Thymic alterations resulting from experimental visceral leishmaniasis in a Syrian hamster (Mesocricetus auratus).

BACKGROUND: The thymus is a lymphoid organ responsible for the development and maturation of T cells, which are part of the Th1, Th2, Th17, and Treg immune responses triggered by visceral leishmaniasis. The maturation and immunological development of T lymphocytes require a bidirectional interaction between the thymic microenvironment of epithelial cells, dendritic cells, and macrophages and the extracellular matrix with differentiating lymphocytes. OBJECTIVES: We evaluated the morphological characteristics and tissue distribution of hematopoietic and stromal cells in the thymuses of hamsters experimentally infected with Leishmania infantum, aiming to gain an insight into the pathophysiology of the disease. METHODS: Fifteen hamsters were subjected to intraperitoneal experimental infection with 107L. infantum promastigotes (MHOM/BR/1972/BH46). The animals were divided into three groups, each comprising five infected hamsters, and were then euthanized 15, 60, and 120 days postinfection. The control groups consisted of three groups of five healthy hamsters euthanized simultaneously with the infected ones. Thymic morphology was evaluated through histopathology and the cell composition through immunohistochemistry. We used antibodies to mark mesenchymal cells (anti-vimentin), epithelial cells (anti-cytokeratin), macrophages (anti-MAC387), B lymphocytes (anti-CD79a), and T lymphocytes (anti-CD3). Immunohistochemistry was also used to mark the parasite in the thymus. RESULTS: Infected and control hamsters showed no difference in thymic morphology and degree of atrophy. After 15 days of infection, CD3 + T lymphocytes in the thymus showed an increase that stabilized over time. At 120 days of infection, we detected a significant decrease in CD79a+ B lymphocytes. The parasite was present in the medullary and corticomedullary regions of 9 out of 15 hamsters. These findings confirm that the presence of a parasite can cause changes in a thymus cell population. However, further studies are needed to evaluate these changes' effects on the immune response of infected animals.

Evaluation of systemic immunity in atypical cutaneous leishmaniasis caused by Leishmania (L.) infantum chagasi.

In some central-American countries, Leishmania (L.) infantum chagasi infection can cause non-ulcerated or atypical cutaneous leishmaniasis (NUCL) in addition to the classic clinical form, visceral leishmaniasis (VL). Little is known about the host-parasite relationship that can contribute to the determination of one or another clinical form. The present study had the objective to evaluate the humoral and cellular immunity in the sera of individuals affected by NUCL to improve the comprehension of this atypical host-parasite interaction. Based on clinical and laboratory diagnosis, serum of 80 individuals was collected to evaluate the cytokines and immunoglobulins profile of NUCL (n = 47), VL patients (n = 5), and negative controls (n = 28). Cytokines were detected using Cytokine Bead Array (CBA) Human Th1/Th2/Th17 kit according to the manufacturer's instructions; class (IgG and IgM), and subclass of (IgG1 and IgG2) immunoglobulins was evaluated by ELISA using specific antigens. The concentration of TNF-α, IFN-γ, IL-2 and IL-4 cytokines in NUCL, VL and control was present below the detection threshold of CBA kit. IL-6, IL-10 and IL-17A cytokines was lower in NUCL compared to LV patients. Regarding to immunoglobulins, NUCL patients produced 4.0 times more IgG than the control, while VL patients produced 6.6 times more; and IgM level was 1.6 times higher in NUCL and 2.6 times in VL patients compared to the control. Concerning the immunoglobulins subclass, only VL patients showed positive reaction for IgG1, and IgG2 did not show positive reaction among the groups. The results showed a weak cellular and humoral systemic immune response in NUCL patients.

Whole-Genome Sequencing of Leishmania infantum chagasi Isolates from Honduras and Brazil.

This work reports on the whole-genome sequencing of Leishmania infantum chagasi from Honduras (Central America) and Brazil (South America).

In situ study of cellular immune response in human cutaneous lesions caused by Leishmania (Viannia) panamensis in Panama.

AIMS: Leishmaniasis is considered a disease with multiple clinical/immunopathological characteristics, depending on the immunity of the host and the species of the parasite. In Panama, the most prevalent species that causes localized cutaneous leishmaniasis (LCL) is Leishmania (Viannia) panamensis, and its immune response is poorly studied. Therefore, we evaluated by immunohistochemistry, the in situ immune response during this infection. METHODS AND RESULTS: Biopsies from Panamanian patients with LCL were collected and processed by histological techniques. Infection by L. (V.) panamensis was demonstrated by isolation in culture and molecular characterization by Hsp70-RFLP. The in situ immune response was assessed by immunohistochemistry. The immune response was characterized by predominance of T cells, mainly CD8 cells that showed positive correlation with IFN-γ and Granzyme B. CD4 cells presented positive correlation with both IFN-γ and IL-13, pointed by mixed cellular immune response. Regulatory response was characterized by FoxP3 cells, which showed positive correlation to IL-10 but not with TGF-ß. CONCLUSIONS: L. (V.) panamensis infection triggers a mixed cellular immune response, characterized by the presence of pro-inflammatory, anti-inflammatory and regulatory elements in the skin lesion of Panamanian patients. These data contribute to a better understanding of the immunopathogenesis of Leishmania Viannia infection in Panama.

Detection of Leishmania infantum DNA in Pintomyia evansi and Lutzomyia longipalpis in Honduras.

BACKGROUND: The two most abundant sand fly species on the Honduran Pacific coast are Lutzomyia (Lutzomyia) longipalpis and Pintomyia (Pifanomyia) evansi. Both species are known vectors of Leishmania (Leishmania) infantum, the etiological agent of visceral leishmaniasis (VL) in the Americas. Although VL and non-ulcerative cutaneous leishmaniasis (NUCL) are endemic on the Pacific versant of the Central American Pacific, the latter is the most frequent manifestation of leishmaniasis there. We evaluated the circulation of Leishmania spp. in the sand fly species on El Tigre Island, an endemic area of NUCL. RESULTS: We collected 222 specimens of six sand fly species. Lu. longipalpis (180 specimens; 81%) and Pif. (Pi.) evansi (35 specimens; 16%) were the most abundant species. L. (L.) infantum DNA was detected in nine of the 96 specimens analyzed; seven of these specimens were identified as Lu. longipalpis, and the remaining two were Pi. evansi, with an infection rate of 9.4% and 2.7%, respectively. CONCLUSION: We present the first record of L. (L.) infantum DNA in Pi. evansi from a NUCL endemic region of Central America. Our results suggest that Pi. evansi could be a secondary vector of L. (L.) infantum in the transmission cycle of leishmaniasis. The detection of natural infections of L. (L.) infantum in sand flies in this region contributes to an understanding of the epidemiology of leishmaniasis in Honduras.

Clinical and Immunological Features of Human Leishmania (L.) infantum-Infection, Novel Insights Honduras, Central America.

Leishmania (Leishmania) infantum is the etiological agent of both American visceral leishmaniasis (AVL) and non-ulcerated cutaneous leishmaniasis (NUCL) in Honduras. Although AVL is the most severe clinical form of infection, recent studies have shown that human immune response to parasite infection can result in a clinical-immunological spectrum. The overall prevalence rate of infection and clinical-immunological profiles of the L. (L.) infantum infection in Amapala municipality, South Honduras was determined. We examined 576 individuals with diagnosis based on combined ELISA (IgG/IgM) and DTH assays. We also used genus-specific kDNA PCR and Hsp70 PCR-RFLP for NUCL cases. Clinical evaluation found 82% asymptomatic and 18% symptomatic individuals. All symptomatic cases (n = 104) showing NUCL were positive for parasites. We identified L. (L.) infantum species in 100% of the skin lesion scrapings and in 90% of the blood samples from NUCL cases studied. A total of 320 asymptomatic individuals were exposed (ELISA+ and/or DTH+), providing an overall L. (L.) infantum prevalence of 73.6%. Clinical, parasitological, and immunological evaluations suggest seven infection profiles, three asymptomatic and four symptomatic. This represents the first report on clinical and immunological features of human L. (L.) infantum-infection in Amapala municipality, Honduras.

Th17 lymphocytes in atypical cutaneous leishmaniasis caused by Leishmania (L.) infantum chagasi in Central America.

Skin lesions in nonulcerated cutaneous leishmaniasis (NUCL) caused by Leishmania (L.) infantum chagasi are characterized by a mononuclear inflammatory infiltrate in the dermis, which is composed mainly of lymphocytes, followed by macrophages, few plasma cells and epithelioid granulomas with mild tissue parasitism. Previous studies have shown that the main population of lymphocytes present in the dermal infiltrate is CD8+ T cells, followed by CD4+ T cells, which are correlated with IFN-γ+ cells. To improve the knowledge of cellular immune responses in NUCL, skin biopsies were submitted to immunohistochemistry using anti-ROR-γt, anti-IL-17, anti-IL-6, anti-TGF-ß, and anti-IL-23 antibodies to characterize the involvement of Th17 cells in the skin lesions of patients affected by NUCL. ROR-γt+ , IL-17+ , IL-6+ , TGF-ß+ and IL-23+ cells were observed in the dermal inflammatory infiltrate of NUCL skin lesions. A positive correlation between CD4+ T-lymphocytes and ROR-γt+ and IL-17+ cells suggests that some of the CD4+ T-lymphocytes in NUCL could be Th17 lymphocytes. Moreover, a positive correlation between ROR-γt+ cells and TGF-ß+ , IL-6+ , IL-17+ and IL-23+ cells could indicate the role of these cytokines in the differentiation and maintenance of Th17 lymphocytes. Our findings improve knowledge of the pathogenesis of this rare and atypical clinical form of leishmaniasis.

Histopathological features of skin lesions in patients affected by non-ulcerated or atypical cutaneous leishmaniasis in Honduras, Central America.

In Honduras visceral leishmaniasis and non-ulcerated or atypical cutaneous leishmaniasis (NUCL) are caused by the species Leishmania (L.) infantum chagasi. NUCL is the most common clinical form in the southern regions of the country, mainly affecting the young. In view of the lack of knowledge about the pathogenesis of the disease pattern caused by L. (L) infantum chagasi in individuals affected by NUCL, the aim of the present study was to describe in detail the histopathological features of the skin lesion caused by the parasite. Biopsies from human NUCL lesions with a positive parasitological diagnosis were collected and processed using standard histological techniques. Paraffin sections stained by haematoxylin and eosin were used to examine the histopathological alterations seen in the skin. The lesions varied between 3 and 5 mm, and the majority of the patients (60%) had a single lesion. Lesions were more frequently seen in females (65%), with an average age of 33.4 years. Microscopically, the skin lesions were characterized by mononuclear inflammatory infiltrate in the dermis composed of lymphocytes, macrophages and a few plasma cells. The intensity of the infiltration varied from discrete to intense. In both cases, the parasitic infection was discrete. Granulomas were present in 60% of cases and were associated with intense inflammation. The data revealed by the histopathological alterations in the skin of individuals affected by NUCL suggest activation of a cellular immune response that potentially controls parasite spreading.

Evaluation of Regulatory Immune Response in Skin Lesions of Patients Affected by Nonulcerated or Atypical Cutaneous Leishmaniasis in Honduras, Central America.

In Honduras, Leishmania (L.) infantum chagasi causes both visceral leishmaniasis (LV) and nonulcerated or atypical cutaneous leishmaniasis (NUCL). NUCL is characterized by mononuclear inflammatory infiltration of the dermis, composed mainly of lymphocytes followed by macrophages with discrete parasitism. Considering that little is known about the pathogenesis of NUCL, the aim of this study was to evaluate the regulatory response in situ in skin lesions of patients affected by NUCL. Biopsies (n = 20) from human cutaneous nonulcerative lesions were collected and processed by usual histological techniques. The in situ regulatory immune response was evaluated by immunohistochemistry using antihuman CD4, FoxP3, IL-10, and TGF-ß antibodies. CD4+, FoxP3+, TGF-ß+, and IL-10+ cells were observed in the dermis with inflammatory infiltration in all studied cases and at higher densities compared to the normal skin controls. A positive and strong correlation was observed between CD4+ and FoxP3+ cells, and a positive and moderate correlation was observed between FoxP3+ and TGF-ß+ but not with IL-10+ cells. The data suggest that T regulatory FoxP3+ cells and the regulatory cytokines, especially TGF-ß, play an important role in the immunopathogenesis of NUCL, modulating a cellular immune response in the skin, avoiding tissue damage, and leading to low tissue parasitic persistence.