Cholera is caused by the Gram-negative bacterium Vibrio cholerae
, and the disease is usually transmitted through contaminated water (1
). V. cholerae
is normally present in coastal and brackish waters worldwide and has been found in countries where the disease is not found in humans. The bacterium can also be transmitted globally in the intestines of asymptomatic carriers. Thus, it is difficult to determine the origin of outbreaks associated with disaster situations where the normal water supply and hygiene measures are disrupted.
More than 200 serogroups of V. cholerae
have been identified, but isolates belonging to serogroup O1 of the “classical” or El Tor biotype have been the most important human pathogen in the last century. Seven different cholera pandemics are believed to have occurred since 1817. The causative agents of the first five pandemics were not cultured, but the sixth pandemic (1899 to 1923) was caused by the classical biotype. El Tor strains were associated with sporadic cases during the sixth pandemic (2
), but in 1961, this biotype was responsible for the seventh pandemic. El Tor and a number of variants have been implicated in numerous outbreaks worldwide and have become prevalent in some countries with limited access to clean water.
On 12 January 2010, a 7.0 MW
earthquake hit Haiti. By 24 January, at least 52 aftershocks had been reported, and an estimated 316,000 people had died, 300,000 were injured and more than one million were homeless. This disaster destroyed the already fragile infrastructure and required international assistance in the form of food, water, and aid workers. On 21 October 2010, the Haitian public health authorities confirmed a cholera outbreak. By 7 July 2011, 386,429 cases, including 5,885 deaths have been reported (3
). The outbreak has also spread to the neighboring Dominican Republic and to Florida and the United States (4
) where sporadic cases have been observed. In the early days of the outbreak, rumors spread that the disease was brought to Haiti by a battalion of Nepalese soldiers serving as United Nations peacekeepers (2
). Though not proven definitively, the putative link to United Nations peacekeepers from Nepal gained global media attention and sparked riots in Haiti that disrupted relief efforts.
Conventional and molecular characterization of bacterial isolates is useful in determining the relationship between strains and can assist in identifying the sources. Traditionally, V. cholerae
strains are classified into serogroups based on their outer membrane O antigen and further subdivided into biotypes based on biochemical testing; however, most outbreaks during the seventh pandemic have been caused by the same serogroup and biotype, El Tor, limiting the utility of these analyses for outbreak investigations. Molecular typing using pulsed-field gel electrophoresis (PFGE) is commonly used to characterize strains but does not always provide sufficient discriminatory power. Single-nucleotide polymorphisms (SNPs) and insertions/deletions have been used to further resolve global transmission of El Tor (9
). Whole-genome sequence typing (WGST) is a powerful tool providing an almost complete picture of genetic polymorphisms for evolutionary and epidemiological investigations (11
A PFGE-based study by the U.S. Centers for Disease Control and Prevention indicated that the Haitian outbreak strain was related to contemporary strains circulating in South Asia and elsewhere (4
). Another study using whole-genome sequencing has similarly shown that the Haitian outbreak strain is more closely related to recent strains from Bangladesh and Mozambique than to a strain from Peru (15
); however, the Peruvian strain used in that study was more than 20 years old, which weakens their conclusions. So far, none of the published studies has included recent Nepalese V. cholerae
isolates to evaluate their relatedness to the Haitian outbreak strain.
Cholera occurs in sporadic cases and outbreaks in Nepal each year. In 2010, a 1,400-case outbreak occurred in midwestern Nepal (http://www.irinnews.org/Report.aspx?ReportID=90231
). The outbreak started around 28 July and was controlled by 13 or 14 August, just prior to the time the Nepalese soldiers left for Haiti. On the request by the public health authorities in Nepal and in our function as a World Health Organization Collaborating Centre, we conducted the current study to determine the genetic diversity of the most contemporary V. cholerae
strains from Nepal. We then compared these data to the publicly available whole-genome sequences of isolates from the recent outbreak in Haiti, as well as those of other available strains.