Ability of immunodiagnostic tests to differentiate between dogs naturally infected with Leishmania infantum and Leishmune®-vaccinated dogs

Abstract

Visceral leishmaniasis (VL) is a serious chronic disease with a lethality rate of up to 10% in humans. In urban areas of Brazil, dogs are the main reservoirs of the etiological agent (Leishmania infantum) of VL, and the Brazilian Ministry of Health recommends the euthanasia of animals that are seropositive in both the immunochromatographic dual path platform rapid test (DPP(®); Bio-Manguinhos) and the enzyme-linked immunosorbent assay (ELISA) with an L. major-like antigen (Bio-Manguinhos). Vaccination is an additional tool in the control of canine VL, but the use of Leishmune(®) (Zoetis Indústria de Produtos Veterinários, São Paulo, SP, Brazil), which contains the fucose mannose ligand (FML) isolated from L. donovani, is not currently recommended by the Brazilian Ministry of Health because vaccinated animals may exhibit positive serology and there are reservations regarding the efficacy of the vaccine. The aims of the present study were: (i) to verify the abilities of the fast agglutination screening test (FAST), the direct agglutination test (DAT), the indirect fluorescent-antibody test (IFAT), the DPP rapid test, and ELISA tests with L. major-like and FML antigens to differentiate between L. infantum-infected and Leishmune(®)-vaccinated dogs, and (ii) to analyze the sensitivities and specificities of the different methods. The reactivities to these tests of Leishmune(®)-vaccinated dogs (n = 71), asymptomatic (n = 20) and symptomatic (n = 20) naturally infected dogs, and unvaccinated healthy control dogs (n = 5) were compared. None of the Leishmune(®)-vaccinated dogs tested seropositive in FAST and DAT, although one dog was reactive to DPP and four dogs to ELISA/L. major-like and IFAT tests. While 69 (97%) of vaccinated dogs reacted to ELISA/FML, only one was seropositive in both ELISA/L. major-like and IFAT tests. Individually, all immunodiagnostic tests presented high specificities and positive likelihood ratios (LR+), and high specificity values were obtained when the tests were considered in pairs. However, sensitivity and LR- values were low for ELISA/L. major-like and IFAT tests individually, and for all pair combinations of tests except for FAST with DPP.

The impact of canine leishmaniosis vaccination with Canileish® in Leishmania infantum infection seroprevalence studies

“…In a country where detection and culling of infected dogs is the control measure established by the Ministry of Health (Ministério da Saúde Brasileiro, 2014), this would pose a risk to healthy vaccinated dogs, which could be mistakenly identified as naturally infected individuals and removed (Marcondes et al, 2013). A more recent study demonstrated that Leishmune ® vaccinated dogs did not test positive with the fast agglutination screening test (FAST) or the direct agglutination test (DAT) (Ribeiro et al, 2015), both based on whole L. donovani promastigote antigen, which could eventually be used as confirmatory diagnostic methods for seropositive vaccinated dogs. Meanwhile, Leishmune® vaccine was withdrawn from the market by the Brazilian Ministry of Agriculture due to lack of effectiveness evidence in phase III trials (MAPA, 2014) and no further cross-reactivity studies were performed.…”

Commercially approved vaccines for canine leishmaniosis: a review of available data on their safety and efficacy

“…In contrast, Marcondes et al (46,47), detected sustained seropositivity up to six months post-vaccination in Leishmune®-immunized dogs, which could not be differentiated by the official diagnostic tests in 11.1-72.2% of cases if the ELISA kit was used, while 5.5-33.3% of vaccinated dogs would be detected by the DPP® test (Dual Path Platform® CVL rapid test, Biomanguinhos, Fiocruz, Rio de Janeiro, Brazil). In samples collected from 71 dogs 45 days after the first Leishmune® annual booster, seropositivity was detected in 5.8% and 1.4% with the official ELISA kit and the DPP® test, respectively (48). De Amorim et al (49) described different humoral immunological profiles for Leishmune®-vaccinated and naturally infected dogs depending on the antigen used [FML vs Leishmania soluble antigen (LSA)], showing that the type of antigen employed interfered with immunoglobulin detection, but without allowing a clear distinction between the two groups of dogs.…”

Possibilities and challenges for developing a successful vaccine for leishmaniasis

“…FML has been characterized as a major antigenic complex of L. donovani and the main antigen in this complex is NH36, an essential enzyme involved in the construction of the parasite’s DNA [ 89 ]. Leishmune is considered a promising tool for the prevention of canine visceral leishmaniasis and, furthermore, its potential as a transmission-blocking vaccine is promising for the control of zoonotic visceral leishmaniasis [ 90 ]. Partially or fully purified Leishmania fractions/subfractions have been widely used in experimental models owing to their excellent immunoprotective properties.…”