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. 2013:20:45.
doi: 10.1051/parasite/2013045. Epub 2013 Nov 20.

Cryptosporidiosis in Haiti: surprisingly low level of species diversity revealed by molecular characterization of Cryptosporidium oocysts from surface water and groundwater

Céline Damiani  1 , Ketty Balthazard-Accou, Elmyre Clervil, Aïssata Diallo, Cécilia Da Costa, Evens Emmanuel, Anne Totet, Patrice Agnamey
Affiliations

Cryptosporidiosis in Haiti: surprisingly low level of species diversity revealed by molecular characterization of Cryptosporidium oocysts from surface water and groundwater

Céline Damiani et al. Parasite. 2013.

Abstract
in English, French

The protozoan parasite Cryptosporidium sp. has emerged as one of the most important water contaminants, causing waterborne outbreaks of diarrhoeal diseases worldwide. In Haiti, cryptosporidiosis is a frequent cause of diarrhoea in children under the age of five years, HIV-infected individuals, and people living in low socioeconomic conditions, mainly due to the consumption of water or food polluted by Cryptosporidium oocysts. The aim of this study was to detect and identify Cryptosporidium oocysts present in 12 water samples collected in Port-au-Prince and 4 water samples collected in Cap Haïtien. Initial detection consisted of immunomagnetic separation - immunofluorescence assay (IMS-IFA), which was confirmed by nested PCR, targeting the most polymorphic region of the 18S rRNA gene in 15/16 samples. Genotyping was performed by PCR-restriction fragment length polymorphism (RFLP) analysis and DNA sequencing. Under our working conditions, neither nested PCR-RFLP nor direct DNA sequencing revealed the expected species diversity, as only Cryptosporidium parvum was identified in the water samples studied. This study highlights the difficulty of detecting mixed populations of Cryptosporidium species in environmental samples.

Le protozoaire parasite Cryptosporidium sp. apparaît comme un polluant hydrique majeur engendrant des épidémies diarrhéiques à travers le monde. En Haïti, la cryptosporidiose est responsable de la plupart des diarrhées chez les enfants âgés de moins de 5 ans, les patients VIH, les personnes vivant dans des conditions socioéconomiques difficiles, surtout suite à la consommation de nourritures et d’eau de boisson polluées par les oocystes de cryptosporidies. Le but de cette étude était de détecter et d’identifier des oocystes de cryptosporidies dans 12 prélèvements d’eau effectués à Port-au-Prince et 4 au Cap Haïtien. La méthode de détection initiale a été une séparation immunomagnétique couplée à l’immunofluorescence, confirmée par méthode de PCR nichée, utilisant des amorces spécifiques de la région la plus polymorphe du gène codant pour 18S rRNA, positive dans 15 des 16 prélèvements. Le génotypage a été réalisé par l’utilisation des enzymes de restriction et le séquençage direct de l’ADN. Dans les conditions de notre étude, ces deux méthodes n’ont pas révélé la diversité d’espèces attendue. En effet, seul Cryptosporidium parvum a été identifié dans tous les prélèvements étudiés. L’étude pointe la difficulté d’obtenir une bonne résolution de mélange d’espèces de cryptosporidies dans les échantillons environnementaux.

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