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Multiplex PCR detection of Cryptosporidium sp, Giardia lamblia and Entamoeba histolytica directly from dried stool samples from Guinea-Bissauan children with diarrhoea
Helsinki Univ Hosp, Div Clin Microbiol, HUSLAB, Helsinki, Finland..
Helsinki Univ Hosp, Div Clin Microbiol, HUSLAB, Helsinki, Finland..
Helsinki Univ Hosp, Div Clin Microbiol, HUSLAB, Helsinki, Finland..
Danderyd Hosp, Dept Infect Dis, Stockholm, Sweden.;Indepth Network, Bandim Hlth Project, Bissau, Guinea Bissau.;Karolinska Inst, Dept Microbiol Tumour & Cell Biol, Stockholm, Sweden..
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2017 (English)In: Infectious Diseases, ISSN 2374-4235, E-ISSN 2374-4243, Vol. 49, no 9, 655-663 p.Article in journal (Refereed) Published
Abstract [en]

Background: In developing countries, diarrhoea is the most common cause of death for children under five years of age, with Giardia lamblia, Cryptosporidium and Entamoeba histolytica as the most frequent pathogenic parasites. Traditional microscopy for stool parasites has poor sensitivity and specificity, while new molecular methods may provide more accurate diagnostics. In poor regions with sample storage hampered by uncertain electricity supply, research would benefit from a method capable of analysing dried stools. Methods: A real-time multiplex PCR method with internal inhibition control was developed for detecting Giardia lamblia, Cryptosporidium hominis/parvum and Entamoeba histolytica directly from stool specimens. Applicability to dried samples was checked by comparing with fresh ones in a small test material. Finally, the assay was applied to dried specimens collected from Guinea-Bissauan children with diarrhoea. Results: The PCR's analytical sensitivity limit was 0.1 ng/ml for G. lamblia DNA, 0.01 ng/ml for E. histolytica DNA and 0.1 ng/ml for Cryptosporidium sp. In the test material, the assay performed similarly with fresh and dried stools. Of the 52 Guinea-Bissauan samples, local microscopy revealed a parasite in 15%, while PCR detected 62% positive for at least one parasite: 44% of the dried samples had Giardia, 23% Cryptosporidium and 0% E. histolytica. Conclusions: Our new multiplex real-time PCR for protozoa presents a sensitive method applicable to dried samples. As proof of concept, it worked well on stools collected from Guinea-Bissauan children with diarrhoea. It provides an epidemiological tool for analysing dried specimens from regions poor in resources.

Place, publisher, year, edition, pages
Taylor & Francis, 2017. Vol. 49, no 9, 655-663 p.
Keyword [en]
Real-time PCR, Giardia, Cryptosporidium, Entamoeba, aetiology, developing country, children, travel, diarrhoea, parasite, stool parasite
National Category
Infectious Medicine
Identifiers
URN: urn:nbn:se:uu:diva-330071DOI: 10.1080/23744235.2017.1320728ISI: 000403158500003PubMedID: 28446068OAI: oai:DiVA.org:uu-330071DiVA: diva2:1150712
Available from: 2017-10-19 Created: 2017-10-19 Last updated: 2017-10-19Bibliographically approved

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