Tamrat Abebe

Local Suppression of T Cell Responses by Arginase-Induced L-Arginine Depletion in Nonhealing Leishmaniasis

The balance between T helper (Th) 1 and Th2 cell responses is a major determinant of the outcome of experimental leishmaniasis, but polarized Th1 or Th2 responses are not sufficient to account for healing or nonhealing. Here we show that high arginase activity, a hallmark of nonhealing disease, is primarily expressed locally at the site of pathology. The high arginase activity causes local depletion of L-arginine, which impairs the capacity of T cells in the lesion to proliferate and to produce interferon-γ, while T cells in the local draining lymph nodes respond normally. Healing, induced by chemotherapy, resulted in control of arginase activity and reversal of local immunosuppression. Moreover, competitive inhibition of arginase as well as supplementation with L-arginine restored T cell effector functions and reduced pathology and parasite growth at the site of lesions. These results demonstrate that in nonhealing leishmaniasis, arginase-induced L-arginine depletion results in impaired T cell responses. Our results identify a novel mechanism in leishmaniasis that contributes to the failure to heal persistent lesions and suggest new approaches to therapy.

Age-related alteration of arginase activity impacts on severity of leishmaniasis.

Background The leishmaniases are a group of vector-borne parasitic diseases that represent a major international public health problem; they belong to the most neglected tropical diseases and have one of the highest rates of morbidity and mortality. The clinical outcome of infection with Leishmania parasites depends on a variety of factors such as parasite species, vector-derived products, genetics, behaviour, and nutrition. The age of the infected individuals also appears to be critical, as a significant proportion of clinical cases occur in children; this age-related higher prevalence of disease is most remarkable in visceral leishmaniasis. The mechanisms resulting in this higher incidence of clinical disease in children are poorly understood. We have recently revealed that sustained arginase activity promotes uncontrolled parasite growth and pathology in vivo. Here, we tested the hypothesis that arginase-mediated L-arginine metabolism differs with age. Methodology The age distribution of patients with visceral or cutaneous leishmaniasis was determined in cohorts of patients in our clinics in endemic areas in Ethiopia. To exclude factors that are difficult to control in patients, we assessed the impact of ageing on the manifestations of experimental leishmaniasis. We determined parasite burden, T cell responses, and macrophage effector functions in young and aged mice during the course of infection. Results Our results show that younger mice develop exacerbated lesion pathology and higher parasite burdens than aged mice. This aggravated disease development in younger individuals does not correlate with a change in T helper cytokine profile. To address the underlying mechanisms responsible for the more severe infections in younger mice, we investigated macrophage effector functions. Our results show that macrophages from younger mice do not have an impaired capacity to kill parasites; however, they express significantly higher levels of arginase 1 than aged mice and promote parasite growth more efficiently. Thus, our results demonstrate that ageing differentially impacts on L-arginine metabolism and subsequent effector functions of physiologically distinct macrophage subsets. Conclusions Here, we show that arginase-mediated L-arginine metabolism is modulated with age and affects the capacity of macrophages to express arginase; the increased capacity to upregulate this enzyme in younger individuals results in a more permissive environment for parasite growth, increased disease severity and pathology. These results suggest that the difference in arginase-mediated L-arginine catabolism is likely to be an important factor contributing to the increased incidence of clinical cases in children. Thus, targeting L-arginine metabolism might be a promising therapeutic strategy against leishmaniasis, especially in children and young adults.

Nasal carriage rate of methicillin resistant Staphylococcus aureus among Dessie Referral Hospital Health Care Workers; Dessie, Northeast Ethiopia

BackgroundStaphylococcus aureus is a common cause of community and hospital acquired infections. One of the important sources of staphylococci for nosocomial infection is nasal carriage among hospital personnel. Emergence of drug resistant strains especially methicillin resistant Staphylococcus aureus is a serious problem in hospital environments. The aim of this study was to determine the nasal carriage rate of methicillin resistant Staphylococcus aureus among Dessie Referral Hospital healthcare-workers in Ethiopia.MethodsA cross sectional study was conducted on a total of 118 healthcare workers. Nasal swabs were collected and cultured on Mannitol Salt Agar. Slide coagulase test was performed. An oxacillin susceptibility test was carried out on Muller Hinton agar using modified Kirby-Bauer disc diffusion method.ResultsOf the 118 healthcare workers, 34 (28.8%) carried S. aureus of which 15 were methicillin resistant. Therefore, 12.7% of all HCWs were identified as MRSA carriers. The rate of methicillin resistance among all S. aureus isolates was 44.1% (15/34). MRSA carriage was particularly high among nurses (21.2%). The highest rate of MRSA carriers (57.1%) were workers of surgical wards.ConclusionsThe high rate of nasal MRSA carriage among healthcare workers found in this study indicates the need for adjusted infection control measures to prevent MRSA transmission in our healthcare setting.

Local increase of arginase activity in lesions of patients with cutaneous leishmaniasis in Ethiopia

Background Cutaneous leishmaniasis is a vector-borne disease that is in Ethiopia mainly caused by the parasite Leishmania aethiopica. This neglected tropical disease is common in rural areas and causes serious morbidity. Persistent nonhealing cutaneous leishmaniasis has been associated with poor T cell mediated responses; however, the underlying mechanisms are not well understood. Methodology/Principal Findings We have recently shown in an experimental model of cutaneous leishmaniasis that arginase-induced L-arginine metabolism suppresses antigen-specific T cell responses at the site of pathology, but not in the periphery. To test whether these results translate to human disease, we recruited patients presenting with localized lesions of cutaneous leishmaniasis and assessed the levels of arginase activity in cells isolated from peripheral blood and from skin biopsies. Arginase activity was similar in peripheral blood mononuclear cells (PBMCs) from patients and healthy controls. In sharp contrast, arginase activity was significantly increased in lesion biopsies of patients with localized cutaneous leishmaniasis as compared with controls. Furthermore, we found that the expression levels of CD3ζ, CD4 and CD8 molecules were considerably lower at the site of pathology as compared to those observed in paired PBMCs. Conclusion Our results suggest that increased arginase in lesions of patients with cutaneous leishmaniasis might play a role in the pathogenesis of the disease by impairing T cell effector functions.

Increased level of arginase activity correlates with disease severity in HIV-seropositive patients.

Infection with human immunodeficiency virus (HIV) results in a chronic infection that progressively impairs the immune system. Although depletion of CD4(+) T cells is frequently used to explain immunosuppression, chronicity of infection and progressive loss of CD4(+) T cells are not sufficient to fully account for immune dysregulation. Arginase-induced l-arginine deprivation is emerging as a key mechanism for the down-regulation of immune responses. Here, we hypothesized that the level of arginase activity increases with disease severity in HIV-seropositive patients. We determined the levels of arginase activity in peripheral blood mononuclear cells from HIV-seropositive patients and uninfected control participants. Our results show that peripheral blood mononuclear cells from HIV-seropositive patients with low CD4(+) T cell counts expressed statistically significantly higher levels of arginase activity, compared with patients with high CD4(+) T cell counts or uninfected control participants. Furthermore, we found a statistically significant correlation between high level of arginase activity and high viral load in HIV-seropositive patients.

Arginase activity in the blood of patients with visceral leishmaniasis and HIV infection.

Background Visceral leishmaniasis is a parasitic disease associated with high mortality. The most important foci of visceral leishmaniasis in Ethiopia are in the Northwest and are predominantly associated with high rates of HIV co-infection. Co-infection of visceral leishmaniasis patients with HIV results in higher mortality, treatment failure and relapse. We have previously shown that arginase, an enzyme associated with immunosuppression, was increased in patients with visceral leishmaniasis and in HIV seropositive patients; further our results showed that high arginase activity is a marker of disease severity. Here, we tested the hypothesis that increased arginase activities associated with visceral leishmaniasis and HIV infections synergize in patients co-infected with both pathogens. Methodology/Principal Findings We recruited a cohort of patients with visceral leishmaniasis and a cohort of patients with visceral leishmaniasis and HIV infection from Gondar, Northwest Ethiopia, and recorded and compared their clinical data. Further, we measured the levels of arginase activity in the blood of these patients and identified the phenotype of arginase-expressing cells. Our results show that CD4+ T cell counts were significantly lower and the parasite load in the spleen was significantly higher in co-infected patients. Moreover, our results demonstrate that arginase activity was significantly higher in peripheral blood mononuclear cells and plasma of co-infected patients. Finally, we identified the cells-expressing arginase in the PBMCs as low-density granulocytes. Conclusion Our results suggest that increased arginase might contribute to the poor disease outcome characteristic of patients with visceral leishmaniasis and HIV co-infection.

Arginase activity - a marker of disease status in patients with visceral leishmaniasis in ethiopia.

The underlying mechanisms resulting in the profound immune suppression characteristic of human visceral leishmaniasis (VL) are not fully understood. Here, we tested the hypothesis that arginase, an enzyme associated with immunosuppression, is higher in patients with VL and contributes to impaired T cell responses. We recruited patients with VL before and after treatment and healthy controls and measured the arginase metabolism in the blood of these individuals. Our results show that arginase activity is significantly higher in the blood of patients with active VL as compared to controls. These high levels of arginase decline considerably once the patients are successfully treated. We identified the phenotype of arginase-expressing cells among PBMCs as neutrophils and show that their frequency was increased in PBMCs of patients before treatment; this coincides with reduced levels of L-arginine in the plasma and decreased expression levels of CD3ζ in T cells.

Hepatitis B virus infection among medical waste handlers in Addis Ababa, Ethiopia

BackgroundHealthcare wastes contain a wide range of microorganisms among which hepatitis B virus (HBV) are the most significant pathogens. No data about the prevalence of HBV among medical waste handlers is available in Addis Ababa, Ethiopia. Therefore; this study was conducted to describe the prevalence of HBV infection among medical waste handlers in Government hospitals of Addis Ababa, Ethiopia.FindingsA cross sectional study was conducted among 252 medical and non-medical waste handlers working in three Government hospitals of Addis Ababa between May to July, 2010. Predesigned and tested questionnaire was used to collect soiociodemographic information. Blood sample was taken from 252 waste handlers and serum was tested for Hepatitis B surface antigen (HBsAg) and anti-Hepatitis core antigen (anti-HBcAg) using Enzyme Linked Immuno Sorbent Assay.Of the 126 Medical Waste Handlers and 126 Non Medical Waste Handler, HBsAg was detected in 8 (6.3%) and 1 (0.8%), and anti-HBcAg in 60 (47.6%) and 40 (31.7%), respectively. Significant differences were observed in the detection rates of HBsAg (OR: 8, 95% CI: 1.02, 63.02; p = 0.01), Anti-HB c Ag (OR: 1.5, 95% CI: 1.1, 2.1; p = 0.01) and either markers (OR: 1.7, 95% CI: 1.2, 2.2; p = 0.001) in medical waste handlers compared to non medical waste handlers. 19.8% were trained to handle medical waste and none was immunized against HBV.ConclusionThis study shows a high prevalence of HBV infection in medical waste handlers compared to non medical waste handlers. Lack of training on how to handle medical waste among medical waste handlers was high.

A clinical isolate of Leishmania donovani with ITS1 sequence polymorphism as a cause of para‐kala‐azar dermal leishmaniasis in an Ethiopian human immunodeficiency virus‐positive patient on highly active antiretroviral therapy

The diagnosis of para‐kala‐azar dermal leishmaniasis (PKDL/VL), either as an immune reconstitution inflammatory syndrome (IRIS)‐associated syndrome or as a complication of visceral leishmaniasis (VL) during human immunodeficiency virus (HIV) co‐infection in patients with or without highly active antiretroviral therapy, represents a challenge for prompt treatment. The aim of this study was to identify the causative Leishmania species and to clarify whether the post‐kala‐azar dermal leishmaniasis (PKDL)‐like lesions appeared as a result of IRIS or not. A 31‐year‐old Ethiopian male patient, with stage IV HIV/acquired immune deficiency syndrome (AIDS), was clinically diagnosed with PKDL/VL. He had developed a generalized maculopapular rash on his face after initiation of highly active antiretroviral therapy. The Leishmania isolate obtained from the skin lesions was analysed by polymerase chain reaction–restriction fragment length polymorphism (PCR‐RFLP) and sequencing of the ribosomal DNA internal transcribed spacer 1 (ITS1) and partial coding sequences of the heat shock protein 70 gene (hsp70). Restriction analysis of the ITS1 PCR product gave a unique RFLP pattern not seen before for any Leishmania isolate. Sequencing of both the ITS1 and hsp70 PCR products identified the causative species as Leishmania donovani, and further revealed the existence of five different sequence variants of the ITS1 among the 10 clones sequenced. The results indicate that PKDL/VL resulted from an infection by L. donovani. The sequence variants of ITS1 might be due to the presence of multiple strains/clones or the existence of intragenomic variations in the multicopy ITS1, or a combination of both.

Evaluation of crude hydatid cyst fluid antigens for the serological diagnosis of hydatidosis in cattle

Echinococcosis is a zoonotic infection caused by adult or larval (metacestode) stages of cestodes belonging to the genus Echinococcus. The purpose of this study was to evaluate the antigenic ability of hydatid cyst fluid antigen for the diagnosis of hydatidosis in cattle using enzyme-linked immunosorbent assay (ELISA) and indirect haemagglutination test (IHA). The source of the antigens for the serological tests was fertile crude cyst fluids collected from naturally infected sheep at the Addis Ababa abattoir. A total of 502 sera were collected from 329 uninfected cattle and 173 hydatid-infected cattle which were confirmed by post-mortem examination. Most cysts were sterile and multiple organ infection predominated. Of 173 infected cattle, 166 (96.0%; confidence interval (CI) 91.8-98.4) were positive using ELISA while 7 (4.0%) were negative. Of 329 sera from uninfected cattle, 274 (83.3%; CI 78.8-87.2) were found to be negative and the remaining 55 (16.7%) were positive by ELISA. Of 173 infected cattle, 151 (87.3%; CI 81.4-91.9) were positive and 22 (12.7%) were negative by IHA. Of 329 negative sera tested using IHA, 266 (80.9%; CI 76.2-85.0) were negative and the remaining 63 (19.1%) were positive. The false positive and negative values of ELISA were 4.0 and 16.7%, respectively, and the corresponding values of IHA were 12.7 and 19.1%. The sensitivity and diagnostic efficiency of IHA were 87.2 and 83.6%, respectively. Crude hydatid cyst fluid antigen seems to have reasonable antigenic properties and hence could be employed for epidemiological surveillance of cattle hydatidosis.