Simultaneous Detection of
 Entamoeba histolytica
 ,
 Giardia lamblia
 , and
 Cryptosporidium parvum
 in Fecal Samples by Using Multiplex Real-Time PCR

Verweij, Jaco J.; Blangé, Roy A.; Templeton, Kate; Schinkel, Janke; Brienen, Eric A. T.; Rooyen, Marianne A.A. van; Lieshout, Lisette van; Polderman, A.M.

doi:10.1128/jcm.42.3.1220-1223.2004

Cited by 357 publications

(307 citation statements)

References 36 publications

(30 reference statements)

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“…To date, a few real -time PCR assays for the specific detection of E. histolytica have been published 17,21,22 however none of them is designed for E. moshkovskii infection. In this study, we describe the development of a real-time PCR for Figure 1. ( A and C ) Amplification in real-time mode for real-time polymerase chain reaction (PCR) and melting curves generated by LightCycler software and differentiation of Entamoeba histolytica , Entamoeba dispar , and Entamoeba moshkovskii .…”

Section: Discussionmentioning

confidence: 99%

Development of Multiplex Real-Time Polymerase Chain Reaction for Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in Clinical Specimens

Hamzah¹,

Petmitr²,

Mungthin³

et al. 2010

The American Journal of Tropical Medicine and Hygiene

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Abstract. Multiplex real -time polymerase chain reaction (PCR) was developed for differential detection of Entamoeba histolytica , Entamoeba dispar , and Entamoeba moshkovskii . Specific primers were designed for all three species, and then differentiation of E. histolytica and E. dispar was achieved simultaneously using a hybridization probe and melting curve analysis, whereas E. moshkovskii was detected with a separate probe under the same condition. This assay detected as little as 0.2 pg of E. histolytica DNA and 2 pg each for E. dispar and E. moshkovskii DNA. Thirty-five clinical samples suspected to be E. histolytica infection by microscopy were tested. The results showed 32 positive samples; four samples were E . histolytica and 28 samples were E. dispar . Interestingly, one E. dispar positive sample showed a mixed infection with E. moshkovskii . This is the first report of E. moshkovskii infection from Thailand and this assay is currently the most rapid and sensitive method to differentiate these human amoebas.

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Section: Discussionmentioning

confidence: 99%

Development of Multiplex Real-Time Polymerase Chain Reaction for Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in Clinical Specimens

Hamzah¹,

Petmitr²,

Mungthin³

et al. 2010

The American Journal of Tropical Medicine and Hygiene

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“…Several real-time quantitative PCR (qPCR) assays for Giardia spp. have subsequently been published (Guy et al, 2003;Verweij et al, 2003Verweij et al, , 2004 Bertrand et al, 2004), plus 3 that are capable of genotyping (Amar et al, 2003; MacKay et al, 2003;Guy et al, 2004). Finally, an assay using multiplex PCR and microarray hybridization has been developed, but it used only cultured trophozoites (Wang and Stenger, 2004).…”

Section: Abstract: Twomentioning

confidence: 99%

Real-Time Pcr/rflp Assay to Detect Giardia intestinalis Genotypes in Human Isolates with Diarrhea in Egypt

Helmy¹,

Abdel-Fattah²,

Rashed³

2009

Journal of Parasitology

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Two; major genotypic assemblages of Giardia intestinalis infect humans; the nested real-time polymerase chain reaction (PCR) was used for targeting the triose phosphate isomerase (tpi) gene to detect and genotype G. intestinalis in human feces in Egypt. Among 97 fecal samples, 30 (31%) were diagnosed as giardiasis by saline wet mount microscopy after staining with Lugol's iodine. The tpi gene was amplified from 41 (42.3%) fecal samples, of which 11 were microscopy-negative specimens. Of the total samples, 24 (58.5%) contained assemblage A group I, and 7 (17.1%) were assemblage A group II from the group of patients complaining of intermittent diarrhea. Eight (19.5%) samples contained assemblage B from patients with persistent diarrhea. Two (5%) samples had a mixture of assemblage A group II and assemblage B. The technique was able to detect as few as 20 trophozoites per PCR on fecal DNA-isolated, microscopy-negative, and quantitative (q)PCR-positive specimens; there was a higher average cycle threshold value than microscopy-positive and qPCR-positive specimens, suggesting that they represented true, low-burden infections. In conclusion, we could genotype G. intestinalis from fresh stool samples in Egypt; in infections commonly presented with intermittent diarrhea, the most prevalent genotype was assemblage A group I. The most vulnerable age group included 10-to 20-yr-old individuals.In developing countries of Asia, Africa, and Latin America, approximately 200 million people have symptomatic giardiasis (Thompson et al., 2000;Yason and Rivera, 2007). Giardia intestinalis is the most prevalent cause of parasitic diarrhea in the developed world, and this infection is also very common in developing countries. Moreover, there is an association of Giardia species and strain assemblages with diarrhea/dysentery (Haque, 2007).In Egypt, Foronda et al. (2008) found that assemblage B was the most prevalent (80%) genotype, with another 15% of the positive samples belonging to assemblage E, and 5% to assemblage A. Transmission of the G. intestinalis cyst to humans occurs mainly after ingestion of contaminated water, autoinfection, and person-to-person contact. Clinical manifestations of symptomatic giardiasis include greasy stools, flatulence, diarrhea, abdominal cramps, epigastric tenderness, as well as steatorrhea accompanied by full-blown malabsorption syndrome (Gardner and Hill, 2001). However, the majority of cases are asymptomatic or minimally symptomatic in immunocompetent individuals (Furness et al., 2000).Species of Giardia can be distinguished on the basis of morphology, ultrastructural features, or the 16S rRNA sequence; there are at least 6 species, i.e., G. intestinalis (5lamblia), G. agilis, G. muris, G. ardeae, G. psittaci, and G. microti (Monis et al., 1999). Among the 6 species, G. intestinalis infects humans and numerous other mammals (Thompson et al., 2000). Isolates of G. intestinalis are classified into 7 assemblages, based on the characterization of the glutamate dehydrogenase, small-subunit rR...

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“…Les résultats de cette étude doivent ouvrir la porte à d'autres analyses, mais aussi d'autres méthodes de détection des unicellulaires, telles la PCR multiple en temps réel (Blessmann et al, 2002 ;Verweij et al, 2004), ou la PCR SHELA, qui combine la technique PCR à l'immunodétection (Britten et al, 1997 ;Furrows et al, 2004).…”

Section: Discussionunclassified

Paléoparasitologie et immunologie. L’exemple d’Entamoeba histolytica

Bailly¹,

Bouchet²

2006

archeosciences

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Simultaneous Detection of Entamoeba histolytica , Giardia lamblia , and Cryptosporidium parvum in Fecal Samples by Using Multiplex Real-Time PCR

Cited by 357 publications

References 36 publications

Development of Multiplex Real-Time Polymerase Chain Reaction for Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in Clinical Specimens

Development of Multiplex Real-Time Polymerase Chain Reaction for Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in Clinical Specimens

Real-Time Pcr/rflp Assay to Detect Giardia intestinalis Genotypes in Human Isolates with Diarrhea in Egypt

Paléoparasitologie et immunologie. L’exemple d’Entamoeba histolytica

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