Diagnostic antigens for visceral leishmaniasis: clarification of nomenclatures

Bhattacharyya, Tapan; Marlais, Tegwen; Miles, Michael A.

doi:10.1186/s13071-017-2120-x

Cited by 8 publications

(6 citation statements)

References 14 publications

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“…As Bhattacharyya et al . pointed out in their recent review, the use of "rk" in the antigen nomenclature is misleading [21]. Eight different antigens included in our analysis are based on genes for kinesin related proteins such as rk39 (Table 2).…”

Section: Resultsmentioning

confidence: 99%

Systematic review on antigens for serodiagnosis of visceral leishmaniasis, with a focus on East Africa

et al. 2019

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Background Accurate and accessible diagnosis is key for the control of visceral leishmaniasis (VL). Yet, current diagnostic tests for VL have severe limitations: they are invasive or not suitable as point of care (POC) test or their performance is suboptimal in East Africa. We analysed the antigens in the VL serodiagnostics development pipeline to identify shortcomings and to propose strategies in the development of an alternative POC test for VL in East Africa. Objectives The objective of this study was to identify and to analyse all antigens for VL serodiagnosis that have been published before 2018 in order to identify candidates and gaps in the pipeline for a new POC test in East Africa. Methods A systematic literature search was performed on PubMed for original research articles on Leishmania -specific antigens for antibody detection of VL in humans. From each article, the following information was extracted: the antigen name, test format and characteristics, its reported sensitivity and specificity and study cohort specifications. Results One hundred and seven articles containing information about 96 tests based on 89 different antigens were included in this study. Eighty six of these tests, comprising 80 antigens, were evaluated in phase I and II studies only. Only 20 antigens, all of which are native, contain a carbohydrate and/or lipid moiety. Twenty-four antigens, of which 7 are non-native, are composed of antigen mixtures. Nineteen tests, comprising 18 antigens, have been evaluated on East African specimens, of which only 2 (rK28 based immunochromatographic test and intact promastigote based indirect fluorescent antibody technique) consistently showed sensitivities above 94 and specificities above 97% in a phase III study and one in a phase II study (dot blot with SLA). Only rK28 is a non-native mixture of antigens which we consider suitable for further evaluation and implementation. Conclusions The development pipeline for an alternative serodiagnostic test for VL is almost empty. Most antigens are not sufficiently evaluated. Non-protein antigens and antigen mixtures are being neglected. We propose to expand the evaluation of existing antigen candidates and to investigate the diagnostic potential of defined non-native carbohydrate and lipid antigens for VL serodiagnosis in East Africa.

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Section: Resultsmentioning

confidence: 99%

Systematic review on antigens for serodiagnosis of visceral leishmaniasis, with a focus on East Africa

et al. 2019

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“…Extracellular proteins are more accessible for binding with host Igs, thus we have included this criterion in our filter. Finally, because intracellular proteins harbouring TRs are amongst the most widely used antigens for VL diagnosis [56], we decided to select such candidates in parallel to extracellular proteins. Although the vast majority of epitopes founds in proteins are discontinuous [57], the use of such prediction algorithms is only meaningful in case the native 3D structure of the proteins is retained, which is not the case upon completion of an SDS-PAGE gel.…”

Section: Discussionmentioning

confidence: 99%

Refining wet lab experiments with in silico searches: A rational quest for diagnostic peptides in visceral leishmaniasis

et al. 2019

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Background The search for diagnostic biomarkers has been profiting from a growing number of high quality sequenced genomes and freely available bioinformatic tools. These can be combined with wet lab experiments for a rational search. Improved, point-of-care diagnostic tests for visceral leishmaniasis (VL), early case detection and surveillance are required. Previous investigations demonstrated the potential of IgG1 as a biomarker for monitoring clinical status in rapid diagnostic tests (RDTs), although using a crude lysate antigen (CLA) as capturing antigen. Replacing the CLA by specific antigens would lead to more robust RDTs. Methodology Immunoblots revealed L. donovani protein bands detected by IgG1 from VL patients. Upon confident identification of these antigens by mass spectrometry (MS), we searched for evidence of constitutive protein expression and presence of antigenic domains or high accessibility to B-cells. Selected candidates had their linear epitopes mapped with in silico algorithms. Multiple high-scoring predicted epitopes from the shortlisted proteins were screened in peptide arrays. The most promising candidate was tested in RDT prototypes using VL and nonendemic healthy control (NEHC) patient sera. Results Over 90% of the proteins identified from the immunoblots did not satisfy the selection criteria and were excluded from the downstream epitope mapping. Screening of predicted epitope peptides from the shortlisted proteins identified the most reactive, for which the sensitivity for IgG1 was 84% (95% CI 60—97%) with Sudanese VL sera on RDT prototypes. None of the sera from NEHCs were positive. Conclusion We employed in silico searches to reduce drastically the output of wet lab experiments, focusing on promising candidates containing selected protein features. By predicting epitopes in silico we screened a large number of peptides using arrays, identifying the most promising one, for which IgG1 sensitivity and specificity, with limited sample size, supported this proof of concept strategy for diagnostics discovery, which can be applied to the development of more robust IgG1 RDTs for monitoring clinical status in VL.

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“…Serum samples (50 μl) were tested with the rK28-based RDT OnSite Leishmania rK39-Plus (CTK Biotech, Inc., USA) following the manufacturer’s instructions. This test is based on the rK28 antigen, a chimeric protein composed by the first two 39 amino acid repeats of the Sudanese kinesin homologue LdK39 flanked by the repeat sequences of HASPB1 (or K26) and the whole open reading frame of HASPB2 (or K9) [ 14 ]. Results were recorded after 15 minutes, and whenever an invalid result was obtained, the test was repeated immediately.…”

Section: Methodsmentioning

confidence: 99%

Sensitive and less invasive confirmatory diagnosis of visceral leishmaniasis in Sudan using loop-mediated isothermal amplification (LAMP)

et al. 2018

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BackgroundConfirmatory diagnosis of visceral leishmaniasis (VL), as well as diagnosis of relapses and test of cure, usually requires examination by microscopy of samples collected by invasive means, such as splenic, bone marrow or lymph node aspirates. This causes discomfort to patients, with risks of bleeding and iatrogenic infections, and requires technical expertise. Molecular tests have great potential for diagnosis of VL using peripheral blood, but require well-equipped facilities and trained personnel. More user-friendly, and field-amenable options are therefore needed. One method that could meet these requirements is loop-mediated isothermal amplification (LAMP) using the Loopamp Leishmania Detection Kit, which comes as dried down reagents that can be stored at room temperature, and allows simple visualization of results.Methodology/Principal findingsThe Loopamp Leishmania Detection Kit (Eiken Chemical Co., Japan), was evaluated in the diagnosis of VL in Sudan. A total of 198 VL suspects were tested by microscopy of lymph node aspirates (the reference test), direct agglutination test-DAT (in house production) and rK28 antigen-based rapid diagnostic test (OnSite Leishmania rK39-Plus, CTK Biotech, USA). LAMP was performed on peripheral blood (whole blood and buffy coat) previously processed by: i) a direct boil and spin method, and ii) the QIAamp DNA Mini Kit (QIAgen). Ninety seven of the VL suspects were confirmed as cases by microscopy of lymph node aspirates. The sensitivity and specificity for each of the tests were: rK28 RDT 98.81% and 100%; DAT 88.10% and 78.22%; LAMP-boil and spin 97.65% and 99.01%; LAMP-QIAgen 100% and 99.01%.Conclusions/SignificanceDue to its simplicity and high sensitivity, rK28 RDT can be used first in the diagnostic algorithm for primary VL diagnosis, the excellent performance of LAMP using peripheral blood indicates that it can be also included in the algorithm for diagnosis of VL as a simple test when parasitological confirmatory diagnosis is required in settings that are lower than the reference laboratory, avoiding the need for invasive lymph node aspiration.

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Diagnostic antigens for visceral leishmaniasis: clarification of nomenclatures

Cited by 8 publications

References 14 publications

Systematic review on antigens for serodiagnosis of visceral leishmaniasis, with a focus on East Africa

Systematic review on antigens for serodiagnosis of visceral leishmaniasis, with a focus on East Africa

Refining wet lab experiments with in silico searches: A rational quest for diagnostic peptides in visceral leishmaniasis

Sensitive and less invasive confirmatory diagnosis of visceral leishmaniasis in Sudan using loop-mediated isothermal amplification (LAMP)

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