Sensitive, Specific, and Rapid Detection of Leishmania donovani DNA by Loop-Mediated Isothermal Amplification

Takagi, Hidekazu; Itoh, Makoto; Islam, Mohammad Zahidul; Razzaque, Abdur; Ekram, Arm Saifuddin; Hashighuchi, Yoshihisa; Noiri, Eisei; Kimura, Eizen

doi:10.4269/ajtmh.2009.09-0145

Cited by 76 publications

(57 citation statements)

References 27 publications

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“…These results also compare favorably with another LAMP assay designed for the specific amplification of L. donovani; this assay was able to amplify 8 of 10 blood samples from parasitologically confirmed patients, a comparable sensitivity to nested PCR. 28 For suspected VL patients, the RT-LAMP assay has the potential to be used as an initial screen of suspected patients with DAT/rK39 positive serology; only patients with negative LAMP will then require an aspirate, saving time, money, and potential infection from aspirate sampling. It is of vital importance that this assay is now tested with suspected VL cases to assess the real sensitivity and specificity as a diagnostic tool.…”

Section: Discussionmentioning

confidence: 99%

Development of a Reverse Transcriptase Loop-Mediated Isothermal Amplification (LAMP) Assay for the Sensitive Detection of Leishmania Parasites in Clinical Samples

Adams¹,

Schoone²,

Ageed³

et al. 2010

The American Journal of Tropical Medicine and Hygiene

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Abstract. Here we describe a generic, reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP) assay, for the identification of Leishmania species from clinical samples. LAMP is an isothermal reaction recently developed as a point-of-care diagnostic tool. Primers were designed in the conserved region of the 18S ribosomal RNA (rRNA) gene; amplification was visualized by the pre-amplification addition of fluorescent detection reagent (FDR) and a simple UV lamp. By using a reverse-transcriptase step, the system detected infections between 10 and 100 parasites per mL. The assay was tested on a range of nucleic acid extracts from Leishmania species, visceral leishmaniasis (VL) patients from Sudan, and cutaneous leishmaniasis (CL) patients from Suriname. The sensitivity of RT-LAMP from the blood of VL patients was 83% ( N = 30) compared with microscopy of bone-marrow and lymph-node aspirates; for CL patients the observed sensitivity was 98% ( N = 43). The potential to use LAMP as a diagnostic tool for leishmaniasis is discussed.

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Section: Discussionmentioning

confidence: 99%

Development of a Reverse Transcriptase Loop-Mediated Isothermal Amplification (LAMP) Assay for the Sensitive Detection of Leishmania Parasites in Clinical Samples

Adams¹,

Schoone²,

Ageed³

et al. 2010

The American Journal of Tropical Medicine and Hygiene

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show abstract

“…This technique has been proven to be an accurate, rapid and simple method, which amplifies the target nucleic acid under isothermal conditions (Notomi et al, 2000). Recently, wide applicability of LAMP in the detection of parasitic protozoa such as Babesia, Plasmodium, Leishmania and Trypanosoma in clinical samples has been demonstrated (Ikadai et al, 2004;Poon et al, 2006;Njiru et al, 2008;Takagi et al, 2009;Thekisoe et al, 2007;Laohasinnarong et al, 2011). Studies have also shown the application of LAMP to survey vectors of infectious diseases (Aonuma et al, 2009;Thekisoe et al, 2010;Nakao et al, 2010); however, these studies used purified DNA as a template for the LAMP assays.…”

Section: Introductionmentioning

confidence: 99%

Development of a loop-mediated isothermal amplification method for rapid mass-screening of sand flies for Leishmania infection

et al. 2014

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Entomological monitoring of Leishmania infection in leishmaniasis endemic areas offers epidemiologic advantages for predicting the risk and expansion of the disease, as well as evaluation of the effectiveness of control programs. In this study, we developed a highly sensitive loop-mediated isothermal amplification (LAMP) method for the mass screening of sand flies for Leishmania infection based on the 18S rRNA gene. The LAMP technique could detect 0.01 parasites, which was more sensitive than classical PCR. The method was robust and could amplify the target DNA within 1 hr from a crude sand fly template without DNA purification. Amplicon

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“…Assays using alternative amplification technologies such as quantitative nucleic acid sequence-based amplification (QT-NASBA) based on amplification of 18S RNA and loop-mediated isothermal amplification (LAMP) of kinetoplast minicircle DNA do not allow discrimination of different Leishmania species (Van der Meide et al 2005;Takagi et al 2009). Commercialized assays that have been developed for direct detection of Leishmania are not able to identify the infecting species.…”

Section: O L E C U L a R E P I D E M I O L O G Y A N D M O L E C U mentioning

confidence: 99%

Molecular approaches for a better understanding of the epidemiology and population genetics ofLeishmania

2010

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S U M M A R YMolecular approaches are being used increasingly for epidemiological studies of visceral and cutaneous leishmaniases. Several molecular markers resolving genetic differences between Leishmania parasites at species and strain levels have been developed to address key epidemiological and population genetic questions. The current gold standard, multilocus enzyme typing (MLEE), needs cultured parasites and lacks discriminatory power. PCR assays identifying species directly with clinical samples have proven useful in numerous field studies. Multilocus sequence typing (MLST) is potentially the most powerful phylogenetic approach and will, most probably, replace MLEE in the future. Multilocus microsatellite typing (MLMT) is able to discriminate below the zymodeme level and seems to be the best candidate for becoming the gold standard for distinction of strains. Population genetic studies by MLMT revealed geographical and hierarchic population structure in L. tropica, L. major and the L. donovani complex. The existence of hybrids and gene flow between Leishmania populations suggests that sexual recombination is more frequent than previously thought. However, typing and analytical tools need to be further improved. Accessible databases should be created and sustained for integrating data obtained by different researchers. This would allow for global analyses and help to avoid biases in analyses due to small sample sizes.

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Sensitive, Specific, and Rapid Detection of Leishmania donovani DNA by Loop-Mediated Isothermal Amplification

Cited by 76 publications

References 27 publications

Development of a Reverse Transcriptase Loop-Mediated Isothermal Amplification (LAMP) Assay for the Sensitive Detection of Leishmania Parasites in Clinical Samples

Development of a Reverse Transcriptase Loop-Mediated Isothermal Amplification (LAMP) Assay for the Sensitive Detection of Leishmania Parasites in Clinical Samples

Development of a loop-mediated isothermal amplification method for rapid mass-screening of sand flies for Leishmania infection

Molecular approaches for a better understanding of the epidemiology and population genetics ofLeishmania

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