RESUMO
OBJECTIVE: Efficient and easy-to-use DNA extraction and purification methods are critical in implementing PCR-based diagnosis of pathogens. In order to optimize the routine clinical laboratory diagnosis of eukaryotic enteric pathogens, we compare, via quantitative PCR cycle threshold (Ct) values, the efficiency of two DNA extraction kits: the semi-automated EZ1® (Qiagen) and the manual QIAamp® DNA Stool Mini Kit (Qiagen), on six protozoa: Blastocystis spp., Cryptosporidium parvum/hominis, Cyclospora cayetanensis, Dientamoeba fragilis, Giardia intestinalis and Cystoisospora belli and one microsporidia: Enterocytozoon bieneusi. RESULTS: Whereas EZ1® (Qiagen) and QIAamp® DNA Stool Mini Kit (Qiagen) yielded similar performances for the detection of Cryptosporidium spp. and D. fragilis, significant lower Ct values (p < 0.002) pointed out a better performance of EZ1® on the five remaining pathogens. DNA extraction using the semi-automated EZ1® procedure was faster and as efficient as the manual procedure in the seven eukaryotic enteric pathogens tested. This procedure is suitable for DNA extraction from stools in both clinical laboratory diagnosis and epidemiological study settings.
Assuntos
DNA de Protozoário/isolamento & purificação , Eucariotos/patogenicidade , Fezes/parasitologia , Reação em Cadeia da Polimerase/métodos , Infecções por Protozoários/diagnóstico , Infecções por Protozoários/parasitologia , Blastocystis/genética , Blastocystis/patogenicidade , Cryptosporidium parvum/genética , Cryptosporidium parvum/patogenicidade , Cyclospora/genética , Cyclospora/patogenicidade , DNA de Protozoário/genética , Eucariotos/classificação , Eucariotos/genética , Giardia lamblia/genética , Giardia lamblia/patogenicidade , Humanos , Microsporídios/genética , Microsporídios/patogenicidade , Técnicas de Diagnóstico Molecular/métodos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
Toscana virus (TOSV), an arthropodborne phlebovirus transmitted by sandflies, can cause febrile illness and meningitis. The vector of TOSV in France was unknown. We detected TOSV RNA in 2 (female Phlebotomus perniciosus) of 61 pools of sandflies captured in southeastern France. Two genotypes of TOSV were identified.
