Identification of Leishmania donovani antigen in circulating immune complexes of visceral leishmaniasis subjects for diagnosis

Abstract

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.

RETRACTED: Leishmania-Specific Promiscuous Membrane Protein Tubulin Folding Cofactor D Divulges Th1/Th2 Polarization in the Host via ERK-1/2 and p38 MAPK Signaling Cascade

“…In the present study, a holistic approach has been proposed for the simultaneous elicitation of both T-and B-cells along with the mediation of long-term immunity against VL infection (16,17). The data of the present study establish the vaccine potential of CIC-derived B-cell epitopes and their combination with a potent T-cell epitope to achieve a desirable immune response in the host (3,14). We also explored the possible involvement of ERK-1/2 and p38 MAPK signaling cascade in the observed host immune cell activation.…”

“…The prevalent circumstances warrant the introduction of potent immune-modulators, which can be used along with a candidate antigen to develop a promising vaccination strategy. Our earlier study revealed the presence of numerous potent Leishmania donovani antigens in the systemic circulation of infected patients (3). Among several identified parasite antigens, the predominant abundance of tubulin folding cofactor D (TFC-D) along with repertoire of its specific antibodies in circulating immune complexes (CICs) suggests its plausible involvement in the activation of parasite-specific B-cell responses (3).…”

Glycoproteins in circulating immune complexes are biomarkers of patients with Indian PKDL: A study from endemic districts of West Bengal, India

“…Formation of ICs is a normal part of the immune defense to remove pathogen. More recently, Jamal et al , 2017 [ 48 ] have demonstrated a 36 fold higher ICs in human VL subjects. In our present study, we observed almost 5 fold higher ICs in human PKDL subjects.…”

“…Among them, distinct seven bands were found to be glycoprotein in nature as demonstrated by PAS silver double stain and lectin blot ( Fig 3D , Fig 4B ). Notably, the 162 kDa, 109 kDa, 70 kDa MW CIC protein bands have similar MW as glycoproteins found on amastigotes and promastigotes of Leishmania donovani [ 60 ], whereas other protein bands like 61 kDa, 55 kDa, 45 kDa and 37 kDa had MW similar to those found in Leishmania antigen [ 48 ]. All the above mentioned seven protein bands were present in immunoblot ( Fig 4A ).…”

Digitoxigenin presents an effective and selective antileishmanial action against Leishmania infantum and is a potential therapeutic agent for visceral leishmaniasis

“…Its diagnosis is accomplished by molecular and/or conventional parasitological techniques, as well as by immunological assays. However, accurate diagnosis depends on the current clinical status and immune response of the infected hosts, with falsenegative and/or false-positive results being frequently Section Editor: Sarah Hendrickx encountered (Sakkas et al 2016;Jamal et al 2017;Van Griensven and Diro 2019).…”