Identification of Canine Visceral Leishmaniasis in a Previously Unaffected Area by Conventional Diagnostic Techniques and Cell-Block Fixation

Abstract

After the report of a second case of canine visceral leishmaniasis (CVL) in São Bento da Lagoa, Itaipuaçu, in the municipality of Maricá, Rio de Janeiro State, an epidemiological survey was carried out, through active search, totaling 145 dogs. Indirect immunofluorescence assay (IFA), enzyme-linked immunosorbent assay (ELISA), and rapid chromatographic immunoassay based on dual-path platform (DPP(r)) were used to perform the serological examinations. The parasitological diagnosis of cutaneous fragments was performed by parasitological culture, histopathology, and immunohistochemistry. In the serological assessment, 21 dogs were seropositive by IFA, 17 by ELISA, and 11 by DPP(r), with sensitivity of 66.7%, 66.7% and 50%, and specificity of 87.2%, 90.2% and 94%, respectively for each technique. The immunohistochemistry of bone marrow using the cell-block technique presented the best results, with six positive dogs found, three of which tested negative by the other parasitological techniques. Leishmania sp. was isolated by parasitological culture in three dogs. The detection of autochthonous Leishmania infantum in Itaipuaçu, and the high prevalence of seropositive dogs confirm the circulation of this parasite in the study area and alert for the risk of expansion in the State of Rio de Janeiro.

Fatores ambientais associados à ocorrência de leishmaniose visceral canina em uma área de recente introdução da doença no Estado do Rio de Janeiro, Brasil

“…CVL prevalence in this study (21.65%) was similar to rates in Rio de Janeiro State both in areas of recent introduction (25.2% in Itaipuaçu, Maricá; 18.1% on Ilha de Marambaia, Rio de Janeiro) and in areas classified as endemic (25% in Barra de Guaratiba, Rio de Janeiro) 15,23,24 , notwithstanding differences in methodologies. Prevalence rates can vary due to the diagnostic test used and the way dogs were recruited for the study 25 The geographic expansion of CVL has been recorded in other states of Brazil, attributed to various factors such as difficulties in eliminating reservoirs, epidemiological diversity of the affected regions, high financial costs of sustaining control measures, the vector's high capacity to adapt to the peridomicile, and insufficient control measures 26 .…”

“…Dogs that tested positive dogs in the DPP assay were sedated to obtain full skin samples from the scapular region and bone marrow aspirates, according to the protocol by Abrantes et al 15 . Skin samples were packed in phosphate buffer saline (PBS) solution, with the addition of antibiotics (penicillin and streptomycin) and an antifungal (fluorocytosine), and the bone marrow samples were stored directly in Novy, MacNeal, Nicole (NNN) biphasic culture, both transported under refrigeration for processing at the Laboratory for Clinical Research and Surveillance in Leishmaniasis (LapClinVigiLeish) at the National Institute of Infectolgy (INI/Fiocruz) for parasitological culture based on the protocol by Silva et al 21 .…”

“…New areas have been identified with the occurrence of canine visceral leishmaniasis (CVL) in various municipalities (counties) in the State of Rio de Janeiro 15,16,17 . However, little is known about the environmental factors involved in the emergence of CVL in these areas.…”

Leishmania infantum Virulence Factor A2 Protein: Linear B-Cell Epitope Mapping and Identification of Three Main Linear B-Cell Epitopes in Vaccinated and Naturally Infected Dogs

“…In the control group (T0) and naturally infected dogs, bone marrow (BM) samples were also collected, according to the methodology of Abrantes et al ( 26 ). Parasitological culture was performed according to the protocol of Ref.…”

Can vaccines against canine visceral leishmaniasis interfere with the serological diagnostics recommended by the Brazilian Ministry of Health?

“…The protocol for bone marrow and blood collections and the procedure for parasitological culture were conducted as previously described by ABRANTES et al (2016).…”