Cysteine protease as a target for nitric oxide in parasitic organisms.
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Animals↗
Biomedical subjects
L Gradoni
Publications and source records attributed to L Gradoni.
Evidence for an impact on the incidence of canine leishmaniasis by the mass use of deltamethrin-impregnated dog collars in southern Italy.
Dogs are the domestic reservoir of Leishmania infantum Nicolle (Kinetoplastida: Trypanosomatidae), the agent of zoonotic human visceral leishmaniasis. In southern Europe, where canine leishmaniasis (CanL) is widespread due to L. infantum, killing seropositive dogs is considered unacceptable and drug treatment has low efficacy in preventing transmission. We made a field evaluation of the efficacy of deltamethrin dog collars in a CanL focus of southern Italy, Mount Vesuvius area of Campania region, where the vector is Phlebotomus perniciosus Newstead (Diptera: Psychodidae), by assessing their impact on the incidence of CanL in an intervention town, compared to that in dogs of control towns where no collars were fitted. During two consecutive transmission seasons, collars were fitted to 350 (1998) and 354 (1999) dogs from San Sebastiano al Vesuvio (70% of the canine population). Control dogs (371 and 264 in the 2 years, respectively) were from four towns of the same area. Before each transmission season, the CanL seroprevalence in the intervention and control towns was evaluated by cross-sectional surveys and found to be similar (about 15% in 1998 and 10% in 1999, respectively). After each transmission period, incidence rates of seroconversions were determined in adult dogs that were serologically negative before the season under evaluation, and in puppies. After the 1998 season, 2.7% of the dogs in the intervention town seroconverted compared to 5.4% in the control towns (50% protection, P = 0.15). After the 1999 season, 3.5% of collared dogs seroconverted compared to 25.8% of control dogs (86% protection, P < 0.001). The increase in seroconversion rates recorded in control dogs suggests an increase in the Leishmania force of infection in the canine reservoir during the 1999 sandfly season, as supported by the concomitant increase of human cases in control towns and in the whole Campania region. Our results suggest that the impact of mass use of deltamethrin-impregnated dog collars on the incidence of CanL may be negligible during low transmission seasons, or probably in low endemic foci, but can be very strong when the force of transmission is high.
Administration, Topical↗
Modulation of the catalytic activity of cruzipain, the major cysteine proteinase from Trypanosoma cruzi, by temperature and pH.
Cysteine proteinases are relevant to several aspects of the parasite life cycle and of parasite-host relationships. Here, a quantitative investigation of the effect of temperature and pH on the total substrate inhibition of cruzipain, the major papain-like cysteine proteinase from Trypanosoma cruzi, is reported. Values of the apparent catalytic and inhibition parameters Km, Vmax, Vmax/Km, and K(i) for the cruzipain-catalysed hydrolysis of N-alpha-benzyloxycarbonyl-L-phenylalanyl-L-arginine-(7-amino-4-methylcoumarin) (Z-Phe-Arg-AMC) and azocasein were determined between 10.0 degrees C and 40.0 degrees C and between pH 4.5 and 8.5. Values of Km were independent of temperature and pH, whereas values of Vmax, Vmax/Km, and K(i) were temperature-dependent and pH-dependent. Over the whole pH range explored, values of logVmax, log(Vmax/Km), and logK(i) increased linearly with respect to T(-1). Values of Vmax and Vmax/Km were affected by the acid-base equilibrium of one temperature-independent ionizing group (i.e. pK(unl)' = pK(lig)' = 5.7 +/- 0.1, at 25.0 degrees C). Moreover, values of K(i) were affected by the alkaline pK shift of one ionizing group of active cruzipain (from pK(unl)" = 5.7 +/- 0.1 to pK(lig)" = 6.1 +/- 0.1, at 25.0 degrees C) upon Z-Phe-Arg-AMC binding. Values of logK(unl)', logK(lig)', and logK(lig)" were temperature-independent. Conversely, values of logK(unl)" were linearly dependent on T(-1). As a whole, total substrate inhibition of cruzipain decreased with increasing temperature and pH. These data suggest that both synthetic and protein substrates can bind to the unique active centre of cruzipain either productively or following a binding mode which results in enzyme inhibition. However, allosteric effect(s) cannot be excluded.
Animals↗
[A Pediatric case of lupoid leishmaniasis diagnosed by PCR].
The purpose of this paper is to describe the clinical features of lupoid leishmaniasis in a child and to underline the use of PRC as a necessary and reliable tool in controversial diagnosis. Lupoid leishmaniasis, also known as chronic or relapsing leishmaniasis, is mainly widespread in the Middle East, where it represents up to 5% of all cutaneous Leishmaniasis. It strongly resembles Lupus Vulgaris, both clinically and histologically, and is therefore not usually diagnosed immediately but after a certain period of time. The amastigotic forms are rare or absent. The cutaneous nodules or plaques can slowly enlarge over the years. The case of an eleven-year-old albanian child living in Durazzo (Albania), suffering since three years with a plaque formed by apple-jelly nodules and scars on his right cheek, is presented. Using PCR, the presence of Leishmania infantum DNA led to a diagnosis of lupoid leishmaniasis. The therapeutic strategy of a combination of oral itraconazole and infiltration of metilglucamina antimoniate has been carried out, with good result, as checked through "telemedicine".
Administration, Oral↗
[Recent findings on the treatment of leishmaniasis].
The increased interest in leishmaniasis is due to the importance of travel medicine and the demonstration of Leishmania as an opportunist agent in immunosuppressed individuals, besides the persistence of traditional endemic foci. During the past ten years there have been important developments in therapy, which include: a) new dosages/combinations of old drugs, resulted from controlled clinical trials; b) new formulations, which have increased the bioavailability and decreased the toxicity of traditional drugs; c) new molecules, developed for other diseases but found effective against Leishmania. However, while it is now possible to cure severe forms of leishmaniasis with 3-5 infusions of new-generation drugs, the prohibitive costs will preclude the access of developing countries to new therapies.
Humans↗
[Concordance of leishmaniasis cutaneous tests in Tunisia].
A cross sectional study aimed to evaluate the effect of antigenic preparation (Leishmania infantum versus Leishmania major) and dose of leishmania antigens (5 x 10(6) versus 2.5 x 10(6) parasites in the same volume) on the reproducibility of delayed type hypersensitivity leishmania skin test. Results showed that among 34 individuals involved from visceral leishmaniasis endemic area. 26 (76.5%) had a positif Leishmania infantum leishmania (L-L. infantum) test and 27 (79.4%) to Leishmania major leishmania (L-L. major). Mean size of cutaneous reaction was 5.94 +/- 2.86 mm for L-L. infantum and 5.41 +/- 3.23 mm for L-L. major, with a significant positive linear association (p < 10-3). Intra-class correlation coefficient was 0.80 (CI95% = [0.64-0.93]) and concordance Kappa (kappa) was 0.57 (CI95% = [0.40-0.74]). Among 153 individuals from zoonotic cutaneous leishmaniasis. 92.9% revealed a positive test for both types of leishmanin (L-L. major full dose versus L-L. major half dose). Mean size of cutaneous reaction was 12.61 +/- 4.65 mm for the reference test and 11.30 +/- 3.95 mm for diluted one, with a positive linear association (p < 10-3). Intra-class correlation coefficient was 0.78 (IC95% = [0.71-0.84]) and concordance Kappa (kappa) was 0.82 (IC95% = [0.73-0.91]). These results demonstrate a limited effect of leishmania antigenic variation and antigen dose on the reproducibility of delayed type hypersensitivity induced by the leishmanin test.
Adolescent↗
Nitric oxide inhibits cruzipain, the major papain-like cysteine proteinase from Trypanosoma cruzi.
Nitric oxide (NO) is a pluripotent regulatory molecule showing, among others, an antiparasitic activity. Moreover, NO inhibits cysteine proteinase action by nitrosylating the Cys catalytic residue. In the present study, the inhibitory effect of the substrate N-alpha-benzyloxycarbonyl-L-phenylalanyl-L-arginine-(7-amino-4-methyl coumarin) and of NO on the catalytic activity of cruzipain, the major papain-like cysteine proteinase from Trypanosoma cruzi (the hemoflagellate protozoan parasite which causes the American trypanosomiasis), is reported. In particular, NO-donors S-nitroso-glutathione (GSNO), (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR-3), 3-morpholinosydnonimine (SIN-1), S-nitroso-acetyl-penicillamine (SNAP), and sodium nitroprusside (SNP) dose-dependently inhibited cruzipain, this effect being likely attributable to the S-nitrosylation of the Cys25 catalytic residue. These results were analyzed in parallel with those concerning the inhibitory effect of the substrate and of NO on the catalytic activity of falcipain, the cruzipain-homologous cysteine proteinase from Plasmodium falciparum. The modulation of the cruzipain and falcipain activity by NO may be relevant in developing new strategies against T. cruzi and P. falciparum in human host. As a whole, the NO-mediated S-nitrosylation of pathogenic viral, bacterial, fungal, and parasitic cysteine proteinases may represent a general mechanism of antimicrobial and antiparasitic host defences.
Animals↗
Nitric oxide inhibits falcipain, the Plasmodium falciparum trophozoite cysteine protease.
Nitric oxide (NO) is a pluripotent regulatory molecule possessing, among others, an antiparasitic activity. In the present study, the inhibitory effect of NO on the catalytic activity of falcipain, the papain-like cysteine protease involved in Plasmodium falciparum trophozoite hemoglobin degradation, is reported. In particular, NO donors S-nitrosoglutathione (GSNO), (+/-)-(E)-p6ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenami de (NOR-3), 3-morpholinosydnonimine (SIN-1), and sodium nitroprusside (SNP) inhibit dose-dependently the falcipain activity present in the P. falciparum trophozoite extract, this effect likely attributable to S-nitrosylation of the Cys25 catalytic residue. The results represent a new insight into the modulation mechanism of falcipain activity, thereby being relevant in developing new strategies for inhibition of the P. falciparum life cycle.
Animals↗
Efficacy of amphotericin B colloidal dispersion in the treatment of Mediterranean visceral leishmaniasis in immunocompetent adult patients.
Twelve immunocompetent adults with Mediterranean visceral leishmaniasis (VL) were treated with amphotericin B colloidal dispersion (ABCD; 2 mg/kg/d for 7 d). All patients showed rapid clinical response without significant adverse events. Two weeks after therapy they were parasitologically cured and no relapses occurred during 6 months. ABCD is a valid alternative in the management of Mediterranean VL in adult patients.
Adult↗
Early suppression of lymphoproliferative response in dogs with natural infection by Leishmania infantum.
Dogs are the domestic reservoirs of zoonotic visceral leishmaniasis caused by Leishmania infantum. Early detection of canine infections evolving to clinically patent disease may be important to leishmaniasis control. In this study we firstly investigated the peripheral blood mononuclear cell (PBMC) response to leishmanial antigens and to polyclonal activators concanavalin A, phytohemagglutinin and pokeweed mitogen, of mixed-breed dogs with natural L. infantum infection, either in presymptomatic or in patent disease condition, compared to healthy animals. Leishmania antigens did not induce a clear proliferative response in any of the animals examined. Furthermore, mitogen-induced lymphocyte proliferation was found strongly reduced not only in symptomatic, but also in presymptomatic dogs suggesting that the cell-mediated immunity is suppressed in progressive canine leishmaniasis. To test this finding, naive Beagle dogs were exposed to natural L. infantum infection in a highly endemic area of southern Italy. Two to 10 months after exposure all dogs were found to be infected by Leishmania, and on month 2 of exposure they all showed a significant reduction in PBMC activation by mitogens. Our results indicate that suppression of the lymphoproliferative response is a common occurrence in dogs already at the beginning of an established leishmanial infection.
Animals↗
The production of the osmiophilic body protein Pfg377 is associated with stage of maturation and sex in Plasmodium falciparum gametocytes.
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Animals↗
Diffuse necrotic hepatic lesions due to visceral leishmaniasis in AIDS.
A rare case of an AIDS patient who developed scattered necrotic involvement of the liver caused by Leishmania infantum is described. Of interest, marked splenomegaly, hypergammaglobulinemia and serum anti-Leishmania antibodies were absent and an incomplete response to therapy was observed. Diagnosis of visceral leishmaniasis (VL) was achieved by the demonstration of numerous amastigotes in both hepatocytes and macrophages on liver biopsy. Hepatic necrotic lesions, which when extensive could lead to acute hepatic failure, possibly reflect an atypical manifestation of liver involvement caused by L. infantum and depend on the immunological impairment which characterizes AIDS patients, thus preventing the formation of granulomas. Our observation confirms that VL can manifest atypical aspects in HIV-positive patients depending on the degree of the immunodeficiency. The frequency and severity of this pathology accounts for the need to list VL among AIDS-defining conditions.
AIDS-Related Opportunistic Infections↗
Comparative efficacy of meglumine antimoniate and aminosidine sulphate, alone or in combination, in canine leishmaniasis.
Thirty-two domestic dogs naturally infected with Leishmania infantum and showing viscero-cutaneous signs of canine leishmaniasis were treated with aminosidine sulphate (11 dogs) meglumine antimoniate (10 dogs) or with a combination of the two drugs (11 dogs) for 21 consecutive days. Clinical and laboratory assessments, made on day 21 and at 2, 4 and 6 months after initiation of treatment, showed that the drug combination gave the best score in terms of clinical efficacy, incidences of early clinical relapse, any clinical relapse or apparent parasitological cure, and reduction in parasite densities in bone-marrow and lymphnode aspirates (even though a lower dose of antimonial was used in the combination than for antimonial monotherapy). For each of the above parameters, however, the higher efficacy of the drug combination was not statistically significant, probably because of the large variations caused by using naturally infected animals of various ages and breeds.
Animals↗
A cloned antigen (recombinant K39) of Leishmania chagasi diagnostic for visceral leishmaniasis in human immunodeficiency virus type 1 patients and a prognostic indicator for monitoring patients undergoing drug therapy.
Serologic assays using crude antigens for the diagnosis of visceral leishmaniasis in human immunodeficiency virus type 1 (HIV)-seropositive patients have been shown to lack sensitivity and specificity, particularly in AIDS patients. Antibodies to a cloned antigen, recombinant (r) K39, of Leishmania chagasi are specific for members of the Leishmania donovani complex and have been shown to indicate active disease in immunocompetent persons. This study demonstrated that antibodies to rK39 were also detectable in HIV-seropositive patients coinfected with Leishmania infantum. Furthermore, the rK39 ELISA was more sensitive than an IFA for detecting L. infantum infections in patients with AIDS. In addition, antibody titers to rK39 in HIV-negative patients infected with L. infantum or L. chagasi declined during treatment with meglumine antimoniate or liposomal amphotericin B. In contrast, most patients who clinically relapsed showed increased antibody titers to rK39. These data demonstrate the diagnostic and prognostic utility of rK39 in detecting active visceral leishmaniasis.
AIDS-Related Opportunistic Infections↗
A retrospective clinical study of canine leishmaniasis in 150 dogs naturally infected by Leishmania infantum.
The clinical and laboratory findings observed in 150 dogs naturally infected by Leishmania infantum, from a large endemic area of southern Italy, are described. There was a gradual onset of clinical signs and the course of the disease was progressive in almost all the cases. The majority of the dogs were mongrels (43.3 per cent), male (64.7 per cent), of medium size (50.6 per cent), three to seven years old (64.7 per cent), and living outdoors (60 per cent). They showed generalised (56.7 per cent) or symmetrical (32 per cent) lymphadenomegaly; the mucous membranes of 87 of the dogs (58 per cent) were pale and moderate or severe splenomegaly was diagnosed in 80 dogs (53.3 per cent); weight loss was observed in 32 per cent of the animals. Skin abnormalities were very common, and included dry exfoliative dermatitis (56 per cent), ulcers (40 per cent) periorbital alopecia ('lunettes') (18 per cent), diffuse alopecia (14 per cent) and onychogryphosis (24 per cent). Ocular signs were observed in 24 dogs (16 per cent) including 16 cases of keratoconjunctivitis (three with keratoconjunctivitis sicca), six cases of moderate uveitis and two cases of panophthalmitis. The acute form of the disease was diagnosed in only six dogs and was characterised by fever and generalised lymphadenomegaly, and by the absence of skin lesions. Another six dogs had severe renal failure without systemic clinical signs of leishmaniasis. The most important laboratory findings were a severe or moderate increase in gammaglobulins, hypoalbuminaemia, hyperproteinemia and anaemia. Cultures or cytology tests for L infantum parasites were positive in 134 of the dogs. Following the standard procedures developed for human lymph node and bone marrow cytology tests, the leishmania density in the dogs varied from 1+ to 2+. Leishmania antibody titres were high (> 1:160) in almost all the dogs. Immunological tests for autoantibodies were positive in 25 of 53 dogs tested in the antinuclear antibody (ANA) test, in 15 of 43 dogs tested in the latex test and in five of 24 dogs tested in the Coombs test.
Animals↗
Treatment of visceral leishmaniasis in children with liposomal amphotericin B.
We used liposomal amphotericin B as first-choice treatment of visceral leishmaniasis in 106 immunocompetent children who acquired the infection in a temperate region of southern Europe (Italy) where Leishmania infantum visceral leishmaniasis is endemic. The aim of the study was to identify the minimum total dose of liposomal amphotericin B needed to cure the infection in children and reduce the period of hospitalization. We conclude that the optimal regimen in immunocompetent children with L. infantum visceral leishmaniasis to be a total dose of 18 mg/kg of liposomal amphotericin B (3 mg/kg per day for 5 days, followed by 3 mg/kg administered as an outpatient regimen on day 10).
Adolescent↗
Epidemiologic surveillance of visceral leishmaniasis in Sicily, Italy.
Visceral leishmaniasis (VL) is endemic in Sicily. Although it is a notifiable disease, there is evidence that the actual number of cases is higher than that reported. In 1987, a regional reference center for active surveillance of VL was established and it recorded a total of 284 cases through 1995, a mean of 31.5 cases/year and about four-fold more than previously reported. Of the 284 cases, 150 (53%) were children (< or = 14 years of age), and of the 134 adults, 39 (29%) were coinfected with human immunodeficiency virus (HIV). The commonest viscerotropic zymodeme of Leishmania infantum, MON 1, was identified in 40 (93%) of 43 HIV-negative and eight (57%) of 14 HIV-positive patients. Among 280 patients evaluated (i.e., all HIV-negative and 35 of 39 HIV-positive subjects), 254 (91%) were treated with meglumine antimoniate alone or in combination with other drugs; 23 (8%) received allopurinol or amphotericin B, either conventional or in liposomal form; and three terminally ill patients were not treated. Among the 245 HIV-negative patients, 236 (96%) were successfully cured, while nine (4%) (seven adults) died during the course of antimonial treatment. None of the 35 HIV-positive patients was definitively cured, although mortality was apparently associated with other opportunistic infections.
Adolescent↗
Visceral leishmaniasis in HIV infected patients: treatment with high dose liposomal amphotericin B (AmBisome).
Visceral leishmaniasis (VL) in patients coinfected with human immunodeficiency virus (HIV) is often atypical, and characteristically relapses after treatment. We treated 10 HIV infected patients (9 men) with parasitologically confirmed VL with liposomal amphotericin B ("AmBisome': L-AMB) at a dose of 4 mg/kg/day on days 1, 2, 3, 4, 5, 10, 17, 24, 31, and 38. Patients were hospitalized for the first 5 days, and were monitored during, and 1 week and 1, 3 and 6 months after, L-AMB therapy. There were no serious adverse events, and L-AMB was well tolerated. 9/10 patients completed therapy, one patient defaulted at day 24. Clinical improvement was seen in all nine patients and the bone marrow aspirate was cleared of visible/culturable parasites in 8/9 patients. During follow-up, one patient defaulted. The seven remaining patients relapsed at 2, 3, 3, 5, 5, 6 and 7 months. Re-treatment with a variety of antileishmanial drugs was unsatisfactory. The time from first diagnosis of VL to death in six patients was 5-40 months (mean 18.8 months). Only one patient remained alive 26 months after treatment. L-AMB is safe and provides a good initial clinical response. Intermittent dosing enables a short period of hospitalization. However, relapse is probably inevitable.
AIDS-Related Opportunistic Infections↗