American cutaneous leishmaniasis: In situ immune response of patients with recent and late lesions

TNF‐α, IFN‐γ, IL‐10, IL‐17, CD68 and CD57 were evaluated in biopsies of patients with American cutaneous leishmaniasis living in Sorocaba, Brazil. The analyses were performed considering the time of lesions from 23 patients with recent lesions (Group I) and 19 patients with late lesions (Group II). All patients were infected with Leishmania (Viannia) braziliensis. Immunostaining cells for CD68, CD57, TNF‐ α, IFN‐γ, IL‐10 and IL‐17 were performed by immunohistochemistry. Except for CD68 and IL‐17, the distribution of in situ for CD57, IL‐10, TNF‐α and IFN‐γ showed that patients with recent lesions expressed higher levels than those with late lesions. The comparison of cytokine expression/group showed that IL‐10 was significantly higher than IL‐17 and IFN‐γ (similar data were shown in IL‐17 compared with TNF‐α), suggesting an immunological balance between inflammatory‐anti‐inflammatory agents. This balance was similar for two groups of patients. In conclusion, these data suggested that (i) patients from Group I had recent lesions (in the beginning of chronic phase) compared to those from Group II and (ii) the modulation of inflammatory response in patients with recent American cutaneous leishmaniasis was correlated with IL‐10 expression in skin lesions preventing the development of mucosal forms. The parasite treatment also prevented the evolution of severe forms.     

T cell responses to crude and defined leishmanial antigens in patients from the Lower Amazon region of Brazil infected with different species of Leishmania of the subgenera Leishmania and Viannia

Amazonian localized cutaneous leishmaniasis (LCL) is caused by parasites of the subgenera Leishmania and Viannia . Respectively, these parasites may cause diffuse cutaneous leishmaniasis (DCL) and mucocutaneous leishmaniasis (MCL). This, together with differing skin test responses, suggests some species-specificity in cell mediated immunity. In this study, T cell responses (proliferative and interferon-γ) to crude and defined antigens were examined in paired samples pre and post chemotherapy. Untreated L. (L.) amazonensis LCL patients showed lower responses to crude leishmanial antigens than the L. (V.) spp. group . L. (V.) braziliensis antigen was a more potent stimulator of T cell responses than L. (L.) amazonensis antigen in all patient groups. Few positive responses were seen to the L. (L.) amazonensis glycoprotein GP46. A substantial proportion of LCL patients did respond to the L. (L.) pifanoi amastigote antigens A2, and the surface membrane glycoprotein P8. DCL patients were poor responders to all leishmanial antigens, except GP46. In contrast, MCL patients were good responders to all antigens except GP46 and A2. A significant rise in the response to P8 and A2 antigen was seen post treatment across all LCL and MCL patients, indicating that these antigens might provide suitable vaccine candidates .     

Familial aggregation of mucosal leishmaniasis in northeast Brazil

To evaluate whether familial clustering occurs in mucosal leishmaniasis (ML), patients with ML (index cases) were randomly selected from medical records at a health post in an endemic area for Leishmania braziliensis infection. Control individuals (index controls) matched by age, gender, and place of residence to index cases were selected. Family members of index cases and controls were compared with respect to environmental factors and the incidence of cutaneous leishmaniasis (CL) and ML. Delayed type hypersensitivity test (DTH) to Leishmania antigen was tested in selected families. Among 289 members of 46 families enrolled, significant differences were found in the frequencies of CL (37% versus 20%) and ML (5% versus 0) in case versus control families, respectively. Families with 2 cases of ML had a higher frequency (29.6%) of DTH-positive individuals than control families (9.4%). These data demonstrate familial clustering of CL, ML, and positive DTH skin tests in an area endemic for L. braziliensis infection.     

Multi-drug resistant A cinetobacter infections in critically injured Canadian forces soldiers

Background

Interferon-gamma is a key cytokine in the protective responses against intracellular pathogens. A single nucleotide polymorphism (SNP) located in the first intron of the human IFN-γ gene can putatively influence the secretion of cytokine with an impact on infection outcome as demonstrated for tuberculosis and other complex diseases. Our aim was to investigate the putative association of IFNG+874T/A SNP with American tegumentary leishmaniasis (ATL) and also the influence of this SNP in the secretion of IFN-γ in vitro.

Methods

Brazilian ATL patients (78 cutaneous, CL, and 58 mucosal leishmaniasis, ML) and 609 healthy volunteers were evaluated. The genotype of +874 region in the IFN-γ gene was carried out by Amplification Refractory Mutational System (ARMS-PCR). Leishmania-induced IFN-γ production on peripheral blood mononuclear cell (PBMC) culture supernatants was assessed by ELISA.

Results

There are no differences between +874T/A SNP frequency in cases and controls or in ML versus CL patients. Cutaneous leishmaniasis cases exhibiting AA genotype produced lower levels of IFN-γ than TA/TT genotypes. In mucosal cases, high and low IFN-γ producers were clearly demonstrated but no differences in the cytokine production was observed among the IFNG +874T or A carriers.

Conclusion

Our results suggest that +874T/A polymorphism was not associated with either susceptibility or severity to leishmaniasis. Despite this, IFNG +874T/A SNP could be involved in the pathogenesis of leishmaniasis by influencing the amount of cytokine released by CL patients, although it could not prevent disease development. On the other hand, it is possible that in ML cases, other potential polymorphic regulatory genes such as TNF-α and IL-10 are also involved thus interfering with IFN-γ secretion.     

IL6 -174 G/C promoter polymorphism influences susceptibility to mucosal but not localized cutaneous leishmaniasis in Brazil

BACKGROUND: Mucosal leishmaniasis (ML) is associated with exaggerated tumor necrosis factor- alpha and interferon- gamma responses and tissue destruction. ML follows localized cutaneous leishmaniasis (CL) caused by Leishmania braziliensis infection. Interleukin (IL)-6 down-regulates T helper (Th) cell type 1 differentiation and drives Th2 cell differentiation. The IL6 -174 G/C polymorphism is associated with proinflammatory diseases and IL-6 regulation. METHODS: The -174 G/C polymorphism was genotyped in population samples and families with CL and ML from Brazil. Genotype frequencies were compared among patients with ML, patients with CL, and 2 control groups by logistic regression and family-based association test (FBAT) analysis. IL-6 levels were measured in macrophages. RESULTS: The C allele was more common in patients with ML than in patients with CL (odds ratio [OR], 2.55 [95% confidence interval {CI}, 1.32-4.91]; P=.005), than in patients who were leishmanin skin-test positive (OR, 2.23 [95% CI, 1.23-4.05]; P=.009), and than in neighborhood control subjects (OR, 2.47 [95% CI, 1.24-4.90]; P=.01). FBAT analysis confirmed an association between allele C and ML under both additive (z=4.295; P=.000017) and dominant (z=4.325; P=.000015) models. Significantly lower levels of IL-6 were measured in unstimulated macrophages from CC individuals than from GG individuals (P=.003) as well as after stimulation with soluble leishmania antigen (P=.009). CONCLUSIONS: IL-6 may regulate type 1 proinflammatory responses, putting individuals with low macrophage IL-6 levels at increased risk for ML.     

Cutaneous leishmaniasis in Switzerland: first experience with species-specific treatment

Background

Different species of the genus Leishmania can cause cutaneous (CL) and mucosal leishmaniasis (ML). PCR-based tests allow a rapid diagnosis and determination of the species, thereby enabling species-oriented treatment. Such treatment procedures have not been evaluated to date.

Methods

Patients presenting with CL and ML between 1999 and 2011 were analysed retrospectively. PCR technology was used to diagnose the disease and identify the protozoan to the species level.

Results

A total of 61 cases were reviewed, including 58 patients with CL and three patients with ML. Treatment was effective in most patients. Treatment failure was reported in six patients with L. panamensis (one fluconazole, one ketoconazole), L. infantum (one excision, one fluconazole), L. tropica (one paromomycin/methylbenzethonium), L. braziliensis (1 paromomycin/methylbenzethonium). In 11 (18 %) patients treatment had to be interrupted due to adverse events, and in eight patients (13 %) a second treatment had to be applied. Treatment with meglumine antimoniate had to be interrupted in six patients, with QTc prolongation the reason for the interruption in three patients.

Conclusions

Species-related, targeted treatment resulted in good responses in CL and ML lesions. Treatment recommendations for L. panamensis were changed from ketoconazole to miltefosine because of new evidence of treatment failures. Meglumine antimoniate should be restricted to species with poor response to alternative medications and should be used with caution in patients older than 60 years because of its toxicity. Treatment in immunosuppressed patients was successful, but relapses were observed when the immune system could not be restored. This is the first report on L. aethiopica from Egypt.     

Case Report: Reactivation of Mucosal and Cutaneous Leishmaniasis in a Renal Transplanted Patient

Mucosal leishmaniasis (ML) is a chronic form of tegumentary leishmaniasis, which causes destructive lesions of nasal, pharyngeal, and laryngeal mucosa. We describe a case of leishmaniasis reactivation with simultaneous cutaneous and mucosal forms in a renal transplanted patient with no history of prior leishmaniasis. Reactivation after renal transplantation was not reported in Brazil. A 67-year-old woman receiving prednisone 20 mg/day, tacrolimus 1 mg/day, and mycophenolic acid 360 mg/day presented with nose edema with erythema and cutaneous lesions. Amastigotes were identified on biopsies and the polymerase chain reaction confirmed Leishmania (Viannia) braziliensis. The patient was treated with liposomal amphotericin B but died 3 weeks after as a result of bacterial septic shock. In conclusion, tegumentary leishmaniasis can reactivate with simultaneous cutaneous and mucosal forms in a renal transplanted patient during the immunosuppressant therapy.     

Evaluation of T-cell subsets in the lesion infiltrates of human cutaneous and mucocutaneous leishmaniasis

We have characterized the T-lymphocytes in the skin lesions of 10 patients with cutaneous leishmaniasis and in the nasal lesions of seven patients with mucosal leishmaniasis, with the immunoperoxidase and monoclonal antibody techniques. There was predominance of cells with helper phenotype (Leu 3A+ 3B) over suppressor phenotype (Leu 2a) in the lesions of both groups. The helper/suppressor (H/S) ratio in the skin lesions was 1.6 +/- 0.5 and in the nasal lesions of mucosal leishmaniasis was 1.7 +/- 0.8. The H/S ratios in the peripheral blood of patients with cutaneous leishmaniasis (2.1 +/- 0.8) and in patients with mucosal leishmaniasis (1.6 +/- 0.8) were comparable and similar to the ratios in the skin and nasal biopsies. The percentage of T-cells and macrophages expressing the Dr antigen in the cutaneous group (69.5 +/- 13.7) was not significantly different from the mucosal patients (90.3 +/- 5.7). We conclude that the immunopathology of the skin lesions in cutaneous leishmaniasis is similar to the nasal lesions of mucosal leishmaniasis.     

Differential immune regulation of activated T cells between cutaneous and mucosal leishmaniasis as a model for pathogenesis

Cutaneous (CL) and mucosal leishmaniasis (ML) are characterized by a predominant type 1 immune response (IFN-gamma and TNF-alpha production) and strong inflammatory response in the lesions with few parasites. This exacerbated type 1 response is more evident in ML as compared to CL. Our main hypothesis is that a differential immune regulation of T cell activation leads to over reactive T cells in ML. In the present study, we investigated immunological factors that could explain the mechanisms behind it by comparing some immune regulatory mechanisms between ML and CL patients: frequency of cells expressing co-stimulatory molecules, apoptotic markers, T cell activation markers; and ability of neutralizing antibodies to IL-2, IL-12 and IL-15 do down-regulate IFN-gamma production in leishmania antigen-stimulated peripheral blood mononuclear cells (PBMC). Interestingly, in CL anti-IL-2 and anti-IL-15 significantly suppressed antigen-specific IFN-gamma production, while in ML only anti-IL-2 suppressed IFN-gamma production. Finally, higher frequency of CD4+ T cells expressing CD28-, CD69+ and CD62L(low) were observed in ML as compared to CL. These data indicate that an exacerbated type 1 response in ML is differentially regulated and not appropriately down modulated, with increased frequencies of activated effectors T cells, maintaining the persistent inflammatory response and tissue damage observed in ML.     

Cutaneous leishmaniasis in British troops following jungle training in Belize

British military personnel account for 45% of cutaneous leishmaniasis (CL) cases seen in the UK and 103 cases from Belize were seen in 1998–2009. Systemic treatment of CL from Belize should not be considered essential in immunocompetent patients because mucosal leishmaniasis very rarely occurs.     

Generic sodium stibogluconate is as safe and effective as branded meglumine antimoniate, for the treatment of tegumentary leishmaniasis in Isiboro Secure Park, Bolivia

Human cutaneous leishmaniasis (CL) and mucous leishmaniasis (ML) are highly endemic in Isiboro Secure Park, which lies in the Bolivian department of Cochabamba--an area where branded meglumine antimoniate (Glucantime) is expensive and poorly distributed. The safety and efficacy of generic sodium stibogluconate (SSG), from Albert David Ltd, was therefore explored, in CL and ML cases from the park, who were treated with 20 mg/kg.day for 20 and 30 days, respectively. A questionnaire recording adverse effects was completed by a physician in each treatment centre. Efficacy of treatment was assessed at the end of treatment and at follow-ups 1 month and 3, 6 and 12 months later. Overall, 146 patients completed treatment with SSG in 2003-2004. No fatalities or severe adverse effects were reported but mild to moderate adverse effects were noted in 41 (28%) of the patients. The incidence of adverse effects was significantly higher among the cases of ML than among the cases of CL. Of the 86 patients with CL who completed 6 months of follow-up, 81 (94.2%) were considered to have been clinically cured; a comparable cohort of 69 CL cases who had been treated with Glucantime in 2001-2002 showed a similar frequency of clinical cure (90%). Generic SSG was shown to be safe and efficacious for the treatment of tegumentary leishmaniasis in Bolivia. Being several times cheaper than Glucantime, it could contribute to improving the access of CL and ML patients to treatment, not only in Bolivia but also in other countries of Latin America.     

MONTENEGRO SKIN TEST AND AGE OF SKIN LESION AS PREDICTORS OF TREATMENT FAILURE IN CUTANEOUS LEISHMANIASIS

A case-control study was conducted to examine the association among the Montenegro skin test (MST), age of skin lesion and therapeutic response in patients with cutaneous leishmaniasis (CL) treated at Evandro Chagas National Institute of Infectious Diseases (INI), Oswaldo Cruz Foundation (FIOCRUZ), Rio de Janeiro, Brazil. For each treatment failure (case), two controls showing skin lesion healing following treatment, paired by sex and age, were randomly selected. All patients were treated with 5 mg Sb⁵⁺/kg/day of intramuscular meglumine antimoniate (Sb⁵⁺) for 30 successive days. Patients with CL were approximately five times more likely to fail when lesions were less than two months old at the first appointment. Patients with treatment failure showed less intense MST reactions than patients progressing to clinical cure. For each 10 mm of increase in MST response, there was a 26% reduction in the chance of treatment failure. An early treatment - defined as a treatment applied for skin lesions, which starts when they are less than two months old at the first appointment -, as well as a poor cellular immune response, reflected by lower reactivity in MST, were associated with treatment failure in cutaneous leishmaniasis.     

Dietary saturated fatty acid and polyunsaturated fatty acid oppositely affect hepatic NOD-like receptor protein 3 inflammasome through regulating nuclear factor-kappa B activation

AIM: To investigate the effect of different dietary fatty acids on hepatic inflammasome activation.METHODS: Wild-type C57BL/6 mice were fed either a high-fat diet or polyunsaturated fatty acid(PUFA)-enriched diet. Primary hepatocytes were treated with either saturated fatty acids(SFAs) or PUFAs as well as combined with lipopolysaccharide(LPS). The expression of NOD-like receptor protein 3(NLRP3) inflammasome, peroxisome proliferator-activated receptor-γ and nuclear factor-kappa B(NF-κB) was determined by real-time PCR and Western blot. The activity of Caspase-1 and interleukine-1β production were measured.RESULTS: high-fat diet-induced hepatic steatosis was sufficient to induce and activate hepatic NLRP3 inflammasome. SFA palmitic acid(PA) directly activated NLRP3 inflammasome and increased sensitization to LPS-induced inflammasome activation in hepatocytes. In contrast, PUFA docosahexaenoic acid(Dh A) had thepotential to inhibit NLRP3 inflammasome expression in hepatocytes and partly abolished LPS-induced NLRP3 inflammasome activation. Furthermore, a highfat diet increased but PUFA-enriched diet decreased sensitization to LPS-induced hepatic NLRP3 inflammasome activation in vivo. Moreover, PA increased but Dh A decreased phosphorylated NF-κB p65 protein expression in hepatocytes.CONCLUSION:Hepatic NLRP 3 inflammasome activation played an important role in the development of non-alcoholic fatty liver disease. Dietary SFAs and PUFAs oppositely regulated the activity of NLRP3 inflammasome through direct activation or inhibition of NF-κB.     

新疆克拉玛依地区几株利什曼原虫分离物的同源性分析

目的：确定新疆克拉玛依皮肤利什曼病患者和硕大白蛉吴氏亚种体内利什曼原虫的种。方法：通过酶切电泳及ＤＮＡ杂交的方法对当地病人体内和蛉体内的利什曼原虫以及参考虫株的ｎＤＮＡ及ｋＤＮＡ的同源性分析，研究克拉玛依病人和蛉体内利什曼原虫的基因型。结果：经ｎＤＮＡ基因型分析，表明病人与蛉体内原虫与婴儿利什曼原虫同源性大。结论：当地皮肤利什曼病的病原体为婴儿利什曼原虫，硕大白蛉吴氏亚种为该病的媒介。     

Leishmania (Viannia) braziliensis: comparative pathology of golden hamsters infected with isolates from cutaneous and mucosal lesions of patients residing in Tres Bracos, Bahia, Brazil.

The histopathology of primary forepaw and metastatic lymph node, spleen, and liver lesions produced in golden hamsters infected with cutaneous leishmaniasis (CL) strains (LTB 111 and LTB558) and mucocutaneous leishmaniasis (MCL) strains (LTB12 and LTB201) of Leishmania (Viannia) braziliensis isolated from patients residing in Tres Bracos, Bahia, Brazil is described. No pathological features providing clear differentiation of the CL and MCL strains were found. Although amastigotes were plentiful early in the development of primary forepaw lesions, they were either absent or could not be identified with certainty in sections of late stage lesions. Similarly, amastigotes were not found in histologic lesions at metastatic sites; however, leishmanial DNA was detected in both early and late stage forepaw lesions and metastatic lesions using Leishmania kinetoplast DNA and the gene coding for gp63 as hybridization probes. The DNA recovered from metastatic lesions was extracted from formalin-fixed paraffin-embedded tissues that had been stored at room temperature for prolonged periods.     

Recombinant Leishmania infantum Heat Shock Protein 83 for the Serodiagnosis of Cutaneous,Mucosal, and Visceral Leishmaniases

Routine serological diagnoses for leishmaniases, except in visceral cases, are performed using whole-parasite antigens. We used enzyme-linked immunosorbent assay (ELISA) to evaluate the performance of Leishmania infantum rHsp83 compared with L. major-like total promastigote antigen in the diagnosis of cutaneous (CL), mucosal (ML), and visceral leishmaniases (VL). ELISA-rHsp83 was significantly more sensitive than ELISA–L. major-like when considering either CL/ML (P = 0.041) or all leishmaniasis patients (P = 0.013). When samples from other infectious disease patients were evaluated for cross-reactivity, ELISA-rHsp83 was more specific than ELISA–L. major-like, specifically for Chagas disease samples (P < 0.001). We also evaluated the anti-rHsp83 antibody titers months after treatment and observed no significant difference in ML (P = 0.607) or CL (P = 0.205). We recommend ELISA–L. infantum-rHsp83 as a routine confirmatory serological assay for the diagnosis of Leishmania infection because of the high sensitivity, the specificity, and the insignificant cross-reactivity with other infectious diseases.     

Intralesional Meglumine Antimoniate for Treatment of Cutaneous Leishmaniasis Patients with Contraindication to Systemic Therapy from Rio de Janeiro (2000 to 2006)

Abstract. We evaluated the effectiveness and safety of intralesional meglumine antimoniate (MA) in 24 not submitted to previous treatment patients with cutaneous leishmaniasis (CL) and with contraindication to systemic therapy. Each treatment consisted of one to four intralesional applications of MA at 15-day intervals. Patients' age ranged from 3 to 90 years; fourteen were females. Intralesional treatment in the absence of any relevant toxicity was successful in 20 (83.3%) patients. Three patients required additional treatment with amphotericin B and one required systemic MA. None of the patients developed mucosal lesions when followed up to 60 months. Intralesional MA is an effective and less toxic alternative treatment of patients with CL and contraindication to systemic therapy.     

The flow-cytometry-based evaluation of cellular immunity in cases of cutaneous leishmaniasis and healthy controls from the endemic area in southern Israel

Only limited data are available on the early immunological events associated with human cutaneous leishmaniasis (CL). In this study, peripheral-blood mononuclear cells were obtained from 66 individuals (34 patients with cutaneous lesions and 32 apparently healthy controls) who had each spent no more than 3 months in the endemic region of Qetzioth, in southern Israel. These cells' responses to Leishmania major antigen were then explored, by the flow-cytometry-based evaluation of blast transformation (BT). The lymphocytes from 17 (50%) of the patients but only one (3%) of the controls displayed BT. When, in an ELISA, most (52) of the subjects were checked for anti-L. major antibodies, none of the 22 controls investigated but 19 (63%) of the 30 patients were found seropositive. Although 14 (47%) of the 30 patients who were checked for antibodies were BT-positive, the seropositive patients were not significantly more or less likely to be BT-positive than the seronegative patients (P<0.919). These data indicate that, in CL, the hosts' cellular and humoral responses develop independently within the first 3 months post-infection, but further investigation is required to confirm this hypothesis.