Kegg Pathway: Cholera - Diarrhea

Appl Environ Microbiol. 2008 Jul 18;
Bag PK, Bhowmik P, Hajra TK, Ramamurthy T, Sarkar P, Majumder M, Chowdhury G, Das SC

V. Cholerae non-O1, non-O139 was isolated from natural surface waters from different sites sampled in Diarrhea endemic zones in Kolkata, India. Twenty-one of these isolates were randomly selected and included for the characterization. The multi serogroup isolates were compared by their virulence traits with a group of clinical non-O1, non-O139 isolates from the same geographic area. Of the 21 environmental isolates, 6 and 14 strains belonged to Heiberg groups I and II respectively. Three of the environmental isolates showed resistance to O/129. All of the non-O1, non-O139 strains were positive for toxR, and except one environmental isolate, none of them were positive for tcpA in the PCR assay. None of the isolates were positive for genes encoding Cholera toxin (ctxA), heat-stable toxin (est), heat-labile toxin (elt), and shiga toxin variants (stx) of Escherichia coli. Additionally, except for one environmental isolate (PC32), all were positive for the gene encoding El Tor hemolysin (hly). Culture supernatants of 86% (18 of 21) of environmental isolates showed a distinct cytotoxic effect to HeLa cells and some of these strains also produced cell rounding factor. Lipase, protease and cell-associated hemagglutination activities, and serum resistance properties of the environmental and clinical isolates did not differ much. However, 7 environmental isolates exhibited very high hemolytic activity (80 to 100%), while none of the clinical strains belonged to this group. Environmental isolates manifested three adherence patterns namely carpet-like, diffuse and aggregative adherence, and clinical isolates showed diffuse adherence on HeLa cells. Of the 11 environmental isolates tested for their enteropathogenic potential, eight (73%) induced positive fluid accumulation (FA >/=100) in mouse model and the reactivities of these isolates were comparable to those of clinical strains of non-O1, non-O139 and toxigenic O139. Comparison in counts of the colonized environmental and clinical strains in mouse intestine showed that the organism of both groups had similar colonizing efficiency. These findings indicate the presence of potentially pathogenic V. Cholerae non-O1, non-O139 strains in surface waters of the studied sites in Kolkata.

Ren Fail. 2008; 30(4): 469-73
Winstanley V, Little MA, Wadsworth C, Cohen P, Martin NM

The McKittrick-Wheelock syndrome is characterized by severe electrolyte and fluid depletion as a result of rectal tumor hypersecretion. Typically, a metabolic acidosis ensues. We report the case of a 58-year-old man who presented with a mixed metabolic acidosis and alkalosis. He was hyponatremic, hypokalemic, and hypochloremic, with acute renal failure on blood testing. Following fluid resuscitation, a predominant alkalemia was observed. The patient was found to be passing 1.5 L of mucous per rectum per day, containing high concentrations of sodium and potassium, similar to that observed in Cholera stool. A large rectal villous adenoma was discovered on sigmoidoscopy, and definitive management was achieved by removal of the tumor. This case provides a demonstration of the ranging metabolic disturbance associated with secretory Diarrhea. Other endogenous and infective causes are discussed, and mechanisms compared with the case described.

Expert Rev Vaccines. 2008 Jul; 7(5): 561-7
Jelinek T, Kollaritsch H

There is currently only one vaccine available that provides protection against Diarrhea caused by Vibrio Cholerae and, to a lesser degree, enterotoxigenic Escherichia coli (ETEC). Adverse events of this oral whole-cell/recombinant B-subunit vaccine have been negligible. Protective efficacy against Cholera is 85%, while protection against the heat-labile toxin of ETEC reaches 67%. There is still a need for data on protection of Western travelers against travelers' Diarrhea in general by Dukoral vaccination. However, current studies show a protective effect of up to 43%. Although the vaccine is only licensed for vaccination against Cholera in most Western countries, there is mounting evidence that the oral Cholera vaccine is a valuable option to those traveling to high-risk endemic areas. Vaccination against Cholera and ETEC should be recommended for at-risk travelers, in particular those with high exposure at their travel destination or high personal risks through fluid loss.

Cell Biol Toxicol. 2008 Jun 19;
Saha S, Chowdhury P, Mazumdar A, Pal A, Das P, Chakrabarti MK

The heat-stable enterotoxin (Y-STa) produced by the pathogenic strains of Yersinia enterocolitica is a causative agent of secretory Diarrhea. We have reported earlier that Y-STa-induced inositol trisphosphate-mediated cytosolic calcium rise occurs in rat intestinal epithelial cells. In the present communication, the involvement of a nuclear calcium store in the action mechanism of Y-STa in rat intestinal epithelial cells has been shown. Calcium imaging with time series confocal microscopy shows that Y-STa stimulates both the nuclear and cytosolic calcium levels in rat intestinal epithelial cells where a rise in nuclear calcium precedes the cytosolic events. Moreover, Y-STa stimulates both cytosolic and nuclear inositol trisphosphate (IP(3)) levels in a time-dependent manner. Western blot and immunocytochemical analysis reveal a higher density of IP(3) receptor type II in the nuclear membrane compared to the cytosol, which may be the cause of an early rise of the nuclear calcium level. Therefore, it is suggested that Y-STa regulates the nuclear and cytosolic calcium signals in a distinct temporal manner in rat intestinal epithelial cells.

Virology. 2008 Jul 20; 377(1): 117-23
Nayak MK, Balasubramanian G, Sahoo GC, Bhattacharya R, Vinje J, Kobayashi N, Sarkar MC, Bhattacharya MK, Krishnan T

Mutation and recombination are recognized as important driving forces of evolution among RNA viruses. An intergenogroup recombinant norovirus strain [Hu/Kol/NLV/L8775/AB290150/2006/India] was detected in the faecal specimen of a 17 year old male, who had suffered from acute watery Diarrhea and severe dehydration. Sequence analysis confirmed that this novel recombinant strain had a polymerase gene fragment that closely resembled a Norovirus (NoV) genogroup-I genotype-3 virus (HuCV/NLV/GI.3/VA98115/AY038598/1998/USA) and a capsid gene resembling NoV genogroup-II genotype-4 virus (NoV/Hu/GII.4/Terneuzen70/EF126964/2006/NL). The crossing over and recombination was observed at nucleotide (nt) 790 of NoV GI VA98115 strain and nt808 of NoV GII Terneuzen70 strain. In both parent strains conserved nucleotide sequence and hairpin structure (DNA secondary structure) were reported at the junction point of ORF1 and ORF2, exhibiting the mechanism of recombination in these viruses. Thus this novel recombinant NoV is another step in evolution among NoVs, indicating that constant surveillance is important to successfully monitor emergence of these strains.

Wkly Epidemiol Rec. 2008 May 2; 83(18): 157-8

Gastroenterology. 2008 Jul; 135(1): 185-193.e1
Lu L, Bao Y, Khan A, Goldstein AM, Newburg DS, Quaroni A, Brown D, Walker WA

BACKGROUND & AIMS: Diarrheal disease is a major cause of morbidity and mortality in infants and children worldwide. Evidence has indicated immature human enterocytes and their interaction with bacteria and enterotoxins may account for the noted increased susceptibility of neonates to Diarrhea. Our aim was to characterize the developmental difference in Cholera toxin (CT)-GM1-mediated endocytosis. METHODS: We used H4 cells (a fetal human small intestinal epithelial cell line), T84 cells, primary cultured mature human small intestinal epithelial cells, and human fetal small intestine xenografts. In addition, hydrocortisone was used as a potent intestinal trophic factor to induce maturation of the human enterocytes. RESULTS: Here we show an increase in CT-caveolae and a decrease in CT-clathrin colocalization in H4/hydrocortisone compared with H4 cells by electron microscopy. In T84 and freshly isolated human small intestinal epithelial cells, a significant amount of GM1 was partitioned into the lipid rafts. In contrast, there was little CT-GM1/lipid raft association in H4 cells. However, hydrocortisone significantly increased GM1/lipid raft association in H4 cells. Furthermore, we noted an increase in the level of phosphatidylcholine, sphingomyelin, and the ratio of phosphatidylcholine/phosphatidylinositol in mature compared with immature enterocytes and that hydrocortisone can accelerate this maturational process. Disruption of phosphatidylinositol transfer protein alpha using small interference RNA showed an increase in GM1/lipid raft association in H4 cells and resulted in a decreased CT response. CONCLUSIONS: Our studies suggest that the developmental change in CT endocytosis is partially caused by an increased GM1-lipid raft association through a maturational change of phospholipid composition on the cell surface of immature enterocytes.

Trop Doct. 2008 Apr; 38(2): 105-7
Rajeshwari K, Gupta A, Dubey AP, Uppal B, Singh MM

V. Cholerae O1 Eltor serotype Ogawa has been causing most of the Cholera outbreaks in India till recently. However this communication reports the occurrence of Vibrio Cholerae O1 Inaba in Delhi in 2005, as a predominant causative organism of Cholera in children. All strains isolated were sensitive to gentamicin and a high level of resistance towards nalidixic acid and amoxicillin was seen. There was no case fatality.

Tidsskr Nor Laegeforen. 2008 Mar 13; 128(6): 724
Riska O

Zhonghua Yi Xue Za Zhi. 2008 Jan 22; 88(4): 265-7
Chen MF, Gao Y, Cong X, Sun CL, Zhu JY, Xi M, Guo XL, Yang X, Li Y, Wei L

OBJECTIVE: To understand the infection and epidemiology of norovirus and rotavirus and enteral adenovirus among adult with sporadic viral gastroenteritis in Beijing and provide theoretical basis for clinical prevention and control. METHODS: Stool specimens were collected from all 312 sporadic outpatient among adult with non-Cholera watery Diarrhea in Infectious Disease Department of Peking University People's Hospital 2005-2006. PAGE were used for detection of rotavirus RNA in stool specimens; R-Biopharm RIDASCREEN norovirus and RIDASCREEN adenovirus were used for detection of norovirus and adenovirus. RESULTS: Rotavirus RNA was not present in all 312 stool specimens; Norovirus was present in 17.6% (22/125) and in 32.4% (11/34) in October; Adenovirus was present in 3.3% (3/92); Mixed infections of norovirus and adenovirus was present in 1 stool specimen. CONCLUSIONS: Norovirus is more common etiologic agents of sporadic acute viral gastroenteritis among adult in Beijing, The infection peak of norovirus is in autumn. Maybe the infection of rotavirus is few.

Transplantation. 2008 Feb 27; 85(4): 656
Sundaram M, Veeraraghavan B, John GT

JNMA J Nepal Med Assoc. 2007 Oct-Dec; 46(168): 175-9
Karki A, Tiwari BR

This retrospective study was conducted during January to September in the year 1997. Three hundred and forty nine stool samples were collected from diarrhoea patients from different places of Kathmandu valley and examined at National Public Health Laboratory (NPHL), Teku, Kathmandu. Acute diarrhoea becomes epidemic in rainy season and is a major public health problem of the city. In this study, people with poor hygiene practice and poor education were infected more than other people. Among the 349 patients with the gastrointestinal disease, 26.0% were found to have bacterial infection. Out of which, 88 (25.1%), one (0.28%), one (0.28%), and one (0.28%) were found to be Vibrio Cholerae 01, Vibrio Cholerae 0139, Shigella dysenteriae and Escherichia coli respectively. Cholera cases were found almost throughout the year in the city though the numbers increased during the rainy season. It was highest during July (34.6%) followed by August (32.35%), September 32% and June (6.89%). The uncommon species of Vibrio i.e. Vibrio Cholerae 0139 was also found in the study. Higher prevalence was found in urban areas (83.52%) than in rural areas (16.48%). Antimicrobial susceptibility testing of bacterial isolates showed that Ciprofloxacin (97.85%) was found to be the most effective antibiotic followed by Tetracycline (92.34%), Erythromycin (92.34%), Norfloxacin (93.34%), Cholramphenicol, Ampicillim, but Cotrimoxazole were found to be resistant to all isolated Vibrio Cholerae.

J Health Popul Nutr. 2007 Sep; 25(3): 278-84
Gutiérrez C, Villa S, Mota FR, Calva JJ

This study assessed whether an oral rehydration solution (ORS) in which glucose is replaced by L-glutamine (L-glutamine ORS) is more effective than the standard glucose-based rehydration solution recommended by the World Health Organization (WHO-ORS) in reducing the stool volume and time to rehydrate in acute diarrhoea. In a double-blind, randomized controlled trial in a Mexican hospital, 147 dehydrated children, aged 1-60 month(s), were assigned either to the WHO-ORS (74 children), or to the L-glutamine ORS (73 children) and followed until successful rehydration. There were no significant differences between the groups in stool output during the first four hours, time to successful rehydration, volume of ORS required for rehydration, urinary output, and vomiting. This was independent of rotavirus-associated infection. An L-glutamine-containing glucose-free ORS seems not to offer greater clinical benefit than the standard WHO-ORS in mildly-to-moderately-dehydrated children with acute non-Cholera diarrhoea.

PLoS ONE. 2008; 3(2): e1587
Ramakrishna BS, Subramanian V, Mohan V, Sebastian BK, Young GP, Farthing MJ, Binder HJ

BACKGROUND: Reduction of gross Diarrhea rate in excess of that seen over time with intravenous therapy and appropriate antibiotics is not usually achieved by oral glucose-electrolyte rehydration therapy for Cholera and Cholera-like Diarrheas. METHODOLOGY AND PRINCIPAL FINDINGS: This prospective randomized clinical trial at a tertiary referral hospital in southern India was undertaken to determine whether amylase resistant starch, substituting for glucose in hypo-osmolar oral rehydration solution, would reduce Diarrhea duration and weight in adults with acute severe dehydrating Diarrhea. 50 adult males with severe watery Diarrhea of less than three days' duration and moderate to severe dehydration were randomized to receive hypo-osmolar ORS (HO-ORS) or HO-ORS in which amylase resistant high amylose maize starch 50g/L substituted for glucose (HAMS-ORS). All remaining therapy followed standard protocol. Duration of Diarrhea (ORS commencement to first formed stool) in hours was significantly shorter with HAMS-ORS (median 19, IQR 10-28) compared to HO-ORS (median 42, IQR 24-50) (Bonferroni adjusted P, P(adj)<0.001). Survival analysis (Kaplan-Meier) showed faster recovery from Diarrhea in the HAMS-ORS group (P<0.001, log rank test). Total Diarrhea fecal weight in grams (median, IQR) was not significantly lower in the HAMS-ORS group (2190, 1160-5635) compared to HO-ORS (5210, 2095-12190) (P(adj) = 0.08). However, stool weight at 13-24 hours (280, 0-965 vs. 1360, 405-2985) and 25-48 hours (0, 0-360 vs. 1080, 55-3485) were significantly lower in HAMS-ORS compared to HO-ORS group (P(adj) = 0.048 and P = 0.012, respectively). ORS intake after first 24 hours was lower in the HAMS-ORS group. Subgroup analysis of patients with culture isolates of Vibrio Cholerae indicated similar significant differences between the treatment groups. CONCLUSIONS: Compared to HO-ORS, HAMS-ORS reduced Diarrhea duration by 55% and significantly reduced fecal weight after the first 12 hours of ORS therapy in adults with Cholera-like Diarrhea. TRIAL REGISTRATION: Current Controlled Trials ISRCTN72841333.

Rev Soc Bras Med Trop. 2007 Nov-Dec; 40(6): 686-9
Filizola LR, Figueirôa AC, Araújo MC, Cavalcanti Vde O, Lima CM, Hofer E

The levels of vibriocidal antibodies were investigated among 41 adults without any past or present history of Diarrhea due to Vibrio Cholerae O1 who were living in the municipality of São Bento do Una, Pernambuco. A Diarrhea outbreak occurred in this locality at the beginning of 2004, involving multiple bacterial agents, including Vibrio Cholerae. The microtitration test was used to investigate the presence of anti-Ogawa and anti-Inaba vibriocidal serum antibodies. Vibriocidal titers e" 1:640 were considered indicative of infection by Vibrio Cholerae O1. The frequency of the reagents was 36 (87.8%) for the Ogawa serovar, which showed that Vibrio Cholerae O1 was possibly circulating during and/or after the Diarrhea epidemic.

J Ethnopharmacol. 2008 Feb 28; 116(1): 194-7
Mathabe MC, Hussein AA, Nikolova RV, Basson AE, Meyer JJ, Lall N

Spirostachys africana Sond. stem bark is used traditionally for the treatment of diarrhoea and dysentery in Limpopo Province of South Africa. Bioassay-guided fractionation of ethanolic extract from bark of Spirostachys africana led to the isolation of four known compounds, two triterpenoids, compound 1 [d-Friedoolean-14-en-oic acid (3-acetyl aleuritolic acid)] and compound 2 (Lupeol), and two diterpenes, compound 3 [ent-2,6alpha-dihydroxy-norbeyer-1,4,15-trien-3-one (diosphenol 2)] and compound 4 (ent-3beta-hydroxy-beyer-15-ene-2-one). Isolated compounds were tested for antibacterial activity using micro-dilution method. Compound 1, exhibited minimum inhibitory concentration (MIC) of 50 microg/ml against Staphylococcus aureus, Salmonella typhy, Vibrio Cholera, Escherichia coli and Shigella dysentery. Compound 2 was not active against all tested microorganisms at 200 microg/ml, which was the highest concentration tested. At this concentration, all four compounds were not active against Shigella sonnei. Cytotoxicity of ethanol crude extracts and isolated compounds from Spirostachys africana was determined using the sodium-2,3-bis-[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) assay on Vero cells. Compounds 2 and 3, isolated from Spirostachys africana, had up to three times higher [50% inhibitory concentration (IC(50) values; 300.9 and 308.9 microg/ml)] than the ethanol crude extracts (102.8 microg/ml) suggesting higher toxicity of the crude extract as compared to these two compounds. In contrast, compounds 1 and 4 were not cytotoxic to Vero cell lines (African green monkey) in vitro at the concentrations tested (IC(50)>400 microg/ml). This is the first report on the antibacterial activity and cytotoxicity of purified compounds from Spirostachys africana.

BMJ. 2008 Feb 2; 336(7638): 266-8
Roy SK, Hossain MJ, Khatun W, Chakraborty B, Chowdhury S, Begum A, Mah-e-Muneer S, Shafique S, Khanam M, Chowdhury R

OBJECTIVE: To investigate the impact of zinc supplementation in children with Cholera. DESIGN: Double blind, randomised, placebo controlled trial. SETTING: Dhaka Hospital, Bangladesh. PARTICIPANTS: 179 children aged 3-14 years with watery diarrhoea and stool dark field examination positive for Vibrio Cholerae and confirmed by stool culture. INTERVENTION: Children were randomised to receive 30 mg elemental zinc per day (n=90) or placebo (n=89) until recovery. All children received erythromycin suspension orally in a dose of 12.5 mg/kg every six hours for three days. MAIN OUTCOME MEASURES: Duration of diarrhoea and stool output. Results 82 children in each group completed the study. More patients in the zinc group than in the control group recovered by two days (49% v 32%, P=0.032) and by three days (81% v 68%, P=0.03). Zinc supplemented patients had 12% shorter duration of diarrhoea than control patients (64.1 v 72.8 h, P=0.028) and 11% less stool output (1.6 v 1.8 kg/day, P=0.039). CONCLUSION: Zinc supplementation significantly reduced the duration of diarrhoea and stool output in children with Cholera. Children with Cholera should be supplemented with zinc to reduce its duration and severity. TRIAL REGISTRATION: Clinical trials NCT00226616.

Structure. 2008 Jan; 16(1): 137-48
Li J, Lim MS, Li S, Brock M, Pique ME, Woods VL, Craig L

The bacterial pathogen Vibrio Cholerae uses toxin-coregulated pili (TCP) to colonize the human intestine, causing the severe Diarrheal disease Cholera. TCP are long, thin, flexible homopolymers of the TcpA subunit that self-associate to hold cells together in microcolonies and serve as the receptor for the Cholera toxin phage. To better understand TCP's roles in pathogenesis, we characterized its structure using hydrogen/deuterium exchange mass spectrometry and computational modeling. We show that the pilin subunits are held together by tight packing of the N-terminal alpha helices, but loose packing of the C-terminal globular domains leaves substantial gaps on the filament surface. These gaps expose a glycine-rich, amphipathic segment of the N-terminal alpha-helix, contradicting the consensus view that this region is buried in the filament core. Our results explain extreme filament flexibility, suggest a molecular basis for pilus-pilus interactions, and reveal a previously unrecognized therapeutic target for V. Cholerae and other enteric pathogens.

Pac Health Dialog. 2005 Sep; 12(2): 17-22
Kirk MD, Kiedrzynski T, Johnson E, Elymore A, Wainiqolo I

In April 2000, a large outbreak of Cholera due to Vibrio Cholerae serotype Ogawa biotype El Tor affected the Island of Pohnpei in the Federated States of Micronesia. A Pacific Public Health Surveillance Network team conducted a case control study in the middle of the epidemic. The aims of the study were to identify individual and household level risk factors for Cholera, and to evaluate public health interventions aimed at controlling the outbreak. A case was a person admitted to the Pohnpei hospital with acute watery diarrhoea in the months of June and July 2000. We used a pre-tested questionnaire to interview cases about exposures in the five days prior to illness and visited their house to collect water samples, observe hygiene, and assess uptake of health education. 100 mL water samples were filtered and cultured for V. Cholerae. We randomly selected neighbouring houses to identify a control that was similar age and sex for each case. Identical observations were made for eligible controls where their household members had not had diarrhoea since the beginning of the epidemic. We stored and analysed data using an Epi Info version 6.04. 53 case control pairs were enrolled into the study. The study identified that storing food outside uncovered, and having a pit latrine as the main toilet were risk factors for Cholera infection. There were also several factors that protected against Cholera infection, including washing hands after using the toilet and before eating, having a container to store safe water, the presence of soap in kitchen and bathroom, the presence of chlorine bleach and two or more hand washing buckets, a working refrigerator/ice box, and toilet inside or near the house and having a flush toilet. In multivariate analysis, having a working refrigerator/ice box (OR 0.19, 95% CI 0.05-0.70) and Clorox present in the house (OR 0.17, 95% CI 0.04-0.81) were strongly protective against illness. Only 13% (14/106) of case households reported disinfecting household water with chlorine. V. Cholerae was isolated from the household water supplies of two controls and one case. During outbreaks of diarrhoeal disease, public health agencies need to aggressively advise affected communities to: disinfect drinking water with clorox bleach, store water in narrow-necked containers, and prepare and store food safely. Health authorities should use multiple strategies inform people about preventive hygiene measures, and implement vaccine campaigns early in outbreaks of Cholera. Improvements in sanitation and hygiene are needed to prevent further Cholera epidemics in the Pacific.

Infect Immun. 2008 Mar; 76(3): 1282-8
Yoon SS, Mekalanos JJ

Vibrio Cholerae is a monoflagellated gram-negative bacterium that causes the severe Diarrheal disease Cholera. In contrast to Salmonella enterica serovar Typhimurium infection, which is accompanied by both acute Diarrhea and high-level inflammation, V. Cholerae infection is largely noninflammatory in human hosts. Bacterial flagella are composed of flagellin, a highly conserved protein that is also a target of the innate immune response. Because the V. Cholerae flagellum is covered by a sheath, we hypothesized that it might be less prone to activation of the innate immune response than nonsheathed flagella, such as those produced by Salmonella serovar Typhimurium. Indeed, compared with Salmonella serovar Typhimurium flagella, V. Cholerae flagella demonstrated significantly reduced NF-kappaB activation in A549 human pulmonary epithelial cells. However, V. Cholerae flagellin monomers, FlaD and FlaC, were almost equally potent with purified FliC, a monomer derived from Salmonella serovar Typhimurium flagella, in NF-kappaB activation. Heat- and acid-induced dissociation assays showed that Salmonella serovar Typhimurium flagella disassembled far more readily into monomeric flagellins than V. Cholerae flagella, suggesting that the differential levels of NF-kappaB activation by V. Cholerae and Salmonella serovar Typhimurium flagella are likely attributable to the difference in their flagellin shedding. Our results suggest that monomer dissociation of V. Cholerae flagella is suppressed likely due to the presence of the sheath and that this unique structural feature of V. Cholerae flagella may have evolved as a strategy to evade flagellin-triggered host innate immune responses in various host species.