Sobre a sensibilidade da cultura de leucócitos circulantes na detecção de Leishmania no sangue periférico de pacientes com leishmaniose tegumentar

Foi investigada a presença de Leishmania, através da cultura de leucócitos circulantes, no sangue periférico de 60 pacientes portadores de leishmaniose tegumentar americana, nas suas diferentes formas clínicas, assim como nas principais fases evolutivas da doença. Biópsias de lesões cutâneas e/ou de mucosa desses pacientes foram obtidas com a finalidade de isolar e caracterizar os parasitas, através da técnica de anticorpos monoclonais. Dos 60 pacientes examinados, foram isoladas 40 amostras de Leishmania das lesões biopsiadas, sendo 5 de Leishmania (V.) brasiliensis, 3 de L. (V.) guyanensis, 1 de L. (V.) lainsoni, 13 de L. (L.) amazonensis e 18 não puderam ser caracterizados a nível específico, porém, reagiram com anticorpos monoclonais do grupo braziliensis. Quanto àpesquisa através das culturas de leucócitos circulantes, esta revelou resultados completamente negativos. Com base nesses achados, os autores concluíram ser pouco consistente atribuir valor à cultura de leucócitos para o diagnóstico da leishmaniose tegumentar.

Leishmaniose cutânea experimental: II - aspectos evolutivos da infecção no primata Cebus apella (Cebidae) pela Leishmania (V.) Braziliensis e L. (L.) Amazonensis

Objetivando avaliar o potencial do primata C. apella como modelo experimental da leishmaniose cutânea, produzida pela L. (V.) braziliensis e L. (L.) Amazonensis , inocularam-se, via intradérmica, 3 X 10(6) de promastigotas dessas leishmanias, em 8 sítios da cauda de 10 espécimens desse primata, 5 deles com a L. (V.) braziliensis e outros 5 com a L. (L.) Amazonensis . Posteriormente, às inoculações, o exame semanal dos animais e biópsias mensais, revelaram os seguintes resultados relativos a cada parasita: a) L. (V.) braziliensis : o período de incubação foi de 15-20 dias; aos 30 dias evidenciaram-se lesões pápulo-eritematosas, que evoluíram para nódulos ao fim de 60 dias; no 3.° mês, notou-se ulceração espontânea destas lesões e, no 4° mês, deu-se o início da reparação das lesões ulceradas, culminando com a cura em um dos animais após 5 meses, em dois após 6 meses, noutro após 7 meses e, no último, após 10 meses. Quanto ao parasitismo nas lesões, foi demonstrado nos 5 animais, até 90 dias; depois disto, somente em 2 até 120 dias e, por fim, até 180 dias apenas naquele que curou depois de 10 meses, b) L. (L.) Amazonensis : o período de incubação foi de 20 dias; aos 30 dias notou- se lesões pápulo-eritematosas, que também evoluíram para nódulos ao fim de 60 dias, porém, a partir do 3.° mês, estas lesões regrediram rapidamente ao fim de 90 dias, quando não mais detectou-se o parasita na pele dos animais. Em relação aos testes de Montenegro, somente 2 dos 5 animais infectados com a L. (V.) braziliensis reagiram ao teste, 60 e 90 dias após as inoculações. Os resultados observados permitiram confirmar a infectividade do C. apella a estas leishmanias e, também, reforçar a indicação desse primata como modelo experimental da leishmaniose cutânea causada por estes parasitas.

Immunocytochemical identification of leishmania and Trypanosoma cruzi amastigotes in situ with homologous and heterologous polyclonal antibodies

The unlabelled antibody peroxidase-antiperoxidase method was used to study the immunocytochemical properties of Leishmania and Trypanosoma cruzi amastigotes in situ after tissues had been submitted to different fixation procedures. Antisera were obtained from rabbits chronically infected with different strains of T. cruzi or immunized with L. mexicana amazonensis and L. braziliensis guyanensis, and were applied on 5 µm thick sections. T. cruzi antigens were well stained by the three anti-T. cruzi sera and the two anti-heis.hmama.sera at optimum dilution between 1:1,000 and 1:2,000, regardless the parasite strain. Differently, the leishmanial antigens were revealed by Leishmania sera only at low dilutions (between 1:60 -1:160), whereas the anti-T. cruzi sera, at these low dilutions, gave rather weak stainings. Although there is no clear explanation for this immunocytochemical "reverse-monodirectional" cross-reactivity between Leishmania and T. cruzi, the present results show that polyclonal antibodies agains Leishmania species, when used for immunocytochemical detection of these parasites in situ, react more strongly with T. cruzi amastigotes than with the homologous amastigotes.

Parasitological diagnosis of mucocutaneous leishmaniasis due to Leishmania b. braziliensis in Bolivia

Parasitological diagnosis, using staned smears, culture and pathological examination of biopsy, was studied in 146 patients infected with mucocutaneous leishmaniasis, in Bolivia and Peru. The most efficient parasite detecting technique appeared to be the smear examination in cutaneous lesions (33 % positive) and the pathology in case of mucous lesions (28 % positive). In both, cutaneous and mucous lesions, the parasites were found most frequently in old lesions.

Glucantime resistant Leishmania promastigotes are sensitive to pentostam

Growth inhibition in vitro tests were used to study the susceptibility to pentostam of different Leishmania strains involved in cutaneous and mucocutaneos leishmaniasis - one glucantime sensitive strain, three naturally glucantime resistant strains and one glucantime resistant line developed by in vitro drug exposure. Contrasting with the high degree , of glucantime resistance, all strains were sensitive to pentostam. These differences suggest that there is some relationship between chemical structure and in vitro activity for these antimonial compounds. These data justify a clinical re-evaluation to compare therapeutic efficacy of glucantime and pentostam in the treatment of leishmaniasis.

Evaluation of a direct immunofluorescent antibody (difma) test using Leishmania genus - specific monoclonal antibody in the routine diagnosis of cutaneous leishmaniasis

A direct immunofluorescent antibody (DIFMA) test using a Leishmania genus- specific monoclonal antibody was evaluated in the routine diagnosis of cutaneous leishmaniasis (CL) in Ecuador. This test was compared with the standard diagnostic techniques of scrapings, culture and histology. Diagnostic samples were taken from a total of 90 active dermal ulcers from patients from areas of Ecuador known to be endemic for cutaneous leishmaniasis. DIFMA was positive in all lesions. It was shown to be significantly superior to standard diagnostic methods either alone or in combination. The sensitivity of DIFMA did not diminish with chronicity of lesions. This test proved to be extremely useful in the routine diagnosis of CL because it is highly sensitive, is easy to use and produces rapid results.

The FML (Fucose Mannose Ligand) of Leishmania donovani: a new tool in diagnosis, prognosis, transfusional control and vaccination against human kala-azar

The Fucose-Mannose Ligand (FML) of Leishmania donovani is a complex glycoproteic fraction. Its potential use as a tool for diagnosis of human visceral leishmaniasis was tested with human sera from Natal, Rio Grande do Norte, Brazil. The FML-ELISA test, showed 100% sensitivity and 96% specificity, identifying patients with overt kala-azar (p < 0.001, when compared to normal sera), and subjects with subclinical infection. More than 20% apparently healthy subjects with positive reaction to FML developed overt kala-azar during the following 10 months. In the screening of human blood donnors, a prevalence of 5% of sororeactive subjects was detected, attaining 17% in a single day. The GP36 glycoprotein of FHL is specifically reconized by human kala-azar sera. The immunoprotective effect of FML on experimental L. donovanii infection was tested in swiss albino mice. The protection scheemes included three weekly doses of FML, supplemented or not with saponin by the subcutaneous or intraperitoneal routes and challenge with 2x 10(7) amastigotes of Leishmania donovani. An enhancement of 80.0 % in antibody response (p<0.001) and reduction of 85.5 % parasite liver burden (p<0.001) was detected in animals immunized with FML saponin, unrespectivety of the immunization route.

Epidemiovigilância das infeções por Leishmania spp. no efetivo cinotécnico da Guarda Nacional Republicana e em mesocarnívoros silvestres

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do Grau de Mestre em Genética Molecular e Biomedicina

Evaluation of Leishmania antigens preparation and storage for use in enzyme immunoassays

The influence of time and temperature on the storage of an alkaline antigen of L.major-like and L.(V.) braziliensis promastigotes added or not of a proteases inhibitor (PMSF) was evaluted by means of an IgG-ELISA. Antibodies in assays using L. major-like antigen stored at -20oC for 6 monsths had a statistically lower geometric mean titer (GMT) and different 95% confidence interval limits (CL) than antigens stored otherwise, as assessed by the "t" statistic. The PMSF L. major-like antigen after storage for 6 months at a temperature of 4oC had the same GMT and 95% CL displayed at time zero as well as when storage for 4 and 6 months at -20oC. Significant diferences were not found when L.(V.) braziliensis antigens were stored at times and temperatures mentioned; the PMSF antigen stored for 2 months at -70oC resulted in a lower serum GMT and 95% CL than any other, as assessed by the "t" statistic. Antigen performance did not show any statistical difference associated to the addition of PMSF within the same species; the largest difference between antigens was that between PMSF-L. (V.) braziliensis and L. major-like without PMSF.

Histoplasmose em região de palato duro simulando lesão causada por Leishmania

Os autores relatam um caso de histoplasmose em indivíduo com suspeita clínica de leishmaniose mucosa. A infecção por Leishmania foi descartada, pela negatividade do teste de Montenegro e ausência do parasita. O diagnóstico de histoplasmose foi confirmado pelo encontro do fungo na lesão e o seu isolamento em Ágar-Sabouraud. O tratamento do paciente com anfotericina B resultou na remissão da lesão.