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Year : 2014  |  Volume : 6  |  Issue : 4  |  Page : 183-184

The first appearance of classical-like phenotype vibrio cholerae in Nepal

Clinical Research Unit, Sukraraj Tropical and Infectious Disease Hospital, Kathmandu, Nepal

Date of Web Publication22-Apr-2014

Correspondence Address:
Sher Bahadur Pun
Clinical Research Unit, Sukraraj Tropical and Infectious Disease Hospital, Kathmandu
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/1947-2714.131248

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How to cite this article:
Pun SB. The first appearance of classical-like phenotype vibrio cholerae in Nepal. North Am J Med Sci 2014;6:183-4

How to cite this URL:
Pun SB. The first appearance of classical-like phenotype vibrio cholerae in Nepal. North Am J Med Sci [serial online] 2014 [cited 2023 Mar 26];6:183-4. Available from: https://www.najms.org/text.asp?2014/6/4/183/131248

Dear Editor,

Vibrio cholerae is the etiology agent of cholera and is mainly spread through contaminated drinking water. V. cholerae O1 and O139 serogroups are known to cause cholera epidemics worldwide. Until the end of the sixth pandemic (1899-1923), all cholera cases were considered to be caused by the V. cholerae O1 of the classical biotype, before the characterization of the O1 El Tor biotype in 1905. The El Tor biotype was implicated in major epidemics approximately 60-years later (1961) with the emergence of the seventh pandemic, [1] although the classical biotype was reportedly identified in Bangladesh in 1988-1989. [2]

From July to August 2012, a total of 503 patients with acute watery diarrhea were hospitalized at Sukraraj Tropical and Infectious Disease Hospital, Kathmandu. Stool specimens were inoculated on thiosulfate citrate bile salt sucrose agar (TCBS) (Hi-media, Mumbai, India) and incubated at 37°C for 24 h. Subcultures were further done on MacConkey Agar (MA) and incubated overnight at 37°C. The colonies were analyzed and biochemical tests were performed. Of these, 21 (4.1%) were culture positive for V. cholerae. Isolates were further serotyped using specific antisera (Denka Seikan, Tokyo, Japan). All the isolates were identified as O1 Ogawa serotype. Biotyping was performed using Polymyxin B (50 U) susceptibility test, Chicken red cell agglutination (CCA), and Voges-Proskauer reaction test (VP test) according to standard recommended procedures. [3],[4] All isolates were negative for VP [Figure 1]a and CCA tests [Figure 1]b but sensitive to Polymyxin B (50 U) [Figure 1]c. Repeat analysis of these assays confirmed that the isolates carried phenotypic traits specific for the classical biotype. Until recently, V. cholerae showing phenotypic traits of the classical biotype was not reported in Nepal. This study, therefore, highlights the first appearance of V. cholerae of the classical biotype in patients with acute diarrhea in Nepal.
Figure 1: (a-c) Laboratory tests showing phenotypic results of the isolates

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Previous studies have shown that V. cholerae strains detected in Nepal were restricted to El Tor phenotype. [5],[6] In a recent study, Nepalese V. cholerae strains were found to be of the El Tor phenotype but carrying the classical cholera toxin. [6] Contrary to expectations, isolates in the present study demonstrated phenotypic characteristics of the classical biotype. A strain designated as "Matlab type II" belonging to the Ogawa serotype having characteristics of the classical biotype was identified in a hospitalized patient with acute diarrhea during a study period of 1991 to 1994 in Bangladesh. [7] Interestingly, this strain was found to have the tcpA gene of the classical biotype, whereas the rstR gene was of the El Tor type. The V. cholerae strains isolated in this study may possess similar phenotypes of both the classical and El Tor strains. Genetic exchange between co-occurring classical and El Tor biotypes could be one of the major factors leading to hybrid or variant V. cholerae strains. V. cholerae classical strains have never been reported in Nepal. Thus, the Nepalese isolates with classical phenotypic properties in this study, therefore, may not be indigenous strains but were presumably introduced from a distant geographic location. At present, it is not known whether Nepalese strains solely consist of classical genome or acquired El Tor genes, as this study investigated phenotypic traits only; it is not known whether the Nepalese strains consist of the classical genotype or whether they have acquired El Tor genes. Genetic testing, therefore, may reveal whether these strains are of the classical biotype or if they carry El Tor-specific alleles. The frequent exchange of genes between classical and El Tor biotypes may promote the emergence of a "classical variant", a potential new variant of V. cholerae, though such designation has not yet been proposed or included in the redefined biotype scheme. [8] Nevertheless, it is possible to hypothesize the emergence of "classical variant" V. cholerae, as genetic exchange between strains carrying traits of both biotypes might occur in the environment, albeit slowly, particularly in areas where cholera is endemic.

During the study period, none of the samples were shown to have El Tor biotypic properties, contrary to previous studies in Nepal. The El Tor strain was not included as a negative control during analysis is another limitation of this study. However, the likelihood of contamination during laboratory analysis was low, as isolates carrying classical phenotypes of V. cholerae have never been reported in Nepal, and reexamination of these strains was performed to confirm the results.

Past studies have shown that classical strains are likely to cause more severe disease compared with El Tor strains, [9] although the role of hybrid type of V. cholerae in pathogenesis is not yet well studied. Although Kathmandu has witnessed both rapid population growth and urbanization over the past decade, progress toward the provision of safe water supplies and improved sanitation facilities remain slow. It can thus be postulated that this newly emerging V. cholerae having classical-like phenotype may lead to severe forms of cholera because there may be little or no immune protection against neither classical nor genetic hybrid strains of V. cholerae in the human population. This, therefore, underscores the immediate need for increased vigilance in monitoring and preparedness for the future outbreaks.

  Acknowledgments Top

The author is grateful to Rojina Maharjan and Dina Shrestha of Kantipur College of Medical Sciences for their help in the laboratory analysis. The author also would like to express sincere gratitude to Professor Richard Culleton of University of Nagasaki for reviewing the manuscript and the anonymous reviewers for their extremely helpful comments and valuable suggestions.

  References Top

1.Barua D. History of cholera. In: Barua D, Greenough WB. 3 rd , ed. Cholera. New York: Plenum Medical Book Co. 1992:1-36.  Back to cited text no. 1
2.Siddique AK, Baqui AH, Eusof A, Haider K, Hossain MA, Bashir I, et al. Survival of classic cholera in Bangladesh. Lancet 1991;337:1125-7.  Back to cited text no. 2
3.Centers for Disease Control and Prevention. Laboratory methods for the diagnosis of Vibrio cholerae, Atlanta, GA, USA. 1994:38-67. Available at http://www.cdc.gov/cholera/Laboratory.html. [Last accessed on 2013 Oct 8].  Back to cited text no. 3
4.Isenberg HD. Clinical Microbiology Procedures Handbook. 2 nd ed. Vol. 1. Washington, ASM Press. 2004:3.8.1.  Back to cited text no. 4
5.Tamang MD, Sharma N, Makaju RK, Sharma AN, Koju R, Nepali R, et al. An outbreak of El tor cholera in Kavre district, Nepal. Kathmandu Univ Med J (KUMJ) 2005;3:138-42.  Back to cited text no. 5
6.Shakya G, Kim DW, Clemens JD, Mall S, Upadhyaya BP, Dumre SP, et al. Phenotypic and genetic characterization of Vibrio Cholerae O1 clinical isolates collected through national antimicrobial resistance surveillance network in Nepal. World J Microbiol Biotechnol 2012;28:2671-8.  Back to cited text no. 6
7.Nair GB, Faruque SM, Bhuiyan NA, Kamruzzaman M, Siddique AK, Sack DA. New variants of Vibrio cholerae O1 biotype El Tor with attributes of the classical biotype from hospitalized patients with acute diarrhea in Bangladesh. J Clin Microbiol 2002;40:3296-9.  Back to cited text no. 7
8.Raychoudhury A, Mukhopadhyay AK, Ramamurthy T, Nandy RK, Takeda Y, Nair GB. Biotyping of Vibrio cholerae O1: Time to redefine the scheme. Indian J Med Res 2008;128:695-8.  Back to cited text no. 8
9.Kaper JB, Morris JG Jr, Levine MM. Cholera. Clin Microbiol Rev 1995;8:48-86.  Back to cited text no. 9


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