PCR detection of Giardia lamblia in stool: targeting intergenic spacer region of multicopy rRNA gene

Ghosh, Sibdas; Debnath, Anjan; Sil, Alok Kumar; De, S. P.; Chattopadhyay, D.; Das, Pradeep

doi:10.1006/mcpr.2000.0302

Cited by 67 publications

(34 citation statements)

References 25 publications

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“…Detection of parasite-specific DNA appears to be more sensitive than microscopy, as has been shown for G. lamblia infections with (real-time) G. lamblia PCR (13,33), for C. parvum infections (24,37), and for amoebic infection with E. histolytica and E. dispar specific (real-time) PCR (22,31). In this study, C. parvum DNA was detected with the multiplex real-time PCR in the course of infection before microscopy became positive in an immunocompromised child with complaints of diarrhea (Table 1).…”

Section: Discussionmentioning

confidence: 93%

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

et al. 2004

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Entamoeba histolytica, Giardia lamblia, and Cryptosporidium are three of the most important diarrhea-causing parasitic protozoa. For many years, microscopic examination of stool samples has been considered to be the "gold standard" for diagnosis of E. histolytica, G. lamblia, and C. parvum infections. Recently, more specific and sensitive alternative methods (PCR, enzyme-linked immunosorbent assay, and direct fluorescent-antibody assay) have been introduced for all three of these parasitic infections. However, the incorporation in a routine diagnostic laboratory of these parasite-specific methods for diagnosis of each of the respective infections is time-consuming and increases the costs of a stool examination. Therefore, a multiplex real-time PCR assay was developed for the simultaneous detection of E. histolytica, G. lamblia, and C. parvum in stool samples. The multiplex PCR also included an internal control to determine efficiency of the PCR and detect inhibition in the sample. The assay was performed on species-specific DNA controls and a range of well-defined stool samples, and it achieved 100 percent specificity and sensitivity. The use of this assay in a diagnostic laboratory would provide sensitive and specific diagnosis of the main parasitic diarrheal infections and could improve patient management and infection control.

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Section: Discussionmentioning

confidence: 93%

“…In recent years, direct fluorescentantibody assay and antigen detection by using enzyme-linked immunosorbent assay (ELISA) have been accepted as costeffective alternative diagnostic methods (12,21). However, PCR-based methods have also showed excellent specificity and sensitivity compared with microscopy as well with antigen detection (13,33).…”

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confidence: 99%

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

et al. 2004

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“…These kits are commonly used in diagnostic laboratories. A number of PCR assays are also available for the detection of Giardia in stool specimens and show excellent sensitivity and specificity compared to microscopy and antigen detection methods (8,10,30).…”

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confidence: 99%

Evaluation of Multiplex Tandem Real-Time PCR for Detection of Cryptosporidium spp., Dientamoeba fragilis , Entamoeba histolytica , and Giardia intestinalis in Clinical Stool Samples

et al. 2011

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The aim of this study was to describe the first development and evaluation of a multiplex tandem PCR (MT-PCR) assay for the detection and identification of 4 common pathogenic protozoan parasites, Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis, from human clinical samples. A total of 472 fecal samples submitted to the Department of Microbiology at St. Vincent's Hospital were included in the study. The MT-PCR assay was compared to four real-time PCR (RT-PCR) assays and microscopy by a traditional modified iron hematoxylin stain. The MT-PCR detected 28 G. intestinalis, 26 D. fragilis, 11 E. histolytica, and 9 Cryptosporidium sp. isolates. Detection and identification of the fecal protozoa by MT-PCR demonstrated 100% correlation with the RT-PCR results, and compared to RT-PCR, MT-PCR exhibited 100% sensitivity and specificity, while traditional microscopy of stained fixed fecal smears exhibited sensitivities and specificities of 56% and 100% for Cryptosporidium spp., 38% and 99% for D. fragilis, 47% and 97% for E. histolytica, and 50% and 100% for G. intestinalis. No cross-reactivity was detected in 100 stool samples containing various other bacterial, viral, and protozoan species. The MT-PCR assay was able to provide rapid, sensitive, and specific simultaneous detection and identification of the four most important diarrhea-causing protozoan parasites that infect humans. This study also highlights the lack of sensitivity demonstrated by microscopy, and thus, molecular methods such as MT-PCR must be considered the diagnostic methods of choice for enteric protozoan parasites.Enteric protozoa continue to be the most commonly encountered parasitic diseases and to cause significant morbidity and mortality throughout both developed and developing regions of the world, affecting millions of people each year (22). While Blastocystis hominis is the most common protozoan parasite detected in stool samples, the major etiological agents of parasitic diarrhea are considered to be Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis (22).E. histolytica is a pathogenic amoeboid protozoan parasite for which humans are the primary reservoir (27). It is a potentially invasive pathogen and the causative agent of amebiasis, with approximately 50 million cases and 100,000 deaths annually (28). The clinical presentation can range from asymptomatic carriage to gastrointestinal disease and invasive disease. E. histolytica is morphologically identical to the nonpathogenic species Entamoeba dispar and Entamoeba moshkovskii, though genetic differences have confirmed their separation into independent species (4, 20). Due to this conserved morphology, stained smears of stool specimens are insufficient for differentiation of the species. Currently, the only way to differentiate the pathogenic E. histolytica from the nonpathogenic strains via microscopy is to visualize ingested red blood cells within the E. histolytica trophozoite; however, this is rarely seen i...

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“…Therefore, not only a high sensitivity but also a high specificity method of genotyping will be required. To achieve this requirement, nested PCR often used for a pathogen diagnosis from fecal sample [7,13] may improve genotyping reliability.…”

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confidence: 99%

Sex Determination of Sika Deer (Cervus nippon yesoensis) Using Nested PCR from Feces Collected in the Field

et al. 2011

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ABSTRACT. We describe a method for determining the sex of sika deer (Cervus nippon yesoensis) from feces collected in the field. Using a nested polymerase chain reaction (nested PCR), partial sequences of the sex determination region of the Y chromosome (SRY) gene and X zinc finger protein (ZFX) gene were amplified. In 19 individuals with sex information, the correct sex was successfully detected and sequences of target amplicons were completely matched between muscle and feces from the rectum. Among 75 fecal samples collected noninvasively in the field, 68-71 samples (90.7-94.7%) yielded successful sex determinations. Using this technique, feces collected in the field would enhance the utility of genetic analysis. As a result, instead of biomaterials, these samples can serve as new or alternative materials. Finally, it can be expected that this technique will contribute to reveal in advanced detail of the population dynamics and genetic diversity that needed to carry out effective population control and to reduce the extinction risk of sika deer.

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PCR detection of Giardia lamblia in stool: targeting intergenic spacer region of multicopy rRNA gene

Cited by 67 publications

References 25 publications

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

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

Evaluation of Multiplex Tandem Real-Time PCR for Detection of Cryptosporidium spp., Dientamoeba fragilis , Entamoeba histolytica , and Giardia intestinalis in Clinical Stool Samples

Sex Determination of Sika Deer (Cervus nippon yesoensis) Using Nested PCR from Feces Collected in the Field

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