Accuracy of qPCR for quantifying Leishmania kDNA in different skin layers of patients with American tegumentary leishmaniasis.

OBJECTIVES: Superficial swab sampling of American tegumentary leishmaniasis (ATL) lesions shows higher amounts of Leishmania than those from biopsy. Subcutaneous involvement is also important in ATL, but parasite quantification according to lesion depth has not been evaluated. We aim to present the best depth at which sampling should be performed for molecular exams of ATL. METHODS: Patients with a clinical presentation compatible with ATL were allocated to ATL and control groups. Qualitative and quantitative qPCR assays were performed using SYBR Green and primers amplifying the kDNA minicircle of Leishmania spp. in different skin layers, including the epidermis, the superior dermis, the inferior dermis, and the hypodermis. RESULTS: Fifty-nine patients were included in this study, including 40 who had been diagnosed with ATL and 19 controls. The number of parasites was greater in samples of the epidermis and superior dermis (159.1 × 106, range 4.0-781.7, and 75.4 × 106, range 8.0-244.5, mean Leishmania parasite equivalents per µg of tissue DNA, respectively) than those in samples of the inferior dermis and hypodermis (54.6, range 8.0-256.6, and 16.8 × 106, range 8.0-24.1, mean Leishmania parasite equivalents per µg of tissue DNA, respectively). The best diagnostic accuracy was achieved in the superior dermis (77.9%) and was significantly greater than that in the hypodermis (63.3%; p 0.039). CONCLUSIONS: We conclude that superficial sampling can retrieve a greater quantity of parasites. Future studies of the role of transepidermal elimination as a mechanism of host defence in ATL must be performed as there is a considerable quantity of Leishmania kDNA in the epidermis.

The influence of treatment on the development of leishmaniasis recidiva cutis: a 17-year case-control study in Midwestern Brazil.

BACKGROUND: The recurrence of American cutaneous leishmaniasis (ACL) in patients experiencing a long-term cure is often called leishmaniasis recidiva cutis (LRC). LRC is considered an unusual form of ACL. OBJECTIVE: This study aims to estimate the incidence of LRC in ACL patients evaluated at a tertiary dermatologic centre in Midwestern Brazil. We also aim to evaluate the association between various treatment regimens and the development of LRC using multivariate analysis in a case-control study. METHODS: We performed a 17-year epidemiological study using data from patients treated at our dermatologic centre from July 1994 to December 2011. A retrospective analysis was then performed to estimate risk and protective factors related to clinical presentation. We also assessed the influence of treatment regimens in the development of LRC. RESULTS: The incidence of LRC among ACL patients was 1.34%. The analysis included 105 patients; 82 patients (78%) were in the control group, and 23 patients (22%) were in the LRC case group. The data analysis indicated that the standard treatment N-methylglucamine antimoniate (N-MA) reduced the development of LRC in bivariate (odds ratio (OR) = 0.34; 95% CI = 0.13-0.91) and multivariate analyses (OR = 0.16; 95% CI = 0.03-0.86; P = 0.03). However, no differences in LRC incidence were observed when the standard treatment N-MA and alternative drugs, such as pentamidine and amphotericin B, were considered (OR = 0.47; 95% CI = 0.16-1.35) CONCLUSION: We conclude that the standard treatment N-MA, as proposed by the Brazilian Ministry of Health, is effective in the prevention of LRC. Although other drugs have shown promising results in LRC, more scientific evidence is needed to assess their efficacy compared with N-MA.

A pilot study comparing low-dose liposomal amphotericin B with N-methyl glucamine for the treatment of American cutaneous leishmaniasis.

BACKGROUND: Cutaneous leishmaniasis is an infectious re-emerging disease that has increased in incidence worldwide. Antimony, a highly toxic drug, remains the first choice therapy to treat it. Liposomal amphotericin B is active against Leishmania and is less toxic than antimony. OBJECTIVE: To compare low-dose liposomal amphotericin B with N-methyl glucamine for the treatment of American cutaneous leishmaniasis. PATIENTS/METHODS: In a controlled open-label trial 35 patients with a localized form of American cutaneous leishmaniasis were included. They were allocated to a first group treated with 1.5 mg/kg/day of liposomal amphotericin B for 5 days, or to a second one treated with 20 mgSbV/kg/day of N-methyl glucamine for 20 days. RESULTS: In the first group, 50% and 81% of patients experienced a clinical cure and clinical improvement respectively. There was a 100% clinical cure in the second group. CONCLUSION: Liposomal amphotericin B seems to be promising and safe for the treatment of American cutaneous leishmaniasis.

Leishmania (Viannia) braziliensis is the main species causing cutaneous leishmaniasis in the Federal District of Brazil

The first autochthonous case of American cutaneous leishmaniasis was reported in the Federal District in 1980, and the species involved in this type of leishmaniasis was unknown. This study aimed to identify the species that causes the disease in the Federal District and to investigate its clinical and epidemiological aspects. Between 2000 and 2007, 71 autochthonous cases of leishmaniasis were reported in the Federal District. Leishmania species were identified by means of direct immunofluorescence reactions using monoclonal antibodies and restriction fragment length polymorphism. The species of 40 (56.33%) out of 71 samples were identified. Thirty-six (90%) were identified as Leishmania (Viannia) braziliensis and four (10%) were identified as Leishmania (Leishmania) amazonensis. In this area, the disease had clinical and epidemiological characteristics similar to those found in other Brazilian regions.

DNA identification of Leishmania (Viannia) braziliensis in human saliva from a patient with American cutaneous leishmaniasis

This study aims to report the amplification of the DNA of Leishmania (V.) braziliensis, using polymerase chain reaction, obtained from the saliva of a patient with American cutaneous leishmaniasis who did not present any lesion in the oral mucosa. Amplification produced fragments of 103 bp, an estimated size employing Leishmania (V.) braziliensis primers (b1 e b2). The present results revealed, for the first time, that the in vitro amplification of Leishmania DNA using samples from the salivary fluid of a patient with American cutaneous leishmaniasis is possible. However, more studies are required with a larger number of participants to evaluate the usefulness of saliva as a non-invasive sample for PCR. The development of such non-invasive technique is necessary for the diagnosis of many diseases in the future, especially infectious and parasitic ones.

Inefficacy of the association N-methyl glucamine and topical miltefosine in the treatment of experimental cutaneous leishmaniasis by Leishmania (Leishmania) amazonensis

Pentavalent antimonial (SbV) is the first treatment for cutaneous leishmaniasis (CL). Other drugs present similar side effects and higher cost. Oral miltefosine is effective to treat kala-azar. The aim of the present study was to compare the efficacy of glucamine (SbV) plus topical miltefosine with glucamine in the treatment of CL. Eighty isogenic C57BL/6 mice were inoculated with Leishmania (Leishmania) amazonensis and divided into two groups: one group was treated with SbV associated with miltefosine, and the other group received SbV plus saline solution. Groups were evaluated according to the diameter of the inoculated foot pad, the culture, and the parasite count using the limiting dilution assay. There was not statistical difference. The efficacy of glucamine in CL treatment did not increase when associated with topical miltefosine.(AU)

Use of Cavia porcellus (Guinea Pigs) as an experimental model for Leishmania (Viannia) braziliensis

Laboratory animals are fundamental to study immunological aspects and the efficacy of new drugs to treat leishmaniasis. However, we do not have practical and good animal models to study leishmaniasis caused by Leishmania (Viannia) braziliensis - L(V)b. In this study, thirty-two experimental animals (Cavia porcellus) were injected in the hind foot with 3x105 promastigote forms of L(V)b. The animals were followed for eight weeks. None of the experimental animals developed lesions or presented the parasite in any of the tests performed (histopathological exam, smears, culture, inoculation in hamsters, and polymerase chain reaction)

Insuficiência renal aguda em leishmaniose tegumentar americana tratada com a associaçäo glucantime e alopurinol / Acute renal insufficiency in mucocutaneous leismaniasis reated with glucantime and allopurinol

Os autores apresentam um caso de leishmaniose tegumentar americana causada por Leishmânia viannia brasiliensis que de um grupo de seis pacientes tratados com a associaçäo glucantime e alopurinol desenvolveu quadro de nefrite intersticial aguda acompanhado de exantema cutâneo. Questionam se näo houve potencializaçäo na interaçäo das drogas com relaçäo ao efeito desejável