Cholerae El Tor Research Articles

Resistance to high temperatures salmonellosis circulating in Kemerovo region

Anti-lactoferrin activity (AlfA) is one of the factors contributing to the persistence of V. cholerae in human body in formation of human vibriocarriers. It is shown that in the AlfA mechanism cholera vibrio lectins are participating as an initial link specifically binding to lactoferrin at the surface of microbial cells, providing for subsequent cleavage of this protein with the help of V. cholerae exoproteases. Lectins are proteins or glycoproteins of non-immune class, capable to reversibly bind to the carbohydrate fraction of glycoconjugates without breaking the covalent structure of any of the recognizable glycosyl ligands. As glycoproteins, lectins ensure the complementary highly specific carbohydrate-protein binding of the active sites of target cells (namely lactoferrin). It is proved that all V. cholerae El Tor and Bengal strains possess hexose-specific lectin receptors (especially mannose). Atoxigenic V. cholerae O1 and O139 strains possess, in addition to hexose receptors, lectins specific to aminosugars.

MLVA-typing of Vibrio choleraе El Tor Strains Isolated in Transbaikal Territory during the Seventh Pandemic

Cholera remains a topical infection in the world, which determines the high risk of importation of the pathogen on the territory of the Russian Federation. MLVA-typing of 135 strains at 5 tandem repeats loci was carried outin order to study the clonal-population structure of the isolated from 1973 to 2016 on the Transbaikal territory V. cholerae. Analysis of the distribution patterns showed that the primary emergence of strains with new atypical for the territory genotypes in 1970s, 1980s, 1990s, occurred in sewage waters with the subsequent detection of the given genotypes generally in Ingoda river and Kenon lake. The strains firstly isolated from Borzya river and Haranor lake and found out at various times in Ingoda river and Kenon lake, Chita, Argun rivers, Haranor lake and Borzya river have been dominating since 2005 in Transbaikal territory. Finding groups of isolates with identical or similar MLVA-profile appeared in cluster complexes evidence of their long-term preservation in specific ecological niches, strains’ allelic profiles undergo minor transformation on the most variable loci in the process of adaptation. Identification of V. cholerae El Tor that variable tandem repeat loci structure materially differs from those of the dominant clones may serve as proof of their invasive origin.

Cholera El Tor in infants below 2 years of age as seen in private and hospitalized patients.

A report was made on cholera El Tor in children under 2 years of age with mute gastroenteritis admitted to the pediatric ward of the Dr. Soetomo Hospital, Surabaya. Twenty-three out of 115 cases (20%) showed positive stool cultures. A similar report was made in the same period on patients with gastroenteritis in private practice. Four out of 108 cases (3.6%) showed positive stool cultures. A brief description of the disease pattern of infantile gastroenteritis was presented. The increasing incidence of cholera, below one year was also discussed.

The "Rose System" in Overcoming the Second Cholera Outbreak in Jakarta from May 25-30, 1976

1.a. During a short cholena outbreak in Jakarta frnom 25th May to 30th May, 1976, 70 children aged from 6 months to 13 years suffering from acute diarrhoea and dehydration, shock or pre-shock were admitted to the provisional Rehydnation Cetntre in the outpatient section of the Departmernt of Child Heatth, University of Indonesia/Dr. Cipto Mangunkusumo General Hospital, Jakarta.b. None died.c. 71.9% of the stool specimens from 32 patients revealed Vibrio Cholera El Tor Ogawa type.2. The methodology used was the so-called "ROSE" system :a. Rehydration with ringer's lactate solution.b. Oralyte in boiled water given ad libi·tum.c. Oralyte solution and intravenous solution given Simultaneously.d. Education to the parents on the importance of giving oralyte solution ad libitum.3. This paper dealt with the second success of the Rose system in facing a cholera outbreak.

Cellular mechanisms underlying the inhibitory effect of flufenamic acid on chloride secretion in human intestinal epithelial cells

Intestinal Cl− secretion is involved in the pathogenesis of secretory diarrheas including cholera. We recently demonstrated that flufenamic acid (FFA) suppressed Vibrio cholerae El Tor variant-induced intestinal fluid secretion via mechanisms involving AMPK activation and NF-κB-suppression. The present study aimed to investigate the effect of FFA on transepithelial Cl− secretion in human intestinal epithelial (T84) cells. FFA inhibited cAMP-dependent Cl− secretion in T84 cell monolayers with IC50 of ∼8 μM. Other fenamate drugs including tolfenamic acid, meclofenamic acid and mefenamic acid exhibited the same effect albeit with lower potency. FFA also inhibited activities of CFTR, a cAMP-activated apical Cl− channel, and KCNQ1/KCNE3, a cAMP-activated basolateral K+ channel. Mechanisms of CFTR inhibition by FFA did not involve activation of its negative regulators. Interestingly, FFA inhibited Ca2+-dependent Cl− secretion with IC50 of ∼10 μM. FFA inhibited activities of Ca2+-activated Cl− channels and KCa3.1, a Ca2+-activated basolateral K+ channels, but had no effect on activities of Na+–K+–Cl− cotransporters and Na+–K+ ATPases. These results indicate that FFA inhibits both cAMP and Ca2+-dependent Cl− secretion by suppressing activities of both apical Cl− channels and basolateral K+ channels. FFA and other fenamate drugs may be useful in the treatment of secretory diarrheas.

EFFECT OF ANTIBACTERIAL PREPARATIONS ON VIBRIO CHOLERAE EL TOR BIOFILMS

Study the effect of antibacterial preparations on biofilms of Vibrio cholerae El Tor. Sensitivity of V cholerae El Tor (6 strains) to various concentrations of antibacterial preparations (doxycycline,.tetracycline, levomycetin, rifampicin, gentamycin, ceftazidime) was determined (MD 4.2.2495-09). Transmission electron microscopy was used for visualization of the effect of preparations on biofilms. The values of minimal inhibiting concentrations of antibacterial preparations against biofilms have increased by 5 - 100 times compared with plankton cultures. Certain smoothing of strands between the bacterial cell and substrate, alteration of vibrios' form, reduction of electron density of the matrix with an increase of its transparency were observed during electron-microscopy of the effect of antibacterial preparations on the biofilm. Study of the effect of antibacterial preparations on biofilms could increase effectiveness of rational antibiotics therapy of infec- tions by selection of preparations that disrupt functioning of microbial communities.

Flufenamic Acid Prevents Vibrio cholerae El Tor Variant‐Induced Diarrhea via AMPK Activation and Suppression of NF‐κB‐Mediated Inflammation

Pathogenesis of cholera diarrhea caused by infection with Vibrio cholerae El Tor variant (EL), a current epidemic strain of Vibrio cholerae that causes severe diarrhea in several regions, involves the nuclear factor kappa B (NF‐κB)‐mediated inflammatory responses including cyclooxygenase‐2 (COX‐2) upregulation and prostaglandin E2 (PGE2)‐induced intestinal fluid secretion and barrier disruption. The aim of this study was to evaluate the anti‐diarrheal efficacy of flufenamic acid (FFA), an non‐steroidal anti‐inflammatory drug (NSAID) that has previously been reported to stimulate AMP‐activated protein kinase (AMPK), a negative regulator of NF‐κB. We found that intraperitoneal injection of FFA (20 mg/kg) effectively reduced EL‐induced intestinal fluid secretion and intestinal barrier leakage in a mouse closed‐loop model of EL infection. In addition, FFA activated AMPK and suppressed p65 NF‐κB nuclear translocation, mRNA expression of proinflammatory cytokines and protein expressions of COX‐2 and inducible nitric oxide synthase (iNOS) in the mouse intestine infected with EL. In human intestinal epithelial cells (T84 cells), FFA induced tight junction assembly via AMPK activation. Mechanistic studies using immunoblot analysis and molecular docking prediction demonstrated that FFA activated AMPK in T84 cells by directly stimulating Ca(2+)/calmodulin dependent kinase kinase beta (CaMKKβ). Taken together, our results indicate that FFA and related compounds may be repurposed for the treatment of cholera caused by EL infection.Support or Funding InformationThis work was supported by grants BRG5980008 from the Thailand Research Fund and Mahidol University and by the Faculty of Science, Mahidol University and Faculty of graduate studies, Mahidol University.

ANTIBIOTIC RESISTANCE OF VIBRIO CHOLERAE O1 EL TOR ISOLATED ON THE TERRITORY OF THE RUSSIAN FEDERATION IN 2006-2015

Aim of the study. The analysis of antibiotic resistance and molecular mechanisms of the persistence of Vibrio cholerae О1 El Tor strains, isolated from patients and environmental objects on the territory of the Russian Federation in 2006-2015. Material and Methods. The susceptibility of 34 V. cholerae El Tor strains to 13 antibacterial drugs was determined by the method of serial dilutions in a rich culture medium (Instructional Guidelines (МUК) 4.2.2495-09). Results. The strains showed resistance to 1-5 antibiotics (levomicetin, streptomycin, nalidixic acid, furazolidone, trimethoprim/sulfamethoxazole) and harbored SXT-element with antibiotic resistance genes. The emergence of quinolone resistance in V. cholerae O1 El Tor strains with accompanying multiple drug resistance limits the choice for drugs for etiotropic therapy and aggravates the possibility of unfavourable cholera outcome.

Replication of Vibrio cholerae classical CTX phage

The toxigenic classical and El Tor biotype Vibrio cholerae serogroup O1 strains are generated by lysogenization of host-type-specific cholera toxin phages (CTX phages). Experimental evidence of the replication and transmission of an El Tor biotype-specific CTX phage, CTX-1, has explained the evolution of V. cholerae El Tor biotype strains. The generation of classical biotype strains has not been demonstrated in the laboratory, and the classical biotype-specific CTX phage, CTX-cla, is considered to be defective with regard to replication. However, the identification of atypical El Tor strains that contain CTX-cla-like phage, CTX-2, indicates that CTX-cla and CTX-2 replicate and can be transmitted to V. cholerae strains. The replication of CTX-cla and CTX-2 phages and the transduction of El Tor biotype strains by various CTX phages under laboratory conditions are demonstrated in this report. We have established a plasmid-based CTX phage replication system that supports the replication of CTX-1, CTX-cla, CTX-2, and CTX-O139. The replication of CTX-2 from the tandem repeat of lysogenic CTX-2 in Wave 2 El Tor strains is also presented. El Tor biotype strains can be transduced by CTX phages in vitro by introducing a point mutation in toxT, the transcriptional activator of the tcp (toxin coregulated pilus) gene cluster and the cholera toxin gene. This mutation also increases the expression of cholera toxin in El Tor strains in a sample single-phase culture. Our results thus constitute experimental evidence of the genetic mechanism of the evolution of V. cholerae.

Flufenamic acid protects against intestinal fluid secretion and barrier leakage in a mouse model of Vibrio cholerae infection through NF-κB inhibition and AMPK activation

Nuclear factor kappa B (NF-κB)-mediated inflammatory responses play crucial roles in the pathogenesis of diarrhea caused by the Vibrio cholerae El Tor variant (EL), which is a major bacterial strain causing recent cholera outbreaks. Flufenamic acid (FFA) has previously been demonstrated to be a potent activator of AMP-activated protein kinase (AMPK), which is a negative regulator of NF-κB signaling. This study aimed to investigate the anti-diarrheal efficacy of FFA in a mouse model of EL infection and to investigate the mechanisms by which FFA activates AMPK in intestinal epithelial cells (IEC). In a mouse closed loop model of EL infection, FFA treatment (20mg/kg) significantly abrogated EL-induced intestinal fluid secretion and barrier disruption. In addition, FFA suppressed NF-κB nuclear translocation and expression of proinflammatory mediators and promoted AMPK phosphorylation in the EL-infected mouse intestine. In T84 cells, FFA induced AMPK activation. Furthermore, FFA promoted tight junction assembly and prevented interferon gamma (IFN-γ)-induced barrier disruption in an AMPK-dependent manner. Biochemical and molecular docking analyses indicated that FFA activates AMPK via a direct stimulation of calcium/calmodulin-dependent protein kinase kinase beta (CaMKKβ) activity. Collectively, our data indicate that FFA represents a class of existing drugs that may be of potential utility in the treatment of cholera caused by EL infection via AMPK-mediated suppression of NF-κB signaling in IEC.

2D-ELECTROPHORESIS IN PROTEIN SPECTRUM CONSTRUCTION FOR EXOPROTEIN FRACTIONS OF PLAGUE AND CHOLERA AGENTS

Objective of the study is to apply 2D-electrophoresis for imaging protein spectrum of exoprotein fractions, as well as identification and comparison of biomarker expression of the key pathogenicity factors in plague and cholera agents. Materials and methods. 2D gels of exoprotein fractions were obtained on the model of Vibrio cholerae Inaba 569B, V. cholerae El Tor M888Ctx+ and Ctx– strains, and also Y. pestis EV NIIEG strain. Capsular antigen F1 and cholera toxin were used as biomarkers of major exoproteins of plague and cholera agents. Results and conclusions. While studying cholera toxin through 2D-electrophoresis on IPG strips with pH 3–10 gradient, the antigen fell into a number of protein spots, two of which were close to A1 domain parameters (MW 20.309 kDa and pI 6.51) and B-subunit monomer (MW 11.091 kDa and pI 7.68). Comparative protein spectrum analysis of the exoprotein fractions of V. cholerae Inaba 569B, V. cholerae El Tor M888 Ctx+ and Ctx– has revealed the spots with similar parameters, excluding the strain V. cholerae M888 Ctx– . Investigation of 2D gels of capsular antigen and exoprotein fractions of Y. pestis EV strain has demonstrated that protein spots corresponding to the parameters of the subunit form and diameter of the antigen are present in 37 °C culture patterns. 28 °C culture sample contains F1 subunit in a far lesser concentrations. On the model of toxigenic V. cholerae Inaba 569B strain, isogenic system of V. cholerae El Tor M888 Ctx+ and Ctx– strains, and Y. pestis EV NIIEG strain, high efficiency of 2D-electrophoresis in protein spectrum construction for exoprotein fractions of the cultures, as well as identification and comparison of biomarker expression of major exoproteins of plague and cholera agents is established.

Холерные вибрионы неО1/неО139 серогрупп в этиологии острых кишечных инфекций: современная ситуация в России и в мире

Cholera vibrios of nonO1/nonO139 serogroups (NAG vibrios) are well known as natural inhabitants of aquatic environments and as agents of acute intestinal infections (AII) with different degree of severity. On the background of the current vehement dissemination of the new highly virulent Vibrio cholerae El Tor strains, the moral and economic damage from NAG vibrios is not that evident, but still, not less real and rather significant. In spite of rare association with large outbreaks, NAG vibrios rank in the etiology of AII all over the world and pose a potential threat for the population health in our country. The paper reviews the present-day morbidity as regards NAG infections in various countries, including Russia; as well as the molecular premises of the agents’ pathogenicity and drug resistance. Analysis of the published data has revealed an utter heterogeneity of NAG populations, circulating in certain territories, by reference to the presence/absence of genetic determinants of pathogenicity, persistence, housekeeping and antibiotic resistance factors. Nevertheless, the joint gene-pool of the population includes rather a wide set of genes and can expand due to importations of strains, which differ from resident ones in the genotypes, from other regions. This brings about the danger of emergence of clones with higher pathogenic and, probably, even epidemic potential driven by gene exchange, and the consequences of their occurrence are unpredictable. Therefore, the NAG vibrios demand proper attention from investigators and sanitary-epidemiological institutions of the Russian Federation.

POSSIBILITIES OF APPLICATION OF MALDI-TOF MASS-SPECTROMETRY FOR STUDY OF CARBOHYDRATE-SPECIFIC RECEPTORS FOR DIAGNOSTIC BACTERIOPHAGE EL TOR

Study mechanisms of interaction of diagnostic bacteriophage El Tor with sensitive strain Vibrio cholerae El Tor 18507 using direct protein profiling, identification of constant and variable proteins, taking part in interaction of the phage and cell, as well as carbohydrate-specific phage receptors. . A commercial preparation of cholera diagnostic bacteriophage El Tor, strain V. cholerae El Tor 18507 were used. Effect of carbohydrates on bacteriophage activity was determined in experiments with phage by a classic and modified by us method. Protein profiles of the studied objects were studied using MSP-analysis method. Sucrose was shown to inhibit lytic activity of bacteriophage. Proteome profiles of El Tor bacteriophage and sensitive indicator strains were studied, identification of constant and variable proteins of the studied objects by MSP Peak-list program was carried out. Analysis of changes of profiles of phage and microbial cell during interaction with sucrose gave a basis for assuming, that sucrose in the mixture of culture-phage enters interaction namely with phage protein receptors, blocking receptors specific for cholera vibrio, that subsequently manifests in a sharp decrease of phage activity against the sensitive strain.

STUDY OF COMPOSITION OF PLANT EXTRACTS, POSSESSING ANTIMICROBIAL EFFECT AGAINST VIBRIO CHOLERAE EL TOR, USING HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

Study the composition of plant extracts using high-performance liquid chromatography. (HPLC) and evaluation of their antimicrobial effect against Vibrio cholerae El Tor. Qualitative and quantitative composition of plant extracts was studied using HPLC. Determination of sensitivity of microorganisms to plant extracts was carried out by diffusion into agar method and serial dilutions method. Antibacterial effect of water, water-alcohol and acetone extracts of roots of Limonium gmelinii L., Berberis vulgaris L. and Glycyrrhiza glabra L. was studied. The most effective methods of extraction of biologically active substances, possessing antimicrobial effect against various strains of V. cholerae El Tor, were determined. The use of HPLC allowed to establish the presence of catechines, alkaloids protoberberines and glycyrrhizic acid in xtracts, possessing antimicrobial effect against V. cholera El Tor strains.

Structural-Functional Analysis of the Genes, Encoding Biosynthesis of Mannose-Sensitive Hemagglutinating Pili of Adhesion in Different Vibrio cholerae El Tor Strains

Objective of the study is to conduct comparative structural and functional analysis of the genes, encoding biosynthesis of man-nose-sensitive hemagglutinating pili in typical V. cholerae El Tor strains and genovariants. Materials and methods. Utilized were typical and genetically altered strains of cholera vibrio, biovar El Tor, isolated in the territory of the Russian Federation between 1970 and 2012 from ambient environment objects and from patients. Applied were microbiological and molecular-genetic methods, as well as analysis of the data on the whole genome sequencing. Results and conclusions. Using PCR assay the presence of mshA gene, encoding basic sub-unit of MSHA pili, has been detected in all of the tested strains. Comparative analysis of the nucleotide sequence of the genes, included into msh cluster, has revealed identity of the nucleotide sequence of mshA gene, both in typical V. cholerae strains and genovariants. Thereat, in strains of genovariants, isolated at the time of their emergence (1988, 1993, 1994), the structure of the other genes in msh operon is identical to typical El Tor vibrios. At the same time, since 1997, in strains of V. cholerae El Tor genovariants, in the sequence of the genes, involved in the assembly and secretion of MSHA pili, identified have been the SNPs, which do not influence biosynthesis of these pili, but their occurrence, probably, was due to adaptation of the genovariant strains to varying living conditions.

Analysis of Vibrio cholerae Genotype Stability at Low Temperature and Nutrients Deficiency

Objective is an experimental study of the low temperature and nutrient deficiency effects on the stability of Vibrio cholerae El Tor genetic loci complex. Materials and methods. The stability of the genetic loci (i.e. the basic pathogenicity genes; genome fragments containing variable tandem repeats and sites of rare cutting restrictase recognition defining VNTR- and PFGE-profiles of the isolates, accordingly; V. cholerae «housekeeping» genes) of the four Vibrio cholerаe El Tor strains was investigated on the experimental microcosm at different stages of incubation. Results and conclusions. The study demonstrated that clones with altered VNTR- and PFGE-profiles (the detection frequency was 4–8·10–2) were being formed when strains were cultivated under nutrient deficiency and low temperature conditions. Neither changes in the structure of «housekeeping» genes nor loss of pathogenicity genes of the mobile genetic elements were revealed. The findings affirm the probability of analyzed V. cholerae loci alteration occurrence under unfavorable environment conditions. Obtained results should be taken into consideration for interpretation of molecular typing data.

The application of bacterophages in laboratory diagnostic of cholera.

The capacity of cholera agent to develop, during the seventh pandemic, the forms differing by attribute of resistance to bacteriophage determines overall complexity of solving problem of their phage-diagnostics. The search of new races of diagnostic cholera phages is actual and in the first instance relates to necessity of confirmation of specific characteristics of cholera agent by efficient, simple and reliable mode. Thereupon, the target was established to search new race of cholera phages being active in relation to phage-resistant to commercial medication of diagnostic phage of strains of cholera comma bacilli of biovar El-Tor. The study used 131 museum resistant to diagnostic cholera phages strain Vibrio cholerae El Tor separated from various sources in 2001-2014. The activity of 25 cholera phages in relation to sampled strains was verified. The most promising for constructing phage medication are three bacteriophages with spectrum of lytic activity as 44%, 53% and 56% correspondingly. Out of number of selected strains only 20 cultures (15%) maintained resistance to all phages. The estimated bacteriophages have different spectrum of lytic activity in relation to cholera comma bacilli El Tor. The new avirulent test-strain was to be found for reproduction and control of cholera diagnostic bacteriophage. So, non-toxigenic culture Vibrio cholerae El Tor 19546 was used. The analysis of received results demonstrated that analyzed cholera phages with high lytic activity can be used in diagnostics of phage-resistant strains of cholera comma bacilli. The actuality of application of phage-diagnostic technique lies in the fact that it requires no expensive equipment and large material expenses.

Peculiarities of Genome Structure of Toxigenic Vibrio cholerae El Tor Inaba Strain, Isolated from a Surface Water Body in the Territory of Rostov-on-Don in 2014

Carried out has been whole-genome sequencing of the Vibrio cholerae 81 El Tor Inaba strain DNA, isolated from Temernik River waters in the territory of Rostov-on-Don in July, 2014. Identified is the presence of hybrid CTX prophage, which contains ctxB gene of the classical type ( ctxB1 allele), as well as rstR gene El Tor type, tcpA gene with the mutations in coding and promoter regions ( tcpET CIRS allele), the null-mutation in rtxA gene, and extended deletion inside the pandemicity island VSP-II. All the stated above peculiarities are characteristic of cholera vibrio genovariants, which have an enhanced epidemic potential and have recently replaced the typical strains of this biovar.

Monoclonal antibodies to thermostable surface antigens of Vibrio cholerae O1 and O139 serogroups

There is delivered the panel of 9 hybridomas producing monoclonal antibodies directed to surface thermo-resistant antigenic determinants of V.cholerae El Tor and O139. By immunoblotting method most of MCA were established to react with the corresponding epitopes of protein nature at the level of 38-42 kDa. There were revealed differences in the specific orientation of the MCA to Vibrio cholerae: the first group of MCA identifies antigenic determinants presented only in strains of V. cholerae O1, O139 with genetic characteristic ctx+ tcp+ and ctx- tcp+, but fails to identify strains ctx- tcp-; the second group of MCA identifies common epitopes for V.cholerae El Tor and O139 regardless of ctx and tcp genes. The new panel of MCA can be used as well for the study of the fine antigenic structure of V. cholerae El Tor and O139 as detection of epidemically dangerous strains genotype ctx+ tcp+ with ELISA methods

Hybrid Vibrio cholerae El Tor lacking SXT identified as the cause of a cholera outbreak in the Philippines.

ABSTRACTCholera continues to be a global threat, with high rates of morbidity and mortality. In 2011, a cholera outbreak occurred in Palawan, Philippines, affecting more than 500 people, and 20 individuals died. Vibrio cholerae O1 was confirmed as the etiological agent. Source attribution is critical in cholera outbreaks for proper management of the disease, as well as to control spread. In this study, three V. cholerae O1 isolates from a Philippines cholera outbreak were sequenced and their genomes analyzed to determine phylogenetic relatedness to V. cholerae O1 isolates from recent outbreaks of cholera elsewhere. The Philippines V. cholerae O1 isolates were determined to be V. cholerae O1 hybrid El Tor belonging to the seventh-pandemic clade. They clustered tightly, forming a monophyletic clade closely related to V. cholerae O1 hybrid El Tor from Asia and Africa. The isolates possess a unique multilocus variable-number tandem repeat analysis (MLVA) genotype (12-7-9-18-25 and 12-7-10-14-21) and lack SXT. In addition, they possess a novel 15-kb genomic island (GI-119) containing a predicted type I restriction-modification system. The CTXΦ-RS1 array of the Philippines isolates was similar to that of V. cholerae O1 MG116926, a hybrid El Tor strain isolated in Bangladesh in 1991. Overall, the data indicate that the Philippines V. cholerae O1 isolates are unique, differing from recent V. cholerae O1 isolates from Asia, Africa, and Haiti. Furthermore, the results of this study support the hypothesis that the Philippines isolates of V. cholerae O1 are indigenous and exist locally in the aquatic ecosystem of the Philippines.