A. Trisciuoglio

Characterization of widespread canine leishmaniasis among wild carnivores from Spain

Visceral Leishmaniasis (VL) is an emerging zoonotic parasitic disease caused by Leishmania infantum in Mediterranean countries, with sand flies (Phlebotomus spp.) as vectors and dogs as the main domestic reservoir. The role of wild carnivores in the epidemiology of leishmaniasis is still controversial. In order to determine the prevalence of natural infection with L. infantum in wild carnivores from Spain, we analyzed 217 samples by PCR and western blotting and used restriction fragment length polymorphism (RFLP) to compare the patterns present in wild carnivores with those of domestic dogs from the same areas. DNA of the parasite was detected in spleen or blood samples from 35 (16.12%) analyzed wild carnivores, including 8 of 39 (20.5%) wolves (Canis lupus), 23 of 162 (14.1%) foxes (Vulpes vulpes), 2 of 7 (28.6%) Egyptian mongooses (Herpestes ichneumon), 1 of 4 genets (Geneta geneta), and 1 of 4 Iberian lynxes (Lynx pardinus). No significant sex or age differences in prevalence were observed in wolves and foxes (P>0.05), but there was a significant difference among regions in foxes (P<0.05). A total of 12 PCR-RFLP patterns were found in foxes, 6 in wolves, 4 in dogs, 2 in Egyptian mongooses and 1 in lynx and genet. RFLP patterns differed between dogs and foxes in the two areas where they could be compared. This is the first study of canine leishmaniasis in wild canids and other carnivores from different regions of Spain by PCR. The prevalence of infection indicates the existence of natural infection in apparently healthy wild carnivore populations, and our results are suggestive of a sylvatic cycle independent of dogs.

Asymptomatic Leishmania infantum Infection in an Area of Northwestern Italy (Piedmont Region) Where Such Infections Are Traditionally Nonendemic

ABSTRACT The prevalence of Leishmania infantum-specific antibodies and asymptomatic infection was assessed in a randomized sample of 526 healthy adults from a continental area of Northwestern Italy where L. infantum is not endemic and where autochthonous cases of visceral leishmaniasis (VL) were recently reported. L. infantum-specific antibodies were detected by Western blotting (WB) in 39 subjects (7.41%), while L. infantum kinetoplast DNA was amplified from buffy coat in 21 out of 39 WB-positive subjects, confirming asymptomatic infection in 53.8% of seropositives. Risk factors significantly associated with WB positivity were uninterrupted residence since childhood in a local rural environment (odds ratio [OR], 3.5; 95% confidence interval [CI], 1.7 to 7.3), daily contact with animals though not exclusively with dogs (OR, 3.7; 95% CI, 1.3 to 10.7), older age (OR, 2.31; 95% CI, 1.2 to 4.5), and agricultural/other outdoor activities (OR, 3.8; 95% CI, 0.99 to 3.7.) Logistic regression analysis showed that uninterrupted residence in a local rural environment and an age of >65 years were the only independent predictors of seropositivity assessed by WB. Follow-up at 24 months did not show evidence of VL in either seropositive or PCR-positive subjects. The detection of a high seroprevalence rate, confirmed as asymptomatic infection by PCR in more than half of the cases, among healthy residents in a continental area of northwestern Italy makes local L. infantum transmission very likely. In a region where VL is considered nonendemic, these findings warrant further epidemiological investigations as well as interventions with respect to both the canine reservoir and vectors, given the possible risks for immunosuppressed patients.

Evaluation of 65% permethrin spot-on and deltamethrin-impregnated collars for canine Leishmania infantum infection prevention

During the 2004 and 2005 sand fly seasons, we evaluated the efficacy of a 65% spot‐on solution of permethrin (Exspot, Schering & Plough) and deltamethrin‐impregnated collar (Scalibor, Intervet) in reducing Leishmania infantum infection, in a canine leishmaniasis (CanL) endemic region (Liguria) in Italy. Immunofluorescent assay (IFA) revealed that three of 120 dogs (2.5%) treated with a 65% spot‐on solution of permethrin, as three of 119 dogs (2.5%) treated with deltamethrin‐impregnated collar have shown seroconversion after sand fly season. On the contrary, seroconversion was 15% in 188 untreated control dogs. Treatment reduced the risk of infection by 84%. The difference in treated dogs and control ones is highly significant (χ2 = 12.4; P = 0.0004). Our results show that treatment with 65% spot‐on solution of permethrin and the deltamethrin‐impregnated collar are effective in reducing the risk of acquiring L. infantum infection.

Piroplasmosis in wildlife: Babesia and Theileria affecting free-ranging ungulates and carnivores in the Italian Alps

BackgroundPiroplasmosis are among the most relevant diseases of domestic animals. Babesia is emerging as cause of tick-borne zoonosis worldwide and free-living animals are reservoir hosts of several zoonotic Babesia species. We investigated the epidemiology of Babesia spp. and Theileria spp. in wild ungulates and carnivores from Northern Italy to determine which of these apicomplexan species circulate in wildlife and their prevalence of infection.MethodsPCR amplification of the V4 hyper-variable region of the 18S rDNA of Babesia sp./Theileria sp was carried out on spleen samples of 1036 wild animals: Roe deer Capreolus capreolus (n = 462), Red deer Cervus elaphus (n = 52), Alpine Chamois Rupicapra rupicapra (n = 36), Fallow deer Dama dama (n = 17), Wild boar Sus scrofa (n = 257), Red fox Vulpes vulpes (n = 205) and Wolf Canis lupus (n = 7). Selected positive samples were sequenced to determine the species of amplified Babesia/Theileria DNA.ResultsBabesia/Theileria DNA was found with a mean prevalence of 9.94% (IC95% 8.27-11.91). The only piroplasms found in carnivores was Theileria annae, which was detected in two foxes (0.98%; IC95% 0.27-3.49). Red deer showed the highest prevalence of infection (44.23%; IC95% 31.6-57.66), followed by Alpine chamois (22.22%; IC95% 11.71-38.08), Roe deer (12.55%; IC95% 9.84-15.89), and Wild boar (4.67%; IC95% 2.69-7.98). Genetic analysis identified Babesia capreoli as the most prevalent piroplasmid found in Alpine chamois, Roe deer and Red deer, followed by Babesia bigemina (found in Roe deer, Red deer and Wild boar), and the zoonotic Babesia venatorum (formerly Babesia sp. EU1) isolated from 2 Roe deer. Piroplasmids of the genus Theileria were identified in Wild boar and Red deer.ConclusionsThe present study offers novel insights into the role of wildlife in Babesia/Theileria epidemiology, as well as relevant information on genetic variability of piroplasmids infecting wild ungulates and carnivores.

Seroprevalence of antibodies to Neospora caninum in urban and rural dogs in north-west Italy.

Sera were collected from 490 dogs from north‐west Italy. One hundred and eighty‐eight dogs were urban, while 302 dogs were rural. Among the latter, 190 were shepherd dogs and 112 were cattle farm dogs. Sera were tested for the presence of antibodies against Neospora caninum using the Neospora agglutination test. Seroprevalence at 1/40, 1/80, 1/160 dilution titres was significantly higher in rural (36.4%, 19.5%, 9.9% respectively) than in urban dogs (20.2%, 10.6%, 4.8% respectively). Seroprevalence did not differ significantly in males and females. In shepherd dogs, prevalence increased according to dogs’ age, thus suggesting a post‐natal exposure by horizontal transmission. The observed higher seroprevalence in rural dogs suggests the importance of lifestyle and alimentary habits (i.e. aborted foetuses, placentas and small mammals) in the acquisition of N. caninum infection. Our results confirm that dogs are exposed to N. caninum and play an important role in the epidemiology of N. caninum.

Epidemiology of Leishmania infantum, Toxoplasma gondii, and Neospora caninum in Rattus rattus in absence of domestic reservoir and definitive hosts

Isolated environments are privileged settings to study transmission of infection. Montecristo is a small island where no wild or domestic carnivores are present. Invasive Black rats Rattus rattus (n=78) were captured and tested by PCR for Leishmania infantum, Toxoplasma gondii and Neospora caninum. We wanted to test, for these parasites, the existence of a sylvatic cycle independent of reservoir or definitive hosts. None of the rats tested positive by PCR for either T. gondii or N. caninum. We recorded a 15.5% prevalence (CI95% 8-26%) of L. infantum in the rats and Phlebotomus mascittii was captured in Montecristo, leading us to identify it as possible vector of the parasite.

Encephalitozoon cuniculi, Toxoplasma gondii and Neospora caninum infection in invasive Eastern Cottontail Rabbits Sylvilagus floridanus in Northwestern Italy

Sylvilagus floridanus is a lagomorph introduced for hunting purposes from North America to Europe where, in certain areas like Northern Italy, its population reached high densities. Brain, kidney and skeletal muscle of 144 Eastern Cottontail Rabbits S. floridanus were examined by PCR for Encephalitozoon cuniculi, Toxoplasma gondii and Neospora caninum. DNA of E. cuniculi was found with a prevalence of 9.72% (CI 95% 0.058-0.156). T. gondii and N. caninum DNA was detected in 2.08% (CI 95% 0.0071-0.0595) and 2.78% (CI 95% 0.0109-0.0692) of the samples examined, respectively. This is the first report of E. cuniculi infection in a lagomorph species other than in its natural host Oryctolagus cuniculus, and this is also the first time N. caninum is found to naturally infect S. floridanus. E. cuniculi, T. gondii and N. caninum infect S. floridanus at low but relevant prevalences, considered the important role that these pathogens could play in both animal and human health.

Evaluation of a Rapid Device for Serological Diagnosis of Leishmania infantum Infection in Dogs as an Alternative to Immunofluorescence Assay and Western Blotting

ABSTRACT In this study, we compared a rapid immunochromatographic test (Speed Leish K; BVT Groupe Virbac, La Seyne sur Mer, France) with an indirect immunofluorescence assay (IFAT) and Western blotting (WB) for the detection of Leishmania infantum antibodies in dogs. A total of 250 serum samples were collected from 125 L. infantum-positive and 125 L. infantum-negative dogs. Among the positive samples, 81 were strongly positive at low IFAT dilutions, while 44 were low-reactivity sera (IFAT titers, 1:40 to 1:80). The sensitivity and specificity of the Speed Leish K were 96.3% and 100%, respectively, compared with those of the IFAT. When IFAT low-reactivity sera (titers, 1:40 or 1:80) were tested with the Speed Leish K, using WB results as a reference, the sensitivities were 93.75% for sera with a 1:80 titer and 73.33% for sera with a 1:40 titer, and the specificity was 100%. The Speed Leish K is easy to use and performs well, so it can be considered a quick and reliable tool for the diagnosis of L. infantum infection in dogs.

Toxoplasma gondii in sympatric wild herbivores and carnivores: epidemiology of infection in the Western Alps

BackgroundToxoplasma gondii is an apicomplexan parasite that is able to infect almost all warm blooded animals. In Europe, the domestic cat is the main definitive host. Worldwide, 6 billion people are infected with this parasite. The goal of our research is to evaluate the prevalence of T. gondii infection in wild animals from a previously unsampled area in Northern Italy where 0.1% of women seroconvert during pregnancy each year.MethodsWe sampled and tested skeletal muscle and central nervous system tissue of 355 wild animals by PCR (n = 121 roe deer Capreolus capreolus, n = 105 wild boar Sus scrofa, n = 94 red fox Vulpes vulpes, n = 22 alpine chamois Rupicapra rupicapra, n = 13 red deer Cervus elaphus).ResultsThe overall prevalence of infection with T. gondii was 10.99% (confidence interval (CI) 95% 8.14%-14.67%). A higher rate of infection was recorded in carnivores and omnivores (red fox 20.21%, CI 95% 13.34%-29.43%; wild boar 16.19%, CI 95% 10.36%-24.41%) compared to ruminants (2.48%, CI 95% 0.85%-7.04% in roe deer; 0.00%, CI 95% 0.00%-22.81% in red deer, and 0.00% alpine chamois (CI 95% 0.00%-14.87%) confirming the importance of tissue cysts in transmitting infection.ConclusionsThe relatively high prevalence of T. gondii DNA in highly consumed game species (wild boar and roe deer) gives valuable insights into T. gondii epidemiology and may contribute to improve prevention and control of foodborne toxoplasmosis in humans.

Nosema ceranae has been infecting honey bees Apis mellifera in Italy since at least 1993

(2013). Nosema ceranae has been infecting honey bees Apis mellifera in Italy since at least 1993. Journal of Apicultural Research: Vol. 52, No. 2, pp. 60-61.