Identification of Leishmania species by next generation sequencing of hsp70 gene.

Leishmaniasis is a major public health problem worldwide. Although next generation sequencing technology has been widely used in the diagnosis of infectious diseases, it has been scarcely applied in identification of Leishmania species. The aim of this study was to compare the efficiency of MinION™ nanopore sequencing and polymerase chain reaction restriction fragment length polymorphism in identifying Leishmania species. Our results showed that the MinION™ sequencer was able to discriminate reference strains and clinical samples with high sensitivity in a cost and time effective manner without the prior need for culture.

Liposomal drug delivery systems for the treatment of leishmaniasis.

Human le ishmaniasis is a vector-borne, neglected infectious disease that is widely distributed in America, Africa, Europe, and Asia. Current therapy is based on old and toxic drugs, including antimonials, aminoglycosides, and amphotericin. As a neglected disease, investment in the development of new therapeutic molecules is scarce. Considering these aspects, the optimization of treatment through novel delivery systems for current therapeutic agents is an attractive alternative. The encapsulation into liposomes of drugs used in treating leishmaniasis increases the concentration of these molecules in macrophages, which may not only increase the chance of cure but also expand their therapeutic spectrum to include resistant Leishmania, as well as reducing toxicity since the drug is less exposed to healthy cells. The classical example is the liposomal formulation of amphotericin B, a well-established therapeutic option that uses liposomes to decrease the progression of renal failure in patients. However, loading other leishmanicidal drugs into liposomes, such as pentavalent antimonials, presents an opportunity for innovative and cheaper therapeutic options for the treatment of human leishmaniasis. This review aims to discuss liposomes as a drug delivery system for leishmanicidal drugs.

Post-kala-azar dermal leishmaniasis and leprosy: case report and literature review.

BACKGROUND: Post-kala-azar dermal leishmaniasis (PKDL) is a dermal complication of visceral leishmaniasis (VL), which may occur after or during treatment. It has been frequently reported from India and the Sudan, but its occurrence in South America has been rarely reported. It may mimic leprosy and its differentiation may be difficult, since both diseases may show hypo-pigmented macular lesions as clinical presentation and neural involvement in histopathological investigations. The co-infection of leprosy and VL has been reported in countries where both diseases are endemic. The authors report a co-infection case of leprosy and VL, which evolved into PKDL and discuss the clinical and the pathological aspects in the patient and review the literature on this disease. CASE PRESENTATION: We report an unusual case of a 53-year-old female patient from Alagoas, Brazil. She presented with leprosy and a necrotizing erythema nodosum, a type II leprosy reaction, about 3 month after finishing the treatment (MDT-MB) for leprosy. She was hospitalized and VL was diagnosed at that time and she was successfully treated with liposomal amphotericin B. After 6 months, she developed a few hypo-pigmented papules on her forehead. A granulomatous inflammatory infiltrate throughout the dermis was observed at histopathological examination of the skin biopsy. It consisted of epithelioid histiocytes, lymphocytes and plasma cells with the presence of amastigotes of Leishmania in macrophages (Leishman's bodies). The diagnosis of post-kala-azar dermal leishmaniasis was established because at this time there was no hepatosplenomegaly and the bone marrow did not show Leishmania parasites thus excluding VL. About 2 years after the treatment of PKDL with liposomal amphotericin B the patient is still without PKDL lesions. CONCLUSION: Post-kala-azar dermal leishmaniasis is a rare dermal complication of VL that mimics leprosy and should be considered particularly in countries where both diseases are endemic. A co-infection must be seriously considered, especially in patients who are non-responsive to treatment or develop persistent leprosy reactions as those encountered in the patient reported here.

Cerebrospinal fluid shunt infection caused by Corynebacterium sp: case report and review.

BACKGROUND: A 36-year-old immunocompetent woman with a posterior fossa arteriovenous malformation (PF-AVM) and hydrocephalus presented with low fever and mental confusion 4 days after ventriculoperitoneal shunting (VPS). METHODS: Cerebrospinal fluid (CSF) and ventricular catheter tip cultures isolated Corynebacterium sp. Similar to previous cases in the literature, species determination was not possible. However, the antibiotic sensitivity profile of this isolate suggested Corynebacterium jeikeium. Conversion to external ventricular drainage (EVD) was done and intravenous vancomycin was administered for 21 days. RESULTS AND CONCLUSIONS: The patient showed progressive improvement. Since the first CSF shunt infection caused by Corynebacterium sp., 16 other cases in the literatures have been reported. Additionally, this study reports the difficulties in recognizing CSF shunt infection caused by this agent and the possible clinical or laboratory patterns as observed in the literature.

Recurrent acquired ocular toxoplasmosis associated with Kyrieleis plaques and documented allergy to sulfonamide-A treatment proposal for two rare conditions.

The aim of this study was to describe a case of a patient with ocular toxoplasmosis, which has resulted in Kyrieleis plaques formation (segmental periarteritis associated with severe inflammation) and later follow-up and alternative treatment due to documented allergy to sulfonamide. A 33-year-old Brazilian woman diagnosed with acute toxoplasmosis, initially treated with sulfonamide, developed a critical cutaneous rash. Cotrimoxazole was changed to clindamycin and pyrimethamine, and prednisone was started. The medication was maintained for 45 days. Four months later, she developed retinal lesions suggestive of toxoplasmosis with Kyrieleis plaques in the upper temporal vessels. Pyrimethamine, clindamycin, and prednisone were initiated until healing. She presented reactivation months later, and a suppressive treatment with pyrimethamine was instituted for one year. This is the first report to use the combination of clindamycin with pyrimethamine in the treatment and recurrence prophylaxis for OT in a documented allergy to sulfonamide.

Case Report: In Situ and Systemic Immune Response to Mucosal Leishmaniasis in an HIV-Infected Patient.

In situ and systemic evaluations of the immune responses of HIV-infected patients to mucosal leishmaniasis have been poorly described. We describe a recently diagnosed HIV-infected patient with mucosal leishmaniasis who was characterized by a CD4 count of 85 cells/mm3 and nasal septum destruction resulting from pruritic and ulcerated nasal mucosa with crust formation and progression over 2 years. In situ and systemic immune evaluations of T cell activation, memory, and exhaustion were conducted using cytofluorometric assays, and sequencing of the Leishmania species was performed. The immune profile of HIV-infected patient with mucosal leishmaniasis shows a mixed Th1/Th2 pattern and an activated and exhausted status.

Case Report: Cutaneous Leishmaniasis in a Rheumatoid Arthritis Patient Receiving Methotrexate.

The immunosuppressive effect of methotrexate has rarely been associated with reactivation of cutaneous leishmaniasis. Here we present a case of a cutaneous leishmaniasis patient with atypical clinical symptoms without splenomegaly but with cutaneous manifestations after treatment of rheumatoid arthritis with methotrexate and blood recovery of the parasite. Next-generation sequencing was used to identify Leishmania infantum chagasi in the patient's blood sample.

Estimation of Leishmania spp. infection in asymptomatic people from Muzaffarpur, Bihar, India by antigen-antibody and skin testing.

Asymptomatic VL is a concern, considering the risk of transmission in highly endemic areas due to human-to-human transmission. The aim of this study was to report the sero-epidemiological prevalence in Bihar, India, a highly endemic area of VL, using the leishmanin skin test (LST) and the direct agglutination test (DAT). This was a cross-sectional study performed in Muzaffarpur, Bihar, India. Relatives of patients with VL were tested by LST and DAT. Other epidemiological data were evaluated and correlated with tests results. Forty individuals (either previous or current patients), and 109 household contacts were studied. There were 36% of male visceral leishmaniasis family members versus 17.57% of females visceral leishmaniasis family members, thus showing more males with symptomatic disease than females (p< 0.01). All visceral leishmaniasis cases had positive DAT tests, but only 37% of past cases were positive on the skin testing. Amongst healthy household contacts, 34% were DAT-positive, whilst 21% were LST-positive. The overall positivity for both assays combined was 44.8% and 23.8% were DAT-positive alone. The finding of high infection prevalence amongst asymptomatic individuals, and the estimation of those at greater risk for overt disease (DAT-positive alone) are important in the development of future disease control policies.

Lipid nanoparticles for amphotericin delivery in the treatment of American tegumentary leishmaniasis.

Leishmaniasis occurs in the five continents and represents a serious public health challenge, but is still a neglected disease, and the current pharmacological weaponry is far from satisfactory. Triglyceride-rich nanoparticles mimicking chylomicrons (TGNP) behave metabolically like native chylomicrons when injected into the bloodstream. Previously we have shown that TGNP as vehicle to amphothericin B (AB) for treatment of fungi infection showed reduced renal toxicity and lower animal death rates compared to conventional AB. The aim of the current study was to test the tolerability and effectiveness of the TGNP-AB preparation in a murine model of Leishmania amazonensis infection. The in vitro assays determined the cytotoxicity of TGNP-AB, AB, and TGNP in macrophages and promastigote forms and the leishmanicidal activity in infected macrophages. The in vivo toxicity tests were performed in healthy mice with increasing doses of TGPN-AB and AB. Then, animals were treated with 2.5 mg/kg/day of AB, 17.5 mg/kg/day of TGNP-AB, or TGNP three times a week for 4 weeks. TGNP-AB formulation was less cytotoxic for macrophages than AB. TGNP-AB was more effective than AB against the promastigotes forms of the parasite and more effective in reducing the number of infected macrophages and the number of amastigotes forms per cell. TGNP-AB-treated animals showed lower hepatotoxicity. In addition, TGNP-AB group showed a marked reduction in lesion size on the paws and parasitic load. The TGNP-AB preparation attained excellent leishmanicidal activity with remarkable lower drug toxicity at very high doses that, due to the toxicity-buffering properties of the nanocarrier, become fully tolerable.

Secondary Prophylaxis with Liposomal Amphotericin B in a Patient with Mucosal Leishmaniasis Undergoing Immunobiological Therapy for Active Ankylosing Spondylitis.

Immunosuppressive treatments for rheumatic diseases present special problems in areas endemic for chronic infectious diseases because of the possibility of reactivation. Leishmaniasis is a significant neglected tropical disease caused by different species of protozoan parasites within the genus Leishmania. Amastigotes live as intracellular parasites in a variety of mammalian cells, most notably within phagocytes such as macrophages, and residual parasites can persist even after treatment and healing of the lesions. We herein report a case of relapsing mucosal leishmaniasis after aggressive immunotherapy for ankylosing spondylitis, with requirement for secondary prophylaxis with amphotericin B to prevent reactivation. This approach can be necessary for patients from endemic areas of tegumentary leishmaniasis, who will undergo aggressive immunotherapy.

Comparative study on liposomal amphotericin B and other therapies in the treatment of mucosal leishmaniasis: A 15-year retrospective cohort study.

BACKGROUND: Liposomal amphotericin B (L-AMB) has been used for mucosal leishmaniasis (ML), but comparative studies on L-AMB and other drugs used for the treatment of ML have not been conducted. The present study aimed to evaluate the outcome of patients with ML who were treated with L-AMB. METHODS: This is a 15-year retrospective study of Brazilian patients with a confirmed diagnosis of ML. The therapeutic options for the treatment of ML consisted of L-AMB, amphotericin B lipid complex (ABLC), deoxycholate amphotericin B (d-AMB), itraconazole, antimonial pentavalent, or pentamidine. Healing, cure rate and adverse effects (AEs) associated with the drugs used to treat this condition were analyzed. RESULTS: In 71 patients, a total of 105 treatments were evaluated. The outcome of the treatment with each drug was compared, and results showed that L-AMB was superior to other therapeutic regimens (P = 0.001; odds ratio [OR] = 4.84; 95% confidence interval [CI] = 1.78-13.17). d-AMB had worse AEs than other treatment regimens (P = 0.001, OR = 0.09; 95% CI = 0.09-0.43). Approximately 66% of the patients presented with AEs during ML treatment. Although L-AMB was less nephrotoxic than d-AMB, it was associated with acute kidney injury compared with other drugs (P <0.05). CONCLUSION: L-AMB was more effective than other therapies for the treatment of ML. However, a high incidence of toxicity was associated with its use. Therapeutic choices should be reassessed, and the development of new drugs is necessary for the treatment of ML.

Local immunological factors associated with recurrence of mucosal leishmaniasis.

Recurrence of mucosal leishmaniasis (ML) is frequent, but the causative mechanisms are unknown. Our aim was to compare cellular and cytokine patterns of lesions from ML that evolved to recurrence or cure in order to determine the risk factor associated with recurrence. Lesions were evaluated by immunohistochemistry before and after therapy, and patients were followed-up for five years. Higher levels of CD4(+) T and IFN-gamma-producing cells were detected in active lesions and decreased after therapy. Macrophages and IL-10 were markedly increased in cured patients. Conversely, CD8(+) T and NK cells were higher in relapsed than in cured cases. Notably, a decrease in these cells in addition to decreased IL-10 and IFN-gamma was also observed after therapy. These data suggest that exacerbated CD8(+) activity, in addition to a poor regulatory response, could underlie an unfavorable fate with regard to ML. These markers may be useful for predicting the prognosis of ML in lesion studies.

Leishmania: origin, evolution and future since the Precambrian.

This brief review discusses the history of leishmaniasis, considering its origin from the Paleoartic, Neoartic or Neotropic. We reassess some of the theories of the likely origin of this protozoan since the beginning of life on Earth, passing through the Mesozoic and continuing to the appearance of humans. The relationship between this parasite or its ancestors, possible vectors and hosts with regard to ecological modifications is discussed. Recent molecular techniques have helped to elucidate some of the evolutionary questions regarding Leishmania, but have also brought doubts about the origin and evolution of this human parasite. PCR has been used for studies in the new discipline of paleoparasitology, helping to elucidate some of the remaining evolutionary questions. Understanding of this global condition is fundamental in determining the best approach to use against the parasite, specifically for the development of an efficient vaccine.

Mucosal leishmaniasis . Current scenario and prospects for treatment.

Leishmaniasis causes significant morbidity and mortality and thus constitutes a serious public health problem. Even though it has long been endemic in developing countries, in recent years the economic globalization and the increased volume of international travel have extended its prevalence in developed countries. In addition, native populations may be exposed to the infection through blood transfusion and the use of blood products produced from infected asymptomatic individuals. Mucosal leishmaniasis (ML) is a chronic form of this infection, which attacks the mucosa. In most cases this form of leishmaniasis results from the metastatic spread of Leishmania (Viannia) braziliensis from cutaneous lesions. It is a healthcare issue because of its wide demographic distribution, its association with significant morbidity levels, and because of the pressing concern that tourists who travel to endemic areas might present the disease even years later. The treatment currently available for ML is based on drugs such as pentavalent antimony-containing compounds, amphotericin B deoxycholate and pentamidine and often guarantees a satisfactory clinical response. Nevertheless, it also frequently provokes serious side effects. This review offers a critical analysis of the drugs now available for the treatment of ML as also of the future prospects for the treatment of the disease.

Hemophagocytic syndrome associated with hepatitis A: case report and literature review.

Virus-Associated Hemophagocytic Syndrome (VAHS) is a severe hematological disorder related to some viral infections. It is an illness characterized by persistent fever, pancytopenia, splenomegaly, hyperferritinemia and, the most important, hemophagocytosis observed in the bone marrow, liver and/or lymph nodes. VAHS associated with hepatitis A virus infection is rarely described, despite the high incidence of this viral infection in the population in general. There is no consensus in the literature regarding the optimal treatment of VAHS. In this article the clinical features, presumed pathogenesis, diagnostic criteria and treatment of VAHS are discussed, including description of cases of VAHS related to hepatitis A virus infection found in the medical literature.

Treatment of mucosal leishmaniasis in Latin America: systematic review.

Mucosal leishmaniasis (ML) is an important endemic disease and public-health problem in underdeveloped countries because of its significant morbidity and mortality. Increases in ecological tourism have extended this problem to developed countries. This form of leishmaniasis, caused by reactivation after primary cutaneous lesion, has a natural history of progressive destruction of the nasal septa and soft and hard palates, causing facial disfiguration and leading to respiratory disturbances. Treatment of ML, based on several therapies, depends on use of toxic compounds, and few drugs have emerged over the past 40 years. Drug resistance has increased, and the cure rate is no better than 70% in the largest studies. Despite these data, there has been no systematic review of therapies used to treat this important tropical disease. The aim of this study is to determine the best drug management for treatment of ML in Latin America based on the best studies offered by the medical literature. The MEDLINE, LILACS, EMBASE, Web of Science, and Cochrane Library databases were searched to identify articles related to ML and therapy. The studies were independently selected by 2 authors. Articles with sufficient data for cure and treatment failures, internal and external validity information, and > 4 patients in each treatment were included. Validation of this systematic review was based on guidelines to guarantee quality; 22 articles met our inclusion criteria. Stibogluconate achieved a 51% cure rate (76/150 patients), and 88% of patients treated with meglumine were cured (121 patients). Pentamidine and amphotericin were as effective as meglumine. Use of itraconazole and other therapies (pentoxifylline, allopurinol, or interferon-gamma) was controversial, and numbers of patients in some studies were insufficient for statistical analysis. Meglumine may be the drug of choice in the treatment of ML, as it offers similar cure rates when compared with amphotericin B and pentamidine. Cost, adverse effects, local experience, and availability of drugs to treat ML are strong points to be considered before determining the best management of this disease, especially in developing countries.

Reactivation of cutaneous and mucocutaneous tegumentary leishmaniasis in rheumatoid arthritis patients: an emerging problem?

Rheumatoid arthritis (RA) is a chronic condition that is frequent in patients living in tropical areas exposed to leishmaniasis. RA therapy involves immunosuppressant drugs such as methotrexate (MTX), monoclonal antibodies (mAbs) and prednisone. We report an unusual presentation of cutaneous (CL) or mucocutaneous leishmaniasis (ML) in RA patients from an endemic area of leishmaniasis. A 51-year-old woman noted a cutaneous ulcer on her left ankle during MTX and prednisone RA therapy. Initially diagnosed as a venous stasis ulcer, the aspirate of the injury revealed the presence of Leishmania DNA. A 73-year-old woman presenting non-ulcerated, infiltrated and painful erythematous nodules inside her nostrils while receiving MTX, anti-TNF mAb, and prednisone for RA, had also the aspirate of injuries showing the presence of Leishmania DNA. Both patients healed after the therapy with liposomal amphotericin. The RA therapy has changed to low-dose prednisone, without further reactivation episodes. Both cases suggest that CL or ML can reactivate after administration of an immunosuppressant for RA treatment. Therefore, immunosuppressive treatments for RA should be carefully prescribed in patients from endemic areas or with a history of CL and ML.

Could kDNA-PCR in Peripheral Blood Replace the Examination of Bone Marrow for the Diagnosis of Visceral Leishmaniasis?

The aim of this study was to evaluate whether the molecular (kDNA-PCR) and parasitological diagnosis in peripheral blood (PB) could replace the invasive and painful bone marrow collection (BM) in the diagnosis of visceral leishmaniasis (VL). PB from suspected VL patients was evaluated by parasitological and molecular techniques using as the gold standard (GS) a combination of clinical, epidemiological, and immunochromatographic test (PB-rK39) results and parasitological examination of BM. Based on the GS, 38 samples from 32 patients were grouped: Group 1, 20 samples of VL cases, and Group 2, 18 samples of non-VL cases. In order to evaluate the parasitological and molecular techniques in PB, the samples were examined. From Group 1, PB kDNA-PCR was positive in 20 samples and in 19 of 20 in BM kDNA-PCR examination. However, the parasitological examination of buffy coat was insensitive, being able to detect only 4 cases from Group 1. All samples from Group 2 were negative. We concluded that, for the diagnosis of visceral leishmaniasis, the parasitological examination of peripheral blood was not useful; however, molecular diagnosis by kDNA-PCR, performed in peripheral blood, could be useful to replace the parasitological examination of bone marrow.