Investigations and comparative detection of Cryptosporidium species by microscopy, nested PCR and LAMP in water supplies of Ordu, Middle Black Sea, Turkey

Abstract

A total of 70 water samples, including tap, river, fountain and well water were collected in the Ordu province, Middle Black Sea, Turkey and investigated for the detection of Cryptosporidium oocysts. The samples were directly screened microscopically for Cryptosporidium oocysts' detection by immunofluorescence test and subsequently DNA was extracted for the molecular detection by loop-mediated isothermal amplification (LAMP) and nested polymerase chain reaction (PCR). Eighteen out of the 70 (25.7%) water samples were found positive for Cryptosporidium spp. by immunofluorescence test and 19 (27.1%) were found positive by LAMP. Nested PCR products were not generated in any of the investigated water samples. A total of 16 randomly selected pellets were spiked with 10 Cryptosporidium oocysts to test the efficiency of the applied method. All the samples were found positive by LAMP for the presence of Cryptosporidium DNA, while the nested PCR assay was positive in only seven (43.75%) out of the 16 examined spiked samples. This is the first report on the occurrence of Cryptosporidium species in environmental and drinking water supplies in the Black Sea area.