Ennio Luz

Lutzomyia whitmani (Diptera: Psychodidae) as vector of Leishmania (V.) braziliensis in Paraná state, southern Brazil

The phlebotomine sandflies in the northern areas of the state of Paraná, Brazil, particularly those in the ‘16a’ health region, were investigated over a 3-year period. Using CDC light traps (with and without hamster bait) and Shannon traps (with lights and horse or human bait), 16 species were collected from seven municipal districts which were known foci for cutaneous leishmaniasis: Arapongas; Apucarana; Cambira; Marumbi; Faxinal; Florestópolis; and Sabáudia. Although the frequency at which each species was collected varied with the collection site, Lutzomyia whitmani predominated (62.0% of all the sandflies collected), followed by Lu. fischeri (13.3%), Lu. pessoai (10.8%), Lu. migonei (8.2%) and Lu. intermedia (2.8%). Lutzomyia monticola, Lu. shanonni, Lu. firmatoi, Lu. lanei, Lu. alphabetica, Lu. misionensis, Lu. correalimai, Lu. cortellezzii, Lu. longipenis, Brumplomyia brumpti and B. nitzulescui together represented the remaining 3.0% of the collected sandflies. Three of the 1961 female sandflies collected and dissected in the municipal district of Cambira, where a recent case of cutaneous leishmaniasis had been registered, were found to have flagellates in their guts. All three were Lu. whitmani. The parasites from each of these infections were successfully isolated in NNN and ‘Tobie and Evans’ media and/or by inoculation into a hind foot of a golden hamster. The results of isoenzyme electrophoresis indicated that all three isolates were of Leishmania (Viannia) braziliensis.

Estudo das características epidemiológicas e clínicas de 332 casos de leishmaniose tegumentar notificados na região norte do Estado do Paraná de 1993 a 1998

Cases of cutaneous leishmaniasis (CL) in northern Parana State have been reported since the 1950s, but towards the end of the 1980s there has been an increase in the number of human infections. From 1993 to 1998, a study was carried out in an attempt to define the epidemiological profile of leishmaniasis in this region. A total of 316 cases of CL were reported from 35 municipal districts and 16 imported cases. Most cases of CL notified (70.8%) were in the age group of 15 to 49 years old and most of these were males (61.2%). Of the 332 positive patients, 66.8% presented single classic lesions and 31.1% multiple lesions. Most of these lesions were located in the lower limbs (47.7%) while 26.7% were in the arms and 16% on the face. Thirty-two Leishmania stocks were isolated and identified by isoenzymatic characterization using 13 enzymatic systems as Leishmania (Viannia) braziliensis.

Molecular diagnosis of leishmaniosis in the Paraná state of southern Brazil

Abstract:  The objective of the present study was to establish a polymerase chain reaction (PCR) technique for the diagnosis of cutaneous and mucocutaneous leishmaniosis from autochthonous cases in the state of Paraná in southern Brazil as well as imported cases. We sought to determine its utility and accuracy compared with smears and present culture methods. To standardize PCR samples, skin and mucosal punch biopsies from human lesions were performed on patients living in different regions of the Paraná state (76 cases) and other endemic areas of Brazil and Argentina (7 cases). For PCR standardization, two pairs of primers (MP1L/MP3H and B1/B2) were utilized for amplification of the conserved sequences in the minicircle of kinetoplast DNA (kDNA) for the Leishmania braziliensis complex. Two other primer pairs (b1/b2 and a1/a2) were species‐specific for L. (V.) braziliensis and L. (V.) amazonensis, respectively. After differential diagnosis, eight patients had clinical diagnosis of the cutaneous ulcer changed to others pathologies such as syphilis, baso‐cellular carcinoma, varicose ulcer, ecthyma and paracoccidioidomycosis. Of the 75 patients with cutaneous (CL) and mucocutaneous (MCL) lesions who provided samples, 47 (46 CL + 1 MCL) were diagnosed with leishmaniosis by smear and 57 (52 LC + 5 MCL) were diagnosed by culture methods. In contrast, our PCR technique presented higher accuracy when compared to the direct examination and culture of parasites. PCR is applicable both for CL where all 61 lesions were diagnosed, and MCL where 12 of 14 lesions were diagnosed. This molecular biology technique is also a faster and more specific diagnostic method compared with present parasitological procedures.

Comparison of serological and parasitological methods for cutaneous leishmaniasis diagnosis in the state of Paraná, Brazil

We evaluated the effectiveness of serological and parasitological methods for cutaneous leishmaniasis (CL) diagnosis in patients from the central region of Paraná state, southern Brazil. Five groups were compared: clinical diagnosis, parasitological diagnosis, communicants, inhabitants of a non-endemic area and carriers of other etiologies. Two antigens were prepared from promastigotes of Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis for indirect immunofluorescence assay, ELISA and immunoblotting. The parasitological approaches detected 79.3% of the patients with a clinical diagnosis; the parasites were identified by PCR as L. (V.) braziliensis. Serological methods showed 95% sensitivity for homologous antigens. Immunoblotting revealed specific proteins for diagnosis of CL and detected 96.6% of the patients when L. (V.) braziliensis was used as an antigen, and 83.3% with L. (L.) amazonensis. This study demonstrated the importance of differential diagnosis for leishmaniasis; the association of two or more indirect methods increased diagnosis sensitivity.

A new focus of cutaneous leishmaniasis in the central area of Paraná State, southern Brazil.

We report a new endemic zone of cutaneous leishmaniasis (CL) in the central area of the State of Paraná (Municipality of Prudentópolis), in southern Brazil. This region was not previously considered endemic for CL, and this work constitutes the first report of CL endemicity there. Leishmaniasis was confirmed by smear, culture, and ELISA. Parasites were isolated and identified by random amplification of polymorphic DNA (PCR-RAPD). Phylogeographical analysis, based on two different criteria, was able to distinguish between RAPD profiles from different geographical regions. In total, 100 patients were diagnosed with leishmaniasis by culture and serology methods. The reported incidence rate was 4.32%. Of the 100 patients, 92% of the patients had single lesions, and 79.98% of these lesions were located on their limbs. The fact that 61% of patients were male rural workers points to an extradomiciliar type of transmission. In houses where human leishmaniasis was diagnosed, 29% of the dogs presented anti-Leishmania antibodies. A total of 1663 phlebotomines, representing 5 species, were captured in the studied area with CDC-like light minitraps. Lutzomyia intermedia s.l. was the most prevalent species (94.40%). The isolated parasites were grouped with Leishmania (V.) braziliensis. The epidemiological implications are discussed in the present article.

Flebótomos da área do reservatório da hidroelétrica de Itaipu, estado do Paraná, Brasil: Diptera, Psychodidae

Durante os anos de 1979 a 1986 foram feitas capturas intra e extra domiciliares e tambem com isca humana, em diversas localidades de 6 municipios situados ao longo da margem brasileira do reservatorio da hidroeletrica de Itaipu. Foram coletadas 8 especies de Lutzomyia, sendo que duas, ja incriminadas como transmissoras da leishmaniose tegumentar, L. intermedia e L. whitmani, sao abundantes na area.

Trypanosoma cruzi: isoenzyme analysis suggests the presence of an active Chagas sylvatic cycle of recent origin in Paraná State, Brazil

Thirty-one trypanosomatid stocks were isolated from various sylvatic hosts and vectors in two different regions from the Paraná State of Brazil. The stocks were analyzed by multilocus enzyme electrophoresis (MLEE) on cellulose acetate plates (22 genetic loci). All stocks were unambiguously attributed to Trypanosoma cruzi, and were found to be closely related to the formerly described zymodeme I (TC1 subgroup of T. cruzi). By comparison with other sylvatic cycles with similar sample sizes (Southern USA, Colombia, French Guiana), genetic variability among these stocks was very limited, with only two variable loci out of 22, and only three different multilocus genotypes. Population structure of T. cruzi in these cycles appears to correspond to a set of very closely related clonal genotypes. This very limited genetic variability could be due to a recent foundation of these populations (founder effect). The implications of an active Chagas sylvatic cycle in this area are discussed.

Larvas de Anopheles (subgênero Kerteszia) Theobald 1905 encontradas em criadouros artificiais, no Estado do Paraná, Brasil

Discute-se encontro de larvas de duas especies de anofelinos do subgenero Kerteszia - An. (Ker.) cruzii e An. (Ker.) bambusicolus - em criadouros artificiais, no Estado do Parana, Brasil.

New isolation of Leishmania enriettii Muniz and Medina, 1948 in Parana State, Brazil, 50 years after the first description, and isoenzymatic polymorphism of the L. enriettii taxon

Three cases of cutaneous leishmaniasis in guinea-pigs from a rural area near Curitiba (Paraná State, Brazil) are reported. The three parasite isolates were characterized by isoenzyme electrophoresis as Leishmania enriettii, of which two distinct zymodemes were observed.

Leishmania Species in Two Regions of Paraná, Brazil: Biochemical Characterization by Isoenzyme Electrophoresis

Cutaneous leishmaniasis is present in two different regions of the State of Parana, (South of Brazil): in the Northern area and the Ribeira River Valley. In 1994 there was an outbreak with a total of 1395 leishmaniosis cases. Here, we report on the isolation and characterization of 52 Leishmania stocks isolated between 1995 and 2000 from humans, dogs, and wild reservoir and phlebotomine hosts. The material, obtained by biopsy (punch no. 4) or by aspiration puncture, was inoculated in two media (NNN and Tobbie & Evans) and maintained at a temperature of 24°C. After isolation, parasites were grown in C.C.S. medium (in Erlenmeyer and bottle of Roux) for protein extract preparation, awaiting identification. Isoenzymatic characterisation was made by electrophoresis in starch thick gel, using 13 enzymatic systems. Leishmania stocks were obtained from active lesions of humans coming from 26 counties in the northern region of Parana and 6 from the Adrianopolis borough (Ribeira River Valley). In the northern area of the state stocks were isolated from 8 dogs (Canis familiaris); 1 stock from Nectomys sp., and 3 stocks were isolated from Lutzomyia whitmani. All stocks were identified as Leishmania (Viannia) braziliensis. Fifteen zymodemes were characterized. Zymodemes LMP1 and LPM2 showed a large distribution in both regions studied. These zymodemes are present from the south of Mexico to the North of Argentina.