Simple method for long-term copro-preservation of Cryptosporidium oocysts for morphometric and molecular analysis

Tropical Medicine & International Health : TM & IH
Somchai JongwutiwesNutaros Chantachum

Abstract

Preservation of Cryptosporidium oocysts in faecal specimens containing 75% ethanol is suitable for subsequent morphometric and molecular analysis. No significant morphologic alteration occurred after storage at ambient temperatures, ranging from 22 to 38 degrees C, for more than 2 years. After washing, sugar floatation and DNA extraction, a nested polymerase chain reaction targeting the small subunit ribosomal RNA gene successfully amplified Cryptosporidium DNA in all 15 isolates examined. The sensitivity of detection by polymerase chain reaction (PCR) was found to be as high as 1.25 oocysts per reaction (mean=3.01, SD=1.14). Importantly, a 2.2-kb of the complete DNA sequence of a gene encoding Cryptosporidium thrombospondin-related adhesive protein (TRAP-C1) was also consistently amplified by PCR in all isolates. The PCR-amplified product can be used as a good template for sequencing. Therefore, this simple procedure should be useful for epidemiological analysis of clinical samples from outbreaks, endemic or sporadic cases of cryptosporidiosis when long-term storage of oocysts is required.

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Citations

Apr 16, 2002·Tropical Medicine & International Health : TM & IH·R Tiangtip, S Jongwutiwes
Feb 18, 2016·Veterinary Medicine International·Gbemisola Magaret OlabanjiGbeminiyi Richard Otolorin
Mar 11, 2016·Medicine·King-Wah ChiuHock-Liew Eng
Jun 3, 2008·Revista Do Instituto De Medicina Tropical De São Paulo·Elenice M N GonçalvesCarlos E P Corbett
Aug 30, 2005·Scandinavian Journal of Infectious Diseases·Marianne Lebbad, Staffan G Svärd
Jan 8, 2019·Vector Borne and Zoonotic Diseases·Antonio Gomez-RiosMatilde Jiménez-Coello
Oct 30, 2004·Scandinavian Journal of Infectious Diseases·Mette Vinther SkriverHenrik Toft Sørensen
Jul 30, 2005·Scandinavian Journal of Infectious Diseases·Mona InsulanderBo Svenungsson
Dec 17, 2009·Environmental Science & Technology·John G MinneryWendy Heiger-Bernays

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