Cibele Baptista

Leishmania (Viannia) braziliensis genotypes identified in lesions of patients with atypical or typical manifestations of tegumentary leishmaniasis: evaluation by two molecular markers.

Analyses of MLEE, RAPD and LSSP-PCR were used to compare the panel of american tegumentary leishmaniasis (ATL) isolates obtained from lesions of patients with rare clinical manifestations of the disease and typical lesions. All of the 34 samples analyzed by MLEE demonstrated similar electromorphic profiles with Leishmania (Viannia) braziliensis reference strain. Through the RAPD analysis, nine genetic profiles (genotypes) were identified. LSSP-PCR corroborates the initial screening and phenetic analysis has grouped the isolates into two major clusters comprising the nine different genotypes. Prevalent genotype defined as LbmtDNAgen1 was detected in the largest number of isolates. There was no association between genotypes and clinical symptoms. However, two different genotypes could be identified in the initial (LbmtDNAGen9) and reactivated lesion (LbmtDNAGen3) of the same patient. Our results support the idea of a less pronounced genotypic diversity among L. (V.) braziliensis circulating in the State of Rio de Janeiro and demonstrate the useful application of these molecular markers in genetics variability studies.

Serological evaluation for detection of anti-Leishmania antibodies in dogs and cats in the district of Santa Rita de Cássia, municipality of Barra Mansa, State of Rio de Janeiro

From a canine case of american cutaneous leishmaniasis in the locality of Santa Rita de Cássia, municipality of Barra Mansa, Rio de Janeiro, a serological survey was performed on 177 dogs and 43 cats. Evaluation of the canine serum samples showed that 10% had a positive reaction in the indirect immunofluorescence test and 10.7% in the enzyme-linked immunosorbent assay. Among the feline serum samples tested, none of the animals had a positive reaction in the indirect immunofluorescence test and only one (2.4%) showed a positive reaction in the enzyme-linked immunosorbent assay. The detection of an autochthonous case of Leishmania braziliensis in Barra Mansa gives warning that a focus of american cutaneous leishmaniasis is possibly becoming established in this region.

Differential Interferon-γ Production Characterizes the Cytokine Responses to Leishmania and Mycobacterium leprae Antigens in Concomitant Mucocutaneous Leishmaniasis and Lepromatous Leprosy

BACKGROUND Tegumentary leishmaniasis and leprosy display similar spectra of disease phenotypes, which are dependent on cell-mediated immunity to specific antigens. Diffuse cutaneous leishmaniasis and lepromatous leprosy represent the anergic end of the spectrum, whereas mucocutaneous leishmaniasis and tuberculoid leprosy are associated with marked antigen-specific cellular immune response. METHODS We characterized and compared the cell-mediated response to Leishmania and Mycobacterium leprae antigens in a patient with an intriguing association of mucocutaneous leishmaniasis with lepromatous leprosy, which are at opposite ends of the immunopathological spectra of these diseases. This was done by performance of skin tests and by assessment of the cell proliferation and cytokine production of peripheral blood mononuclear cells (PBMCs). RESULTS Strong skin-test reactions and PBMC proliferation were observed in response to Leishmania antigens but not to M. leprae antigens. The stimulation of PBMCs with Leishmania and M. leprae antigens induced comparable levels of tumor necrosis factor- alpha , interleukin-5, and interleukin-10. However, the interferon- gamma response to Leishmania antigens was remarkably high, and that to M. leprae antigens was almost nil. CONCLUSIONS We found that concomitant leprosy and tegumentary leishmaniasis can produce opposite polar forms associated, respectively, with absent or exaggerated cell-mediated immune responses to each pathogen. This suggests that independent mechanisms influence the clinical outcome of each infection. Moreover, interferon- gamma appears to play a major role in the clinical expression of these intracellular infections.

Occurrence of Trypanosoma caninum in areas overlapping with leishmaniasis in Brazil: what is the real impact of canine leishmaniasis control?

Trypanosoma caninum is a parasite of the Trypanosoma genus recently described in the natural infection of dogs in the municipality of Rio de Janeiro, Brazil. Suspecting the existence of a natural cycle and the circulation of this new species, the objective of this study was the taxonomic identification of samples of Trypanosoma spp. isolated from dogs in different Brazilian regions. Parasites were solely obtained from skin fragments culture and characterized by nested-PCR targeting the partial sequence of 18S rRNA gene and PCR products were sequenced. Thirty-three samples, obtained in São Paulo, Minas Gerais, Goiás, Mato Grosso and Rio de Janeiro states were analyzed. PCR and sequencing showed that the isolates were genetically identical or closely similar and confirmed T. caninum identity. This report broadens the geographical distribution of T. caninum in Brazil and discusses the impact of the presence of this parasite in areas of canine leishmaniasis occurrence.

Isolation of Trypanosoma caninum in domestic dogs in Rio de Janeiro, Brazil.

SUMMARY The domestic dogs involvement with different members of the Trypanosomatidae family has been the focus of several studies due to this animals close proximity to man. Recently this animal has been infected by a new Trypanosoma species (T. caninum), described in Rio de Janeiro and 19 similar isolates were later obtained. The objective of this study was to identify these isolates. All samples were isolated from intact skin cultures and analysed morphologically, by biochemical isoenzyme electrophoresis assays and by several molecular PCR assays. Additionally, anti-Leishmania sp. antibodies were assessed using the indirect Immunofluorescence Antibody Test (IFAT) in all animals. The methodologies employed to identify the isolates, including partial nucleotide sequences of 18S rRNA gene, indicated patterns identical to T. caninum and patterns different from the other species, including T. cruzi and T. rangeli samples. A phylogenetic tree constructed with the partial 18S ribosomal sequence shows that T. caninum is clustered with T. pestanai. Ten (52.6%) animals presented anti-Leishmania sp. antibodies with titres varying from 1:40 to 1:320. Thus, the hypothesis that this protozoan has disseminated among the dogs in Rio de Janeiro must be considered. The importance of a correct diagnosis in those animals and the possible consequences in the areas where visceral leishmaniasis is found are discussed here.

American Tegumentary Leishmaniasis in Older Adults: 44 Cases Treated with an Intermittent Low‐Dose Antimonial Schedule in Rio de Janeiro, Brazil

American tegumentary leishmaniasis (ATL) is a disease affecting the skin and mucosae caused by protozoans of the genus Leishmania transmitted by the bite of female sandflies. Cutaneous leishmaniasis (CL) presents mainly as skin ulcers at exposed body sites. Mucosal leishmaniasis (ML) manifests as chronic and destructive lesions of the nasal, oral, pharyngeal, and laryngeal tissues.1 Pentavalent antimonials are the first-line treatment for ATL. Reports of pentavalent antimonial toxicity include renal tubular dysfunction; cardiac, hepatic, pancreatic, and hematological alterations; and even death.2–6 Adverse effects (AEs) are frequent, and interruption is sometimes needed in patients aged 60 and older, even those receiving low-dose treatment. Observing that lesions continued to heal during withdrawal, it was decided to evaluate the safety and efficacy of an intermittent low-dose meglumine antimonate (MA) regimen for ATL in the elderly.....

American cutaneous leishmaniasis caused by Leishmania (Viannia) braziliensis resistant to meglumine antimoniate, but with good response to pentamidine: a case report

This is a case report of a Brazilian soldier with cutaneous leishmaniasis. The lesion relapsed following two systemic treatments with meglumine antimoniate. The patient was treated with amphotericin B, which was interrupted due to poor tolerance. Following isolation of Leishmania sp., six intralesional infiltrations of meglumine antimoniate resulted in no response. Leishmania sp promastigotes were again isolated. The patient was submitted to intramuscular 4 mg/kg pentamidine. Parasites from the first and second biopsies were identified as Leishmania (Viannia) braziliensis; those isolated from the first biopsy were more sensitive to meglumine antimoniate in vitro than those isolated from the second biopsy. No relapse was observed.

Evaluation of Leishmania (Leishmania) chagasi strains isolated from dogs originating from two visceral leishmaniasis-endemic areas in Brazil using multilocus enzyme electrophoresis

INTRODUCTION Domestic dogs are the most important reservoir in the peridomestic transmission cycle of Leishmania (Leishmania) chagasi. The genetic variability of subpopulations of this parasite circulating in dogs has not been thoroughly analyzed in Brazil, even though this knowledge has important implications in the clinical-epidemiological context. METHODS The objective of this study was to evaluate and compare the phenotypic variability of 153 L. chagasi strains isolated from dogs originating from the municipalities of Rio de Janeiro (n = 57) and Belo Horizonte (n = 96), where the disease is endemic. Strains isolated only from intact skin were selected and analyzed by multilocus enzyme electrophoresis using nine enzyme systems (6PG, GPI, NH1 and NH2, G6P, PGM, MDH, ME, and IDHNADP). RESULTS The electrophoretic profile was identical for all isolates analyzed and was the same as that of the L. chagasi reference strain (MHOM/BR/74/PP75). Phenetic analysis showed a similarity index of one for all strains, with the isolates sharing 100% of the characteristics analyzed. CONCLUSIONS The results demonstrate that the L. chagasi populations circulating in dogs from Rio de Janeiro and Belo Horizonte belong to a single zymodeme.

In Vitro Sensitivity of Paired Leishmania (Viannia) braziliensis Samples Isolated before Meglumine Antimoniate Treatment and after Treatment Failure or Reactivation of Cutaneous Leishmaniasis

This study evaluated the in vitro sensitivity of paired Leishmania braziliensis samples isolated from the same patient before pentavalent antimonial treatment (Sample A) and after treatment failure or cutaneous leishmaniasis reactivation (Sample B) in patients undergoing intralesional administration or injections (5 mgSbV/kg/d) of meglumine antimoniate. Fourteen samples from 7 patients were studied. After 24 h of drug exposure, 50% lethal dose (LD50) values for promastigotes ranged from 0.37 mg/mL to 5.86 mg/mL for samples obtained before treatment (A) and 0.89 mg/mL to 7.80 mg/mL for samples obtained after treatment (B). After 48 h, LD50 values ranged from 0.37 mg/mL to 5.75 mg/mL and 0.70 mg/mL to 7.68 mg/mL for A and B samples, respectively. After 48 h, LD50 values for amastigotes ranged from 11.7 to 44.3 μg/mL for A samples and 13.7 to 52.7 μg/mL for B samples. Of 7 patients, 1 discontinued treatment and 6 were cured after retreatment with amphotericin B (4 cases) or meglumine antimoniate (2 cases). Overall the B samples had higher LD50 values than A samples; however the difference was not significant. These results do not support the hypothesis that low-dose and intralesional treatments induce selection of resistant parasites in vitro and suggest that other factors may influence therapeutic outcome in patients with poor response to initial treatment.

Evaluation of Genetic Polymorphism of Leishmania (V.) braziliensis Isolates Obtained from the Same Patient before and after Therapeutic Failure or Reactivation of Cutaneous Lesions

The aim of this study was to investigate genetic polymorphism in Leishmania braziliensis population previously typed through isoenzyme electrophoresis, isolated from the same patient in two different moments: (A) before the beginning of treatment and (B) after treatment failure to meglumine antimoniate or reactivation after successful initial treatment. Fifteen pairs of isolates were assessed using the polymorphic molecular marker LSSP-PCR and following the phenetic analysis. The genetic profiles of the 30 samples were grouped in four clusters. Only two patients presented total identity in the A and B isolates. Most isolates presented similarity coefficients varying from 0.63 to 0.91. In this group of patients genetic polymorphisms could be observed indicating low similarity between the pairs of isolates. The results demonstrate the existence of genetic polymorphism between the samples isolated before treatment and after reactivation or treatment failure, suggesting a possible differentiation of the structure of the original parasite population which could be involved in the mechanisms of resistance to treatment or reactivation of lesions in the ATL. This phenomenon is important, although other factors also could be involved in this context and are discussed in this paper.