Reliability of serological methods for detection of leishmaniasis in Portuguese domestic and wild reservoirs

A direst agglutination test (DAT) and an immunofluorescence (IFAT) were compared for detection of Leishmania infantum infection in 43 dogs and five foxes from Alto-Douro and Arrabida, two known endemic areas in Portugal. In four dogs with proved canine leishmaniasis, both DAT and IFAT showed positive readings (titres ò1:320 and ò1:128). Of 34 samples collected form apparently healthly dogs, ten were positive by both serological tests and eight were serologically positive by one test or the other. Three foxes out of five captured in this area, scored titres indicative of leishmaniasis in both DAT and IFAT. The concordance between DAT and IFAT in all collected samples (48) was 81.25 per cent. Considering these and previous studies in the adjancent Mediterranean areas, the seroprevalence of L. infantum infection in the canine and vulpine populations appear to be high magnitude.

Reliability of serological methods for detection of leishmaniasis in Portuguese domestic and wild reservoirs.

A direct agglutination test (DAT) and an immunofluorescence technique (IFAT) were compared for detection of Leishmania infantum infection in 43 dogs and five foxes from Alto-Douro and Arrábida, two known endemic areas in Portugal. In four dogs with proved canine leishmaniasis, both DAT and IFAT showed positive readings (titres > or = 1:320 and > or = 1:128). Of 34 samples collected from apparently healthy dogs, ten were positive by both serological tests and eight were serologically positive by one test or the other. Three foxes out of five captured in this area, scored titres indicative of leishmaniasis in both DAT and IFAT. The concordance between DAT and IFAT in all collected samples (48) was 81.25%. Considering these and previous studies in the adjacent Mediterranean areas, the seroprevalence of L. infantum infection in the canine and vulpine populations appear to be of high magnitude.

Serodiagnosis of canine visceral leishmaniasis in Portugal: comparison of three methods.

Sera collected in Portugal from 43 dogs were screened for specific antibodies to Leishmania donovani antigens. Three different techniques were compared: an indirect immunofluorescence assay (IFA), a direct enzyme-linked immunosorbent assay (ELISA) and a competitive-ELISA (C-ELISA) using two species-specific monoclonal antibodies, D2 and D13. By IFA, 22 of the sera examined showed positive reactions, compared with 26 by ELISA or 27 by C-ELISA. There was no direct correlation observed between the serum titre by IFA and the strength of the reaction in ELISA or inhibition in C-ELISA. However, a good correlation was observed between sera identified as positive (95.5%) by all three techniques. Western blotting on leishmanial membranes showed that common antigens with Mr of 26,000 and 70-84,000 were recognized by all infected dog sera, regardless of the serum titre. In large scale studies, ELISAs are preferred to IFA for the rapid diagnosis of canine visceral leishmaniasis because of their greater simplicity.

Canine leishmaniasis: pathological and ecological factors influencing transmission of infection.

Canine leishmaniasis was studied in 1,823 dogs from the Lisbon metropolitan region. The breeds most affected were doberman and German shepherd, independent of sex and use. Young adult (12.2%) and older dogs (14.7%) had higher prevalences of infection. Parasitological confirmation of serological diagnosis was higher in dogs with indirect fluorescent antibody test titer greater than or equal to 1:512, indicating that parasitological patency is a late event. Exposure of Leishmania in lymph nodes is more efficient for parasitological confirmation (75.4% of cases). Frequent signs of disease were enlarged lymph nodes and onychogriphosis. However, 53.8% of the dogs with significant antibody titers (greater than or equal to 1:128) showed no symptom, suggesting that canine leishmaniasis has a prolonged asymptomatic period. This study confirmed the importance of the dog as the reservoir of visceral leishmaniasis.

Hepatozoonosis in foxes from Portugal.

Hepatozoon, sp. is described for the first time in foxes (Vulpes vulpes silacea) in Portugal. Of 301 foxes examined, 143 (48%) were infected. The gametocyte was the predominant stage of the life cycle and was found in every organ except the bone marrow, where schizonts were the most abundant stage. The morphological similarity of this parasite's gametocytes to Hepatozoon canis is emphasized.

Kala-azar in Portugal. V. The sylvatic cycle in the enzootic endemic focus of Arrabida.

Leishmania parasites were found in three of 43 foxes examined in the Arrabida region of the Setúbal peninsula in Portugal, this brings the total number of cases of vulpine leishmaniasis to four in 71 animals examined in this area, a prevalence rate of 5.63%. The three parasitized foxes found in this study were all symptomless and NNN cultures inoculated with tissue extracts were positive for all three animals. The immunofluorescence antibody test (IFAT) was positive with titres greater than or equal to 1/128 in the three infected foxes. The isoenzyme electrophoretic profile obtained from these positive cultures showed the strains to be identical with others isolated in Portugal from man, dog and fox, as well as with strains from elsewhere in the Mediterranean (zymodeme 1). The prevalence rate of 5.63% is probably sufficient to maintain endemicity. The presence of a semi-autonomic sylvatic cycle in the area seems likely. Hepatozoon was found in 28 foxes (65.11%).