Application of recombinant proteins for serodiagnosis of visceral leishmaniasis in humans and dogs

Visceral leishmaniasis [VL] is a zoonotic disease caused by leishmania species. Dogs are considered to be the main reservoir of VL. A number of methods and antigen-based assays are used for the diagnosis of leishmaniasis. However, currently available methods are mainly based on direct examination of tissues for the presence of parasites, which is highly invasive. A variety of serological tests are commonly applied for VL diagnosis, including indirect fluorescence antibody test, enzyme-linked immunosorbent assay [ELISA], dot-ELISA, direct agglutination test, Western-blotting, and immunochromatographic test. However, when soluble antigens are used, serological tests are less specific due to cross-reactivity with other parasitic diseases. Several studies have attempted to replace soluble antigens with recombinant proteins to improve the sensitivity and the specificity of the immunodiagnostic tests. Major technological advances in recombinant antigens as reagents for the serological diagnosis of VL have led to high sensitivity and specificity of these serological tests. A great number of recombinant proteins have been shown to be effective for the diagnosis of leishmania infection in dogs, the major reservoir of L. infantum. Although few recombinant proteins with high efficacy provide reasonable results for the diagnosis of human and canine VL, more optimization is still needed for the appropriate antigens to provide high-throughput performance. This review aims to explore the application of different recombinant proteins for the serodiagnosis of VL in humans and dogs

An epidemiological comparative study on diagnosis of rodent leptospirosis in Mazandaran Province, northern Iran / 한국역학회지

OBJECTIVES: Leptospirosis is a zoonosis caused by leptospires, in which transmission occurs through contact with contaminated biological fluids from infected animals. Rodents can act as a source of infection for humans and animals. The disease has a global distribution, mainly in humid, tropical and sub-tropical regions. The aim of this study was to compare culture assays, the microscopic agglutination test (MAT), polymerase chain reaction (PCR), and nested PCR (n-PCR), for the diagnosis of leptospirosis in rodents in Mazandaran Province, northern Iran. METHODS: One hundred fifty-one rodents were trapped alive at 10 locations, and their urine and kidney samples were collected and used for the isolation of live Leptospira. The infecting serovars were identified and the antibody titres were measured by MAT, using a panel of 20 strains of live Leptospira species as antigens. The presence of leptospiral DNA was evaluated in urine and kidney samples using PCR and n-PCR. RESULTS: No live leptospires were isolated from the kidney and urine samples of the rodents. Different detection rates of leptospirosis were observed with MAT (21.2%), PCR (11.3%), and n-PCR (3.3%). The dominant strain was Leptospira serjoehardjo (34.4%, p=0.28), although other serotypes were also found. The prevalence of positive leptospirosis tests in rodents was 15.9, 2.6, and 2.6% among Rattus norvegicus, R. rattus, and Apodemus sylvaticus, respectively. CONCLUSIONS: Leptospirosis was prevalent in rodents in Mazandaran Province, northern Iran. MAT was able to detect leptospires more frequently than culture or PCR. The kidney was a more suitable site for identifying leptospiral DNA by n-PCR than urine. Culture was not found to be an appropriate technique for clinical diagnosis.

Lead exposure changes gastric motility in rats: role of nitric oxide [no]

Abdominal colic, constipation and delay in gastric emptying are symptoms of lead poisoning, but there is scant information about the effect of lead on gastric motility. In the present study, we investigated the effect of lead acetate on gastric motility in rats. Animals were divided into nine groups [n=8]; four groups were exposed to lead acetate solution [1%] for 1, 2, 3, and 4 weeks [Pb1, Pb2, Pb3, and Pb4 groups, respectively]. Sodium acetate solution was given to another four groups for 1, 2, 3, and 4 weeks [Na1, Na2, Na3, and Na4 groups, respectively] and the control group had free access to tap water. Gastric motility was measured in the basal and acetylcholine [Ach]-stimulated states using a physiograph instrument. Nitric oxide metabolite of gastric tissue was determined by Griess micro-assay. There were no significant differences between basal and Ach-stimulated gastric motility in Pb1, Pb2, Na1, and Na2 groups. However, it was significantly greater in Pb3 and Pb4 groups when compared with Na3 and Na4 groups in both basal and Ach-stimulated states [P<0.05]. In addition, nitric oxide metabolite of gastric tissue was more in all Pb groups in comparison with their Na counterparts [P<0.05]. We found that lead exposure could affect gastric motility via the nitric oxide pathway

Molecular analysis of A2-genes encoding stage-specific s antigen-like proteins among isolates from iranian cutaneous and visceral leishmaniasis

Leishmania can lead to a broad spectrum of diseases, collectively known as leishmaniasis. The A2 gene/ protein family could be one of the most eligible candidate factors of virulence in visceral leishmaniasis [VL]. The previous results confirmed that in Leishmania infantum, several A2 proteins are abundantly expressed by the amastigote, but not the promastigote stage. As there are no data available on the pattern of A2 gene / protein in Iranian Leishmania isolates of either cutaneous leishmaniasis [CL] or VL; the current study aimed to investigate molecular analysis of A2 gene in leishmania species among field isolates of Iran. An A2 gene was identified by sequencing of crude PCR products resulting from 20 samples of CL and 10 samples of VL isolates from Iranian patients. The results indicated the A2 gene in CL is only a single copy of 153 bp encoding a protein of 51 amino acids, as opposed to A2 of VL species with multi-copy genes of varying length. A2 sequences in Iranian L. major strains represented a homology with stage-specific S antigen-like protein [A2] of L major and L infantum. Moreover, A2 sequences in Iranian L. tropica strains have homology with A2 protein of L. major and L. tropica. It is concluded that A2 is an antigen candidate for vaccine development and diagnosis purposes and that A2 sequences are conserved among field isolates

effects of cholestasis and cirrhosis on gastric acid and pepsin secretions in rat: involvement of Nitric Oxide

The liver has major role in the organism homeostasis, interactions with other systems, synthesis and metabolism of bile production, drug detoxification and hormone inactivation. Cholestasis can be defined as an impairment of the bile flow which can lead to hepatocytes necrosis and finally cirrhosis. Some studies reported a gastric acid secretion reduction in cirrhotic subjects, while others reported normal production gastric acid secretion. Our aim was to evaluate the effects of cholestasis and cirrhosis on gastric acid and pepsin secretions and its possible mechanism in rat. Male Wistar rats were randomly divided into five groups [n=8]: control, cholestasis, sham cholestasis, cirrhosis and sham cirrhosis. Laparotomy was done under general anesthesia and then bile duct ligation [BDL] was performed. After 2 and 4 weeks in cholestasis and cirrhosis groups respectively, gastric content was collected by wash-out technique. Basal and stimulated acid an pepsin secretions were measured by using titration and the Anson method respectively in all groups. In order to measure stimulated acid and pepsin secretions, pentagastrin [25 micro g/kg, i.p.] was used. Nitric Oxide [NO] metabolites of gastric tissue were determined by Griess microassy method. Acid and pepsin secretions were significantly reduced in cholestatic and cirrhotic rats in comparison with control and sham groups [P< 0.01]. NO metabolite of gastric tissue was significantly increased in cholestatic and cirrhotic rats [P> 0.01]. Reducing of gastric acid and pepsin output in cholestatic and cirrhotic rats may be due to increasing in NO content of gastric tissue

Nitric oxide functions: An emphasis on its diversity in infectious diseases

Nitric oxide is a short-lived mediator, which can be induced in a variety of cell types and produces many physiologic and metabolic changes in target cells. It is important in many biological functions and generated from L-arginine by the enzyme nitric oxide synthase. Nitric oxide conveys a variety of messages between cells, including signals for vasorelaxation, neurotransmission and cytotoxicity. In macrophages, nitric oxide synthase activity appears slowly after exposure of the cells to cytokines and bacterial products, is sustained, and functions independently of calcium and calmodulin. The cytokine- inducible nitric oxide synthase [iNOS] is activated by several immunological stimuli, leading to the production of large quantities of nitric oxide which can be cytotoxic. To date, there have been conflicting reports concerning the clinical significance of nitric oxide in infections. Some authors have proposed that nitric oxide contributes to the development of severe and complicated cases, while others have argued that nitric oxide has a protective role. The aim of this review is to evaluate the functions of nitric oxide production toward oxidative stress induced by infections or inflammations. It is indicated that NO is an important, but possibly not essential contributor in the control of acute phase of infections and it is only part of an immunopathological chain against pathogens. The anti-microbial function does not relate only to nitric oxide action or its related molecules, a combination of nitric oxide and immune factors is required to resolve pathogenic microorganisms. Consequently, the NO theory in infectious diseases may lead to the novel ideas for therapy and prevention

Anti-leishmanial Effects of Trinitroglycerin in BALB/C Mice Infected with Leishmania major via Nitric Oxide Pathway

This study investigated whether trinitroglycerine (TNG) as nitric oxide (NO) releasing agent had anti-leishmanial effects and mediated pathology in BALB/c mice infected with Leishmania major. Cutaneous leishmaniasis (CL), a zoonotic infection caused by leishmania protozoa is still one of the health problems in the world and in Iran. NO is involved in host immune responses against intracellular L. major, and leishmania killing by macrophages is mediated by this substance. Moreover, application of CL treatment with NO-donors has been recently indicated. In our study, TNG was used for its ability to increase NO and to modify CL infection in mice, in order to evaluate NO effects on lesion size and formation, parasite proliferation inside macrophages, amastigote visceralization in target organs, and NO induction in plasma and organ suspensions. Data obtained in this study indicated that TNG increased plasma and liver-NO, reduced lesion sizes, removed amastigotes from lesions, livers, spleens, and lymph nodes, declined proliferation of amastigotes, hepatomegaly, and increased survival rate. However, TNG reduced spleen-NO and had no significant effects on spelenomegaly. The results show that TNG therapy reduced leishmaniasis and pathology in association with raised NO levels. TNG had some antiparasitic activity by reduction of positive smears from lesions, livers, spleens, and lymph nodes, which could emphasize the role of TNG to inhibit visceralization of L. major in target organs.

Pathogenicity variations of susceptibility and resistance to leishmania major MRHO/IR/75/ER strain in BALB/c and C57BL/6 mice

To compare the pathogenicity differences in two susceptible Balb/c and resistant C57b1/6 mice infected with Leishmania major MRHO/IR/75/ER as a prevalent strain of zoonotic cutaneous leishmaniasis in Iran. Mice were assigned into four groups as control and infected BALB/c and C57BL/6 mice. Experimental leishmaniasis was initiated by [s. c] injection of the 2x10[6] L. major promastigotes into the basal tail of infected groups. The development of lesions was determined weekly by measuring the two diameters. After 10 weeks, all mice were killed humanly, target tissues including lymph node, spleen and liver from each mouse were removed, weighted, and their impression smears were prepared. Proliferation of amastigotes inside macrophages, pathogenicity signs in two susceptible, resistant hosts was varied, and these variations were depended on mice strain. Host immunity may modify clinical signs and could affect the proliferation of amastigotes inside macrophages, the size of lesions, the survival rates, the degree of hepatomegaly and splenomegaly and the percentage of amastigotes in lesion, liver, spleen, lymph node and brain smears