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Noninvasive Diagnosis of Visceral Leishmaniasis: Development and Evaluation of Two Urine-Based Immunoassays for Detection of Leishmania donovani Infection in India.

ABSTRACT: Visceral Leishmaniasis (VL), a severe parasitic disease, could be fatal if diagnosis and treatment is delayed. Post kala-azar dermal leishmaniasis (PKDL), a skin related outcome, is a potential reservoir for the spread of VL. Diagnostic tests available for VL such as tissue aspiration are invasive and painful although they are capable of evaluating the treatment response. Serological tests although less invasive than tissue aspiration are incompetent to assess cure. Parasitological examination of slit-skin smear along with the clinical symptoms is routinely used for diagnosis of PKDL. Therefore, a noninvasive test with acceptable sensitivity and competency, additionally, to decide cure would be an asset in disease management and control.We describe here, the development of antibody-capture ELISA and field adaptable dipstick test as noninvasive diagnostic tools for VL and PKDL and as a test of cure in VL treatment. Sensitivity and specificity of urine-ELISA were 97.94% (95/97) and 100% (75/75) respectively, for VL. Importantly, dipstick test demonstrated 100% sensitivity (97/97) and specificity (75/75) in VL diagnosis. Degree of agreement of the two methods with tissue aspiration was 98.83% (? = 0.97) and 100% (? = 1), for ELISA and dipstick test, respectively. Both the tests had 100% positivity for PKDL (14/14) cases. ELISA and dipstick test illustrated treatment efficacy in about 90% (16/18) VL cases when eventually turned negative after six months of treatment.ELISA and dipstick test found immensely effective for diagnosis of VL and PKDL through urine samples thus, may substitute the existing invasive diagnostics. Utility of these tests as indirect methods of monitoring parasite clearance can define infected versus cured. Urine-based dipstick test is simple, sensitive and above all noninvasive method that may help not only in active VL case detection but also to ascertain treatment response. It can therefore, be deployed widely for interventions in disease management of VL particularly in poor resource outskirts.

SUBMITTER: Ejazi SA

PROVIDER: S-EPMC5065134 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Noninvasive Diagnosis of Visceral Leishmaniasis: Development and Evaluation of Two Urine-Based Immunoassays for Detection of Leishmania donovani Infection in India.

Ejazi Sarfaraz Ahmad SA Bhattacharya Pradyot P Bakhteyar Md Asjad Karim MA Mumtaz Aquil Ahmad AA Pandey Krishna K Das Vidya Nand Ravi VN Das Pradeep P Rahaman Mehebubar M Goswami Rama Prosad RP Ali Nahid N

PLoS neglected tropical diseases 20161014 10

<h4>Background</h4>Visceral Leishmaniasis (VL), a severe parasitic disease, could be fatal if diagnosis and treatment is delayed. Post kala-azar dermal leishmaniasis (PKDL), a skin related outcome, is a potential reservoir for the spread of VL. Diagnostic tests available for VL such as tissue aspiration are invasive and painful although they are capable of evaluating the treatment response. Serological tests although less invasive than tissue aspiration are incompetent to assess cure. Parasitolo ...[more]

PMID: 27741241

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Project description:The unreliability of most of the existing antibody-based diagnostic kits to discriminate between active and treated VL cases, relapse situation and reinfection are a major hurdle in controlling the cases of Kala-azar in an endemic area. An antigen targeted diagnostic approaches can be an attractive strategy to overcome these problems. Hence, this study was focused on identifying the Leishmania antigens, lies in circulating immune complex (CICs), can be used for diagnostic as well as prognostic purposes. The present study was conducted on peripheral blood samples of 115 human subjects, based on isolation of CICs. The SDS-PAGE patterns showed an up-regulated expression of 55 kDa and 23 kDa fractions in an antigens obtained from CICs of all clinical and parasitologically proven untreated visceral leishmaniasis patients before treatment (VL-BT), which ensured absolute sensitivity. However, light expressions of these bands were observed in some VL treated cases. To ascertain the prognostic value, 2D expression profiles of circulating antigens were carried out, which revealed 3 upregulated and 12 induced immunoreactive spots. Out of these, ten prominent spots were excised and subjected for enzymatic digestion to generate peptides. Mass spectrometry (MS) analysis successfully explored 20 peptides derived from kinase, kinesin, acetyl Co-A carboxylase, dynein heavy chains (cytoplasmic and axonemal/flagellar), 60S ribosomal protein, nucleoporin protein, RNA polymeraseII, protease gp63, tubulin, DNA polymerase epsilon subunit, GTP-binding protein and tyrosyl-methionyl t-RNA synthetase-like protein and 19 hypothetical protein of unknown function. Presence of L. donovani proteins in circulating antigens were further validated using anti-Ld actin and anti-α tubulin antibody. Besides, MS derived peptides confirmed its reactivity with patients' sera. Therefore, these shortlisted potential antigens can be explored as antigen-based diagnostic as well as prognostic kit.

Dataset Information

Noninvasive Diagnosis of Visceral Leishmaniasis: Development and Evaluation of Two Urine-Based Immunoassays for Detection of Leishmania donovani Infection in India.

Publications

Noninvasive Diagnosis of Visceral Leishmaniasis: Development and Evaluation of Two Urine-Based Immunoassays for Detection of Leishmania donovani Infection in India.

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