Molecular detection of Schistosoma japonicum in infected snails and mouse faeces using a real-time PCR assay with FRET hybridisation probes

Thanchomnang, Tongjit; Intapan, Pewpan M.; Sri-Aroon, Pusadee; Lulitanond, Viraphong; Janwan, Penchom; Sanpool, Oranuch; Maleewong, Wanchai

doi:10.1590/s0074-02762011000700008

Cited by 10 publications

(11 citation statements)

References 24 publications

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“…Real-time PCR assays using fluorescence resonance energy transfer hybridization probes combined with melting curve analysis (FRET-PCR) have been used to detect S. japonicum infection in snails and have proved successful. This is reflected in the capability of detecting a cercaria artificially introduced into a pool of 10 noninfected snails, highlighting the possibility of using this method in epidemiological surveys of snail intermediate hosts (323). In addition, PCR with an oligochromatographic dipstick for the detection of amplicons has been shown to reduce the extensive technological requirements for real-time PCR and has also been used effectively as a simple, rapid method for snail diagnosis (324,325).…”

Section: Detection Of Infected Intermediate Snail Hostsmentioning

confidence: 99%

Advances in the Diagnosis of Human Schistosomiasis

et al. 2015

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SUMMARY Schistosomiasis is a major neglected tropical disease that afflicts more than 240 million people, including many children and young adults, in the tropics and subtropics. The disease is characterized by chronic infections with significant residual morbidity and is of considerable public health importance, with substantial socioeconomic impacts on impoverished communities. Morbidity reduction and eventual elimination through integrated intervention measures are the focuses of current schistosomiasis control programs. Precise diagnosis of schistosome infections, in both mammalian and snail intermediate hosts, will play a pivotal role in achieving these goals. Nevertheless, despite extensive efforts over several decades, the search for sensitive and specific diagnostics for schistosomiasis is ongoing. Here we review the area, paying attention to earlier approaches but emphasizing recent developments in the search for new diagnostics for schistosomiasis with practical applications in the research laboratory, the clinic, and the field. Careful and rigorous validation of these assays and their cost-effectiveness will be needed, however, prior to their adoption in support of policy decisions for national public health programs aimed at the control and elimination of schistosomiasis.

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Section: Detection Of Infected Intermediate Snail Hostsmentioning

confidence: 99%

Advances in the Diagnosis of Human Schistosomiasis

et al. 2015

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“…Mean infection intensity was 3.0 EPG (range 0-88). Of the 11 Kato-Katz positive individuals, seven had only 1-3 eggs detected (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24). At the village level, S. japonicum infections were detected by Kato-Katz and the hatching test in two of the four villages, and by PCR in three of the four villages (Table 1).…”

Section: Resultsmentioning

confidence: 99%

“…No additional amplified bands were observed in the samples from individuals with STH infections. DISCUSSION PCR-based detection methods for S. japonicum have been evaluated using experimental animal models 17,18,20,22 ; however, PCR-testing of clinical samples from humans and fieldcollected samples from other mammalian hosts has been limited. 19,21 Here, we have demonstrated a highly stable and sensitive PCR method able to detect schistosomiasis infection in a region where schistosomiasis had reemerged and infection intensity was low (mean infection intensity: 3.0 EPG).…”

Section: Resultsmentioning

confidence: 99%

“…[12][13][14][15][16] Schistosoma japonicum PCR techniques have also been described. [17][18][19][20][21][22] However, the use of PCR techniques to screen field-collected samples has been limited. 19,21 Conventional PCR rather than quantitative real-time PCR (qPCR) was tested because of the limited need for a quantitative description of infections in a low-prevalence setting and because conventional PCR requires fewer resources than qPCR, making it a more accessible tool for resourceconstrained settings.…”

Section: Introductionmentioning

confidence: 99%

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Field Evaluation of a PCR Test for Schistosoma japonicum Egg Detection in Low-Prevalence Regions of China

Fung

Xiao²,

Wang³

et al. 2012

The American Society of Tropical Medicine and Hygiene

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Abstract. Sensitive Schistosoma japonicum detection methods are needed to progress from schistosomiasis control to elimination. The sensitivity of the Kato-Katz thick smear and miracidium hatching tests decrease with infection intensity and serological tests cannot always identify current infections. We evaluated a fecal polymerase chain reaction (PCR) assay to detect S. japonicum infection in 106 humans and 8 bovines in China. PCR was highly sensitive, detecting S. japonicum DNA at 0.5 eggs/g of stool. Comparing PCR examination of a single stool sample to the miracidium hatching test using three consecutive stool samples, more humans were hatching test positive (20%) than PCR positive (15%). However, two individuals were PCR positive in a village where no infections were detected by coprological methods. The sensitivity of PCR makes it a promising tool for schistosomiasis diagnostics and screening, although egg shedding variability and stool sample size present challenges for any detection method in low-transmission areas.

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“…(e.g., conventional PCR, 7 random amplified polymorphic DNA-PCR, 5 and multiplex PCR 17 ). As an effective molecular method, a real-time fluorescence resonance energy transfer PCR (real-time FRET PCR) has been developed as a diagnostic tool for the detection and species differentiation of various parasites (e.g., the detection of Opisthorchis viverrini, 16 Clonorchis sinensis, 16 and Schistosoma japonicum 18 ). In the current study, a rapid and high throughput real-time FRET PCR assay combined with melting curve analysis was developed for the alternative detection of P. heterotremus eggs in the feces of experimentally infected cats.…”

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confidence: 99%

Application of a real-time fluorescence resonance energy transfer polymerase chain reaction assay with melting curve analysis for the detection of Paragonimus heterotremus eggs in the feces of experimentally infected cats

et al. 2013

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ArticleParagonimiasis is an important food-borne parasitic zoonosis caused by a lung fluke belonging to the genus Paragonimus. 2,10 It is estimated that more than 20 million people are infected worldwide, 21 and it has been calculated that 293 million people are at risk. 9 Human beings and other mammals are infected by ingesting raw crustaceans containing metacercariae 22 and usually present with signs and symptoms in the lower respiratory tract (i.e., cough and hemoptysis).2 Paragonimus westermani is the most widely distributed species in Asia and the most important human pathogen in China, Korea, and Japan. 2,10 In Southeast Asia, however, P. heterotremus is the most important pathogen, and confirmed human cases have been found. 2,10 To date, at least 7 lung fluke species have been documented in Thailand: Paragonimus heterotremus, P. siamensis, P. westermani, P. bangkokensis, P. macrorchis, P. harinasutai, and P. pseudoheterotremus. 3,19,20 However, only P. heterotremus and P. pseudoheterotremus are proven human pathogens. 6 The standard diagnosis of paragonimiasis is based on the demonstration of the presence of the eggs of Paragonimus spp. in the sputum (by alkaline decontamination and centrifuge sedimentation technique) and/or feces (by formalin-ether concentration technique).1,22 However, species identification 497944V DIXXX10.1177/1040638713497944Real-time PCR for the differential detection of ParagonimusTantrawatpan et al. research-article2013From the Research and Diagnostic Center for Emerging Infectious Diseases (Tantrawatpan, Intapan, Thanchomnang, Sanpool, Janwan, Lulitanond, Maleewong), the Departments of Parasitology (Intapan, Sanpool, Janwan, Maleewong) Abstract. Paragonimus heterotremus is a medically important lung fluke that causes human and animal paragonimiasis in Southeast Asia, including Thailand. In the current study, a real-time fluorescence resonance energy transfer polymerase chain reaction (real-time FRET PCR) with melting curve analysis was developed and evaluated to detect P. heterotremus eggs in the feces of experimentally infected cats. The detection limit of this method for the P. heterotremus DNA sequence was 3 × 10 2 copies of the positive control plasmid and 10 -3 ng of P. heterotremus genomic DNA. The assay system could detect 10 eggs of P. heterotremus per gram of cat feces. No fluorescence signal was observed when DNA purified from 16 other organisms or genomic DNA from cats and human beings were tested. Real-time FRET PCR yielded positive results for all fecal samples from 17 P. heterotremus-infected cats and showed a negative relationship (r = -0.852, P < 0.001) between the number of parasite eggs in feces and the number of PCR cycles. The assay could detect genomic DNA from P. heterotremus, P. westermani, P. macrorchis, P. siamensis, P. harinasutai, and P. bangkokensis and can differentiate P. heterotremus from the other 5 species. The 6 Paragonimus species examined were divided into 4 groups by melting peak analysis. This assay can be useful for the detection of, a...

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Molecular detection of Schistosoma japonicum in infected snails and mouse faeces using a real-time PCR assay with FRET hybridisation probes

Cited by 10 publications

References 24 publications

Advances in the Diagnosis of Human Schistosomiasis

Advances in the Diagnosis of Human Schistosomiasis

Field Evaluation of a PCR Test for Schistosoma japonicum Egg Detection in Low-Prevalence Regions of China

Application of a real-time fluorescence resonance energy transfer polymerase chain reaction assay with melting curve analysis for the detection of Paragonimus heterotremus eggs in the feces of experimentally infected cats

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