Evaluation of the diagnostic potential of CD1a immunohistochemistry for visceral leishmaniasis

Abstract

Visceral Leishmaniasis is a public health problem caused by protozoans of the genus Leishmania. K39 serological test is commonly used in the initial investigation, with high specificity, but variable sensitivity. Amastigotes can be identified by optical microscopy, however, the differential diagnosis with cellular debris or other intracellular parasites is necessary. Recent studies have raised the possibility of using immunohistochemistry in the diagnosis of visceral leishmaniasis with labeling of amastigotes by the anti-CD1a antibody. This retrospective study was based on 38 samples from patients with visceral leishmaniasis whose diagnoses were confirmed by myelogram and/or k39 testing, aside from positive (N=13) and negative biopsies (N=25), 2 samples from patients with false positive biopsies for visceral leishmaniasis and 8 samples from patients with histoplasmosis diagnosis. The histological slides were evaluated for the presence of amastigotes and their Modified Ridley Parasitic Index. The samples were submitted to immunohistochemical reactions using the anti-CD1a antibody with MTB1 and O10 clones. Immunohistochemical reactions with MTB1 and O10 clones had low sensitivity in this study. However, all bone marrow samples were previously decalcified with nitric acid which is probably a deleterious treatment for immunohistochemical reactions in this site. Excluding these samples, we obtained 58.33% sensitivity and 100% specificity with the MTB1 clone. Despite the intermediate sensitivity, the immunohistochemistry for the CD1a marker with clone MTB1 can be useful in the differential diagnosis of visceral leishmaniasis, helping to discriminate leishmania amastigotes from other pathogens with similar morphology and cellular debris in different samples, except in bone marrow biopsies previously decalcified with nitric acid.

Figure 1. Amastigotes within macrophages are observed on the left side, in a duodenal sample with a modified Ridley parasite index of 6 (H&E, 1000x). A slide of this same sample is shown with the immunohistochemical reaction using the clone MTB1 of CD1a (1000x) on the right side. At the bottom, on the right corner, an amastigote is shown in detail, stained by immunohistochemistry in a pattern of peripheral membrane-like reinforcement, leaving the unstained nucleus in the center, and presenting with a reinforcement in one pole, apparently because of the kinetoplast staining (Digitally magnified. Original 1,000X magnification).