PCR primers designed for new world Leishmania: A systematic review

Conter, Carolina Cella; Mota, Camila Alves; Santos, Barbara Andreo dos; Braga, Laís de Souza; Terron, Mariana de Souza; Navasconi, Taísa Rocha; Fernandes, Andrea Claudia Bekner Silva; Demarchi, Izabel Galhardo; Castro, Kárin Rosi Reinhold de; Aristides, Sandra Mara Alessi; Lonardoni, Maria Valdrinez Campana; Teixeira, Jorge Juarez Vieira; Silveira, Thaís Gomes Verzignassi

doi:10.1016/j.exppara.2019.107773

Cited by 12 publications

(12 citation statements)

References 143 publications

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“…For this purpose, L. major and L. donovani parasites were marked with DsRed and RFP, respectively. Polymerase chain reaction (PCR) is being increasingly used for leishmaniasis diagnosis, characterization, and identification of Leishmania species, due to the high sensitivity for the detection and identification of Leishmania species (reviewed in [47]). Quantitative PCR is a commonly used method for determining the quantity of Leishmania DNA in various tissues of laboratory animals experimentally infected with Leishmania parasites, asymptomatic humans from endemic areas, and also for testing putative reservoir hosts [18,[48][49][50].…”

Section: Discussionmentioning

confidence: 99%

Central Asian Rodents as Model Animals for Leishmania major and Leishmania donovani Research

et al. 2020

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The clinical manifestation of leishmaniases depends on parasite species, host genetic background, and immune response. Manifestations of human leishmaniases are highly variable, ranging from self-healing skin lesions to fatal visceral disease. The scope of standard model hosts is insufficient to mimic well the wide disease spectrum, which compels the introduction of new model animals for leishmaniasis research. In this article, we study the susceptibility of three Asian rodent species (Cricetulus griseus, Lagurus lagurus, and Phodopus sungorus) to Leishmania major and L. donovani. The external manifestation of the disease, distribution, as well as load of parasites and infectiousness to natural sand fly vectors, were compared with standard models, BALB/c mice and Mesocricetus auratus. No significant differences were found in disease outcomes in animals inoculated with sand fly- or culture-derived parasites. All Asian rodent species were highly susceptible to L. major. Phodopus sungorus showed the non-healing phenotype with the progressive growth of ulcerative lesions and massive parasite loads. Lagurus lagurus and C. griseus represented the healing phenotype, the latter with high infectiousness to vectors, mimicking best the character of natural reservoir hosts. Both, L. lagurus and C. griseus were also highly susceptible to L. donovani, having wider parasite distribution and higher parasite loads and infectiousness than standard model animals.

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Section: Discussionmentioning

confidence: 99%

Central Asian Rodents as Model Animals for Leishmania major and Leishmania donovani Research

et al. 2020

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“…Currently, methods for detecting Leishmania infection include microscopy (Carneiro et al, 2017 ), in vitro culturing (Ates et al, 2013 ), isolation of experimental animals (Loria-Cervera and Andrade-Narvaez, 2014 ), dermal diagnostic tests (Weigle et al, 1991 ), xenodiagnosis (Sadlova et al, 2015 ), and molecular approaches (Galluzzi et al, 2018 ; Sundar and Singh, 2018 ; Conter et al, 2019 ). Each method has its own advantages and disadvantages.…”

Section: Introductionmentioning

confidence: 99%

Leishmaniasis Diagnosis via Metagenomic Next-Generation Sequencing

Chen

Fan

Gao

et al. 2020

Front. Cell. Infect. Microbiol.

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Leishmaniasis is a vector-borne disease caused by Leishmania. Although the incidence of leishmaniasis in China is currently low, it has not been completely eradicated. In 2019, visceral leishmaniasis was diagnosed in three patients using bone marrow microscopic examination and metagenomic next-generation sequencing (mNGS). The bone marrow mNGS results from the three patients indicated that 99.9, 99.6, and 30.3% of non-human reads matched the Leishmania genome, and plasma mNGS results from one of the patients revealed that 46.2% of non-human reads matched the Leishmania genome. In the second patient's plasma, no Leishmania sequences were detected by plasma mNGS, and the third patient's plasma was unavailable. The pathogen in all three patients was identified as Leishmania infantum. Leishmania amastigotes were observed by microscopic examination of bone marrow smears in all three patients, but were not found in peripheral blood smears. This indicates that the sensitivity of mNGS is higher than that of smear microscopy and that mNGS can be used to identify Leishmania at the species level. All three patients were elderly male farmers, two from Shanxi and one from Beijing. All three patients had splenomegaly and pancytopenia. Originally, these patients were misdiagnosed and treated for extended periods in other hospitals. Diagnoses of visceral leishmaniasis took place 6, 2, and 2 months after the onset of symptoms in the three patients. In conclusion, this study confirms that bone marrow mNGS can be used to quickly and accurately confirm a diagnosis in patients with suspected leishmaniasis.

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“…The PCR can amplify up to one single parasite in human blood [30], which relates to clinical disease [58]. However, in our study, the SSU PCR had a lower sensitivity than the kDNA PCRs as has been reported before [19,20].…”

Section: Plos Neglected Tropical Diseasesmentioning

confidence: 45%

“…Studies done in Old World CL countries demonstrated high sensitivities for kDNA targeting PCRs ranging between 91.7% up to 100% [10,20,50,59]. The kDNA minicircle sequence is by far the most often used target in studies on visceral [26,32,58] or New World leishmaniasis [19,60]. With over 10,000 copies of minicircles per parasite, this PCR is more sensitive than the SSU [57] and ITS PCRs [10].…”

Section: Plos Neglected Tropical Diseasesmentioning

confidence: 99%

“…PCRs designed in either conventional or real time formats can target different regions of the Leishmania genome for parasite detection at the genus, complex or species level [19,20]. Typically, these PCRs target nuclear and ribosomal DNA, like the small subunit (SSU) 18S ribosomal RNA or internal transcribed spacer (ITS) ribosomal regions [21,22] or the miniexon spliced leader (SL) gene repeat [23,24].…”

Section: Introductionmentioning

confidence: 99%

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Evaluation of conventional and four real-time PCR methods for the detection of Leishmania on field-collected samples in Ethiopia

et al. 2021

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In most low-resource settings, microscopy still is the standard method for diagnosis of cutaneous leishmaniasis, despite its limited sensitivity. In Ethiopia, the more sensitive molecular methods are not yet routinely used. This study compared five PCR methods with microscopy on two sample types collected from patients with a suspected lesion to advise on optimal diagnosis of Leishmania aethiopica. Between May and July 2018, skin scrapings (SS) and blood exudate from the lesion spotted on filter paper (dry blood spot, DBS) were collected for PCR from 111 patients of four zones in Southern Ethiopia. DNA and RNA were simultaneously extracted from both sample types. DNA was evaluated by a conventional PCR targeting ITS-1 and three probe-based real-time PCRs: one targeting the SSU 18S rRNA and two targeting the kDNA minicircle sequence (the ‘Mary kDNA PCR’ and a newly designed ‘LC kDNA PCR’ for improved L. aethiopica detection). RNAs were tested with a SYBR Green-based RT-PCR targeting spliced leader (SL) RNA. Giemsa-stained SS smears were examined by microscopy. Of the 111 SS, 100 were positive with at least two methods. Sensitivity of microscopy, ITS PCR, SSU PCR, Mary kDNA PCR, LC kDNA PCR and SL RNA PCR were respectively 52%, 22%, 64%, 99%, 100% and 94%. Microscopy-based parasite load correlated well with real-time PCR Ct-values. Despite suboptimal sample storage for RNA detection, the SL RNA PCR resulted in congruent results with low Ct-values. DBS collected from the same lesion showed lower PCR positivity rates compared to SS. The kDNA PCRs showed excellent performance for diagnosis of L. aethiopica on SS. Lower-cost SL RNA detection can be a complementary high-throughput tool. DBS can be used for PCR in case microscopy is negative, the SS sample can be sent to the referral health facility where kDNA PCR method is available.

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PCR primers designed for new world Leishmania: A systematic review

Cited by 12 publications

References 143 publications

Central Asian Rodents as Model Animals for Leishmania major and Leishmania donovani Research

Central Asian Rodents as Model Animals for Leishmania major and Leishmania donovani Research

Leishmaniasis Diagnosis via Metagenomic Next-Generation Sequencing

Evaluation of conventional and four real-time PCR methods for the detection of Leishmania on field-collected samples in Ethiopia

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