The recombinant protein rSP03B is a valid antigen for screening dog exposure to Phlebotomus perniciosus across foci of canine leishmaniasis.

The frequency of sandfly-host contacts can be measured by host antibody levels against sandfly salivary proteins. Recombinant salivary proteins are suggested to represent a valid replacement for salivary gland homogenate (SGH); however, it is necessary to prove that such antigens are recognized by antibodies against various populations of the same species. Phlebotomus perniciosus (Diptera: Psychodidae) is the main vector of Leishmania infantum (Trypanosomatida: Trypanosomatidae) in southwest Europe and is widespread from Portugal to Italy. In this study, sera were sampled from naturally exposed dogs from distant regions, including Campania (southern Italy), Umbria (central Italy) and the metropolitan Lisbon region (Portugal), where P. perniciosus is the unique or principal vector species. Sera were screened for anti-P. perniciosus antibodies using SGH and 43-kDa yellow-related recombinant protein (rSP03B). A robust correlation between antibodies recognizing SGH and rSP03B was detected in all regions, suggesting substantial antigenic cross-reactivity among different P. perniciosus populations. No significant differences in this relationship were detected between regions. Moreover, rSP03B and the native yellow-related protein were shown to share similar antigenic epitopes, as canine immunoglobulin G (IgG) binding to the native protein was inhibited by pre-incubation with the recombinant form. These findings suggest that rSP03B should be regarded as a universal marker of sandfly exposure throughout the geographical distribution of P. perniciosus.

Temporal trends in canine leishmaniosis in the Balearic Islands (Spain): a veterinary questionnaire. Prospective canine leishmaniosis survey and entomological studies conducted on the Island of Minorca, 20 years after first data were obtained.

Leishmaniosis is present in the Mediterranean region of Europe, where Leishmania infantum is responsible for the disease, dogs are the main reservoir, and sand flies of the Phlebotomus genus, subgenus Larroussius, are proven vectors. Some areas, including Minorca in the Balearic Islands, are considered free of the disease, despite the presence of vectors. However, in the context of the current expansion of canine leishmaniosis in parts of Europe, an epidemiological study using a veterinary questionnaire was carried out to establish the current situation of the disease in the Balearic Islands. While 50% of veterinarians thought that the incidence of canine leishmaniosis had not changed over time, 26.2% perceived an increasing trend, mainly those from Minorca, where most of the veterinarians polled (88.1%) considered the new diagnosed cases as autochthonous. A cross-sectional serological study performed in this island gave a seroprevalence rate of 24%. Seroprevalence among animals of local origin and with no history of movements to endemic areas was 31%. The presence of autochthonous canine leishmaniosis in Minorca was not correlated with an increase in vector density. The environmental and climatic factors that influenced the distribution and density of Phlebotomus perniciosus on the island and the possible causes of the apparent emergence of canine leishmaniosis in Minorca are discussed.

Ongoing outbreak of visceral leishmaniasis in Bologna Province, Italy, November 2012 to May 2013.

An increased number of autochthonous visceral leishmaniasis (VL) cases has recently been reported in Bologna Province in northern Italy. Over six months from November 2012 to May 2013, 14 cases occurred, whereas the average number of cases per year was 2.6 (range: 0-8) in 2008 to 2012. VL was diagnosed in a median of 40 days (range: 15-120) from disease onset. This delay in diagnosis shows the need for heightened awareness of clinicians for autochthonous VL in Europe.

The burden of visceral leishmaniasis in Italy from 1982 to 2012: a retrospective analysis of the multi-annual epidemic that occurred from 1989 to 2009.

Starting from 1989 Italy experienced an increase of visceral leishmaniasis (VL) cases over a baseline of 10 to 30 cases reported annually. The number of cases peaked in 2000 and 2004 with more than 200 cases/year, and subsequently declined to reach on average one third of the 2000 peak value in the period after 2010. A retrospective analysis from 1982 to 2012 showed that the multi-annual epidemic consisted of major components including (i) an outbreak involving infants and immunocompetent adults in parts of the Campania region (southern peninsular Italy) and that appears to have declined naturally, (ii) a second outbreak affecting human immunodeficiency virus (HIV)-infected individuals throughout the country, that declined owing to the use of highly active antiretroviral therapies (HAART), (iii) a generalised increase of VL cases in immunocompetent individuals and patients affected by associated conditions other than HIV from endemic regions of peninsular and insular Italy (other than Campania), which was due to a geographical spreading of VL foci, with no major case-clusters or outbreak features. A minor component consisted in the appearance of a few autochthonous cases in formerly non-endemic areas, starting from the early 1990s. Epidemic determinants and reasons for VL decline in the Campania region remain largely unexplained, despite the information available on canine reservoir and phlebotomine vectors in Italy.

Clinical features of cardio-renal syndrome in a cohort of consecutive patients admitted to an internal medicine ward.

INTRODUCTION: Cardiorenal syndrome (CRS) is a disorder of the heart and kidney whereby interactions between the 2 organs can occur. We recorded the clinical features of CRS in patients consecutively admitted to an Internal Medicine ward. PATIENTS AND METHODS: We retrospectively analyzed the anthropometric, history, clinical, biochemical and treatment characteristics in 438 out of 2,998 subjects (14.6%) admitted to our unit (from June 2007 to December 2009), diagnosed with CRS, according to Acute Dialysis Quality Initiative (ADQI) recommendations. Estimated glomerular filtration (eGFR) was calculated using several equations: MDRD (Modification of Diet in Renal Disease; 2 variations GFR(MDRD186), GFR(MDRD175)), Mayo, Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) and Cockroft-Gault. RESULTS: Mean age was 80±8 years, 222 (50.6%) were males, 321 (73.2%) were smokers, 229 (52.2%) were diabetic, 207 (47.2%) had a history of acute myocardial infarction, 167 (38.1%) had angina, 135 (30.8%) were affected by cerebrovascular disease, 339 (77.3%) had peripheral arterial disease. CRS was type 1 in 211 cases (48.2%), type 2 in 96 (21.9%), type 3 in 88 (20.1%), type 4 in 29 (6.6%) and type 5 in 14 (3.2%). eGFR, calculated by different formulae, ranged between 31 and 36 ml/min/1.73 m(2). GFR was lower in CRS type 3 than in the other types, and the values ranged between 24 and 27 ml/min/1.73 m(2). Mean hospital length-of-stay (LOS) was 9.8±6.3 days. Diuretics were the most prescribed medication (78.7%); only 5 patients underwent haemodialysis. CONCLUSIONS: CRS is common, especially in the elderly. CRS Type 1 was the prevalent subset and patients had stage 3-4 renal insufficiency. Results obtained from the GFR equations were similar although the Mayo equation tended to overestimate the eGFR.

Randomized, double-blind comparison of effects of abiciximab bolus only vs. on-label regimen on ex vivo inhibition of platelet aggregation in responders to clopidogrel undergoing coronary stenting.

BACKGROUND: On top of aspirin, an abciximab bolus-only regimen results in a 30% drop in platelet inhibition at 6 h as compared with the on-label regimen. The concomitant administration of high loading dose clopidogrel, by bridging with abciximab bolus, may sustain suppression of platelet activity over time. OBJECTIVES: To investigate the non-inferiority of abciximab bolus-only and concomitant high loading dose clopidogrel vs. abciximab bolus + infusion with respect to the inhibition of platelet aggregation (IPA) as determined by light transmission aggregometry. PATIENTS/METHODS: Seventy-three patients with non-ST segment elevation acute coronary syndromes underwent double-blind randomization to abciximab bolus followed by a 12-h placebo infusion and concomitant 600-mg clopidogrel vs. abciximab bolus + a 12-h infusion and 300 mg of clopidogrel. IPA was determined by light transmission aggregometry throughout 24 h. Clopidogrel poor responsiveness was defined as ≥ 50% 5 µmol L⁻¹ ADP-induced maximum platelet aggregation. RESULTS: In clopidogrel responders (n = 68), IPA after 20 µmol L⁻¹ ADP at 4 h was 89% ± 13% in the bolus-only arm vs. 92% ± 14% in the bolus + infusion arm (P = 0.011 for non-inferiority). IPA after 5 or 20 µmol L⁻¹ ADP and 5 or 15 µmol L⁻¹ TRAP and the proportion of patients showing ≥ 80% IPA did not differ at any time point, irrespective of clopidogrel responsiveness status. Thirty-day outcomes were similar, whereas hemoglobin (0.91 ± 0.8 vs. 0.5 ± 0.7 g dL⁻¹ ; P = 0.01) and platelet count mean drop (41.7 ± 57 vs. 18.6 ± 34 109 L⁻¹; P = 0.042) were significantly reduced in the bolus-only arm. CONCLUSIONS: Withholding abciximab post-bolus infusion in patients receiving high loading dose clopidogrel does not impair platelet inhibition throughout 24 h, and has the potential to improve the safety profile of the drug at reduced costs.

Seasonal phenology, host-blood feeding preferences and natural Leishmania infection of Phlebotomus perniciosus (Diptera, Psychodidae) in a high-endemic focus of canine leishmaniasis in Rome province, Italy.

A 2-year survey aimed to study seasonal phenology, host-blood feeding preferences and Leishmania infections of Phlebotomus perniciosus, was carried out in a high-endemic focus of canine leishmaniasis in Rome province, Italy. Sandfly densities were monitored by sticky traps, while CDC light traps and hand collections were used for individual sandfly analyses. Four species were identified, three belonging to Phlebotomus (P. perniciosus, Phlebotomus mascittii and Phlebotomus papatasi) and one to Sergentomyia (Sergentomyia minuta) genera. In sticky traps, P. perniciosus (53.5%) and S. minuta (46.0%) were the prevalent species, whereas specimens collected by CDC light traps and hand collections consisted mostly of P. perniciosus. The study showed a markedly different sandfly density between 2003 and 2002 collections (5024 specimens versus 644 specimens, respectively), which was attributed to different climatic conditions. In 2003 adults were active during 7 months (May-November) with a typical bimodal distribution, while in 2002 their activity was shorter (5 months, June-October) without clear density peaks. In this low-density year, the prevalence of P. perniciosus was significantly higher in domestic than in wild resting sites. Blood meal tests showed that P. perniciosus had no preferences for any of the two Leishmania susceptible hosts, being the forage ratio rate 0.7 and 1.0 for man and dog, respectively. A significant number of P. perniciosus was found with avian (60.3%) or ovine (24.2%) blood, that was in relation with the habitats where fed specimens were collected. The rate of specimens of P. perniciosus without blood and found positive for Leishmania DNA, detected by a sensitive nested-PCR technique, was much higher (27.6%) than the promastigote infection rate determined by microscopy in the dissected females (1.4%). The intense Leishmania transmission in the study area was confirmed by the high prevalence of anti-leishmanial antibodies in dogs (33.3%).

Failure of a multi-subunit recombinant leishmanial vaccine (MML) to protect dogs from Leishmania infantum infection and to prevent disease progression in infected animals.

We report results of a Phase III trial of the multi-subunit recombinant Leishmania polyprotein MML for the protection of dogs against infection by Leishmania infantum. The antigen, also known as Leish-111f, is the first antileishmanial human vaccine entered Phase I clinical testing. The study was performed in a leishmaniasis endemic area of southern Italy. Three groups of 15 Leishmania-free beagle dogs each, received 3 monthly injections with vaccines A (MML+MPL-SE adjuvant), B (sterile saline = control) and C (MML+Adjuprime adjuvant), respectively, before transmission season 2002. The surviving dogs received a second three-dose vaccine course 1 year later. The dogs were naturally exposed to sandfly bites for 2.5 months in 2002, and for 5 months in 2003. Every 2 months post vaccination, dogs were examined by clinical and immunological evaluation, and by specific serology, microscopy, culture and PCR. A weak lymphoproliferative response to MML was seen in A and C groups throughout the study period. One year after the first vaccine course, the cumulative incidence of leishmanial infections was 40% in group A, 43% in group B and 36% in group C. Two-year post-vaccination (1 year after the second vaccine course) the cumulative incidence was 87% in group A (with three symptomatic cases), 100% in group B (with no symptomatic cases) and 100% in group C (with two symptomatic cases). The efficacy of the MML vaccine as an immunotherapeutic agent for the prevention of disease progression (subpatent infection-->asymptomatic patent infection-->symptomatic patent infection) was evaluated through follow-up of dogs found infected prior to the second vaccination. Among 15 infected animals, progression to a subsequent stage of infection was found in 5/6 dogs of group A, 3/6 of group B and 2/3 of group C. We conclude that vaccination with MML is not effective to prevent leishmaniasis infection and disease progression in dogs under field conditions.

Development of recombinant chimeric antigen expressing immunodominant B epitopes of Leishmania infantum for serodiagnosis of visceral leishmaniasis.

Wild canids and domestic dogs are the main reservoir of zoonotic visceral leishmaniasis (VL) caused by Leishmania infantum (syn.: Leishmania chagasi). Serological diagnosis of VL is therefore important in both human and dog leishmaniasis from a clinical and epidemiological point of view. Routine diagnosis of VL is traditionally carried out by immunofluorescent antibody test (IFAT), which is laborious and difficult to standardize and to interpret. In the last decade, however, several specific antigens of Leishmania infantum have been characterized, allowing the development of a recombinant-based immunoassay. Among them, the whole open reading frame encoding K9 antigen, the gene fragment encoding the repetitive sequence of K26, and the 3'-terminal gene fragment of the kinesin-related protein (K39sub) were previously evaluated as diagnostic markers for canine leishmaniasis and proved to be independent in their antibody reactivity. Since sensitivity of serological test is usually higher in multiple-epitope format, in this study the relevant epitopes of K9, K26, and K39 antigens were joined by PCR strategy to produce the chimeric recombinant protein. The resulting mosaic antigen was found highly expressed in Escherichia coli and efficiently purified by affinity chromatography. Antigenic properties of this recombinant antigen were evaluated by indirect enzyme-linked immunosorbent assay (ELISA) using a panel of human and dog sera previously characterized by parasitological and/or serological techniques. Chimeric ELISA showed 99% specificity in both human (n = 180) and canine (n = 343) control groups, while sensitivity was higher in canine VL (96%, n = 213) than in human VL (82%, n = 185). Accordingly, concordance between IFAT and canine chimeric ELISA (k = 0.95, 95% confidence interval = 0.93 to 0.98) was higher than between IFAT and human chimeric ELISA (k = 0.81, 95% confidence interval = 0.76 to 0.87). Results suggest the potential use of this new antigen for routine serodiagnosis of VL in both human and canine hosts.

[Change in human visceral leishmaniasis treatment in Italy: retrospective study of 630 patients]. / Cambiamento nella terapia della leishmaniosi viscerale umana in Italia: studio retrospettivo su 630 pazienti.

Since the 1940s meglumine antimoniate (MA) has been the only first-line drug for visceral leishmaniasis (VL) treatment in Italy. From 1991 through 1994, several patients of all ages, representing 1/3 of all immunocompetent VL patients reported during that period, were enrolled in clinical trials of liposomal amphotericin B (L-AmB), which led to a novel, safe, short course of VL treatment as an alternative to MA. In the same period, other lipid-associated AmB drugs were registered in Italy for the treatment of fungal infections, i.e., AmB colloidal dispersion (ABCD) and AmB lipid complex (ABLC). A retrospective analysis was performed on data collected at the Unit of Protozoology of Istituto Superiore di Sanità, Rome, to assess whether changes have occurred in first-line drug regimens adopted in Italy for routine VL treatment, during the 1995-2002 period. The sample consisted of immunocompetent individuals clinically suspected for VL, in whom the disease was confirmed by the examination of serum and bone marrow specimens sent to the Unit by hospitals from throughout the country. Relevant information on patients was then recorded, which included drug regimens used and post-therapy results. We recorded treatment information for 630 patients, representing a large proportion (55.5%) of 1,135 immunocompetent individuals with VL reported in Italy from 1995 through 2002. About half were children (306). Every year, patients were referred by 19 to 42 hospitals, with a range of 1 to 30 patients per hospital. MA was the first-line drug used in 159 patients (25.2%). However, the proportion of MA-treated patients has steadily decreased from 55.9% in 1995 to 1.0% in 2002. We recorded the failure of MA therapy in 16 patients (10.1%), who were successfully retreated with a L-AmB regimen. The rate of MA failures significantly increased in recent years, from 5.3% in 1995 to 36.4% in 2000 (p = 0.01). AmB drugs have been the only alternative drugs used in the remaining 471 patients (74.8%). L-AmB accounted for most regimens (441, 93.6%). The proportion of patients treated with any AmB-based drugs increased from 44.1% in 1995 to 99.0% in 2002. Drug treatment was unsuccessful in 15 patients (3.2%), who were successfully retreated with a high-dose L-AmB regimen. This rate was significantly lower than the MA failure rate (p = 0.001). Results have shown a countrywide change in therapy over the period considered. A traditionally effective, but moderately toxic drug (MA) has been almost fully replaced by a new compound (L-AmB) with negligible toxicity, in an epidemiologic context of disease reemergence. Furthermore, short courses of 6 to 7 days, as required for lipid-associated AmB, are highly cost-effective if compared with 21- to 28-day courses needed for standard MA treatment.

[Infantile visceral leishmaniasis in the Campania region, Italy: experience from a Paediatric Referral Centre]. / Leishmaniosi viscerale infantile in Campania: l'esperienza del Centro Pediatrico di Riferimento.

In the first half of the 20th century, visceral leishmaniasis (VL) was a common infantile syndrome in coastal territories of the Campania region of Italy. After World War II, the incidence dropped to a few cases/year for three decades; in late 1980s the disease reemerged among both children and adults. To face the VL recrudescence, a Paediatric Reference Centre was established at the Santobono-Pausilipon hospital in Naples, for the clinical diagnosis, care and drug treatment of all infantile VL cases occurred in the Campania region. Rapid laboratory diagnosis was secured by a Diagnostic Reference Centre established at the Istituto Superiore di Sanità. Here, we report on the epidemiological and parasitological features of all cases referred to the Centre in the past 15 years. From 1990 to March 2004, a total of 255 cases were diagnosed and treated at the Centre. The Figure shows the yearly trend of patients (min. 3 cases in 1990 and 1991, max. 30 cases in 2000). There were 135 males (52.9%); the age ranged 4 months-14 years, but 189 patients (74.1%) were < or = 3 years old. The majority of the patients (189, 74.1%) were from the Naples province, with a cluster of 102 cases (40% of total patients) from the towns and districts surrounding Vesuvius. Twenty-seven cases (10.6%) were from the town of Maddaloni, Caserta province, whereas 15 cases (5.9%) were from coastal villages of the Salerno province. Only 1 and 2 cases were from Benevento and Avellino provinces, respectively. All patients but seven, who have been treated with antimonial drugs in the 1990-1993 period, were successfully treated with a liposomal amphotericin B regimen. From bone-marrow aspirate samples, 138 Leishmania cultures were obtained in EMTM and Sloppy Evans' media, of which 134 have been typed by the electrophoretic analysis of 13 isoenzymes. Two zymodemes (Z) of L. infantum were routinely identified over the study period, ZMON-1 (the commonest zymodeme in the Mediterranean area) and ZMON-72, variant from MON-1 in PGM mobility and detected only in our region. The latter, identified in 61 patients (45.5%), was found exclusively distributed in towns of the Vesuvius area and in Maddaloni until 1996, but in recent years it appears to have spread to other areas of the Naples (including the island of Ischia) and Caserta provinces, but not to Salerno province. In conclusion, the VL macrofocus of the Naples-Caserta area is probably responsible for the highest number of infantile cases among any VL macrofoci described in southern Europe. Considering the limited efforts paid to control the canine reservoir, rapid diagnosis and appropriate treatment of patients still remain the first-line control measures aimed at reducing the health impact of the disease.

Evaluation of the Leishmania recombinant K39 antigen as a diagnostic marker for canine leishmaniasis and validation of a standardized enzyme-linked immunosorbent assay.

Canine infections with Leishmania infantum are important as a cause of serious disease in the dog and as a reservoir for human visceral leishmaniasis (VL). Accurate diagnosis of canine infections is essential to the veterinary community and for VL surveillance programs. A standardized ELISA using a purified recombinant antigen (rK39) specific to VL was compared to the immunofluorescent antibody test (IFAT) as the standard. The ELISA was developed, optimized and evaluated using sera from 6368 dogs. The standardized ELISA and IFAT results were highly concordant. The timing and pattern of ELISA and IFAT seroconversion in dogs followed prospectively after natural infections were very similar. Antibodies reacting with rK39 were more common in asymptomatic canine infections than reported for subclinical human VL. The rK39 ELISA is a relatively simple and rapid assay for assessing the infection status of dogs, and is an alternative to IFAT, especially when screening large numbers of samples.

[Prognostic systems in intensive care: TRISS, SAPS II, APACHE III]. / Scale prognostiche in terapia intensiva: TRISS, SAPS II, APACHE III.

BACKGROUND: To validate the accuracy of SAPS II, APACHE III and TRISS for the prediction of mortality in Intensive Care Unit (ICU) at polytrauma patients admission. The outcome of multiple trauma patients is often linked to the degree of physiologic dysfunction and to the extension of anatomic lesions, the age of the patient and the lesion mechanism. METHODS: The study population consisted of 93 cases of multiple injured patients hospitalised at the ICU of the Padua hospital from October 1998 to October 1999; the term polytraumatized patient is referred to patients who have multiple lesions of which at least one potentially endangers, immediately or in a short term, their life. These cases were evaluated with the APACHE III, SAPS II, Revised Trauma Score and Injury Severity Score. The predictive power of each system was evaluated by using decision matrix analysis to compare observed and predicted outcome with a decision criterion of 0.50 and 0.40 for risk of hospital death. RESULTS: All trauma score systems under study showed high accuracy rates, above all if they are used with a 40% positive test. CONCLUSIONS: The prognostic scales used in this study showed a good correlation between expected and observed cases, particularly with TRISS and APACHE III systems. The APACHE III system seems to be the most reliable of the different methods analysed. These prognostic systems are seldom or occasionally used in the ICU, in Padua and in the whole of Italy, so Italian data are not suitable to be compared to international ones. Due to urgency, the importance of the evaluation scales is often underestimated, but even if they require time and attention, they surely can be useful in the evaluation of the treatment, and not only of a polytraumatized patient.

NO donors inhibit Leishmania infantum cysteine proteinase activity.

Nitric oxide (NO) releasing drugs (e.g., glyceryl trinitrate) were successfully used in the treatment of cutaneous leishmaniasis in man. In the present study, the effect of NO donors on the catalytic activity of the cysteine proteinase from promastigotes of Leishmania infantum, an agent of Old World visceral and cutaneous leishmaniases, is reported. In particular, one equivalent of NO, released by the NO donors S-nitrosoglutathione, glyceryl trinitrate, (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide, 3-morpholinosydnonimine, S-nitrosoacetylpenicillamine and sodium nitroprusside, inhibited one equivalent of the parasite cysteine proteinase. As expected, NO-deprived compounds did not affect the catalytic activity of the parasite cysteine proteinase. Furthermore, the absorption spectrum of the (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide-treated inactive L. infantum enzyme displayed a maximum in the 330-350 nm wavelength range. The reducing agents dithiothreitol and L-ascorbic acid completely prevented parasite cysteine proteinase inhibition by NO, fully restored the catalytic activity, and reversed the NO-induced absorption spectrum of the inactive enzyme. Moreover, S-nitrosoacetylpenicillamine displayed a leishmanicidal effect, inhibiting the cysteine proteinase activity in vivo. As expected, the NO-deprived compound N-acetylpenicillamine did not affect significantly the parasite viability and the enzyme activity in vivo. These data suggest that the L. infantum cysteine proteinase undergoes NO-mediated S-nitrosylation, thereby representing a possible mechanism of antiparasitic host defence.

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.

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.

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.

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.

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.