Differential diagnosis of cutaneous leishmaniasis by immunoassays with density-controlled presentation of glycan biomarkers - Leishmaniasis is an infectious vector-borne disease caused by a number of different species of the protozoan parasite
Leishmania. There are different forms of leishmaniasis, the most common being cutaneous leishmaniasis (CL), which is
associated with large, open skin ulcers that may heal over many months, but can lead to complications due to secondary
infections. CL affects 12 million people worldwide, particularly in Northern Africa, the Middle East, Asia, and in Latin
America. Mostly due to military deployment to endemic regions (e.g., Afghanistan and Iraq), CL is also on the rise in the
U.S. Proper diagnostic tools (PCR) are often not available in developing countries, and the presentation of CL can be easily
confused with other skin conditions. Even though a number of drugs – most of which are highly toxic– are available to treat
CL, there is no perfect drug with equal efficacy for all CL-causing Leishmania species. For example, ketoconazole is
effective for CL caused by L. major, but it is almost entirely ineffective against L. tropica. Thus, there is a need not only for
reliable diagnostic tools for the accurate diagnosis of CL, but also for the differentiation between CL infections by different
Leishmania species, which is important to determine drug treatment regimens. Leishmania spp. express unusual glycans
on their cell surfaces that are foreign to humans, which may elicit a strong antibody (IgG) response. Specifically, it is known
that the glycoinositol phospholipids (GIPLs) of L. major contain unusual α-galactopyranosyl (αGal) and β-galactofuranosyl
(βGalf)-residues. Likewise, L. tropica expresses lipophosphoglycans (LPGs) that contain β-arabinose (βAra), suggesting
that L. major and L. tropica could be serologically distinguishable. Our preliminary data show that the reactivities of sera
from CL patients against certain synthetic αGal and/or βGalf epitopes differ depending on the Leishmania species that
causes the underlying CL infection. However, some antibody cross-reactivities are also observed. Whitesides and others
have demonstrated that the density at which glycan antigens are presented has a major influence on antibody recognition
and specificity. We hypothesize that αGal- and βGalf-containing glycans that exist in the GIPLs of L. major, and βAra-
containing glycans that exist in the LPGs of L. tropica, cause a strong IgG response in CL patients. We further hypothesize
that varying the densities at which certain immunodominant glycotopes are displayed will identify conditions at which these
glycans show large differentials in enzyme-linked immunosorbent assay (CL-ELISA) between CL patients and the control
group, and between CL patients with L. major and L. tropica infections. The specific aims are to synthesize αGal-, βGalf-, or
βAra containing partial structures of the cell surface glycans of L. major and L. tropica, and to identify those that are strongly
recognized by antibodies in the sera of CL patients that are infected with L. major or L. tropica using CL-ELISA. The most
promising glycans will be subjected to CL-ELISA at different glycan densities. The goal is to identify conditions under which
a glycan shows large differentials between CL patients and the control group, and also between CL patients with L. major or
L. tropica infection. Can the newly discovered CL biomarkers indeed achieve a differential diagnosis between L. major and
L. tropica infections? Their specificity will be put to a test by screening the individual sera of a large cohort of CL patients
with an active L. major or L. tropica infection by CL-ELISA.
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