDM-1-producing strains were isolated in 2008 and were resistant to all antibiotics tested except for tigecycline and colistin. All three isolates were positive for bla(OXA-1) group, bla(CTX-M-1) group (bla(CTX-M-15)) and bla(SHV) genes, whilst two isolates were positive for 16S rRNA methylase (armA) and qnr (qnrB) genes. One isolate was positive for the bla(CMY) gene and one for the rmtB gene. The bla(NDM-1) gene was located on a conjugative plasmid of ~23-24 MDa. The PFGE patterns of the isolates were different from each other. This study highlights the occurrence of NDM-1-producing organisms in Bangladesh in 2008. The clonal diversity of the isolates and the transferability of bla(NDM-1) plasmids suggest a wider distribution of NDM-1-producing bacteria in Bangladesh.

Characteristics of Multidrug Resistant Shigella and Vibrio cholerae O1 Infections in Patients Treated at an Urban and a Rural Hospital in Bangladesh.

We determined the frequency of multidrug resistant (MDR) infections with Shigella spp. and Vibrio cholerae O1 at an urban (Dhaka) and rural (Matlab) hospital in Bangladesh. We also compared sociodemographic and clinical features of patients with MDR infections to those with antibiotic-susceptible infections at both sites. Analyses were conducted using surveillance data from the International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), for the years 2000-2012. Compared to patients with antibiotic-susceptible for Shigella infections, those in Dhaka with MDR shigellosis were more likely to experience diarrhea for >24 hours, while, in Matlab, they were more likely to stay inhospital >24 hours. For MDR shigellosis, Dhaka patients were more likely than those in Matlab to have dehydration, stool frequency >10/day, and diarrheal duration >24 hours. Patients with MDR Vibrio cholerae O1 infections in Dhaka were more likely than those in Matlab to experience dehydration and stool frequency >10/day. Thus, patients with MDR shigellosis and Vibrio cholerae O1 infection exhibited features suggesting more severe illness than those with antibiotic-susceptible infections. Moreover, Dhaka patients with MDR shigellosis and Vibrio cholerae O1 infections exhibited features indicating more severe illness than patients in Matlab.

Differential host immune responses to epidemic and endemic strains of Shigella dysenteriae type I.

Shigella dysenteriae type 1 causes devastating epidemics in developing countries with high case-fatality rates in all age-groups. The aim of the study was to compare host immune responses to epidemic (T2218) and endemic strains of S. dysenteriae type 1. Shigellacidal activity of serum from rabbits immunized with epidemic or endemic strains, S. dysenteriae type 1-infected patients, and healthy adult controls from Shigella-endemic and non-endemic regions was measured. Immunogenic cross-reactivity of antibodies against Shigella antigens was evaluated by Western blot analysis. Oxidative burst and phagocytic responses of monocytes and neutrophils to selected S. dysenteriae type 1 strains were assessed by flow cytometry. Rabbit antisera against epidemic strain were less effective in killing heterologous bacteria compared to endemic antisera (p=0.0002). Patients showed an increased serum shigellacidal response after two weeks of onset of diarrhoea compared to the acute stage (3-4 days after onset) against their respective homologous strains; the response against T2218 and heterologous endemic S. dysenteriae type 1 strains was not significant. The serum shigellacidal response against all the S. dysenteriae type 1 strains was similar among healthy controls from endemic and non-endemic regions and was comparable with the acute stage response by patients. Compared to endemic strains of S. dysenteriae type 1, T2218 was significantly resistant to phagocytosis by both monocytes and neutrophils. No obvious differences were obtained in the induction of oxidative burst activity and cathelicidin-mediated killing. Cross-reactivity of antibody against antigens present in the epidemic and endemic strains showed some differences in protein/peptide complexity and intensity by Western blot analysis. In summary, epidemic T2218 strain was more resistant to antibody-mediated defenses, namely phagocytosis and shigellacidal activity, compared to endemic S. dysenteriae type 1 strains. Part of this variation may be attributed to the differential complexity of protein/peptide antigens.

Burden of typhoid and paratyphoid fever in a densely populated urban community, Dhaka, Bangladesh.

BACKGROUND: We conducted blood culture surveillance to estimate the incidence of typhoid and paratyphoid fever among urban slum residents in Dhaka, Bangladesh. METHODS: Between January 7, 2003 and January 6, 2004, participants were visited weekly to detect febrile illnesses. Blood cultures were obtained at the clinic from patients with fever (≥38°C). Salmonella isolates were assayed for antimicrobial susceptibility. RESULTS: Forty Salmonella Typhi and eight Salmonella Paratyphi A were isolated from 961 blood cultures. The incidence of typhoid fever was 2.0 episodes/1000 person-years, with a higher incidence in children aged<5 years (10.5/1000 person-years) than in older persons (0.9/1000 person-years) (relative risk=12, 95% confidence interval (CI) 6.3-22.6). The incidence of paratyphoid fever was 0.4/1000 person-years without variation by age group. Sixteen S. Typhi isolates were multidrug-resistant (MDR). All S. Paratyphi isolates were pan-susceptible. The duration of fever among patients with an MDR S. Typhi infection was longer than among patients with non-MDR S. Typhi (16±8 vs. 11±4 days, p=0.02) and S. Paratyphi (10±2 days, p=0.04) infections. CONCLUSIONS: Typhoid fever is more common than paratyphoid fever in the urban Bangladeshi slum; children<5 years old have the highest incidence. Multidrug resistance is common in S. Typhi isolates and is associated with prolonged illness. Strategies for typhoid fever prevention in children aged<5 years in Bangladesh, including immunization, are needed.

Variation of toxigenic Vibrio cholerae O1 in the aquatic environment of Bangladesh and its correlation with the clinical strains.

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.