Prevalence and incidence of canine visceral leishmaniasis and its clinical-immunological features in an endemic area of the Brazilian Amazon.

BACKGROUND: A cohort study for 2 years period analysed the prevalence, incidence and clinical-immunological features of canine Leishmania (L.) chagasi-infection in 316 mongrel dogs in a visceral leishmaniasis-endemic area in Pará State, Brazil. OBJECTIVE/METHODS: Diagnosis of infection was performed by the indirect fluorescent antibody test (IFAT-IgG), the leishmanin skin test (LST) and a parasite search (from the popliteal lymph node aspiration) at the beginning of the study and at 6, 12 and 24 months intervals. RESULTS: IFAT/LST revealed three immune profiles of infection: (I) IFAT(+) /LST(-) (81), (II) IFAT(-) /LST(+) (17) and (III) IFAT(+) /LST(+) (13). Prevalence of profiles I, II and III were 25.6, 5.4 and 4.1%, and an overall prevalence 35.1%. Incidence of profiles I, II and III were 5.4, 0.3 and 0.0%, and an overall incidence 5.7% dogs per month. Incidence at the age ranges <1 year, ≥1 year, <7 years and ≥7 years evidenced a highest rate in the age range <1 year (6.6% dogs per month). Parasitological diagnosis was positive in 19% dogs at the prevalence (85.7% profile I), and in 11% at the incidence (100% profile I). The clinical picture of 179 infected dogs showed 145 (81%) of profile I (82% subclinical); 21 (11.7%) of profile II (100% subclinical); and 13 (7.3%) of profile III (84.6% subclinical). Conversion from subclinical to sick dogs was higher (p < 0.05) in profile I (40.2%) than in profiles II (5.8%) and III (9%). Immunological conversion showed that only 3.2% of profile I dogs (prevalence) converted to LST(+) (two at the end of the first 6 months and 1 after 24 months), while 82.3% of profile II dogs converted to IFAT(+) (11 in the first 6 months, whereas three after 12 months). A 100% death rate was observed in dogs from profile I alone. CONCLUSION: These results reinforce the need of adopting preventive strategies against CVL as early as in the first semester of the dog's life.

Phlebotomines (Diptera: Psychodidae) from a Urban Park of Belém, Pará State, Northern Brazil and Potential Implications in the Transmission of American Cutaneous Leishmaniasis.

In urban ecotourism parks, the life cycle of American cutaneous leishmaniasis (ACL) agents can remain established, where phlebotomines may comprise potential risks for visitors. The present study aimed to survey the phlebotomine fauna of a forest park 'Bosque Rodriques Alves-Jardim Botânico da Amazônia' (BRAJBA), in the urban area of Belém, Brazil. The park was monthly surveyed in 2018 using CDC light traps placed in ground and canopy strata. Leishmania spp. isolated from dissected females were characterized by polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP) analysis. Fluctuations in specimen capture were correlated with rainfall. Nyssomyia antunesi (Coutinho, 1939) was predominant for all surveyed ecotopes and capture methods in both areas. Females of Ny. antunesi resting on tree bases were observed attempting to bite researchers during early morning. One Bichromomyia flaviscutellata (Mangabeira, 1942) and one Trichophoromyia brachipyga (Mangabeira, 1942) were found naturally infected by flagellates. Only the strain from Th. brachipyga was isolated and characterized as Leishmania (Viannia) lainsoni Silveira, Shaw, Braga and Ishikawa, 1987. Monthly fluctuations of the three most abundant species, Ny. antunesi, Trichophoromyia ubiquitalis (Mangabeira, 1942) and Th. brachypiga, had statistically significant negative correlations with rainfall. The present study provided further information to better understand ACL ecology in the Belém urban area, where the urban parks surveyed appeared to offer potential risk of contracting the disease, thus requiring environmental management. These observations highlighted the need for including Ny. antunesi, Bi. flaviscutellata, Th. ubiquitalis, and Th. brachypiga in the priority list for continuous entomological surveillance.

A cross-sectional study on canine Leishmania (L.) infantum chagasi infection in Amazonian Brazil ratifies a higher prevalence of specific IgG-antibody response than delayed-type hypersensitivity in symptomatic and asymptomatic dogs.

This was a cross-sectional study which analyzed the prevalence and the clinical and immunological spectrum of canine Leishmania (L.) infantum chagasi infection in a cohort of 320 mongrel dogs living in an endemic area of American visceral leishmaniasis in the Amazonian Brazil by using, mainly, the indirect fluorescence antibody test (IFAT-IgG) and the delayed-type hypersensitivity (DTH), and the parasite research by the popliteal lymph node aspiration. The IFAT and DTH reactivity recognized three different immune response profiles: (1) IFAT((+))/DTH((-)) (107 dogs), (2) IFAT((-))/DTH((+)) (18 dogs), and (3) IFAT((+))/DTH((+)) (13 dogs), providing an overall prevalence of infection of 43% (138/320). Thus, the specific prevalence of IFAT( (+) )/DTH( (-) ) 33.4% (107/320) was higher than those of IFAT( (-) )/DTH( (+) ) 5.6% (18/320) and IFAT( (+) )/DTH( (+) ) 4.0% (13/320). Moreover, the frequency of these profiles among 138 infected dogs showed that the IFAT( (+) )/DTH( (-) ) rate of 77.5% (107/138) was also higher than those of 13.0% (18/138) of IFAT( (-) )/DTH( (+) ) and 9.5% (13/138) of IFAT( (+) )/DTH( (+) ) rates. The frequency of asymptomatic dogs (76%-105) was higher than those of symptomatic (16.6%-23) and oligosymptomatic ones (7.4%-10). A total of 16 (11.6%) L. (L.) i. chagasi isolates were obtained from infected dogs, all from the IFAT( (+) ) /DTH( (-) ) profile: 41% (9/22) from symptomatic, 33.3% (3/9) from oligosymptomatic, and 5.2% (4/76) from asymptomatic dogs. These findings strongly suggested that despite the higher frequency of asymptomatic dogs (76%-105), the majority (72.4%-76) was characterized by the IFAT( (+) ) /DTH( (-) ) profile with a doubtful immunogenetic resistance against infection.

Characterization in vivo and in vitro of a strain of Leishmania (Viannia) shawi from the Amazon Region.

The present study analyses complement resistance, cell surface carbohydrates expression, lipidic composition and morphology in vivo and in vitro, of Leishmania (Viannia) shawi, a parasite identified in the Amazon region, Pará state, in 1989. We demonstrated that promastigotes in the stationary (STAT) growth phase are more resistant to complement lysis than in the logarithmic (LOG) growth phase. Ultrastructural analyses and imidazol technique showed accumulation of lipids in STAT growth phase promastigotes, which was confirmed by biochemical approach. Light and electron microscopy of skin lesion in hamster footpads caused by promastigotes in STAT growth phase, 90 days post inoculation, showed amastigotes inside of macrophage and free in the tissue surrounded by collagen fibers as well as extensive inflammatory reaction with tissue destruction. We also demonstrated, using lectins by agglutination assays and flow cytometry, the presence of fucose, mannose and/or glucose carbohydrate residues on the surface of LOG and STAT promastigotes. The results constitute the first characterization essay combining biochemical and morphological approaches dedicated to LOG and STAT growth phase promastigotes of L. (V) shawi contributing for a better knowledge of this poorly studied species of the New World.

Leishmania (Leishmania) amazonensis-induced cutaneous leishmaniasis in the primate Cebus apella: a model for vaccine trials.

A primate model of leishmaniasis was developed with the objective of future vaccine testing. Lesion development and immunological parameters were studied upon primary and secondary infections. Seven Cebus apella were injected subcutaneously with 2 x 10(6) Leishmania (Leishmania) amazonensis promastigotes. Erythematous nodules appeared 19-29 days p.i., which disappeared 100 days p.i. Four months later, six of the monkeys were challenged with the same inoculum; three of them developed erythematous nodules after 7 days p.i., with ulcer formation in two of these subjects. The lesions were short-lived and all were cured 40 days post challenge. Anti-Leishmania IgG antibodies were detected and they increased after the challenge infection. Leishmania antigen-induced lymphoproliferation was found 1 month post-primary infection, which coincided with IFN-gamma production and lesion development. It decreased to control levels afterwards, but at the time of the challenge dose, it was significantly above the initial level. After the challenge infection, it first increased then decreased sharply at 40 days post-challenge, coinciding with the healing of the lesion. It increased again to a higher level at 60 days post-challenge. Leishmania (Leishmania) amazonensis-infection in C. apella did not induce complete protection against a secondary infection with a homologous parasite although specific antibody production and lymphoproliferation with IFN-gamma production were observed. This fact indicates that vaccine has to be better than infection in the induction of protective immunity, and raises a question on in vitro parameters that should be considered as a counterpart of expected protection induced by vaccine candidate. In addition, we conclude that this is a useful primate model for the evaluation of candidate vaccines.

Asymptomatic human carriers of Leishmania chagasi.

In Brazil, programs based on elimination of infected dogs have not curtailed the spread of visceral leishmaniasis (VL), suggesting that other reservoirs of infection exist. Persons with active VL can infect the sand fly vector, but in endemic areas, persons with asymptomatic infections, whose infectivity to sand flies is unknown, are far more numerous. In this study, a polymerase chain reaction-based assay detected kinetoplast DNA of Leishmania chagasi in the blood of eight of 108 asymptomatic persons living with patients with recently diagnosed VL. These eight persons had low or unmeasurable levels of IgG antibodies to Leishmania, demonstrating the insensitivity of serology for subclinical infection. All eight persons had positive leishmanin skin test results, as did 70% of persons living in households of persons with active VL. Even if a small proportion of such asymptomatic persons are infective to sand flies, they represent a formidable reservoir of infection in endemic areas.