IgG seroprevalence of Toxoplasma gondii and Leishmania infantum in leprosy patients: Implications for screening and management of co-infections.

Parasitic co-infections are common in developing countries and can interfere with leprosy treatment, leading to an increased risk of inflammatory leprosy reactions. This study assessed serum immunoglobulin G (IgG) levels against Toxoplasma gondii and Visceral Leishmaniasis (VL) antigens in 270 leprosy patients from Brazilian states. Regarding the respective cut-offs, the prevalence of IgG seropositivity for T. gondii and VL were 21.05 % and 47.36 % in the leprosy-negative group, and 77.7 % and 52.6 % in the leprosy-positive group. Of the 270 leprosy patients, 158 (58.5 %) presented with inflammatory leprosy reactions. Of those, 72 (59.5 %) had neuritis, 35 (48.6 %) had reverse reactions, and 28 (38.9 %) had ENL in both Brazilian states. Leprosy patients with anti-Leishmania IgG seropositivity were 3.25 times more likely to develop neuritis (95 % C.I.: 1.187 - 9.154; p = 0.019). These findings are particularly relevant for clinical settings where both leprosy and parasitic diseases are prevalent and could provide essential guidance for detecting and addressing complications arising from parasitic co-infections in leprosy patients, thereby improving clinical management strategies.

Immunogenic mapping of rDyn-1 and rKDDR-plus proteins and selection of oligopeptides by immunoblotting for the diagnosis of Leishmania infantum-infected dogs.

Endemic in Brazil, visceral leishmaniasis (VL) is a zoonotic infection that is among the most important parasitic diseases transmitted by vectors. Dogs are the main reservoirs of canine leishmaniasis (CanL) and their identification is used in some countries as part of disease prevention and control measures in the canine and human population. In this context, serological tests are necessary, composed of antigens capable of correctly identifying infected dogs, minimizing the number of false-negative cases. This study aimed to identify more immunoreactive peptides derived from two previously described whole proteins (rDyn-1 and rKDDR-plus) and compare their performance to the control antigens rK39 and the crude extract for the detection of dogs infected with L. infantum, especially the asymptomatic ones. The three selected peptides and a mixture of them, along with the rDyn-1, rKDDR-plus, rK39, and crude extract antigens were evaluated using indirect ELISA with sera samples from 186 dogs with CanL, being asymptomatic (n = 50), symptomatic (n = 50), co-infected (n = 19), infected with Babesia sp. (n = 7), Ehrlichia sp. (n = 6), T. cruzi (n = 20) and uninfected (n = 34). The results showed that the rDyn-1 protein and the peptide mixture had the highest sensitivity (100% and 98.32%, respectively) and specificity (97.01 and 98.51, respectively). A high degree of kappa agreement was found for rDyn-1 protein (0.977), mixed peptides (0.965), rKDDR-plus protein (0.953), K-plus peptide 1 (0.930) and Dyn-1 peptide (0.893). The mixture of peptides showed the highest likelihood (65.87). The ELISA using the mixture of peptides and the rDyn-1 protein showed high performance for CanL serodiagnosis. More mix combinations of the peptides and additional extended field tests with a larger sample size are recommended.

The increased presence of repetitive motifs in the KDDR-plus recombinant protein, a kinesin-derived antigen from Leishmania infantum, improves the diagnostic performance of serological tests for human and canine visceral leishmaniasis.

Visceral leishmaniasis (VL) is caused by protozoa belonging to the Leishmania donovani complex and is considered the most serious and fatal form among the different types of leishmaniasis, if not early diagnosed and treated. Among the measures of disease control stand out the management of infected dogs and the early diagnosis and appropriate treatment of human cases. Several antigens have been characterized for use in the VL diagnosis, among them are the recombinant kinesin-derived antigens from L. infantum, as rK39 and rKDDR. The main difference between these antigens is the size of the non-repetitive kinesin region and the number of repetitions of the 39 amino acid degenerate motif (6.5 and 8.5 repeats in rK39 and rKDDR, respectively). This repetitive region has a high antigenicity score. To evaluate the effect of increasing the number of repeats on diagnostic performance, we designed the rKDDR-plus antigen, containing 15.3 repeats of the 39 amino acid degenerate motif, besides the absence of the non-repetitive portion from L. infantum kinesin. Its performance was evaluated by enzyme-linked immunosorbent assay (ELISA) and rapid immunochromatographic test (ICT), and compared with the kinesin-derived antigens (rKDDR and rK39). In ELISA with human sera, all recombinant antigens had a sensitivity of 98%, whereas the specificity for rKDDR-plus, rKDDR and rK39 was 100%, 96% and 71%, respectively. When evaluated canine sera, the ELISA sensitivity was 97% for all antigens, and the specificity for rKDDR-plus, rKDDR and rK39 was 98%, 91% and 83%, respectively. Evaluation of the ICT/rKDDR-plus, using human sera, showed greater diagnostic sensitivity (90%) and specificity (100%), when compared to the IT LEISH (79% and 98%, respectively), which is based on the rK39 antigen. These results suggest that the increased presence of repetitive motifs in the rKDDR-plus protein improves the diagnostic performance of serological tests by increasing the specificity and accuracy of the diagnosis.

Leishmania infantum recombinant kinesin degenerated derived repeat (rKDDR): A novel potential antigen for serodiagnosis of visceral leishmaniasis.

Visceral leishmaniasis (VL) or kala-azar, the most severe form of leishmaniasis, can lead to death if not properly diagnosed and treated. Correct identification of infected patients and reservoirs is vital for controlling the spread of leishmaniasis. Current diagnostic kits for leishmaniasis show high sensitivity and specificity, but can also result in false negatives and cross reactions with related parasitic infections. New diagnostic methods with greater accuracy are urgently needed for diagnosis of leishmaniasis. In this study, we aimed to uncover a new highly effective antigen for the diagnosis of visceral leishmaniasis in dogs and humans, aiming to improve the accuracy compared with those of current methods of diagnosis. Initially, in-silico epitope prediction analyses identified several potential B-cell epitopes in the repetitive region of Leishmania infantum kinesin, which co-localized with predicted structural disordered regions, suggesting high potential for antigenicity. Based on this analysis, 8.5 genomic motifs, which encode the repetitive sequence of 39 degenerate amino acids, were selected for recombinant expression. BLASTn analysis of this repetitive region indicated that it is absent in the T. cruzi parasite, which is closely related to Leishmania, indicating the specificity of this region. This potentially antigenic protein, named recombinant kinesin degenerated derived repeat (rKDDR), was recombinantly expressed in Escherichia coli BL21-Star using the pET28a-TEV expression vector. We then evaluated the performance of rKDDR in correctly diagnosing Leishmania infection and compared this new assay with currently used diagnostic tests for leishmaniasis. rKDDR showed greater sensitivity and specificity in correctly diagnosing leishmaniasis both in human (sensitivity 92.86% and specificity 100%) and canine (sensitivity 88.54% and specificity 97.30%) sera compared with those of rK39 (human: sensitivity 90.48% and specificity 97.92%; canine: sensitivity 78.13% and specificity 90.09%). In addition, the rKDDR-ELISA outperformed the EIE-LVC kit, which is the serologic kit recommended by the Brazilian Ministry of Health for the diagnosis of canine visceral leishmaniasis. These results indicate that rKDDR is a highly promising candidate for diagnosis of visceral leishmaniasis, and is more accurate than the currently used gold-standard antigens.