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Abstract Background Dogs are considered the main domestic animals that may be a reservoir for Leishmania infantum, the agent of zoonotic visceral leishmaniasis (ZVL) in several countries of the world. The dog may host other Leishmania species, but its epidemiological role in the maintenance and spreading of these parasites is not completely elucidated. Zoonotic cutaneous leishmaniasis (ZCL), caused by Leishmania major, affects thousands of people every year and is particularly diffused in many countries of North Africa and Middle East Asia. In ZCL endemic countries, few reports of L. major-positive dogs have been reported, probably because most human cases occur in poor rural areas where the social role of the dog and its medical management is not well considered. The aim of the present study is to better understand the possible involvement of domestic dogs in the epidemiology of ZCL. Methods Our research focused on a well-established endemic focus of ZCL, in the area of Echrarda, Kairouan Governorate, central Tunisia. A total of 51 dogs with no or mild clinical signs of vector borne diseases were selected in small villages where human cases of ZCL are yearly present. All dogs were sampled for the Leishmania spp. diagnosis, by using the following procedures: blood sample for serology and buffy coat quantitative polymerase chain reaction (qPCR), popliteal fine needle aspiration, and cutaneous biopsy punch for lymph node and skin qPCR. Results The results demonstrated a high percentage (21.6%) of dogs positive at least at one or more test; the most sensitive technique was the lymph node qPCR that detected 8/11 positive dogs. Nine, out of the eleven positive dogs, resulted as infected by Leishmania infantum; ITS1-PCR-sequencing allowed Leishmania major identification in the remaining two cases, both from the popliteal lymph node samples, which can suggest a possible visceral spread of a cutaneous Leishmania species in the dog. Interestingly, one of the two L. major-positive dogs was living in the same house where 6-year-old children showed cutaneous lesions referred to as ZCL. Conclusions To our knowledge, this is the first report of L. major-positive dogs in Tunisia, the epidemiological role of which remains under investigation. Graphical Abstract

In the field of veterinary medicine, the detection of parasite eggs in the fecal samples of livestock animals represents one of the most challenging tasks, since their spread and diffusion may lead to severe clinical disease. Nowadays, the scanning procedure is typically performed by physicians with professional microscopes and requires a significant amount of time, domain knowledge, and resources. The Kubic FLOTAC Microscope (KFM) is a compact, low-cost, portable digital microscope that can autonomously analyze fecal specimens for parasites and hosts in both field and laboratory settings. It has been shown to acquire images that are comparable to those obtained with traditional optical microscopes, and it can complete the scanning and imaging process in just a few minutes, freeing up the operator's time for other tasks. To promote research in this area, the first AI-KFM challenge was organized, which focused on the detection of gastrointestinal nematodes (GINs) in cattle using RGB images. The challenge aimed to provide a standardized experimental protocol with a large number of samples collected in a well-known environment and a set of scores for the approaches submitted by the competitors. This paper describes the process of generating and structuring the challenge dataset and the approaches submitted by the competitors, as well as the lessons learned throughout this journey.

Abstract Background Cystic echinococcosis (CE), caused by the larval stage of Echinococcus granulosus sensu lato, is a zoonotic parasitic disease of economic and public health importance worldwide, especially in the Mediterranean area. Canids are the main definitive hosts of the adult cestode contaminating the environment with parasite eggs released with feces. In rural and peri-urban areas, the risk of transmission to livestock as well as humans is high because of the free-roaming behavior of owned/not owned dogs. Collecting data on animal movements and behavior using GPS dataloggers could be a milestone to contain the spread of this parasitosis. Thus, this study aims to develop a comprehensive control strategy, focused on deworming a dog population in a pilot area of southern Italy (Campania region) highly endemic for CE. Methods Accordingly, five sheep farms, tested to be positive for CE, were selected. In each sheep farm, all shepherd dogs present were treated every 2 months with praziquantel. Furthermore, 15 GPS dataloggers were applied to sheep and dogs, and their movements were tracked for 1 month; the distances that they traveled and their respective home ranges were determined using minimum convex polygon (MCP) analysis with a convex hull geometry as output. Results The results showed that the mean daily walking distances traveled by sheep and dogs did not significantly differ. Over 90% of the point locations collected by GPS fell within 1500 mt of the farm, and the longest distances were traveled between 10:00 and 17:00. In all the sheep farms monitored, the area traversed by the animals during their daily activities showed an extension of < 250 hectares. Based on the home range of the animals, the area with the highest risk of access from canids (minimum safe convex polygon) was estimated around the centroid of each farm, and a potential scheme for the delivery of praziquantel-laced baits for the treatment of not owned dogs gravitating around the grazing area was designed. Conclusions This study documents the usefulness of geospatial technology in supporting parasite control strategies to reduce disease transmission. Graphical Abstract

Abstract Background The zoonotic hookworms Ancylostoma caninum and Uncinaria stenocephala are widespread soil-transmitted helminths in dogs in Europe. Given the veterinary and public health importance of hookworms in dogs and the recent changes in the molecular epidemiology of some species, there is a need to continuously monitor the epidemiological and molecular prevalence of these parasites also at the “local” level. The present study aimed to update the epidemiological scenario of hookworm infections in both owned and stray dogs in southern Italy and to discriminate between different hookworm species (A. caninum and U. stenocephala) through molecular analyses. For this purpose, a retrospective analysis was performed over 10 years (2011–2021), including a total of 7008 owned dogs and 5642 stray dogs referred to our laboratory for copromicroscopic examinations. Moreover, 72 faecal samples, from dogs naturally infected by hookworms, were used to discriminate between A. caninum and U. stenocephala using two PCR protocols. Prior to molecular analyses, a subsample of 40/72 positive faecal samples was used for morphometric investigations on hookworm eggs. Results The results of the ten-year retrospective analysis (2011–2021) showed an overall prevalence of hookworm infection of 9.16%, specifically 5.1% in owned dogs and 14.2% in stray dogs. Logistic regression showed a significant association between positivity to hookworms and the variable “puppies” both in stray (13.84%; OR = 2.4) and owned (7.07%; OR = 2.2) dogs. The results of molecular analyses showed that positivity was confirmed only in 21/72 samples, specifically, 6 samples using protocol A and 19 with protocol B. Sequencing revealed 15 samples positive to U. stenocephala and 6 to A. caninum. Conclusions The findings of this study showed a high prevalence of hookworm infections in dogs in southern Italy, updating the epidemiological scenario of the last decade. Moreover, the results of the study revealed the first identification of hookworm species in dogs in Italy by molecular studies, highlighting that U. stenocephala is more prevalent than A. caninum.

Abstract Background Fasciolosis (Fasciola hepatica) and paramphistomosis (Calicophoron daubneyi) are two important infections of livestock. Calicophoron daubneyi is the predominant Paramphistomidae species in Europe, and its prevalence has increased in the last 10–15 years. In Italy, evidence suggests that the prevalence of F. hepatica in ruminants is low in the southern part, but C. daubneyi has been recently reported at high prevalence in the same area. Given the importance of reliable tools for liver and rumen fluke diagnosis in ruminants, this study evaluated the diagnostic performance of the Mini-FLOTAC (MF), Flukefinder® (FF) and sedimentation (SED) techniques to detect and quantify F. hepatica and C. daubneyi eggs using spiked and naturally infected cattle faecal samples. Methods Briefly, negative bovine faecal samples were artificially spiked with either F. hepatica or C. daubneyi eggs to achieve different egg count levels: 10, 50 and 100 eggs per gram (EPG) of faeces. Moreover, ten naturally infected cattle farms from southern Italy with either F. hepatica and/or C. daubneyi were selected. For each farm, the samples were analysed individually only with MF technique and as pools using MF, FF and SED techniques. Bayesian latent class analysis (LCA) was used to estimate sensitivity and accuracy of the predicted intensity of infection as well as the infection rate in the naturally infected farms. Results The outcome of this study showed that the highest number of eggs (F. hepatica and C. daubneyi) recovered was obtained with MF, followed by FF and SED in spiked infected samples at 50 and 100 EPG, while at lower infection levels of 10 EPG, FF gave the best results. Moreover, the sensitivity for all the techniques included in the study was estimated at > 90% at infection levels > 20 EPG for both F. hepatica and C. daubneyi eggs. However, MF was the most accurate of the three techniques evaluated to estimate fluke infection intensity. Nevertheless, all three techniques can potentially estimate infection rate at farm level accurately. Conclusions Optimization and standardization of techniques are needed to improve the FEC of fluke eggs. Graphical Abstract

Toxoplasmosis, caused by the protozoan Toxoplasma gondii, is one of the main food-, water- and soil-borne zoonotic disease worldwide. Over the past 20 years many papers were published on the transmission of T. gondii by marine animals, including mollusks, which can concentrate the oocysts and release them. Sporulated oocysts may remain viable and infective for 18 months in seawater. Therefore, raw or undercooked bivalve mollusks pose a risk to humans. This study aimed to apply and validate for the first time a very sensitive digital droplet polymerase chain reaction (ddPCR) protocol to detect and quantify T. gondii DNA in mussels. Four concentration levels: 8000 genomic copies (gc)/μL, 800 gc/μL, 80 gc/μL, 8 gc/μL of a T. gondii reference DNA were tested. DNA was extracted from 80 pools of mussels (Mytilus galloprovincialis). Forty pools were contaminated with T. gondii reference DNA and used as positive controls, while 40 pools were used as negative controls. DdPCR reaction was prepared using a protocol, previously developed by the authors, for detection of T. gondii in meat. Amplification was obtained up 8 gc/μL. All infected replicates resulted positive, as well as no droplets were detected in negative controls. The droplets produced in the reaction ranged from 8,828 to 14,075 (average 12,627 droplets). The sensitivity and specificity of ddPCR were 100% (95%CI = 94.3–99.9). In addition, 100 pools of mussels collected in the Gulf of Naples were used to validate the protocol. Of these 16% were positive (95% CI = 9.7–25.0) for T. gondii. Samples were also tested by real-time PCR and no positive samples were found. Data obtained from ddPCR showed good identification of negative and positive samples with higher specificity and efficiency than real-time PCR. This tool could be very useful for a rapid sensitive detection of low DNA concentrations of T. gondii in mussels, reducing the risk of toxoplasmosis in humans.

Nowadays, the exclusive use of commercial anthelmintics for the treatment of gastrointestinal nematode infections in ruminants is less sustainable due to anthelmintic resistance, as well as the problem of drug residues in animal products and the environment. Therefore, an integrated therapeutic approach is needed, including the search for alternatives to synthetic anthelmintic drugs. The aim of this study was to evaluate the possibility of using the essential oil of peppermint (Mentha x piperita L.) in the control of gastrointestinal nematodes in sheep. For this purpose, the in vitro and in vivo anthelmintic efficacy of this oil and the toxic effects on the hosts were examined. In the in vitro egg hatch test, ovicidal activity varied from 21.0–90.3% depending on the concentration of essential oil used (0.0125, 0.025, 0.049, 0.195, 0.781, 3.125, 12.5, and 50 mg/mL). To some extent, anthelmintic efficacy was confirmed in the in vivo fecal egg count reduction test at a mean dose of 150 mg/kg, with an average reduction of nematode eggs of 26.9 and 46.0% at Days 7 and 14 after treatment, respectively. Furthermore, no toxic effects of applied oil were observed on sheep behavior, kidney, or liver function. The main compounds identified by gas chromatography–mass spectrometry analyzes were menthol (32.6%), menthone (22.0%), menthyl-acetate (10.0%), and isomenthone (9.39%). Due to their complex chemical compositions, numerous bioactive ingredients, and natural origin, herbal formulations represent a potentially valuable alternative for the control of gastrointestinal nematodes in sheep. In this context, the results of the present study showed that peppermint essential oil is one of the promising candidates. Further studies should be performed to collect more data on the safety profile of M. piperita EO in treated animals to find the most appropriate formulation for use in field conditions and to test it against resistant gastrointestinal nematode populations.

In southern Italy, the number of autochthonous cases of Dirofilaria immitis in dogs has increased considerably. This also occurs in the Campania region, particularly in coastal areas, where infections with D. immitis and Dirofilaria repens have been reported more frequently. Therefore the aim of the present study was to better investigate the occurrence of Dirofilaria spp. in a local dog shelter in Castel Volturno (Campania region, southern Italy). Briefly, a total of 260 blood samples were analysed for identification of microfilariae (mff) and detection of Dirofilaria immitis antigen. Dogs were classified according to their age (1–3 years; 4–6 years; 7–11 years; > 11 years) and length of stay in the shelter at the time of sampling (dogs that entered in the shelter in the last 4 months; dogs housed in the shelter for more than 4 months up to 2 years; dogs housed for more than 2 years). The modified Knott’s test revealed that 195 dogs (75.0%) were positive for circulating mff of Dirofilaria spp. Specifically, 104/260 (40.0%) dogs were positive for D. immitis and 91/260 (35.0%) were positive for D. repens. In addition, 72/260 (27.7%) dogs had both D. immitis and D. repens mff. Antigen testing revealed that 78/260 (30.0%) dogs were positive for D. immitis. However, 26/104 (25.0%) of the dogs with D. immitis mff were antigen-negative. The overall k concordance between the modified Knott’s test and the antigenic test was ≤0.2 (poor) (p = 0.000). The results of the logistic regression model showed a significant association between Dirofilaria exposure and the period of time the dogs had spent in the shelter at the time of sampling. Dogs housed in the shelter for 4 months (group 1) and between 4 months and 2 years (group 2) had higher Dirofilaria positivity than dogs in group 3 (housed for more than 2 years) (80.4% vs. 79.6% vs. 62.4%, respectively). Moreover, male dogs and older dogs (between 7 and 11 years of age) were more likely to be infected with Dirofilaria spp.

Abstract Background Cystic echinococcosis (CE) is a worldwide parasitic zoonosis caused by the larval stage of Echinococcus granulosus sensu lato affecting livestock, particularly sheep and goats. However, often this parasitosis is underestimated. For this reason, this study aimed to evaluate the epidemiological features and spatial distribution of CE in sheep and goats slaughtered in a hyperendemic Mediterranean area. Methods A survey was conducted in the Basilicata region (southern Italy) from 2014 to 2019. A total of 1454 animals (1265 sheep and 189 goats) from 824 farms were examined for hydatid cyst detection by visual inspection, palpation and incision of target organs. All the CE cysts were counted and classified into five morphostructural types (unilocular, multiseptate, calcified, caseous and hyperlaminated). Molecular analysis was performed on 353 cysts. For spatial analysis, a kriging interpolation method was used to create risk maps, while clustering was assessed by Moran’s I test. Results CE prevalence of 72.2% (595/824) and 58.4% (849/1454) was observed at the farm and animal levels, respectively, with higher values in sheep (62.9%) than goats (28.0%). The liver and lungs were the most frequently infected organs in both sheep and goats. Most of recovered cysts were of the calcified and multiseptate morphotypes. All the isolates were identified as E. granulosus sensu stricto (genotypes G1–G3). Spatial distribution showed a moderate clustering of positive animals. Conclusion The findings of this study can be used to better understand the eco-epidemiology of echinococcosis and to improve CE surveillance and prevention programs in regions highly endemic for CE. Graphical abstract

In order to characterize the inflammatory phenotype of livers of sheep naturally infected by cystic echinococcosis, 100 sheep livers have been macroscopically assessed for the presence of hydatid cysts and sampled for histopathological and molecular analysis. According to gross and microscopic examination, livers were subsequently classified into three groups: normal liver (Group A), liver with the presence of fertile hydatid cysts (Group B), and liver with the presence of sterile hydatid cysts (Group C). Immunohistochemical analyses were accomplished using primary antibodies anti-Iba1, anti-CD3, anti-CD20, anti-TGF-β, and anti-MMP9. Finally, real-time PCR was performed in order to estimate the concentration levels of tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ), interleukin (IL)-12, IL-10, and TGF-β. Immunohistochemical analysis showed a diffuse immunolabelling of mononuclear cells for Iba-1 and TGF-β and a higher amount of CD20+ B cells compared to CD3+ T cells in both Groups B and C. The expression levels of Th-1-like immune cytokines TNF-α, INF-γ, and IL-12 did not show significant statistical differences. However, we found a significant increase in expression levels of Th-2 immune cytokines TGF-β and IL-10 in Groups B and C compared to Group A. Taken together, our findings suggest that macrophages have a predominant role in the local immune response to cystic echinococcosis. Moreover, we can speculate that Th2 immunity may be dominant, corroborating the idea that B cells are decisively essential in the control of the immune response during parasitic infection and that the immunomodulatory role of IL-10 and TGF-β may ensure the persistence of the parasite within the host.

Abstract Background The worldwide increased difficulty to combat gastrointestinal nematode (GIN) infection in sheep, due to progressing anthelmintic resistance (AR), calls for an enhanced and standardized implementation of early detection of AR. This study provides a snapshot of the current AR status against benzimidazoles and macrocyclic lactones in southern Italy, generated with standardized techniques. Methods On 10 sheep farms, the efficacy of albendazole (ALB) and either eprinomectin (EPR) or ivermectin (IVM) was evaluated based on the faecal egg count reduction test (FECRT) performed with the Mini-FLOTAC. For each tested drug, 40 sheep were rectally sampled at D0 and sampled again 14 days after the treatment (D14). The FECRT was calculated from individual samples and pooled samples which consist of 5 individual samples. Efficacy was classified as ‘reduced, ‘suspected’ and ‘normal’. Coprocultures were set for D0 and D14 faecal samples of each group. From farms with FECR < 95%, an in vitro egg hatch test (EHT) and a follow-up FECRT using fenbendazole (FBZ) were conducted. Results Based on the FECR, high efficacy (from 95.7% to 100%) was observed for ALB and IVM in eight farms (Farms 3–10). On Farm 1 and Farm 2, the efficacy for the macrocyclic lactones was classified as ‘normal’, but ‘reduced’ efficacy was observed for ALB on Farm 1 (FECR = 75%) and ‘suspected’ efficacy on Farm 2 (FECR = 93.3%) with the predominant GIN genus Trichostrongylus followed by Haemonchus at D14. The FEC results of pooled samples strongly correlated with those of individual samples, for FEC at D0 (r s = 0.984; P < 0.0001) and at D14 (rs = 0.913; P < 0.0001). The classifications of efficacy in Farm 1 (FECR = 86.0%) and Farm 2 (FECR = 93.0%) in the follow-up FECRT with FBZ coincide with the main FECRT trial. The in vitro EHT confirmed AR in both farms (Farm 1: 89%; Farm 2: 74%). Conclusions In regions like southern Italy, where the negative impacts from AR have played a minor role, efficient monitoring of AR is important in order to evaluate potential risks and being able to promptly respond with countermeasures.

Abstract Background Faecal egg count (FEC) techniques are commonly used to detect gastrointestinal nematodes (GINs) in cattle and to determine anthelmintic efficacy/resistance through the faecal egg count reduction test (FECRT). Mini-FLOTAC is one of the techniques recommended for a standardised FEC/FECRT of helminth eggs in cattle. However, only one paper evaluated the recovery rate of GIN eggs by Mini-FLOTAC (compared to McMaster and modified-Wisconsin method) in cattle, using only a level of contamination of 200 eggs per gram (EPG) of faeces and using GIN eggs collected from goat faeces to spike faecal samples from cattle. To further study the recovery rate of GIN eggs from cattle faeces, this study was conducted in two laboratories, one in Belgium and one in Italy to evaluate the sensitivity, accuracy, precision and reproducibility of the Mini-FLOTAC and McMaster techniques (at two reading levels: grids and chambers) for the detection of GIN eggs in spiked bovine faecal samples. Methods In both countries, spiked cattle faecal samples with five different levels of egg contamination (10, 50, 100, 200 and 500 EPG) of GINs were used. The study was performed in both laboratories by the same expert operator and using the same standard operating procedures (SOPs) for the Mini-FLOTAC and McMaster techniques. Sensitivity, accuracy and precision were calculated for each technique and for each level of contamination. Statistical analyses were performed to evaluate differences in performance between the two techniques. Results Mini-FLOTAC had a higher sensitivity (100% at all EPG levels for Mini-FLOTAC vs 0–66.6% for McMaster chambers and grids at levels

Abstract Background Ticks and tick-borne diseases are increasingly recognized as a cause of disease in dogs worldwide. The epidemiology of ticks and tick-transmitted protozoa and bacteria has changed due to the spread of ticks to urban and peri-urban areas and the movement of infected animals, posing new risks for animals and humans. This countrywide study reports information on distribution and prevalence of pathogens in ticks collected from privately-owned dogs in Italy. We analyzed 2681 Ixodidae ticks, collected from 1454 pet dogs from Italy. Specific PCR protocols were used to detect i) Piroplasms of the genera Babesia and Theileria, ii) Gram-negative cocci of the family Anaplasmataceae and iii) Borrelia burgdorferi sensu lato. Sequencing of positive amplicons allowed for species identification. Results Babesia/Theileria spp. DNA was detected in 435 homogeneous tick-pools (Minimum Infection Rate (MIR) = 27.6%; 95% confidence interval (CI) = 25.4–29.8%) with higher prevalence in Ixodes ricinus and Rhipicephalus sanguneus group. The zoonotic B. venatorum was the most prevalent species (MIR = 7.5%; 95% CI = 6.3–9.0%). Anaplasma and Ehrlichia species were detected in 165 tick-pools (MIR = 10.5%; 95% CI = 9.3–11.8%) and specifically, A. phagocytophilum was identified with MIR = 5.1% (95% CI = 4.1–6.3%). Borrelia burgdorferi s.l. and B. afzelii were detected with MIR = 0.4% (95% CI = 0.2–0.8%) and MIR = 0.3% (95% CI 0.1–0.7%) respectively. Conclusions Zoonotic pathogens B. venatorum and A. phagocytophilum were the most frequently detected in ticks collected from privately-owned dogs which might be used as markers of pathogens presence and distribution.

Soil-transmitted helminth (STH) infections are among the most common neglected tropical diseases worldwide causing high morbidity and mortality rates in endemic areas. Preventive chemotherapy (PC) programmes and health education are recommended by the World Health Organization (WHO) to reduce the impact of STH in endemic countries. Following our role as WHO collaborating centre (WHO CC ITA-116), we have developed a WebGIS and a dataset to support PC programmes to monitor the impact of STH control. This vHealth presentation shows the potentiality of these tools in improving communication among WHO’s regional and country offices, Ministries of Health, pharmaceutical industries and other partners.

The development of anthelmintic resistance in sheep gastrointestinal nematodes (GINs) requires novel strategies for the sustainable control of these parasites. This study aimed to evaluate the anthelmintic efficacy of the Origanum vulgare (L.) essential oil (EO) against sheep GINs and to evaluate the possibility of its use in control practice. The in vitro egg hatch test was conducted at eight different concentrations (50, 12.5, 3.125, 0.781, 0.195, 0.049, 0.025, and 0.0125 mg/mL) of the tested EO. For the in vivo fecal egg count reduction test, the EO of O. vulgare was administrated orally at a mean single dose of 150 mg/kg to sheep from two farms in Southern Italy, whereby potential toxic effects to the hosts were also evaluated. In the egg hatch test, the inhibition of egg hatchability varied from 71.3% to 93.7%, depending on the concentration used. The high anthelmintic potential was confirmed in the fecal egg count reduction test with an average reduction of nematode eggs in feces of 43.2% and 60.1% on days 7 and 14 after treatment, respectively. In addition, no toxic effects were noticed during the clinical examination of sheep or by observing blood count and liver or kidney function test results. The obtained results suggest the strong activity of the O. vulgare EO against sheep GINs, probably due to a high percentage of carvacrol (76.21%), whereby it can be considered safe for sheep at the dose tested in vivo. Therefore, it is suitable for use in veterinary practice as a part of an integrated strategy for the control of sheep GINs.

Abstract Background Faecal egg counts (FEC) and the FEC reduction test (FECRT) for assessing gastrointestinal nematode (GIN) infection and efficacy of anthelmintics are rarely carried out on ruminant farms because of the cost of individual analyses. The use of pooled faecal samples is a promising method to reduce time and costs, but few studies are available for cattle, especially on the evaluation of different pool sizes and FECRT application. Methods A study was conducted to assess FEC strategies based on pooled faecal samples using different pool sizes and to evaluate the pen-side use of a portable FEC-kit for the assessment of FEC on cattle farms. A total of 19 farms representing 29 groups of cattle were investigated in Italy and France. On each farm, individual faecal samples from heifers were collected before (D0) and two weeks after (D14) anthelmintic treatment with ivermectin or benzimidazoles. FEC were determined individually and as pooled samples using the Mini-FLOTAC technique. Four different pool sizes were used: 5 individual samples, 10 individual samples, global and global on-farm. Correlations and agreements between individual and pooled results were estimated with Spearman’s correlation coefficient and Lin’s concordance correlation coefficients, respectively. Results High correlation and agreement coefficients were found between the mean of individual FEC and the mean of FEC of the different pool sizes when considering all FEC obtained at D0 and D14. However, these parameters were lower for FECR calculation due to a poorer estimate of FEC at D14 from the faecal pools. When using FEC from pooled samples only at D0, higher correlation and agreement coefficients were found between FECR data, the better results being obtained with pools of 5 samples. Interestingly, FEC obtained on pooled samples by the portable FEC-kit on-farm showed high correlation and agreement with FEC obtained on individual samples in the laboratory. This field approach has to be validated on a larger scale to assess its feasibility and reliability. Conclusions The present study highlights that the pooling strategy and the use of portable FEC-kits on-farm are rapid and cost-effective procedures for the assessment of GIN egg excretion and can be used cautiously for FECR calculation following the administration of anthelmintics in cattle.

Abstract Background Giardia spp. is a protozoan pathogen and is the most common enteric parasite of domestic animals and humans. Assays for detecting infection in fecal samples using direct or indirect examinations are important tools for diagnosing the disease. The objective of the present study was to compare the cost-effectiveness of immunoassays and FLOTAC technique for diagnosing Giardia spp. infection in dogs. Results Fecal samples from 80 positive stray dogs were tested for the presence of copro-antigens of Giardia spp. using the direct immunofluorescence assay (IFA), a rapid enzyme-linked immunosorbent assay (ELISA) and the FLOTAC double technique. All methods were performed in accordance with the instructions reported in the original description for each technique. The results showed that ELISA can be run in less time than IFA and almost at the same time of the FLOTAC technique. Among the tests used in this study, FLOTAC had the lowest cost per correct diagnosis, compared with immunoassays. Conclusions The results from this cost-effectiveness analysis, in combination with the sensitivity and specificity of the FLOTAC technique, suggest that the FLOTAC technique can be use in the routine diagnosis of Giardia spp. infection in dogs.

Abstract Animals reared in restricted environments are highly susceptible to gastrointestinal infection by helminths and protozoa and therefore zoos are characterized as being parasite-rich environments. Successful implementation of control programs of these parasites in zoo environment depends upon precise and rapid diagnosing of gastrointestinal infections. The aim of this study was to demonstrate the role of the Mini-FLOTAC technique in combination with Fill-FLOTAC for rapidly diagnosing parasitic infections in zoo mammals. Fecal samples were collected from 70 animals in four different zoos located in central and southern Italy. All the samples were analyzed using Mini-FLOTAC in combination with Fill-FLOTAC. Out of the 70 pooled samples examined, 80% (24/30) were positive for at least one parasite. Among the gastrointestinal nematodes, Strongyles were the most frequent (40%), followed by Trichuris spp. (23.3%), Parascaris spp. (13.3%) and Capillaria spp. (3.3%). Among the protozoa, Blastocystis spp., Giardia spp. and Eimeria spp. were detected in 6.6%, 3.3% and 3.3%, respectively. These results show that Mini-FLOTAC in combination with Fill-FLOTAC can be used, not only for rapidly diagnosing parasitic infections in zoo mammals, but also for monitoring control programs in which large numbers of fecal samples need to be examined rapidly and reliably.

Infected dogs are considered the main domestic animal reservoirs for Leishmania infantum parasite. Infectiousness to competent phlebotomine vectors has been associated with many factors, the main being the severity of the disease exhibited by infected dogs. This study examines the relationship between different clinical parameters and the infectiousness to colonized Phlebotomus perniciosus sand flies having a blood meal on dogs. Data obtained in the present study come from an untreated group of Leishmania sick dogs submitted to xenodiagnosis for the evaluation of a spot on insecticide solution. Seventeen dogs were diagnosed as affected by leishmaniasis through clinical examination, immunofluorescence antibody test (IFAT) serology, and loop-mediated isothermal amplification (LAMP). The disease severity (clinical score) was staged by using a numeric value derived from eight clinical and parasitological parameters. Xenodiagnosis was performed on caged dogs exposed for 1.5 h to sand-fly bites. The following parameters related to sand flies were examined: blood feeding (% of blood engorged females), promastigote detection (% of promastigote-positive sand flies), promastigote burden, and the promastigote stage maturation (potential transmissibility rate). Statistical relationship between the clinical score and entomological parameters was investigated, as well as the possible correlation between each clinical and laboratory parameters and sand fly infection/infectivity. The severity of clinical score may influence the blood feeding by, and the probability of promastigote detection in, sand flies; skin lesions seem to be the main factor that influences the rate of blood feeding. Promastigote burden is related to IFAT titer, skin lesions, and clinical score. All entomological parameters are strongly related among them. This study confirms that both P. perniciosus infection and infectivity are influenced by a dog's clinical condition.

A field trial performed in-home conditions was conducted on 24 dogs naturally infected with Giardia, in order to compare the efficacy of fenbendazole and metronidazole. Animals were allocated in groups randomly in order to obtain two groups of 12 dogs each with similar parasitic loads of Giardia cysts: dogs in Group A were treated with fenbendazole (Panacur®, Intervet Italia Srl) administered at the dose of 50 mg/kg orally once a day for 5 consecutive days, dogs in Group B were treated with metronidazole (Flagyl®, Zambon Italia Srl) administered orally at the dose of 50 mg/kg, once a day for 5 consecutive days. All the dogs that were shedding Giardia cysts after the first treatment (Day 0) were retreated (either at Day 7 or at Day 14 or at Day 21) until a negative result was obtained with the same treatment. Additionally, all the dogs were re-examined at Day 50. All the dogs were tested for the presence of Giardia cysts using a fecal flotation method (FLOTAC). The percent efficacy of the treatments (A and B) was calculated at each sampling point (Days 7, 14, 21, and 50) as reduction in mean Giardia cysts. After the first therapy, on day 7, 4/12 (33.3%) dogs tested positive for Giardia cysts in the Group A and 5/12 (41.7%) in the Group B. Efficacies at (Days 7, 14, 21, and 50) of the treatments against Giardia infection were 80.9, 94, 100, and 97% in the Group A and 70.8, 99, 100, and 97.1% in the Group B. Statistically significant differences were not observed between the efficacy of Fenbendazole and Metronidazole against infection by G. duodenalis (P = 0.686). Molecular analysis revealed full homology (i.e., 100% with JN416550) with the canine specific assemblage D in six positive dogs. Different hypotheses might explain the re-appearance of the Giardia cysts in some dogs after treatment, e.g., re-infection from the home environment, the correct medication given by the owners, the diet, as well as treatment failure, but also biological issues related to the intermittent excretion of Giardia cysts.

Helminth infections, mainly by gastrointestinal nematodes (GIN), are one of the main concerns for animal health, welfare and productivity in grazing ruminant livestock worldwide. The use of a sensitive, precise, accurate, low-cost, and easy-to-perform copromicroscopic technique is of pivotal importance to perform reliable fecal egg count (FEC) and fecal egg count reduction test (FECRT), in order to determine the need of anthelmintic treatment, but also anthelmintic efficacy or resistance. This approach is fundamental to a correct and efficient control of GIN. Unfortunately, in worldwide ruminant farm practice, repeated anthelmintic treatments are carried out, without prior diagnosis of infection, contributing to the spread of Anthelmintic Resistance (AR). Tackling this phenomenon, improving mainly the GIN diagnosis and AR status in farm animals, is a priority of the European COST Action “COMBAR—COMBatting Anthelmintic Resistance in Ruminants” and of the STAR-IDAZ International Research Consortium on Animal Health. One of the specific objectives of the COMBAR Working Group 1 (WG1) is to conduct an European market analysis of new diagnostics and develop a business plan for commercial test introduction, leveraging technical know-how of participants. Since the Mini-FLOTAC in combination with the Fill-FLOTAC may be considered a good candidate for a standardized FEC and FECRT in the laboratory, as well as directly in the field, the aim of this study was to conduct SWOT (Strength—Weaknesses—Opportunities—Threats) and PESTEL (Political, Economic, Social, Technological, Environmental, and Legal) analyses of these tools in 20 European countries involved in the COMBAR WG1, in order to identify the opportunities, barriers, and challenges that might affect the Mini-FLOTAC and Fill-FLOTAC commercialization in Europe.

Coccidiosis due to Eimeria spp. are widespread parasitic infections in cattle and water buffaloes and may impair health, welfare, and production of these livestock species. The aims of this study were (i) to investigate the prevalence and seasonal dynamics of eimeriosis and (ii) to characterize the Eimeria species in large ruminants in a Mediterranean area, in order to plan effective control strategies. Parasitological data were obtained from a 10-year surveillance program (2010–2019) on 3,631 farms (2,089 buffalo and 1,542 cattle farms) sampled in central and southern Italy. Pooled fecal samples were analyzed using the FLOTAC technique with an analytic sensitivity of 2 oocysts per gram of feces (OPG) utilizing a saturated sodium chloride flotation solution (specific gravity = 1.200). Eimeria species identification was performed by morphometric analysis after a one week incubation of oocysts in a 2.5% potassium dichromate solution. The results showed high prevalence of Eimeria (up to 100%) in both cattle and buffaloes in the 10 years of surveillance, even if a slight reduction was reported in the last three years. The overall prevalence of eimeriosis was 91.7% (95% confidence interval, 95% CI = 90.2–93.1) in cattle farms and 81.5% (95% CI = 79.8–83.1) in water buffalo farms. The mean OPG value was 66.8 (min = 2; max = 8,065) in cattle and 55.9 (min = 2; max = 15,415) in water buffaloes, but this difference was not statistically significant (p > 0.05). In total, nine species of Eimeria were found in cattle the most prevalent being Eimeria bovis, E. ellipsoidalis, E. cylindrica, and E. zuernii, whereas in water buffaloes eight species of Eimeria were found, the most prevalent being E. ellipsoidalis, E. auburnensis, E. bovis, and E. zuernii. Mixed infections were common in both ruminant species. The seasonal pattern showed a higher prevalence of eimeriosis in cattle in spring (86.9%) whereas in buffalo farms the prevalence was higher in winter (82.3%) and summer (82.4%). In conclusion, the 10-year surveillance program indicates that eimeriosis is common in cattle and water buffaloes and therefore continuous effective control strategies are needed.

Herbal products such as essential oils may play a promising role in the treatment of infections caused by gastrointestinal nematodes (GINs). The aim of this study was to evaluate the in vitro potential of 11 essential oils (EOs) and one binary combination of isolated EO compounds, as well as the in vivo anthelmintic efficacy of two EO formulations. Four GIN genera were identified in the coproculture examination: Haemonchus, Trichostrongylus, Teladorsagia and Chabertia. The in vitro egg hatch test (EHT) was performed at six different concentrations (50, 12.5, 3.125, 0.781, 0.195 and 0.049 mg/mL) for each EO, whereas in the in vivo faecal egg count reduction test (FECRT), each EO sample was diluted in sunflower oil and orally administrated at a dose of 100 mg/kg to the different group of animals. In the EHT, the EOs of Origanum vulgare, Foeniculum vulgare, Satureja montana, Satureja hortensis and two types of Thymus vulgaris were the most effective. The dominant compounds of these EOs were carvacrol, thymol, anethol, p-cymene and γ-terpinene, indicating their importance for the anthelmintic activity. In the FECRT, both T. vulgaris EO type 1 and linalool:estragole combination show an anthelmintic potential with a mean effect on FECR of approximately 25%. The results suggest the possible role of tested EOs as anthelmintic agents in sheep farms, although further in vivo tests are needed.

Abstract Background The geographical distribution of ticks on companion animals needs to be monitored to develop and plan effective control measures, as suggested by the European Scientific Counsel on Companion Animal Parasites. The aim of this study was to conduct the first Italian national survey of tick distribution on privately owned dogs. Methods The study was performed over 20 months (February 2016 - September 2017) and involved 153 veterinary practices in 64 different provinces covering 17/20 (85%) Italian regions. Participating practitioners were asked to examine five different dogs per month at random and complete a questionnaire for each dog. Differences in tick infestation associated with: sex, age and hair length (long and short); the dog’s habitat (indoor or outdoor/kennel); and the dog’s environment (urban or rural/sylvatic) were evaluated. The attachment site of ticks on the dog was also recorded. Acaricide efficacy was evaluated for the subset of dogs for which complete information on product used, date of sampling and date of last ectoparasiticide treatment was available. Results Of the 3026 dogs examined, 1383 (45.7%) were carrying at least one tick. Overall, 2439 tick samples were collected and a total of 14 tick species identified. Rhipicephalus sanguineus group were the most predominant ticks (63.6%), followed by Ixodes ricinus (30.6%) and I. hexagonus (5.6%). Twenty-four dogs had mixed tick infestations. Long-haired dogs had a higher tick infestation risk as did dogs with outdoor and rural/sylvatic lifestyles. Ticks were located on the head (37.4%), the neck (28.8%), the muzzle (15.5%) and the back (15.3%). A higher prevalence of Rhipicephalus was found in the interdigital spaces (10.8%) compared to Ixodes (0.2%). Finally, ectoparasiticide treatments were found significantly protective against tick infestation, especially orally administered formulations. Conclusions Privately owned dogs in Italy have a high prevalence (45.7%) of infestation with ixodid ticks and this risk varies by dog phenotype and lifestyle.

Abstract Background Nematode infections in horses are widespread across the world. Increasing levels of anthelmintic resistance, reported worldwide in equine parasites, have led to the creation of programs for the control of nematodes based on faecal egg counts (FEC). To improve nematode egg counting in equine faecal samples and establish whether the matrix of equine faeces or the eggs affect the counts, the analytical sensitivity, accuracy and precision of Mini-FLOTAC (combined with Fill-FLOTAC), McMaster and Cornell-Wisconsin techniques were compared. Known numbers of eggs extracted from equine or ovine faeces were added to egg free ovine and equine faeces to give counts of 10, 50, 200 and 500 eggs per gram (EPG) of faeces. Results The Cornell-Wisconsin significantly underestimated egg counts and McMaster showed a low analytical sensitivity, revealing 100% of sensitivity only for concentrations greater than 200 EPG. EPG values detected by Mini-FLOTAC did not differ significantly from expected counts at any level of egg density. Conclusions Mini-FLOTAC combined to Fill-FLOTAC which provides an accurate method of weighing without need for a balance and filtering out debris, could be used for FEC on the farm as well as in the laboratory.

Animal Assisted Interventions (AAIs) programmes have been considered useful in different settings, such as hospital, therapeutic, educational and assisted living environments. In these contexts, all animals, and particularly dogs, should be subjected to appropriate health controls to prevent a potential risk of transmission of zoonotic agents. Domestic dogs are reservoirs of many zoonotic pathogens including several gastrointestinal parasites (protozoa and helminths). Therefore, the aim of the present study was to investigate the presence of the protozoan Giardia duodenalis and zoonotic gastrointestinal nematodes (geohelminths) in dogs hosted in a dog educational centre in the city of Naples (southern Italy) where the animals were trained to AAI. Between April and June 2016, 74 dog faecal samples were analysed using the FLOTAC dual technique to detect G. duodenalis cysts and other parasitic elements. Out of the 74 faecal samples examined, 18 (24.3%; 95% CI = 15.4–35.9) were positive for parasitic elements. Specifically, 8 were positive for G. duodenalis (44.4%; 95% CI = 22.4–68.7). In addition, some co-infections were also found: one sample (5.6%; 95% CI = 0.3–29.4) resulted positive to both Toxocara canis and Trichuris vulpis and two samples (11.1%; 95% CI = 1.9–36.1) were positive to both G. duodenalis and Ancylostomidae. Given that children, young adults and immunocompromised individuals are among the main users of the AAIs, specific guidelines targeting G. duodenalis and other gastrointestinal zoonotic parasites should be formulated in order to develop effective control and prevention strategies and reduce the zoonotic risk favoured by the human-dog interaction.

Although the distribution of Babesia spp. and Hepatozoon canis is well known in Romania, there is still a marked lack of information in many places of the country. This study aimed to investigate the occurrence of these haemoparasites in symptomatic dogs and in their ticks in Iasi, eastern Romania. Ninety owned dogs were subjected to clinical examination at the Faculty of Veterinary Medicine of Iasi and all detectable ticks (58 ticks from 15 dogs) were collected. Additionally, 124 ticks collected from the coat of other dogs (no. = 23) were included. Three Babesia species were found in dogs: Babesia canis (94.4%), Babesia vogeli (3.3%), and Babesia rossi (2.2%). All the dogs resulted negative for H. canis. The ticks were identified as follows: Ixodes ricinus (64%), Dermacentor reticulatus (33%), and Rhipicephalus sanguineus group (3%). B. canis (Minimum Infection Rate; MIR = 81%), B. vogeli (MIR = 3%), and Babesia microti-like piroplasm (MIR = 1%) were found in ticks. Moreover, 15 ticks were positive for H. canis, 6 were co-infected with B. canis, and 1 with B. microti-like piroplasm. This is the first molecular identification of B. rossi in two symptomatic dogs from Romania, although further studies are needed to investigate the vector competence of other ticks from Europe.

Rumen flukes (Calicophoron daubneyi) represent a growing threat to the animal health, productivity and welfare of ruminants. The present study aimed to assess the spatial distribution of C. daubneyi infections in ruminants and to develop a predictive model of the environmental suitability for rumen flukes in a Mediterranean area. A cross-sectional coprological survey was conducted in 682 sheep, 73 goat and 307 cattle farms located in the Basilicata region (southern Italy). Faecal samples collected were analysed using the FLOTAC technique. Geographical Information Systems (GIS) and statistical models were developed to determine environmental risk factors and to delimitate the areas at highest risk of infections in small ruminants. The results showed 7.9% (95% CI 6.05–10.27) of sheep farms, 2.7% (95% CI 0.48–10.44) of goat farms and 55.0% (95% CI 49.62–60.99) of cattle farms were infected by C. daubneyi. The areas with high predicted risk were situated in the western part of the region. The soil texture, land use and the presence of streams and brooks were the variables statistically significant (p < 0.05) in explaining the C. daubneyi distribution in the study area. The study confirms the importance of geospatial technology in supporting parasite control strategies in livestock and demonstrates that a combined use of different geostatistical techniques can improve the prediction of the C. daubneyi infection risk in ruminants.

Abstract Background Analyses of environmental correlates of the composition of gastrointestinal parasite communities in black howler monkeys (Alouatta pigra) have been hindered by inadequate calibration techniques of detection and quantification methods of the parasites. Here we calibrate samples and compare the likelihood of parasite detection using two flotation techniques, FLOTAC and Mini-FLOTAC, and compare flotation solution, preservation method and dilution ratio for egg detection and counts of the most common parasites (Controrchis spp. and Trypanoxyuris spp.) in howler monkeys. Results For samples preserved in 5% formalin, the Mini-FLOTAC technique was the best option for qualitative and quantitative copro-microscopic analysis. This technique displays an 83.3% and 100% detection of Controrchis spp. and Trypanoxyuris spp. infections, respectively. For the trematode Controrchis spp., more eggs per gram of feces (EPG) were recorded with the flotation solution (FS) #7 (zinc sulfate; specific gravity SG = 1.35) at 1:20 and 1:25 dilution than other methods. By contrast, for the nematode Trypanoxyuris spp., the best results were recorded with FS1 (sucrose and formaldehyde; SG = 1.20) at 1:10 dilution. Conclusions We recommend the Mini-FLOTAC technique for general use with parasite analysis on frugivore/folivores like the howler monkey, especially if many samples are analyzed. The technique has a high detection rate and the best EPG counts, allowing the qualitative and quantitative analysis of parasite load among the species or populations without the need for specialized equipment.

Canine vector-borne diseases (CVBDs) are a spectrum of diseases caused by different pathogens transmitted by blood-feeding arthropoda. The aim of this study was to investigate leishmaniosis, babesiosis, and filarial infections in dogs with three different lifestyles (hunting, stray, and sheep dogs) in Molise, the smallest region of southern Italy, where data available about these parasitic infections are very scant. A cross-sectional survey was conducted on 318 hunting, 180 stray, and 218 sheep dogs. Immunofluorescence antibody test, blood smear, molecular techniques and Knott's test were performed to detect Leishmania infantum, Babesia spp. and filarial nematodes. Association between positivity to CVBDs, age, sex, and living conditions was evaluated. An overall prevalence of 12.3% of CVBDs caused by L. infantum (10.2%), B. canis canis (0.3%) and filarial nematodes (2.1%) was detected. Three dogs showed co-infections of L. infantum and B. c. canis (0.1%) or Acanthocheilonema reconditum (0.3%). A significantly association was found only for filarial infection in hunting dogs. These parasites were reported also in dogs without clinical signs. It is very important to plan effective control programs for CVBDs to guarantee not only the health and welfare of pets, but also the public safety, because some of mentioned parasites are of zoonotic importance.

Canine leishmaniosis (CanL) is caused by protozoans of the genus Leishmania and characterized by a broad spectrum of clinical signs in dogs. Early diagnosis is of great importance in order to perform an appropriate therapy and to prevent progression towards severe disease. The aim of this study was to compare a point-of-care molecular technique, i.e., the loop-mediated isothermal amplification (LAMP), with a real-time polymerase chain reaction (Rt-PCR), and three serological techniques, i.e., immunofluorescence antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA), and a rapid SNAP Leishmania test, to develop an integrated approach for the diagnosis of CanL. Sixty dogs were chosen after physical examination and collection of blood and sera samples, fine-needle aspiration of lymph nodes, and conjunctival swabs were performed. Lymphadenopathy (82.3%), as well as clinicopathological alterations of total proteins (70.6%), were the most frequent signs. Forty-one (68.3%) samples resulted positive at least to one technique. IFAT resulted in the best serological diagnostic method (specificity = 100%, sensitivity = 97.2%), detecting a higher number of positive samples than those revealed by other techniques. Among the samples used for molecular analysis, fine-needle aspiration of lymph nodes was revealed as the best sample source. LAMP showed a substantial agreement (κ = 0.80; p

One hundred twenty-four water buffaloes (Bubalus bubalis) originating from 9 farms located in southern Italy were tested to investigate simultaneously, for the first time, the seroprevalence of the protozoa Neospora caninum, Toxoplasma gondii and Besnoitia besnoiti by ELISA tests and to evaluate the clinical findings potentially associated to the presence of these aborting parasitic pathogens. Twenty-five of 124 buffaloes (20.2%) were positive for N. caninum, while 17/124 (13.7%) for T. gondii. No buffalo showed specific antibodies for B. besnoiti. Nineteen of 124 animals (15.3%) were found seropositive for both T. gondii and N. caninum. The univariate statistical analysis showed that the seroprevalence of N. caninum is significantly associated with abortion and presence of retained foetal membranes, while the seroprevalence of T. gondii is significantly associated with an increase of days open. The logistic regression models showed that the co-infection by N. caninum and T. gondii strengthened the abortive effects (OR = 7.330) and showed further negative effects on the parameter embryonic death (OR = 2.607). The outcome revealed herein represents a high exposure of N. caninum and T. gondii in water buffaloes with reproduction disorders that deserves attention for both economic reasons, animal health and welfare.

Abstract Background Kennel dogs are at the high risk of infections with intestinal and extra-intestinal helminths. Therefore, regular parasitological surveillance, appropriate treatment strategies and high quality standard of hygiene are required to guarantee the health and welfare of kennel dogs. The aim of the present study was to evaluate the efficacy of helminth control in kennels using different broad-spectrum anthelmintics that are routinely used in canine veterinary practice. Particular attention was given to the field efficacy and ease-of-use of each product. Methods The study was conducted in 3 public kennels in the Campania region (southern Italy). Eighteen boxes from each of the three kennels were selected for treatment based on faecal egg counts (FECs) at Day −30. The treatments were conducted using tablets containing combinations of: i) milbemycin oxime and praziquantel (Milbemax®); ii) pyrantel embonate, febantel and praziquantel (Drontal Plus Flavour®), and; iii) pyrantel pamoate, oxantel pamoate and praziquantel (Nemex® POP). All dogs were treated on Day 0 and sampled on Days 0, 7, 14 and 21 for copromicroscopic analyses. The FLOTAC dual technique on pooled samples was used with efficacy determined by reduction in FECs. Results At Day −30 intestinal nematodes (hookworms, Toxocara, Trichuris) and cestodes (Dypilidium caninum) were found. Milbemax® showed 100% efficacy against all the helminths in all the kennels. Drontal Plus Flavour® was 100% effective against hookworms in all the kennels but gave lower efficacy against T. canis (range = 97.1-100%) and T. vulpis (range = 95.6-100%). Nemex® POP was also 100% effective against hookworms in all kennels but less effective against T. canis (range = 95.7-100%) and T. vulpis (range = 95.7-100%). All three drugs were 100% effective against D.caninum. Conclusions Milbemax®was fully effective against all the helminthes and was palatable and ease-of-use for all the dogs. It is the optimum choice for the treatment and prevention of helminth infections in kennel dogs under field conditions.

Animal-assisted interventions (AAIs) usually contribute to the well-being and health of users/patients, but it is essential that the animals involved in these activities do not represent a source of zoonoses. This systematic review focused on the evaluation of the potential risk of the transmission of Toxocara by dogs’ fur, considering their involvement as the main animal species in AAIs. Three databases were considered: MEDLINE/PubMed, Scopus, and Web of Science, and the PRISMA guidelines were used. Out of 162 articles found, 14 papers were identified as eligible for inclusion in the review. Although the findings were very heterogeneous, they showed that regular parasitological surveillance to plan effective control programs is strongly needed to guarantee the health of pets and consequently the public health, according to the concept of One Health. Since AAIs involve patients and/or users potentially susceptible, it is very important to appropriately treat dogs enrolled in these interventions after an accurate diagnosis of parasitic zoonoses.

A survey was conducted in the eastern part of Romania to assess the prevalence and geographical distribution of Dirofilaria immitis in dogs. Plasma samples were collected from 458 stray dogs hosted in shelters in 8 counties and tested serologically for the presence of heartworm. In addition, 45 blood samples from dogs of a shelter in Galati city were examined by the modified Knott and multiplex polymerase chain reaction (PCR) techniques. The immmunological assay showed a heartworm infection prevalence of 8.9% in the dogs. Optical density results for positive samples ranged between 0.217 and 2.683. Geographical information systems (GIS) were used to produce overlays of distribution maps of D. immitis prevalence and predictive maps based on temperature suitability. High prevalence of D. immitis was found in the central East up to the northern border of the country, i.e. Galati county (60%), followed by the counties of Vaslui (12.0%) and Iasi (7.7%). Out of 45 samples examined using the Knott test, 23 were positive for circulating microfilariae (51.1%), while 19 dogs were positive for D. immitis and 4 for both D. immitis and D. repens with the multiplex PCR test. The high prevalence for D. immitis shown in dogs in the Southeast (Galati, 42.2%) also by multiplex PCR gave strong support to the results achieved by the serological tests. The present study confirms the ability of GIS to predict the distribution and epidemiology of dirofilariosis in different geographical territories as has been already demonstrated by the empirical epidemiological data obtained at the continental, national and intraregional levels.

Toxoplasmosis, an important cause of reproductive failure in sheep, is responsible for significant economic losses to the ovine industry worldwide. Moreover, ovine meat contaminated by the parasite Toxoplasma gondii is considered as a common source of infection for humans. The aim of this study was to develop point and risk profiling maps of T. gondii seroprevalence in sheep bred in Campania Region (Southern Italy) and analyse risk factors associated at the flock-level. We used serological data from a previous survey of 117 sheep flocks, while environmental and farm management information were obtained from an analysis based on geographical information systems and a questionnaire purveyance, respectively. An univariate Poisson regression model revealed that the type of farm production (milk and meat vs only meat) was the only independent variable associated with T. gondii positivity (PT. gondii oocysts, so the high within-flock seroprevalence might derive from this management factor. However, further studies are needed to better assess the actual epidemiological situation of toxoplasmosis in sheep and to clarify the factors that influence its presence and distribution.

Cystic echinococcosis (CE), caused by the larval stages of the tapeworm Echinococcus granulosus, is known to be one of the most important parasitic infection in livestock worldwide and one of the most widespread zoonoses known. In the present study, we used a geographical information system (GIS) to study the spatial structure of livestock (cattle, water buffaloes and sheep) populations to gain a better understanding of the role of sheep as reservoir for the transmission of CE to cattle and water buffaloes. To this end, a survey on CE in cattle and water buffaloes from the Campania region of southern Italy was conducted and the geo-referenced results linked to the regional farm geo-referenced data within a GIS. The results showed a noteworthy prevalence of CE in cattle and water buffalo farms (overall prevalence = 18.6%). The elaboration of the data with a GIS approach showed a close proximity of the bovine and/or water buffalo CE positive farms with the ovine farms present in the study area, thus giving important information on the significance of sheep and free-ranging canids in the transmission cycles of CE in relation to cattle and water buffaloes. The significantly higher prevalence found in cattle as compared to water buffalo farms (20.0% versus 12.4%) supports the key role of sheep in the CE transmission; indeed, within the 5 km radius buffer zones constructed around the cattle farms positive for CE, a higher number of (potentially infected) sheep farms were found compared to those found within the buffer zones around the water buffalo farms. Furthermore, the average distances between the sheep and cattle farms falling in the same buffer zones were significantly lower than those between the sheep and water buffalo farms. We emphasize that the use of GIS is a novel approach to further our understanding of the epidemiology and control of CE and we encourage other groups to make use of it.