Survey of ectoparasites affecting dog and cat populations living in sympatry in Gamo Zone, Southern Ethiopia.

BACKGROUND: Ectoparasites of dogs and cats are implicated to be responsible for life-threatening anaemia, allergic dermatitis and pruritic and non-pruritic skin disorders. In Ethiopia, few studies have been conducted on the arthropods of dogs and cats. OBJECTIVES: In order to shed light on some of these aspects, a survey was conducted to investigate the presence of ectoparasites in dogs and cats living in sympatry in the urban and rural areas of Gamo Zone, Ethiopia. METHODS: A total of 297 dogs and 110 cats were examined for ectoparasites, and questionnaires were employed to obtain information concerning owner's knowledge about arthropods and vector-borne diseases (VBDs). RESULTS: The overall prevalence of ticks, fleas and lice in dogs was 36.7%, 69.7% and 4.7%, respectively. Similarly, on cats, an overall prevalence of 2.7% ticks and 21.8% fleas was recorded. On dogs, fleas (Ctenocephalides felis 69.4%, Echidnophaga gallinacea 1.3%, Ctenocephalides canis 1.0% and Pulex irritans 0.3%), ticks (Amblyomma variegatum 22.9%, Rhipicephalus sanguineus 14.1%, Haemaphysalis leachi 8.8%, Rhipicephalus praetextatus 4.0% and Rhipicephalus pulchellus 3.4%) and lice (Heterodoxus spiniger 4.0% and Trichodectes canis 0.7%) were identified. Likewise, on cats, fleas (C. felis [15.5%] and E. gallinacea [7.3%]) and ticks (H. leachi [2.7%]) were identified. The abundance of C. felis was significantly higher (p < 0.001) on dogs, whereas in cats, the abundance of E. gallinacea was significantly higher (p = 0.002) than the other ectoparasites. On dogs, a significantly higher prevalence of Rh. sanguineus was recorded in urban areas (<0.001) and on dogs which live in indoor environments (p = 0.003) than on dogs which live in other environments. On the other hand, the prevalence of A. variegatum in rural areas and midland agroecology was significantly higher (p < 0.001). The prevalence of H. leachi was significantly higher in midland (p < 0.001) and on adult dogs (p = 0.001). Overall, fleas were more prevalent in rural (p = 0.029) than in urban areas, and female dogs were with higher infestation than the male (p = 0.047) dogs; C. felis was prevalent in female (p = 0.038) dogs than males. Overall, 88.3% owners in the study area had no knowledge about ectoparasites and VBDs of dogs and cats. Majority of the owners (64.8%) attest that they had never visited veterinary clinics. CONCLUSIONS: In conclusion, the data presented in the present study provide additional knowledge on the importance of ectoparasites of dogs and cats and are believed to contribute in awareness creation and strengthening of veterinary services of dogs and cats of the study area.

Faecal egg count reduction test in goats: Zooming in on the genus level.

The faecal egg count reduction test (FECRT) is the most widely used method to assess treatment efficacy against gastrointestinal nematodes (GIN). Information on genera composition of the GIN community is not available with this test and it is commonly obtained by identifying cultured third-stage larvae (L3) or through molecular assays in the post-treatment survey, but results provided are usually only qualitative or semi-quantitative. The updated WAAVP guidelines now recommend assessing anthelmintic efficacy for each GIN genus/species separately (genus-specific FECRT), but this approach is poorly employed in Europe and in goats especially. For this reason, four FECRT trials were conducted using oxfendazole and eprinomectin in two Italian goat farms. Samples were processed individually using the McMaster technique and then pooled to create two samples from faeces of 5 animals each. Pooled samples were analysed using the McMaster and cultured for seven days at 26°C to obtain L3s. The genus-specific FECRT was based on larval identification, integrating coproculture and FEC results. Larvae were identified as Haemonchus, Trichostrongylus, Teladorsagia, Oesophagostomum / Chabertia and Bunostomum. Molecular assays (a multiplex real-time PCR and two end-point PCRs) were also implemented on pooled samples to support the morphological identification. The Spearmann Rho test confirmed a high correlation between the two approaches (Rho = 0.941 and Rho = 0.914 respectively for Haemonchus and Trichostrongylus, the two most common genera). Both oxfendazole and eprinomectin were effective in one farm, while none in the other farm (FECR = 75.9% and 73.3% respectively). In the second farm, the genus-specific FECRT highlighted a different response to treatment among genera: oxfendazole lacked efficacy against both Haemonchus and Trichostrongylus spp., eprinomectin only against Haemonchus, while all other genera were susceptible to both drugs. This study brings new attention on the importance of adopting a genus-specific approach to identify and quantify differences in susceptibility to anthelmintics among genera in goats, providing support for FECRT interpretation, anthelmintic resistance evaluation and evidence-based GIN control.

Parasitological transitions: selected outcomes from the XXXII Congress of the Italian Society for Parasitology.

Founded in 1959, the Italian Society of Parasitology (SoIPa) includes nearly 200 researchers and professionals in the fields of medicine, veterinary medicine, biotechnology, epidemiology and environmental sciences. The diversity of its members, in a historical and continuous collaboration with other international scientific societies, embodies a broad and multidisciplinary field such as parasitology. Since 1959, SoIPa has organized a biennial congress, covering all aspects of general parasitology with participants from all over Italy, Europe and beyond, involved in a dynamic and multi-faceted scientific framework of contributions and symposia. The present Special Issue (SI) contains 6 review papers and 1 research article, focussed on emerging topics presented and discussed during some of the symposia organized within the XXXII SoIPa Congress, held in Naples from 27th June to 30th June 2022. These review papers reflect several emerging subjects (i.e. 'Italian network on Neglected Tropical Diseases', 'Wildlife parasites and citizen science', 'Comparing approaches to parasitological issues', 'Unusual perspectives on the role of parasites') with the aim to explore the new role that parasitologists can play in the future society, working together to promote dialogue on science-informed decisions to support the so-called 'twin green and digital transition'.

Wild ungulates as sentinels of flaviviruses and tick-borne zoonotic pathogen circulation: an Italian perspective.

BACKGROUND: Vector-borne zoonotic diseases are a concerning issue in Europe. Lyme disease and tick-borne encephalitis virus (TBEV) have been reported in several countries with a large impact on public health; other emerging pathogens, such as Rickettsiales, and mosquito-borne flaviviruses have been increasingly reported. All these pathogens are linked to wild ungulates playing roles as tick feeders, spreaders, and sentinels for pathogen circulation. This study evaluated the prevalence of TBEV, Borrelia burgdorferi sensu lato, Rickettsia spp., Ehrlichia spp., and Coxiella spp. by biomolecular screening of blood samples and ticks collected from wild ungulates. Ungulates were also screened by ELISA and virus neutralization tests for flaviviral antibody detection. RESULTS: A total of 274 blood samples were collected from several wild ungulate species, as well as 406 Ixodes ricinus, which were feeding on them. Blood samples tested positive for B. burgdorferi s.l. (1.1%; 0-2.3%) and Rickettsia spp. (1.1%; 0-2.3%) and showed an overall flaviviral seroprevalence of 30.6% (22.1-39.2%): 26.1% (17.9-34.3%) for TBEV, 3.6% (0.1-7.1%) for Usutu virus and 0.9% (0-2.7%) for West Nile virus. Ticks were pooled when possible and yielded 331 tick samples that tested positive for B. burgdorferi s.l. (8.8%; 5.8-11.8%), Rickettsia spp. (26.6%; 21.8-31.2%) and Neoehrlichia mikurensis (1.2%; 0-2.4%). TBEV and Coxiella spp. were not detected in either blood or tick samples. CONCLUSIONS: This research highlighted a high prevalence of several tick-borne zoonotic pathogens and high seroprevalence for flaviviruses in both hilly and alpine areas. For the first time, an alpine chamois tested positive for anti-TBEV antibodies. Ungulate species are of particular interest due to their sentinel role in flavivirus circulation and their indirect role in tick-borne diseases and maintenance as Ixodes feeders and spreaders.

Epidemiology and identification of Eimeria species affecting poultry in East Gojjam Zone, North West Ethiopia.

BACKGROUND: Despite the expansion of modernized poultry farming in Ethiopia, the presence of high prevalence of Eimeria species is the bottleneck in the sector causing high morbidity and mortality rate in poultry. OBJECTIVES: The objectives of this study were to estimate the prevalence and identify Eimeria species and investigate the major risk factors. METHOD: A cross-sectional study was conducted from November 2019 to April 2020 in East Gojjam Zone, North West Ethiopia. A total of 384 chickens were used. Both floatation and McMaster coprological techniques were employed. Univariate and multinomial logistic regression was used to calculate the odds ratio for the associated risk factors. Analysis of variance was used to analyse differences in Eimeria oocyst counts among the groups. RESULTS: Overall prevalence of Eimeria species in poultry from the study area was 26.5%. Age (OR = 0.25, p = 0.001), management system (OR = 12.44, p = 0.001) and production system (OR = 0.37, p = 0.001) were found significantly (p < 0.05) associated with the risk of Eimeria species in poultry. The mean Eimeria oocyst count was significantly different by age and management system (F = 6.526, p = 0.002), (F = 5.369, p = 0.005), respectively. The mean Eimeria oocyst count was significantly greater in 6-12 weeks (p = 0.004) and <6 weeks of age (p = 0.025). A total of 6 Eimeria species were identified. Eimeria tenella (46.07%), Eimeria necatrix (24.5%) and Eimeria acervulina (8.82%) were the most common Eimeria species encountered. CONCLUSION: The prevalence of Eimeria species was higher in poultry in North West Ethiopia. Therefore, tailor-made intervention is required to mitigate risk factors and reduce the prevalence of Eimeria species in poultry from the study area.

Corrigendum: Comparing copromicroscopy to intestinal scraping to monitor red fox intestinal helminths with zoonotic and veterinary importance.

[This corrects the article DOI: 10.3389/fvets.2022.1085996.].

Comparing pooled and individual samples for estimation of gastrointestinal strongyles burden and treatment efficacy in small ruminants.

Monitoring endoparasite burden (FEC) and treatment efficacy (FECR) is a key element of sustainable parasite control. However, the costs of the analysis often discourage their implementation by farmers and veterinary practitioners. Pooling samples is considered to be a good alternative to reduce time and monetary costs, but limited data are available on the use of pooled samples in small ruminants, especially for goats. In this study, data collected over the years in sheep and goat farms were analyzed, and results obtained from individual and pooled analysis were compared for the purposes of FEC and FECR assessment. A total of 801 individual and 134 pooled samples (composed of 3-12 individual samples) were included. For FECR testing, 2 pools of 5 samples each were created per trial and the same animals were sampled at day 0 (D0 - treatment day) and 14 days after (D14). Samples were analyzed by McMaster technique (limit of detection 20 EPG). Results from pooled and individual FEC were not significantly different (Wilcoxon signed-rank test) and correlation (Spearman's rank test) was high for all sub-categories, although agreement (Lin's concordance correlation) was often classified as poor. Results were not influenced by the pool size (<6 or ≥6). Interpretation of treatment efficacy between the two methods was comparable for all sheep trials, while it differed for goats in 4 out of 10 trials. Wilcoxon signed-rank test indicated a non significant difference between pooled and individual FECR. However, correlation and agreement between FECR were considerably better for sheep compared to goats, for which they were very limited, despite the correlation between FEC at D0 and D14 was always high. According to our results, pooled FECR can be a good option but the absence of 95 %CI represents a major drawbacks in the interpretation of results. Further studies on the topic for goats are needed.

Control of gastrointestinal helminths in small ruminants to prevent anthelmintic resistance: the Italian experience.

Helminth infections are ubiquitous in grazing ruminants and cause significant costs due to production losses. Moreover, anthelmintic resistance (AR) in parasites is now widespread throughout Europe and poses a major threat to the sustainability of modern ruminant livestock farming. Epidemiological data on the prevalence and distribution of gastrointestinal nematodes, cestodes and liver- and rumen-flukes in Italian small ruminants are outdated and fragmentary. However, anthelmintics are commonly used to control these infections and often without prior diagnosis. Compared to other European countries, few reports of AR in small ruminants against levamisole, ivermectin and benzimidazoles have been published in Italy, but recent studies suggest that this phenomenon is spreading. Increased and integrated research efforts in developing innovative approaches to control helminth infections and AR are needed and must be tailored to the peculiarities of each context in order to be effectively implemented. This manuscript provides an overview on helminth prevalence and distribution, sustainable treatment strategies and integrated control approaches in small ruminants in Italy. The implementation of targeted treatments and targeted selective treatments is discussed based on different parameters, such as fecal egg count, degree of anaemia (FAMACHA© method), milk production and body condition score. In addition, several Italian studies have also investigated the implementation of alternative strategies such as the use of different natural bioactive compounds or genetic selection for resistance and resilience to helminth infections. These concrete solutions for helminth management in small ruminant farms in the country are reported and discussed, representing a valid example for other Mediterranean countries.

Distribution and Prevalence of Anaplasmataceae, Rickettsiaceae and Coxiellaceae in African Ticks: A Systematic Review and Meta-Analysis.

In Africa, ticks continue to be a major hindrance to the improvement of the livestock industry due to tick-borne pathogens that include Anaplasma, Ehrlichia, Rickettsia and Coxiella species. A systemic review and meta-analysis were conducted here and highlighted the distribution and prevalence of these tick-borne pathogens in African ticks. Relevant publications were searched in five electronic databases and selected using inclusion/exclusion criteria, resulting in 138 and 78 papers included in the qualitative and quantitative analysis, respectively. Most of the studies focused on Rickettsia africae (38 studies), followed by Ehrlichia ruminantium (27 studies), Coxiella burnetii (20 studies) and Anaplasma marginale (17 studies). A meta-analysis of proportions was performed using the random-effects model. The highest prevalence was obtained for Rickettsia spp. (18.39%; 95% CI: 14.23-22.85%), R. africae (13.47%; 95% CI: 2.76-28.69%), R. conorii (11.28%; 95% CI: 1.77-25.89%), A. marginale (12.75%; 95% CI: 4.06-24.35%), E. ruminantium (6.37%; 95% CI: 3.97-9.16%) and E. canis (4.3%; 95% CI: 0.04-12.66%). The prevalence of C. burnetii was low (0%; 95% CI: 0-0.25%), with higher prevalence for Coxiella spp. (27.02%; 95% CI: 10.83-46.03%) and Coxiella-like endosymbionts (70.47%; 95% CI: 27-99.82%). The effect of the tick genera, tick species, country and other variables were identified and highlighted the epidemiology of Rhipicephalus ticks in the heartwater; affinity of each Rickettsia species for different tick genera; dominant distribution of A. marginale, R. africae and Coxiella-like endosymbionts in ticks and a low distribution of C. burnetii in African hard ticks.

Epidemiological Survey on Tick-Borne Pathogens with Zoonotic Potential in Dog Populations of Southern Ethiopia.

Dogs are known to host several tick-borne pathogens with zoonotic potential; however, scant information is available on the epidemiology of these pathogens in low-income tropical countries and in particular in sub-Saharan Africa. With the aim of investigating a wide range of tick-borne pathogens (i.e., Rickettsia spp., Anaplasma spp., Erhlichia spp., Borrelia spp., Hepatozoon spp. and Babesia spp.), 273 blood samples were collected from dogs in selected districts of Ethiopia and analyzed by real-time and/or end-point PCR. The results of the study showed that Hepatozoon canis was the most prevalent pathogen (53.8%), followed by Anaplasma phagocythophilum (7.0%), Babesia canis rossi (3.3%), Ehrlichia canis (2.6%) and Anaplasma platys (2.2%). Furthermore, five samples tested positive for Borrelia spp., identified as Borrelia afzelii (n = 3) and Borrelia burgdorferi (n = 2), and two samples for Rickettsia spp., identified as Rickettsia conorii (n = 1) and Rickettsia monacensis (n = 1). The finding of Anaplasma phagocythophilum and different species of the genera Borrelia and Rickettsia with zoonotic potential was unexpected and alarming, and calls for further investigation on the roles of dogs and on the tick, species acting as vector in this specific context. Other pathogens (Hepatozoon canis, Babaesia canis rossi, Anaplasma platys, Ehrlichia canis) are already known to have an important impact on the dogs' health but have minor zoonotic potential as they were rarely or never reported in humans. Dogs from rural areas were found to be at higher risk for different pathogens, probably due to the presence of other wild canids in the same environment. The findings of the present study contribute to a better knowledge of the epidemiology of tick-borne pathogens, which is relevant to human and animal health.

Epidemiology of Gastrointestinal Parasites of Cattle in Three Districts in Central Ethiopia.

Parasitic diseases are a major impediment to livestock production worldwide. However, knowledge about the epidemiology of gastrointestinal parasites in many low-income countries is still limited. An epidemiological survey on these parasites in traditionally reared cattle was performed in central Ethiopia (West Shewa Zone), from September 2019 to November 2021. Overall, 691 samples were analyzed qualitatively (flotation technique) and quantitatively (McMaster technique) to detect helminth eggs and Eimeria oocysts. Furthermore, coprocultures were conducted on pooled samples to identify the genera of gastrointestinal strongyles (GIS). The difference in prevalence according to agroecology, season, age, sex and body condition score (BCS) was investigated using univariable tests and through a multivariable logistic regression analysis, whereas abundance values were interpreted using a descriptive approach. The highest prevalence values were observed for GIS (50.2%) and Eimeria spp. (36.0%), whereas Moniezia spp. (16.3%), Strongyloides spp. (5.1%) and Schistosoma spp. (4.2%) showed lower values. Trichuris spp. and Toxocara vitulorum were sporadically found. A coproculture revealed that Haemonchus spp. (34.6%), Trichostrongylus spp. (25.9%) and Bunostomum spp. (19.1%) were the most frequent genera of GIS. A poor BCS was strongly associated with the occurrence of GIS, while seasonal variations were detected for Eimeria spp., GIS, Moniezia spp. and Schistosoma spp., with a higher prevalence and burden during the rainy season, and agroecology also strongly influenced different parasitic taxa. This present study increases the knowledge about the epidemiological features of gastrointestinal parasites in the context of a low-income country, suggesting a more tailored approach for their control. The use of a BCS as an indicator for the selective treatment of highly infested animals is supported by our findings, introducing a possible way to prevent anthelmintic resistance in areas where basic diagnostic services are rarely used by farmers.

Pearl formation associated with gymnophallid metacercariae in Mytilus galloprovincialis from the Northwestern Adriatic coast: Preliminary observations.

Gymnophallids of the genus Parvatrema are small trematodes infecting waterbirds in their adult stage. Several species of clams and mussels have been found to act as first and second intermediate hosts, in which the trematode larval stages induce the formation of pearls. In this study, a wild population of Mytilus galloprovincialis was sampled along the Northwestern coast of the Adriatic Sea to evaluate the origin and extent of visible pearls. Parasitological investigations, including morphological and molecular analyses, and histopathology were carried out on a representative sample of mussels (n = 158) from June to September 2021. The overall prevalence of infection reached 75.3 %, and the intensity of infection ranged from a few trematodes to thousands per mussel, mostly occurring in the mantle and surrounded by variable numbers of conchiolin layers. Morphological studies allowed classification of the metacercariae as belonging to the genus Parvatrema, and the pairwise comparison of the obtained sequences, encompassing the internal transcribed spacer (ITS) region, revealed a genetic similarity (96.8 %) to Parvatrema duboisi. However, the phylogenetic analysis demonstrated the independent clustering of the obtained ITS sequences compared to other available Parvatrema species. For the relevant commercial impact that pearl formation may have on farmed mussels, ecological and epidemiological aspects of this infection would deserve further investigation in the area.

Evidence of Dirofilaria immitis in Felids in North-Eastern Italy.

Dirofilaria immitis is a mosquito-borne nematode, causing heartworm (HW) disease in wild and domestic canids. HW can also affect felids with different clinical patterns from asymptomatic pictures to sudden death, making the monitoring and diagnosis complicated. Canine HW is endemic in North-eastern Italy; however, very little information has been recorded for felids. This study aims to provide new information on HW in felids in North-eastern Italy. Two hundred and six domestic cats from Veneto, Friuli-Venezia Giulia, Trentino Alto-Adige regions (North-eastern Italy), nine captive felids from zoological parks from Veneto, and nineteen European wildcats from Friuli Venezia Giulia were recruited. Sera/plasma was analysed for the detection of anti-HW antibodies (Ab) and HW antigens (Ag); positive blood samples were molecularly analysed, targeting the HW DNA (5S-rRNA gene). Twelve out of two hundred and six (5.8%) cats presented with Ab, and three out of two hundred and six (1.5%) presented with Ag, mainly those from the Veneto region, already known as a canine HW-endemic area. Among Ab-positive cats, two were from Belluno, a mountain province previously considered free, suggesting the expansion of HW into the northern areas. No cats were positive for both Ab and Ag. Three out of nineteen (15.8%) wildcats were Ag-positive, constituting the first HW report in Italy. No captive felids were positive. Dirofilaria immitis DNA was not amplified in positive samples, suggesting the low sensitivity of PCR on blood. This study provides new data on the occurrence of HW in domestic cats and wildcats in North-eastern Italy.

Molecular characterization of Rickettsia spp., Bartonella spp., and Anaplasma phagocytophilum in hard ticks collected from wild animals in Benin, West Africa.

Tick and tick-borne pathogens constitute a growing veterinary and public health concern around the world. Ticks are considered natural reservoirs for tick-borne related pathogens and are equally responsible for the spread of infections in animals as well as humans. In this study, the presence of Rickettsia, Bartonella, and Anaplasma phagocytophilum was investigated in hard ticks collected from reptiles, birds, and wild mammalian animals. The sample collection was carried out between December 2020 and September 2021. Adult ticks (male and female) were collected from wild animals in six districts of southern Benin. Molecular analysis was used to verify the presence of pathogens in all the ticks collected from wild animals. A total of 504 ticks were collected and grouped into 115 different tick pools. The PCR analysis detected 19 out of 115 tick pools which turn out positive for Rickettsia spp. and 9/115 tick pools positive for Bartonella spp., while Anaplasma phagocytophilum DNA was not detected in any tick. Several of the tick species collected from our studied reptiles/wild mammalian animals could be potential sources of zoonotic pathogens when subjected to further investigation. Therefore, stringent attention should be established for tick infestation of reptiles/wild mammalian animals in order to put in place proper control and prevention measures for tick-borne diseases in the wild, which could serve as reservoirs in the infestation of domestic animals/humans in the event of any possible contact.

Description of a Sarcoptic Mange Outbreak in Alpine Chamois Using an Enhanced Surveillance Approach.

Since 1995, the Alpine chamois (Rupicapra r. rupicapra) population of the Dolomites has been affected by sarcoptic mange with considerable management concerns. In this study, 15 years (2006-2020) of passive surveillance and demographic data were analyzed in order to describe a mange outbreak. Furthermore, an enhanced passive surveillance protocol was implemented in order to evaluate the efficiency of ordinary vs. enhanced surveillance protocol in identifying dead chamois in the field and in reaching a correct diagnosis. Our results confirm the role of mange as a determining factor for chamois mortality, while stressing the importance of a wider view on the factors affecting population dynamics. The enhanced passive surveillance protocol increased the probability of carcass retrieval and identification of the cause of death; however, its adoption may be too costly if applied for long periods on a wide scale. Passive surveillance, in both ordinary and enhanced surveillance protocol, should encompass the use of other strategies in the future to study the eco-epidemiology of the disease in wild Caprinae.

A Comparative Study of Mini-FLOTAC With Traditional Coprological Techniques in the Analysis of Cetacean Fecal Samples.

Free-ranging cetaceans are considered sentinels for the marine ecosystem's health. New and non-invasive methods have been set up for the collection of fecal samples from free-ranging big whales at sea, permitting to gain an excellent epidemiological picture of parasitic infections in wild populations. To select the best protocol to be used for copromicroscopic examination in cetaceans stool samples, we evaluated the sensitivity of two commonly used techniques, i.e., a sedimentation-floatation method and the Mini-FLOTAC through validation by helminth isolation from the digestive tract. For this aim, gastrointestinal content and fecal samples were collected during necropsy from 44 cetaceans, including bottlenose dolphins (Tursiops truncatus), striped dolphins (Stenella coeruleoalba), sperm whales (Physeter macrocephalus), Risso's dolphins (Grampus griseus), Cuvier's beaked whales (Ziphius cavirostris), pilot whale (Globicephala melas), and fin whales (Balaenoptera physalus). Helminths were recovered through washing and filtering of the gastrointestinal contents and morphologically identified. Copromicroscopic examinations were performed on formalin-preserved fecal samples, using a sodium nitrate, sodium thiosulphate, and sucrose solution (s.g. = 1.450) for both methods. Helminths belonging to 9 taxa (i.e., the trematodes Synthesium tursionis, Synthesium delamurei, Campula palliata, Braunina cordiformis, Pholeter gastrophilus, the nematode Anisakis sp., cestodes of the family Tetrabothriidae and the acanthocephalan Bolbosoma sp.) were isolated. Eggs referable to the same taxa, with the exception of cestodes, were found in copromicroscopic analyses. Sensitivity of the Mini-FLOTAC method appeared higher or equal for all taxa, proving superior to the sedimentation-flotation method for the detection of all except Anisakis sp. The concordance of the two tests indeed revealed a moderate to perfect agreement (kappa values 0.42-1). Not excluding the limitations inherent to the techniques themselves, explanations for false-negative results at copromicroscopy could be linked to parasite-related factors, including prepatent infections, low parasitic burdens, or intermittent egg shedding. Notwithstanding these limitations, this study evidenced that the Mini-FLOTAC protocol approximates more accurately the composition of the gastrointestinal helminthic community of cetaceans from copromicroscopic examination, providing at the same time a quantitative estimation.

A highly endemic area of Echinococcus multilocularis identified through a comparative re-assessment of prevalence in the red fox (Vulpes vulpes), Alto Adige (Italy: 2019-2020).

Surveillance of Echinococcus multilocularis at the edge of its range is hindered by fragmented distributional patterns and low prevalence in definitive hosts. Thus, tests with adequate levels of sensitivity are especially important for discriminating between infected and non-infected areas. In this study we reassessed the prevalence of E. multilocularis at the southern border of its distribution in Province of Bolzano (Alto Adige, northeastern Alps, Italy), to improve surveillance in wildlife and provide more accurate estimates of exposure risk. We compared the diagnostic test currently implemented for surveillance based on coproscopy and multiplex PCR (CMPCR) to a real-time quantitative PCR (qPCR) in 235 fox faeces collected in 2019 and 2020. The performances of the two tests were estimated using a scraping technique (SFCT) applied to the small intestines of a subsample (n = 123) of the same foxes as the reference standard. True prevalence was calculated and the sample size required by each faecal test for the detection of the parasite was then estimated. True prevalence of E. multilocularis in foxes (14.3%) was markedly higher than reported in the last decade, which was never more than 5% from 2012 to 2018 in the same area. In addition, qPCR showed a much higher sensitivity (83%) compared to CMPCR (21%) and agreement with the reference standard was far higher for qPCR (0.816) than CMPCR (0.298) meaning that for the latter protocol, a smaller sample size would be required to detect the disease. Alto Adige should be considered a highly endemic area. Routine surveillance on definitive hosts at the edges of the E. multilocularis distribution should be applied to smaller geographic areas, and rapid, sensitive diagnostic tools using directly host faeces, such as qPCR, should be adopted.

High Prevalence of Tick-Borne Zoonotic Rickettsia slovaca in Ticks from Wild Boars, Northeastern Italy.

Tick-borne rickettsiae are emerging pathogens that are becoming widespread in Europe. Rickettsiae are endemic in Italy, but epidemiological data are currently scarce. This study aimed to improve our knowledge about rickettsial infections in tick and wild boar populations. Blood and ticks were collected from 102 wild boars in 2010 and 2018. Ticks were also collected from the vegetation in the area. All of the samples were examined using real-time PCR targeting the gltA gene to detect Rickettsia DNA. Positivity was confirmed by PCR amplifying the gltA and/or ompB genes. A total of 254 ticks and 89 blood samples were analyzed. Zoonotic rickettsiae were detected in the ticks but not in the blood samples. Rickettsia slovaca (R. slovaca) was the most prevalent in ticks and was found in 23.7% of Dermacentor marginatus (D. marginatus) and in 3.4% of Ixodes ricinus (I. ricinus). Other zoonotic species were identified, such as Rickettsia monacensis, which was detected in 12% of I. ricinus ticks, and Rickettsia helvetica which was found in 3.4% of questing I. ricinus ticks and in 1.1% of D. marginatus collected from wild boars. This study highlights a high prevalence of zoonotic rickettsiae, particularly that of R. slovaca, in northeastern Italy. As rickettsioses are underreported and underdiagnosed in human medicine, both clinicians and researchers should pay more attention to this topic.

Monitoring and detection of new endemic foci of canine leishmaniosis in northern continental Italy: An update from a study involving five regions (2018-2019).

Canine leishmaniosis (CanL) is an emerging zoonosis caused by Leishmania infantum and transmitted in southern Europe by phlebotomine sand flies of the subgenus Phlebotomus (Larroussius). Endemic foci of CanL have been recorded in northern continental Italy since early 1990s and attributed to the northward expansion of vector populations due to climatic changes in association with travelling/relocated infected dogs from the southern Mediterranean littoral. In this study, further spread of endemic Leishmania foci was monitored during 2018-2019 in five regions (Aosta Valley, Piedmont, Lombardy, Veneto and Friuli-Venezia Giulia), with focus to territories where investigations were not performed, or they have been inconclusive. Clinical cases of CanL identified by local veterinary practitioners and confirmed by reference diagnosis centers were regarded as autochthonous if their origin from, or travel to, areas endemic for CanL were excluded in the previous ≥2 years. Around these index cases, i) serosurveys for L infantum were carried out where indicated, ii) sampling from potential autochthonous cases in healthy or clinically-suspected resident dogs was intensified by collaborating veterinary practitioners, and iii) suitable sites were investigated for the presence of competent phlebotomine vectors. Fifty-seven municipalities whose enzootic status of CanL was unreported before 2018, were identified as endemic. The stability of 27 foci recorded over the past decade, was also confirmed. Competent phlebotomine vectors, mainly Phlebotomus perniciosus, were collected for the first time in 23 municipalities. The newly recorded endemic municipalities appear to be distributed over a west-to-east decreasing gradient: 30 in Piedmont, 21 in Lombardy, 4 in Veneto and 2 in Friuli-Venezia Giulia. As regards Veneto, it should be noted that a relatively restricted territory was investigated as several municipalities of the region had already been surveyed and detected as endemic for CanL in the past. Cold climate conditions of the easternmost region of Friuli-Venezia Giulia bordering non-endemic territories of Slovenia, are probably less favorable to L infantum transmission.

Comparing copromicroscopy to intestinal scraping to monitor red fox intestinal helminths with zoonotic and veterinary importance.

The red fox acts as reservoir for several helminthic infections which are of interest for both public and animal health. Huge efforts have been made for the assessment of the sensitivity of coprological tests for the detection of Echinococcus multilocularis, while less attention has been paid to other helminthic species. This study aimed at assessing the performance of two copromicroscopic techniques in the detection and prevalence estimation of gastrointestinal helminths in the red fox. Helminths were isolated from the small intestines of 150 red foxes from Bolzano province, Italy, with a scraping, filtration and counting technique (SFCT) and morphologically identified. Rectal contents were collected and submitted to simple flotation (FT) and, only for Taenids, a method based on the concentration of eggs and identification with multiplex PCR (CMPCR). Using SFCT as a reference standard, we estimated the sensitivity of the copromicroscopic tests. Three species of nematodes (namely, Toxocara canis, Uncinaria stenocephala and Pterygodermatites sp.) and five species of cestodes (E. multilocularis, Taenia crassiceps, T. polycantha, Hydatigera taeniaeformis, Mesocestoides sp.) were identified with SFCT, whereas eggs referable to the same taxa were detected with fecal diagnostics, except for Pterygodermatites sp. and Mesocestoides sp. The sensitivity of FT was low for all taxa, ranging from 9.8 to 36.3%, with lower values for Taeniidae. CMPCR was confirmed to perform better for the detection of Taeniidae eggs (23.5%) and the multiplex PCR on retrieved eggs was effective in the identification of the species. A meta-analysis of literature also suggested that our results are consistent with existing data, indicating that copromicroscopy tends to underestimate the prevalence of helminthic infections. The extent of such underestimation varies with taxon, being higher at high prevalence levels, in particular for cestodes. Irregular dynamics of egg shedding, and routine deep freezing of red fox feces may explain the frequency of false negatives with copromicroscopy. Low sensitivity of copromicroscopic tests should be accounted for when estimating prevalence and when defining the correct sample size for the detection of the parasites.