The Origin of the Haitian Cholera Outbreak Strain

BACKGROUND—Although cholera has been present in Latin America since 1991, it had not been epidemic in Haiti for at least 100 years. Recently, however, there has been a severe outbreak of cholera in Haiti. METHODS—We used third-generation single-molecule real-time DNA sequencing to determine the genome sequences of 2 clinical Vibrio cholerae isolates from the current outbreak in Haiti, 1 strain that caused cholera in Latin America in 1991, and 2 strains isolated in South Asia in 2002 and 2008. Using primary sequence data, we compared the genomes of these 5 strains and a set of previously obtained partial genomic sequences of 23 diverse strains of V. cholerae to assess the likely origin of the cholera outbreak in Haiti. RESULTS—Both single-nucleotide variations and the presence and structure of hypervariable chromosomal elements indicate that there is a close relationship between the Haitian isolates and variant V. cholerae El Tor O1 strains isolated in Bangladesh in 2002 and 2008. In contrast, analysis of genomic variation of the Haitian isolates reveals a more distant relationship with circulating South American isolates. CONCLUSIONS—The Haitian epidemic is probably the result of the introduction, through human activity, of a V. cholerae strain from a distant geographic source. (Funded by the National Institute of Allergy and Infectious Diseases and the Howard Hughes Medical Institute.) The outbreak of cholera that began in Haiti in late October 2010 illustrates the continued public health threat of this ancient scourge.1 Cholera, an acutely dehydrating diarrheal disease that can rapidly kill its victims, is caused by Vibrio cholerae, a gram-negative bacterium.2 This disease, which is usually transmitted through contaminated water, can and has spread in an explosive fashion. In the weeks since cases were first confirmed in the Artibonite province of Haiti on October 19, 2010, the disease has reached all 10 provinces in Haiti and has spread to the neighboring Dominican Republic on the island of Hispaniola. Of Copyright © 2010 Massachusetts Medical Society. Address reprint requests to Dr. Schadt at [email protected].. Drs. Chin, Sorenson, Harris, and Robins contributed equally to this article. Disclosure forms provided by the authors are available with the full text of this article at NEJM.org. NIH Public Access Author Manuscript N Engl J Med. Author manuscript; available in PMC 2011 July 6. Published in final edited form as: N Engl J Med. 2011 January 6; 364(1): 33–42. doi:10.1056/NEJMoa1012928. N IH PA Athor M anscript N IH PA Athor M anscript N IH PA Athor M anscript the more than 93,000 persons who have been sickened from the outbreak, more than 2100 have died, according to the Haitian Ministry of Public Health and Population (www.mspp.gouv.ht/site/index.php), and it is thought that the epidemic has not yet peaked.3 Cholera epidemics had not been reported in Haiti for more than a century, and the origin of the Haitian V. cholerae outbreak has been the subject of some controversy.4 Traditionally, V. cholerae strains are classified into serogroups on the basis of the structure of an outer-membrane O antigen and into biotypes on the basis of a variety of biochemical and micro-biologic tests. The ongoing seventh pandemic of cholera is caused by the V. cholerae El Tor biotype of serogroup O1 (El Tor O1),5 which has replaced the previous “classical” biotype and has spread globally since its appearance in Indonesia in 1961. It reached the Americas in 1991, beginning in Peru and then spreading throughout much of South America and Central America, where it has since become endemic6; however, the strains of V. cholerae El Tor O1 that are now endemic in South America and Central America had not previously been reported to have caused cholera on Hispaniola. Analyses carried out by Haitian and U.S. laboratories have indicated that the current outbreak strain in Haiti is also V. cholerae El Tor O1 and thus is related to strains that are causing the ongoing seventh pandemic of cholera. Both genetic and phenotypic diversity have arisen among circulating strains of V. cholerae El Tor O1, reflecting the acquisition, loss, or alteration of mobile genetic elements (for this and other key terms, see the Glossary), including CTX phage, which bears the genes encoding cholera toxin7; genomic islands8; and SXT-family integrative and conjugative elements, which often en code resistance to several antibiotics.9 Single-nucleotide variations (SNVs) and insertions and deletions have also been detected in the core V. cholerae genome. 10,11 Such heterogeneity has been used to group strains and to model and understand their transmission around the globe10,11 and is most comprehensively captured by sequencing genomic DNA. Second-generation DNA-sequencing technologies, although greatly productive, require a week or more to generate DNA sequence at high coverage and produce reads that are much shorter than those produced with first-generation sequencing technologies — making it difficult to characterize DNA variation in repeat regions.12 Thirdgeneration single-molecule real-time sequencing involves direct observation of the DNA polymerase while it synthesizes a strand of DNA; thus, it is much faster than previously developed methods and provides a comparatively long read length.13,14 We therefore used a third-generation, single-molecule, real-time DNA sequencing method13,14 to determine the genome sequences of two Haitian V. cholerae isolates and three additional V. cholerae clinical isolates from other regions of the world, allowing us to determine the probable origin of the cholera outbreak strain in Haiti.