Tassia Cristina Bello de Vasconcelos

Canine susceptibility to visceral leishmaniasis: A systematic review upon genetic aspects, considering breed factors and immunological concepts

Dogs have different susceptibility degrees to leishmaniasis; however, genetic research on this theme is scarce, manly on visceral form. The aims of this systematic review were to describe and discuss the existing scientific findings on genetic susceptibility to canine leishmaniasis, as well as to show the gaps of the existing knowledge. Twelve articles were selected, including breed immunological studies, genome wide associations or other gene polymorphism or gene sequencing studies, and transcription approaches. As main results of literature, there was a suggestion of genetic clinical resistance background for Ibizan Hound dogs, and alleles associated with protection or susceptibility to visceral leishmaniasis in Boxer dogs. Genetic markers can explain phenotypic variance in both pro- and anti-inflammatory cytokines and in cellular immune responses, including antigen presentation. Many gene segments are involved in canine visceral leishmaniasis phenotype, with Natural Resistance Associated Macrophage Protein 1 (NRAMP1) as the most studied. This was related to both protection and susceptibility. In comparison with murine and human genetic approaches, lack of knowledge in dogs is notorious, with many possibilities for new studies, revealing a wide field to be assessed on canine leishmaniasis susceptibility research.

Cytokine and iNOS profiles in lymph nodes of dogs naturally infected with Leishmania infantum and their association with the parasitic DNA load and clinical and histopathological features

In South America, visceral leishmaniasis is a zoonotic disease with severe evolution characteristics in humans, and dogs are its main reservoir. In this context, this study aimed to evaluate the clinical status of dogs from a Brazilian endemic area naturally, at Barra Mansa municipality, infected with Leishmania infantum, in conjunction with their histopathological profile and, in order to determine possible markers of susceptibility or resistance to the disease, parasitic DNA load, cytokine and iNOS mRNA expression profiles were investigated in lymph nodes. High levels of IFN-ɣ and IL-6 mRNA were detected. Both IFN-ɣ and IL-6 mRNA were associated with disorganization of the corticomedullary region. IFN-ɣ and TNF-α mRNA were associated with the absence of follicular hyperplasia. The regulatory pathway was remarkable with IL-10 mRNA detection and its significant association with the severity of the disease. Plasmacytosis and sinus histiocytosis were associated with high loads of parasitic DNA, but there was no significant association between the parasite DNA load and animal clinical alterations. Since high parasitic loads were found in animals with or without symptoms, clinical examination cannot be considered as a criterion for disease susceptibility assessment.

Validation of the Dual-path Platform chromatographic immunoassay (DPP® CVL rapid test) for the serodiagnosis of canine visceral leishmaniasis

BACKGROUND Visceral leishmaniasis is a major public health challenge in South America, and dogs are its main urban reservoir. OBJECTIVE Validation of the canine Dual-path Platform immunoassay for canine visceral leishmaniasis (DPP® CVL) for a sample set composed of 1446 dogs from different Brazilian endemic areas. METHODS A well-defined reference standard by means of parasitological culture, immunohistochemistry, and histopathology was used. Animals were classified as asymptomatic, oligosymptomatic, or symptomatic. Sensitivity and specificity were assessed as a single set and in clinical groups. A reproducibility assessment of the tests was conducted using the Kappa (κ) index at three different laboratories (A, B, and C). FINDINGS Overall, 89% sensitivity and 70% specificity were obtained for the entire sample set. Analysis of the clinical groups showed a gradual decrease in the sensitivity and an increase in the specificity with the reduction of clinical signs in the dogs that were assessed, reaching a sensitivity of 75% (42.8-94.5%) among asymptomatic dogs and lower specificity of 56% (46.2-66.3%) among symptomatic dogs. Inter-laboratory agreement was substantial (κAB= 0.778; κAC= 0.645; κCB= 0.711). MAIN CONCLUSIONS The test performance is somewhat dependent on canine symptomatology, but such influence was less evident than in previous studies. Favourable results for sensitivity and specificity can be obtained even in asymptomatic animals; however, caution is needed in these evaluations, and the results suggest that the immunochromatographic test may be further improved for better investigation in asymptomatic dogs. The results obtained confirm the usefulness of DPP® CVL for application in serological surveys.

Serological Diagnosis of Feline Tegumentary Leishmaniasis by Indirect Immunofluorescence (IFI) and Enzyme-Linked Immunosorbent Assay (ELISA) in an Endemic Area in Brazil

Background: American tegumentary leishmaniasis (ATL) is a serious public health problem, and the participation of domestic cats in its epidemiological process has not yet been fully elucidated. Therefore, the standardization of monitoring methodologies of cat populations becomes important for the generation of information on the disease. In Brazil, ATL presents a wide geographical distribution, being Leishmania (Viannia) braziliensis its etiologic agent of ATL in Rio de Janeiro. The main objective of the present study was investigate the presence of specific anti-Leishmania (Viannia) braziliensis Immunoglobulin G (IgG) in 34 cats from an ATL endemic area in this municipality. Materials, Methods & Results: Sera from three cats from the study area naturally infected with Leishmania (Viannia) braziliensis were used as positive control. Analyses were performed with antigen preparations using indirect immunofluorescence (IFI) and enzyme-linked immunosorbent assay (ELISA). We found prevalence rates of 20.6% (7/34) in the IFI and 14.7% (5/34) in the ELISA. Specificity was 87.1% for the IFI and 93.5% for the ELISA, and both tests presented sensitivity of 100%. Concordance between the used tests was assessed as moderate. Discussion: By conducting a feline serological survey in an endemic area for ATL we provide information on the involvement of this species in such epidemiological process. In this context, the participation of the cat in cycles of leishmaniases has been highlighted by several authors; infection of the phlebotomine by parasites originated from this vertebrate host can already be demonstrated by xenodiagnosis and several cases of feline infection have been reported in endemic regions of several countries such as Brazil, Spain, France, Portugal, Greece, Iran, Israel and Italy. Therefore, the standardization of diagnostic techniques that support the screening and monitoring of such populations becomes of utmost importance. In fact, the use of the IFI and ELISA serological techniques, showing sensitivity and NPV of 100%, confirm the use of such tools in epidemiological surveys for the detection of anti-L. (V.) braziliensis antibodies in domestic cats, and these methodologies are frequently employed in canine surveys showing good results. The serological titers found in this study for Leishmania (V.) braziliensis are similar to those of studies carried out with cats in endemic areas of visceral leishmaniasis in Spain. This similarity shows that the titers can be high even when L. (V.). braziliensis is the etiological agent. It is worth mentioning that these values are significant when compared with those of canine studies carried out in ATL endemic regions. Ultimately, the sample refers to an ATL endemic region located in Brazil, one of the countries with the highest endemicity levels of cutaneous leishmaniasis in the world and, therefore, an important area for the implementation and evaluation of diagnostic techniques for the generation of data on the disease and its possible hosts. Thus, the present study provides information on the occurrence of ATL by Leishmania (V.) braziliensis in domestic cats; demonstrates that the methodologies employed were favorable to the use of such techniques as tools in epidemiological surveys for the diagnosis of feline tegumentary leishmaniasis; and paves the way for further studies on the subject.

Avaliação da confiabilidade entre dois observadores em exames citopatológico e imunocitoquímico de aspirado de medula óssea no diagnóstico da leishmaniose visceral canina

Visceral leishmaniasis is a zoonosis in which the dog appears as the main source of infection in urban areas. Its diagnosis is complex and the cytopathological exam is a fast and cheap alternative to parasite direct visualization and its sensitivity can be increased by immunocytochemistry, though with a higher cost. The accuracy of such methods is dependent on the microscopists experience and therefore, this study evaluated the reliability of such techniques between two observers, from bone marrow aspirates of 50 dogs from an endemic area for the disease. The parasitological culture in Novy-MacNeal-Nicolle medium was used as the reference standard. Among the main findings, the sensitivities obtained by observers I and II were respectively 62.5% and 37.5%, while specificities were 81.1% and 100%. On immunocytochemistry evaluation, the sensitivity was 0% for both evaluators and the specificity 97.3% and 100%. The agreement between evaluators was weak (κ = 0.167) for the cytopathological test and it could not be evaluated for immunocytochemistry, for which there was no detection by the evaluator II. The agreements among the diagnostic methods and the standard reference for the observer I were reasonable (κ = 0.364) for cytopathological examination and bad (κ = -0.041) for immunocytochemistry. For observer II, such agreement could be assessed only for the cytopathological test, being moderate (κ = 0.497). The results point to the possible expertise difference between evaluators, with the evaluator II demonstrating greater experience when interpreting the citopathological test. Although there was the expected sensitivity increase with immunocytochemistry, the technique used in this study was not effective for the diagnosis of infection, regardless of the evaluator.