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Abstract Background Rhipicephalus sanguineus sensu lato (s.l.) and Ctenocephalides felis are among the most important year-round ectoparasites of dogs. The persistent efficacy of one treatment with fluralaner injectable suspension (Bravecto® 150 mg/ml powder and solvent for suspension for dogs, referred to as Bravecto® injectable) was investigated in a negative-controlled, randomised, partially blinded 12-month laboratory study. Methods A total of 20 dogs were randomly allocated to two equal groups (treatment and control). Treatment-group dogs were injected subcutaneously on study day 0 with the investigational veterinary product at the recommended dose of 15 mg fluralaner/kg body weight (0.1 mL/kg), whereas the control group dogs received saline solution (0.1 mL/kg). Each dog was infested with 50 (25 female, 25 male) adult R. sanguineus s.l. and 100 adult C. felis 2 days before treatment, 5 and 28 days after treatment, and then once monthly for a 12-month period. Live tick and flea counts were performed 48 h after treatment or subsequent infestation, respectively. Efficacy was determined by comparing arithmetic means of the treatment group tick and flea counts with those of the control group. Infestation was considered adequate if at least 25.0% of ticks and 40.0% of fleas were recovered from at least six dogs in the control group at the respective assessment times. Results Adequate R. sanguineus s.l. and C. felis infestations of control group dogs were observed at each time point. Arithmetic mean treatment group values were significantly lower than those of the control group at all time points. The immediate efficacy when treating existing infestations of R. sanguineus s.l. and C. felis (infestation 2 days before treatment), was 49.7% and 89.7%, respectively. The persistent efficacy against post-treatment re-infestations was 94.4–100% against R. sanguineus s.l. and 92.2–100% against C. felis. Seven dogs in the control group developed flea allergy dermatitis due to the repeated re-infestations over the study period, whereas no dogs in the treatment group were affected. No clinically relevant side effects were observed over the entire study period. Conclusions The fluralaner injectable suspension (Bravecto® injectable) provides 1 year of efficacy against R. sanguineus s.l. and C. felis infestations in dogs following a single treatment, allowing once-yearly treatment, which can significantly improve owner compliance with year-round protection of dogs. Graphical Abstract

Abstract Background Infections with (tricho-)strongyles, Dictyocaulus viviparus or Fasciola hepatica have been shown to reduce milk production in dairy cows. However, the current published studies focused on one single helminth infection by neglecting helminth co-infections and their possible (additive) effects on host performance. Hence, for the first time, we investigated differences in the impact of patent helminth co-infections versus mono-infections on milk production parameters in individual cows. Methods A total of 1583 dairy cows from 27 herds were included in this study. Faecal samples were examined in 2015 and 2021/2022 to determine the number of eggs/larvae per gram faeces for (tricho-)strongyles, D. viviparus, F. hepatica and rumen flukes. The cows were classified as non-infected, mono-infected and co-infected. Linear mixed models were applied to analyse the association between infection status (non-infected vs. mono-infected vs. co-infected) with milk yield, milk protein and milk fat content by including potential confounders. Results Infections with (tricho-)strongyles, D. viviparus, F. hepatica and rumen flukes were detected in 100%, 28.6%, 50.0% and 21.4% of herds, and 27.4%, 2.6%, 10.8% and 0.8% of faecal samples in 2015, while 100%, 0.0%, 86.7% and 60.0% of herds and 52.3%, 0.0%, 13.3% and 26.8% of faecal samples were positive in 2021/2022. Co-infections with two or more helminth taxa were detected in 74.4% of herds and 5.0% of faecal samples in 2015, and in 93.3% of herds and 21.7% of faecal samples in 2021/2022. The correlations between strongyle EPG, D. viviparus LPG and F. hepatica EPG were significantly positive in 2015. Significantly higher mean EPGs were identified in 2015 in faecal samples presenting co-infections with F. hepatica and one or two other helminth taxa than in faecal samples presenting F. hepatica mono-infections (P = 0.013). Although expected, the infection status (mono- or co-infected) had no significant impact on milk yield, milk protein and milk fat content in the linear mixed model analyses based on individual faecal examinations. Conclusions Patent helminth co-infections had no additive detrimental impact on milk production parameters in the present study. This might be a result of presumably low worm burdens, but should be confirmed in future studies. Graphical Abstract

IntroductionInfections with Fasciola hepatica and Ostertagia ostertagi impinge upon dairy cow health and welfare and represent a major economic factor in livestock industry. Control measures largely rely on the use of anthelminthic drugs. However, reports of anthelmintic resistance necessitate sustainable approaches. Farmer characteristics such as attitude and personality are crucial for the implementation of control strategies and on-farm practices.MethodsIn the present study, the HEXACO (Honesty-Humility, Emotionality, eXtraversion, Agreeableness, Conscientiousness, Openness to experience) model of personality structure, which conceptualizes human personality, was used to evaluate the relationship of farmer aspects with on-farm bulk tank milk seropositivity for F. hepatica and O. ostertagi. Moreover, information on farm structure, housing, management, and farmers’ attitude was collected in a face-to-face interview. Farm-level seropositivity for F. hepatica and O. ostertagi was predicted via elastic net regression.ResultsOut of 193 farms housing 8,774 cows in the German Federal State of Bavaria, 47 farms (24.4%) were seropositive for F. hepatica, 77 farms (39.9%) for O. ostertagi, and 42 farms (21.8%) for both endoparasites. The model for F. hepatica seropositivity selected the covariates pasture access, O. ostertagi seropositivity, higher farmer conscientiousness, and organic farming as relevant predictors. Seropositivity for O. ostertagi was predicted by F. hepatica seropositivity, pasture access, organic farming, and farmers being neutral regarding their satisfaction with animal health on their farm. Higher values for the HEXACO factors extraversion and emotionality were inversely associated with O. ostertagi seropositivity.DiscussionThe present work emphasizes the importance of farmer traits in regard to animal health and parasite occurrence. For the effective acceptance and implementation of sustainable control strategies for livestock helminth infections, it is crucial to consider these aspects to holistically address the challenges of managing parasitic diseases. Moreover, tailored communication strategies can be developed incorporating the understanding of individual stockman characteristics and subsequently ensuring encouragement of stakeholders.

Abstract Background Aelurostrongylus abstrusus is one of the most important respiratory nematodes of felines. Infections may lead to respiratory clinical signs with varying severity or even death, emphasizing the need for preventive treatment of cats with outdoor access to circumvent patent infections. Methods Therefore, the preventive efficacy of a spot-on formulation of 280 mg/ml fluralaner and 14 mg/ml moxidectin (Bravecto® Plus spot-on solution for cats, MSD) against A. abstrusus was evaluated in a negative controlled, randomized and partially blinded efficacy study with 28 purpose-bred cats in a non-terminal design. In three different treatment regimes, the minimum recommended dose of 40 mg fluralaner and 2.0 mg moxidectin/kg bodyweight (BW) was administered once at 12, 8 or 4 weeks (study group G1, G2 and G3, respectively) prior to experimental infection with 300 third-stage A. abstrusus larvae, while G4 served as placebo-treated control. Results From 30 to 46 days post infection (dpi; SD 114 to 130), faeces were sampled to monitor first-stage larvae (L1) excretion for efficacy determination. Secondary efficacy criteria, including respiratory parameters, serological antibody levels and computed tomography (CT) findings, were assessed once before enrolment (SD −7 to −1) and before infection (SD 75 to 83). After infection, CT evaluation was performed once at 47–50 dpi (SD 131 to 134), and respiratory parameters and antibody levels were regularly assessed twice or once a week, respectively (1 up to 78 dpi, SD 85 up to 162). All animals in the control group excreted L1 by 33–37 dpi and remained positive throughout the study period from 41 to 46 dpi (SD 125 to 130). In the treatment groups, only one animal each of G1 and G2 excreted L1 at two consecutive days, and four cats of G1, two of G2 and three of G3 were positive on single occasions. While the geometric mean (GM) of the maximum number of excreted L1 per 5 g of faeces was 7380.89 in the control group (G4), GMs were significantly lower in the treatment groups with 1.63 in G1, 1.37 in G2 and 0.79 in G3. Thus, based on GMs, the reduction in excreted L1 exceeded 99.9% in all three treatment groups. Based on CT severity scores, all lungs of the animals of the control group showed severe pulmonary changes post infection, whereas lungs of the cats of the treatment groups were either unaltered (4 animals), mildly (11 animals), or moderately altered (5 animals). Moreover, seroconversion was observed in all cats of the control group, but not in those of the treatment groups. Conclusions The combination of diagnostic methods used in this non-terminal study yielded coherent and reliable results. A single administration of Bravecto® Plus spot-on solution for cats was well tolerated and effective in the prevention of aelurostrongylosis for at least 12 weeks. Graphical Abstract

Abstract Background Soil-transmitted helminths (STH) infect more than a quarter of the world’s human population. In the absence of vaccines for most animal and human gastrointestinal nematodes (GIN), treatment of infections primarily relies on anthelmintic drugs, while resistance is a growing threat. Therefore, there is a need to find alternatives to current anthelmintic drugs, especially those with novel modes of action. The present work aimed to study the composition and anthelmintic activity of Combretum mucronatum leaf extract (CMLE) by phytochemical analysis and larval migration inhibition assays, respectively. Methods Combretum mucronatum leaves were defatted with petroleum ether and the residue was extracted by ethanol/water (1/1) followed by freeze-drying. The proanthocyanidins and flavonoids were characterized by thin layer chromatography (TLC) and ultra-high performance liquid chromatography (UPLC). To evaluate the inhibitory activity of this extract, larval migration assays with STH and GIN were performed. For this purpose, infective larvae of the helminths were, if necessary, exsheathed (Ancylostoma caninum, GIN) and incubated with different concentrations of CMLE. Results CMLE was found to be rich in flavonoids and proanthocyanidins; catechin and epicatechin were therefore quantified for standardization of the extract. Data indicate that CMLE had a significant effect on larval migration. The effect was dose-dependent and higher concentrations (1000 µg/mL) exerted significantly higher larvicidal effect (P 1000 µg/mL). Likewise, CMLE was active against most infective larvae of soil-transmitted ruminant GIN, except for Cooperia punctata. Trichostrongylus colubriformis was most sensitive to CMLE (IC50 = 2.1 µg/mL) followed by Cooperia oncophora (IC50 = 27.6 µg/mL), Ostertagia ostertagi (IC50 = 48.5 µg/mL), Trichostrongylus axei (IC50 = 54.7 µg/mL), Haemonchus contortus (IC50 = 145.6 µg/mL), and Cooperia curticei (IC50 = 156.6 µg/mL). Conclusions These results indicate that CMLE exhibits promising anthelmintic properties against infective larvae of a large variety of soil-transmitted nematodes. Graphical Abstract

Understanding the impact of parasites on wildlife populations is an important aspect of conservation management. However, research on ectoparasites in wildlife can be difficult, as examinations of live animals which are not habituated to human handling are often impossible. The European wildcat (Felis silvestris) is a strictly protected wildlife species whose population has been recovering in Germany in recent decades. Several studies from different European countries have investigated the parasitological status of European wildcat populations. However, most of these studies assessed endoparasite infections, whereas ectoparasite infestations have often been neglected. To fill this knowledge gap for wildcats in Germany, 131 dead found specimens were examined for ectoparasites by macroscopic and microscopic examination of the fur and the ear canals. Infestation with ectoparasites was present in 84.0% (110/131) of the wildcats. Ticks showed the highest prevalence with 72.5% (95/131) of wildcats infested, with 49.6% (65/131) infested with Ixodes ricinus and 36.6% (48/131) with Ixodes hexagonus/canisuga. A total of 27.5% (36/131) of the wildcats were positive for at least one flea species. Of the nine different flea species identified by morphology and/or molecular analyses, Ceratophyllidae were most common (16.8% [22/131]), with Ceratophyllus sciurorum confirmed on 12.2% (16/131) and Nosopsyllus fasciatus on 1.5% (2/131) animals, followed by Pulex irritans (5.3% [7/131]), Spilopsyllus cuniculi (3.8% [5/131]), Chaetopsylla spp. (3.1% [4/131]) (2/131 Chaetopsylla trichosa and 1/131 Chaetopsylla globiceps), Ctenocephalides felis (1.5% [2/131]), Archaeopsylla erinacei (1.5% [2/131]) and Ctenophthalmus baeticus (0.8% [1/131]). Further, 23.7% (31/131) of the wildcats harboured mites, identified as Trombicula autumnalis in 12.2% (16/131) and Otodectes cynotis in 4.8% (6/124) of cases. The only louse species detected was Felicola hercynianus with a prevalence of 2.3% (3/131). Infestation intensities ranged from 1 to 86 ticks, 1–49 fleas, 1–1896 mites, and 1–92 F. hercynianus per wildcat. This study demonstrates that a variety of ectoparasites infests wildcats in Germany, but they do not seem to have a serious impact on the general health of wildcats, as judged by the hosts' mostly good or very good nutritional condition. In addition, the potential risk to domestic cats (Felis catus) and humans posed by the wildcats’ ectoparasites, appears to be low but present.

Blood parasites of the genus Leucocytozoon commonly infect many bird species worldwide and are particularly prevalent in birds of prey. As a vector-borne parasitic disease, the infection occurrence overlaps with that of the dominant vectors: blackflies (Diptera, Simuliidae). These blood-sucking insects are dependent on habitats with flowing freshwaters for the development of their larval stages. We investigated the correlation between the proximity to flowing waters and Leucocytozoon infection probability in common buzzard (Buteo buteo) broods, as well as the occurrence of adult blackflies directly at the nests. In addition, we investigated the survival of captured simuliids in relation to host infection intensity. In total in 2019, we examined 112 different nests, including 297 common buzzard nestlings, with a Leucocytozoon prevalence of 56.6% among the nestlings and of 80.3% at brood level. We found no significant association of Leucocytozoon infection probability with nestling age, the distance to the nearest stream and the sum of the length of streams within a radius of 200 and 1000 m around each nest. The number of blackflies caught around the nest showed a tentative correlation with the probability of Leucocyozoon infection of the nestlings. Among the subsample of 218 blackfly individuals that survived day one after capture, survival averaged 6.2 days. Our results suggest that Leucocytozoon transmission is complex and requires consideration of many factors, related to habitat and vector prevalence, especially given their temporal variation.

Madagascar's lemur populations are declining in dwindling habitats due to anthropogenic expansion and changing climatic conditions. Gastrointestinal parasites can be important indicators to assess the health status of threatened species. However, parasites, hosts and the environment are connected in complex interactions. The present study aimed to disentangle the impact of seasonal and several host-specific factors (sex, species, age, reproductive status, and body mass) on endoparasitism in two small-bodied, co-occurring lemur species (Microcebus murinus and Microcebus ravelobensis) in the Ankarafantsika National Park. Helminth prevalence and egg shedding intensity was investigated via copromicroscopic examination of 810 fecal samples that were obtained from 178 individuals across an 11-month period with a longitudinal approach via repeated captures in a 30.6 ha forest area. Both mouse lemur hosts shed seven morphologically distinct egg types (assigned to Subulura baeri, unidentified Enterobiinae, Spirura sp., Lemuricola sp., two Hymenolepididae spp., one unidentified ascarid). Postmortem examination of two deceased individuals enabled assignment of adult worms to egg morphotypes of S. baeri, Spirura sp. and one Hymenolepididae sp., supported by molecular analysis. A significant seasonal variation was observed in the occurrence of the three most common helminth species S. baeri (total prevalence 71%), unidentified Enterobiinae (46%) and Spirura sp. (38%), with a higher likelihood of infection with advancing dry season. Neither host species, sex nor reproductive status had a significant effect on gastrointestinal helminth infections. Host body mass showed pronounced seasonal changes but did not differ significantly between infected and non-infected individuals. The pathogenic effects of gastrointestinal helminths therefore likely remained within compensable limits in the studied mouse lemur populations. Our findings highlight the prominent influence of seasonal changes on helminth communities. The results of combined morphologic and genetic approaches can furthermore help to overcome limitations of parasite identification via copromicroscopy by linking egg morphology to DNA sequences.

BackgroundThe standard diagnosis of Ascaris lumbricoides and other soil-transmitted helminth (STH) infections relies on the detection of worm eggs by copromicroscopy. However, this method is dependent on worm patency and shows only limited accuracy in low-intensity infection settings. We aimed to decipher the diagnostic accuracy of different antibodies using various Ascaris antigens in reference to copromicroscopy and quantitative PCR (qPCR), four months after national STH preventative chemotherapy among school children in western Kenya.MethodologySTH infection status of 390 school children was evaluated via copromicroscopy (Kato-Katz and mini-FLOTAC) and qPCR. In parallel, Ascaris-specific antibody profiles against larval and adult worm lysates, and adult worm excretory-secretory (ES) products were determined by enzyme-linked immunosorbent assay. Antibody cross-reactivity was evaluated using the closely related zoonotic roundworm species Toxocara cati and Toxocara canis. The diagnostic accuracy of each antibody was evaluated using receiver operating curve analysis and the correspondent area under the curve (AUC).Principal findingsAscaris was the predominant helminth infection with an overall prevalence of 14.9% (58/390). The sensitivity of mini-FLOTAC and Kato-Katz for Ascaris diagnosis reached only 53.5% and 63.8%, respectively compared to qPCR. Although being more sensitive, qPCR values correlated with microscopic egg counts (R = -0.71, PConclusionIgG1 antibody responses against adult Ascaris-ES products hold a promising potential for complementing the standard fecal and molecular techniques employed for monitoring Ascaris infections. This is of particular importance in the context of deworming programs as the antibody diagnostic accuracy was independent of egg counts.

Vector-borne diseases (VBDs) are considered as (re-)emerging, but information on the transmission cycles and wildlife reservoirs is often incomplete, particularly with regard to urban areas. The present study investigated blood samples from European hedgehogs (Erinaceus europaeus) presented at wildlife rehabilitation centres in the region of Hanover. Past exposure to B. burgdorferi sensu lato (s.l.) and tick-borne encephalitis virus (TBEV) was assessed by serological detection of antibodies, while current infections with Borrelia spp., Anaplasma phagocytophilum, Rickettsia spp., Neoehrlichia mikurensis, Bartonella spp., Babesia spp. and Spiroplasma ixodetis were investigated by (q)PCR. Of 539 hedgehogs tested for anti-Borrelia antibodies, 84.8% (457/539) were seropositive, with a higher seropositivity rate in adult than subadult animals, while anti-TBEV antibodies were detected in one animal only (0.2%; 1/526). By qPCR, 31.2% (168/539) of hedgehog blood samples were positive for Borrelia spp., 49.7% (261/525) for A. phagocytophilum, 13.0% (68/525) for Bartonella spp., 8.2% for S. ixodetis (43/525), 8.0% (42/525) for Rickettsia spp. and 1.3% (7/525) for Babesia spp., while N. mikurensis was not detected. While further differentiation of Borrelia spp. infections was not successful, 63.2% of the A. phagocytophilum infections were assigned to the zoonotic ecotype I and among Rickettsia spp. infections, 50.0% to R. helvetica by ecotype- or species-specific qPCR, respectively. Sequencing revealed the presence of a Rickettsia sp. closely related to Rickettsia felis in addition to a Bartonella sp. previously described from hedgehogs, as well as Babesia microti and Babesia venatorum. These findings show that hedgehogs from rehabilitation centres are valuable sources to identify One Health pathogens in urban areas. The hedgehogs are not only exposed to pathogens from fleas and ticks in urban areas, but they also act as potent amplifiers for these vectors and their pathogens, relevant for citizens and their pets.

For years, there has been an increasing interest in natural alternatives to the conventional coccidiostats applied as feed additives, which have been used for decades to prevent coccidiosis in poultry and fattening rabbits. This study aimed to compare the possible anticoccidial effects of oregano oil to the established substance diclazuril in growing rabbits. The control group (CG) received a non-supplemented basal compound feed, to which either diclazuril (1 mg/kg; DG) or oregano oil (75 mg/kg; OG) was added. In each of the three trials, subgroups of 50 rabbits each were assigned to one of the three experimental groups (CG, DG and OG). Natural Eimeria infection was monitored weekly by fecal oocyst counts and Eimeria species identification following sporulation. Additionally, the performance parameters were determined at the middle and the end of the trials, and the deceased rabbits were subjected to necropsy. Neither oocyst excretion nor the performance parameters differed significantly between the three experimental groups. Eimeria media, Eimeria magna, Eimeria perforans and Eimeria exigua were identified as the occurring species. The highest animal losses (16.0%) occurred in the OG, while the losses were 12.7% in the DG and 12.0% in the CG. However, these differences were not statistically significant. Overall, neither diclazuril nor oregano oil was superior to the non-supplemented feed. This underlines the importance of diagnostics, as this study’s results indicate that in the absence of the highly pathogenic Eimeria species, economic rabbit rearing and fattening is achievable without the use of coccidiostats.

Abstract Background Changing geographical and seasonal activity patterns of ticks may increase the risk of tick infestation and tick-borne pathogen (TBP) transmission for both humans and animals. Methods To estimate TBP exposure of dogs and cats, 3000 female I. ricinus from these hosts were investigated for Anaplasma phagocytophilum and Borrelia species. Results qPCR inhibition, which was observed for ticks of all engorgement stages but not questing ticks, was eliminated at a template volume of 2 µl. In ticks from dogs, A. phagocytophilum and Borrelia spp. prevalence amounted to 19.0% (285/1500) and 28.5% (427/1500), respectively, while ticks from cats showed significantly higher values of 30.9% (464/1500) and 55.1% (827/1500). Accordingly, the coinfection rate with both A. phagocytophilum and Borrelia spp. was significantly higher in ticks from cats (17.5%, 262/1500) than dogs (6.9%, 104/1500). Borrelia prevalence significantly decreased with increasing engorgement duration in ticks from both host species, whereas A. phagocytophilum prevalence decreased only in ticks from dogs. While A. phagocytophilum copy numbers in positive ticks did not change significantly over the time of engorgement, those of Borrelia decreased initially in dog ticks. In ticks from cats, copy numbers of neither A. phagocytophilum nor Borrelia spp. were affected by engorgement. Borrelia species differentiation was successful in 29.1% (365/1254) of qPCR-positive ticks. The most frequently detected species in ticks from dogs were B. afzelii (39.3% of successfully differentiated infections; 70/178), B. miyamotoi (16.3%; 29/178), and B. valaisiana (15.7%; 28/178), while B. afzelii (40.1%; 91/227), B. spielmanii (21.6%; 49/227), and B. miyamotoi (14.1%; 32/227) occurred most frequently in ticks from cats. Conclusions The differences in pathogen prevalence and Borrelia species distribution between ticks collected from dogs and cats may result from differences in habitat overlap with TBP reservoir hosts. The declining prevalence of A. phagocytophilum with increasing engorgement duration, without a decrease in copy numbers, could indicate transmission to dogs over the time of attachment. The fact that this was not observed in ticks from cats may indicate less efficient transmission. In conclusion, the high prevalence of A. phagocytophilum and Borrelia spp. in ticks collected from dogs and cats underlines the need for effective acaricide tick control to protect both animals and humans from associated health risks. Graphical Abstract

Abstract Background Hyalomma marginatum and H. rufipes are two-host tick species, which are mainly distributed in southern Europe, Africa to central Asia but may also be found in Central and Northern Europe through introduction by migratory birds. Methods Ticks were collected while feeding or crawling on animals and humans, or from the environment, in different regions in Germany, between 2019 and 2021 in a citizen science study and from 2022 to 2023 in the wake of this study. Results From 2019 to 2023, a total of 212 Hyalomma adult ticks were detected in Germany. This included 132 H. marginatum and 43 H. rufipes ticks sent to research institutions and 37 photographic records that were only identified to genus level. The number of detected ticks varied over the years, with the highest number of 119 specimens recorded in 2019, followed by 57 in 2020. Most of the specimens were collected from horses, while some were collected from other animals, humans or found crawling on human clothes or other objects inside or outside houses. The screening of 175 specimens for Crimean-Congo hemorrhagic fever virus and of 132 specimens for Babesia/Theileria spp. by PCR gave negative results, while human-pathogenic Rickettsia were detected in 44% (77/175) of the total samples. Subsequent amplicon sequencing and phylogenetic analysis of representative samples determined the species of 41 Rickettsia aeschlimannii and one R. slovaca sequences. Conclusions Analysis of climatic factors indicated a significantly higher probability of Hyalomma occurrence at locations with higher average spring temperature during the years 2019 and 2020 compared to randomly generated pseudo-absence locations. Dry and hot conditions probably facilitated Hyalomma nymphs’ survival and molting into adults during these years. Graphical Abstract

Abstract Background The European hedgehog (Erinaceus europaeus) is known for high levels of ectoparasitism that not only represents a health risk for the animals themselves, but also for pet animals and humans as hedgehogs are frequently taken into human care. In the present study, patterns of ectoparasite infestation were assessed in hedgehogs taken into care at northern German animal rehabilitation centres. Methods Ectoparasites (ticks, fleas and mites) of 498 hedgehogs were collected over a period of 3 years from July 2018 to May 2021. Species were identified based on morphological characteristics and also via amplification and sequencing of the partial cytochrome c oxidase subunit 2 (COX-2) gene for fleas of the family Ceratophyllidae. Seasonal changes in infestation patterns as well as correlations with animal age, body weight and health status were assessed using generalised linear models. Results Infestation with ticks, fleas and mites occurred throughout the year. Overall, 86.5% (431/498) of the examined hedgehogs were infested with ticks, 91.4% (455/498) with fleas and 17.7% (88/498) with mites. Ixodes ricinus and Ixodes hexagonus/Ixodes canisuga were the most common tick species detected, with the additional occurrence of one Ixodes frontalis. Significant seasonal changes were observed for I. ricinus, but not for I. hexagonus/I. canisuga. Additionally, I. ricinus nymph prevalence declined significantly as of 2020, probably as a consequence of the climate change-related drought as of 2018. In hedgehogs with flea infestations, Archaeopsylla erinacei, Ceratophyllus sciurorum, Nosopsyllus fasciatus and Ctenocephalides felis were identified. In all cases of mite infestation, Caparinia tripilis was detected, in addition to specimens of the family Macronyssidae and free-living mites of the family Acaridae. Statistical analyses showed correlations regarding the factors month, year, body weight and age, but no correlation was evident regarding the health status of the animals. Conclusions With a detected infestation rate of 98.6%, almost all of the examined hedgehogs were infested with at least one ectoparasite species. The seasonal activity patterns of the different ectoparasite species together with the complex annual cycle of hedgehogs lead to different seasonal patterns in ectoparasite prevalence and infestation intensities. Due to the risk of transmission of zoonotic pathogens as well as the possible negative impact on the host itself, hedgehogs should be treated against ectoparasites when taken into care facilities. Graphical Abstract

Abstract Toxoplasma (T.) gondii is an obligate intracellular parasite with a worldwide distribution. Congenital infection can lead to severe pathological alterations in the brain. To examine the effects of toxoplasmosis in the fetal brain, pregnant guinea pigs are infected with T. gondii oocysts on gestation day 23 and dissected 10, 17 and 25 days afterwards. We show the neocortex to represent a target region of T. gondii and the parasite to infect neural progenitor cells (NPCs), neurons and astrocytes in the fetal brain. Importantly, we observe a significant reduction in neuron number at end-neurogenesis and find a marked reduction in NPC count, indicating that impaired neurogenesis underlies the neuronal decrease in infected fetuses. Moreover, we observe focal microglioses to be associated with T. gondii in the fetal brain. Our findings expand the understanding of the pathophysiology of congenital toxoplasmosis, especially contributing to the development of cortical malformations.

Abstract Background The ticks Ixodes ricinus and Dermacentor reticulatus are two of the most important vectors in Europe. Climate niche modelling has been used in many studies to attempt to explain their distribution and to predict changes under a range of climate change scenarios. The aim of this study was to assess the ability of different climate niche modelling approaches to explain the known distribution of I. ricinus and D. reticulatus in Europe. Methods A series of climate niche models, using different combinations of input data, were constructed and assessed. Species occurrence records obtained from systematic literature searches and Global Biodiversity Information Facility data were thinned to different degrees to remove sampling spatial bias. Four sources of climate data were used: bioclimatic variables, WorldClim, TerraClimate and MODIS satellite-derived data. Eight different model training extents were examined and three modelling frameworks were used: maximum entropy, generalised additive models and random forest models. The results were validated through internal cross-validation, comparison with an external independent dataset and expert opinion. Results The performance metrics and predictive ability of the different modelling approaches varied significantly within and between each species. Different combinations were better able to define the distribution of each of the two species. However, no single approach was considered fully able to capture the known distribution of the species. When considering the mean of the performance metrics of internal and external validation, 24 models for I. ricinus and 11 models for D. reticulatus of the 96 constructed were considered adequate according to the following criteria: area under the receiver-operating characteristic curve > 0.7; true skill statistic > 0.4; Miller’s calibration slope 0.25 above or below 1; Boyce index > 0.9; omission rate

Abstract Background The lungworm Aelurostrongylus abstrusus infects wild and domestic feline species worldwide and is considered a primary respiratory parasite of cats. Definitive diagnosis is based on the identification of first-stage larvae (L1s) released in faeces approximately 5 to 6 weeks after infection. More recently, serology has been shown to be a diagnostic alternative for A. abstrusus infection in cats. The present study aimed at evaluating the diagnostic performance of serological antibody detection compared to faecal examination for A. abstrusus infection in a population of cats with known infection status from endemic areas in Italy and to identify factors (larval scores, age, co-infections with other helminths) that may influence test sensitivity and specificity of serology. Methods All cats resulting positive using the Baermann technique (n = 78) were tested with the A. abstrusus ELISA. An additional 90 serum samples from cats living in three geographical areas with infection prevalence > 10%, but that resulted negative on Baermann, were also tested. Results Among 78 cats copromicroscopically positive for L1s of A. abstrusus (Group 1), 29 (37.2%) were seropositive in ELISA. Of the 90 cats from Group 2 (cats living in three geographical areas in Italy with A. abstrusus prevalence > than 10%, but negative on Baermann examination), 11 (12.2%) were positive on ELISA. The overall seroprevalence was 23.8%. There was no statistical difference either between average optical density (OD) values of cats excreting > 100 L1s vs. cats excreting

The ornate dog tick Dermacentor reticulatus, vector of Babesia canis, has shown a considerable range expansion in several European countries. Previously, only few areas in Germany were recognised as endemic for B. canis, but a marked increase in autochthonous canine babesiosis cases and spread to new areas has been noted recently. To better assess the current risk for dogs, the present study screened 5913 specimens of D. reticulatus from all over Germany, collected in the frame of a Citizen Science study during 2019–2023. Moreover, 343 Dermacentor marginatus ticks were also included. Babesia detection was achieved by quantitative real-time PCR (qPCR). Positive samples were confirmed by sequencing. Moreover, a MGB-probe-based triplex qPCR was established to detect and distinguish between the canine Babesia spp. relevant in Europe, i.e. B. canis, Babesia vogeli and Babesia gibsoni. Overall, B. canis DNA was detected in five D. reticulatus specimens (0.08%). Two of the B. canis-positive ticks originated from areas previously known as endemic for canine babesiosis, namely from the area of Freiburg im Breisgau, federal state of Baden-Wuerttemberg, and from the district St. Wendel, federal state of Saarland. Three further B. canis-positive ticks were detected in districts not yet recognised as endemic, one each in the district of Mansfeld-Suedharz, federal state of Saxony-Anhalt, the district of Ravensburg, federal state of Baden-Wuerttemberg and in the city of Fürth, federal state of Bavaria. However, the tick in Fürth was found on a dog who had returned from a trip to the Breisgau region on the previous day, indicating translocation of the specimen out of this well-known endemic focus. The geographical distribution of the positive samples shows that B. canis is currently spreading in Germany, particularly via dogs travelling within the country, increasing the infection risk throughout the country. Important measures to contain a further spread of the pathogen include comprehensive year-round tick prophylaxis with licensed acaricides, not only to protect the individual pet, but also the entire dog population. Moreover, screening of dogs entering Germany from B. canis-endemic countries is required and any treatment should aim at pathogen elimination by use of appropriate imidocarb dosages.

The hard tick Ixodes ricinus transmits a variety of zoonotic pathogens, including Babesia divergens, the most common cause of bovine babesiosis in northern Europe. In endemic areas, cattle are rarely clinically affected, as animals up to the age of nine months are resistant against relevant clinical disease and develop protective premunity. However, outbreaks in immunologically naïve herds may lead to considerable losses. Such an outbreak with a high mortality rate occurred in 2018 on a northern German beef production farm, as previously reported. The present study provides an update on the epidemiological situation and management strategy of the farm. In spring 2022, blood samples were taken from 46 animals for PCR and serological testing before pasture turnout. Although no clinical cases had been noticed since 2019, B. divergens DNA was detected by quantitative real-time PCR (qPCR), followed by amplification and sequencing of the 18S rRNA gene, in 6.5% (3/46) of cattle blood samples. Presence of anti-B. divergens antibodies was confirmed in 26.1% (12/46) of animals, while further 10.9% (5/46) had a borderline antibody titre. The antibody status of 23 of these animals had already been determined in 2018 and/or 2020, revealing fluctuating titre patterns indicative of repeated pathogen exposure. Moreover, 457 questing I. ricinus specimens collected on the farm’s pastures and 83 I. ricinus specimens detached from cattle were screened for Babesia spp. DNA by qPCR, followed by 18S rDNA amplification and sequencing. Endemisation of B. divergens was confirmed by 0.9% (4/457) positive questing I. ricinus, while the ticks detached from cattle were Babesia-negative. The farm’s management strategy includes annual metaphylactic treatment with imidocarb dipropionate during the main tick exposure period in spring. However, the antibody titre fluctuations and the persistent infections at the end of the housing period indicate that the absence of clinical disease is primarily due to a rising level of premunity. Metaphylactic treatment with imidocarb seems to be a suitable management option to protect newly acquired immunologically naïve animals. The endemisation of B. divergens is also of public health significance, as the pastures are located close to a tourist destination in a popular hiking area.

Sucking lice live in intimate association with their hosts and often display a high degree of host specificity. The present study investigated sucking lice of the genus Lemurpediculus from six mouse lemur (Microcebus) and two dwarf lemur (Cheirogaleus) species endemic to the island of Madagascar, considered a biodiversity hotspot. Louse phylogenetic trees were created based on cytochrome C oxidase subunit I (COI), elongation factor 1α (EF1α) and internal transcribed spacer 1 (ITS1) sequences. While clustering according to host species was generally observed for COI and ITS1, suggesting high host specificity of the examined lice, EF1α sequences alone did not distinguish between lice of different Microcebus species, possibly due to rather recent divergence. As bootstrap support for basal tree structure was rather low, further data are necessary to resolve the evolutionary history of louse-mouse lemur associations. Three new species of sucking lice are described: Lemurpediculus zimmermanni sp. Nov. From Microcebus ravelobensis, Lemurpediculus gerpi sp.nov. from Microcebus gerpi, and Lemurpediculus tsimanampesotsae sp. nov. from Microcebus griseorufus. These new species are compared with all known congeneric species and identifying features are illustrated for all known species of Lemurpediculus.

Equine piroplasmosis (EP) is caused by Theileria (T.) equi and/or Babesia (B.) caballi. The aim was to assess the percentage of positive test results for EP in horses in Europe and to identify risk factors for pathogen contact/infection. This study included results from PCR and competitive enzyme-linked immunosorbent assay testing requested by European veterinarians between 2008 and 2021. Binary bivariate logistic regression was used to analyze risk factors. A total of 4060 horses were included. PCR testing was positive in 9.7% (154/1589), serology for T. equi in 15.2% (393/2591) and for B. caballi in 6.8% (175/2578). The odds of positive serology increased by 6.8% (B. caballi, p = 0.008) and 9.5% (T. equi, p < 0.001) each year. Regionality had a statistically significant impact on PCR (Eastern p = 0.047/OR = 1.605; Southern p = 0.029/OR = 1.451; Central p = 0.007/OR = 0.617) and serological testing for T. equi (Southern p < 0.001/OR = 2.521; Central p < 0.001/OR = 0.537; Northern p = 0.003/OR = 0.462), as well as breeds on seroprevalence of B. caballi (heavy horses: p = 0.016/OR = 2.239) and T. equi (ponies: p = 0.007/OR = 0.340; warmbloods: p = 0.025/OR = 1.602). In conclusion, there was a significant geographical impact on the results of PCR and serology, consistent with known vector habitats. The rising numbers of horses tested serologically positive highlights the importance of surveillance.

Tick-borne encephalitis virus (TBEV) is a tick-transmitted flavivirus, which can infect humans and animals, sometimes even with a fatal outcome. Since many decades, TBEV is endemic in southern Germany, while only sporadic occurrence has been noted in northern parts of the country so far. Nevertheless, autochthonous human clinical cases are increasing in the federal state of Lower Saxony in north-western Germany, and several natural foci of TBEV transmission have recently been detected in this federal state. In order to shed more light on the current distribution of TBEV in Lower Saxony, the present study examined blood samples from wild and domestic animals for antibodies against TBEV. Overall, samples from 4,085 animals were tested by ELISA, including wild boar (N = 1,208), roe deer (N = 149), red deer (N = 61), fallow deer (N = 18), red foxes (N = 9), nutria (N = 9), raccoon dogs (N = 3), raccoons (N = 3), badgers (N = 1), European pine martens (N = 1), horses (N = 574), sheep (N = 266), goats (N = 67), dogs (N = 1,317) and cats (N = 399). Samples with an ELISA result of ≥60 Vienna units (VIEU)/ml were subjected to confirmatory serum neutralization tests (SNT). In total, 343 of 4,085 (8.4%) animals tested positive for anti-TBEV-IgG by ELISA, of which 60 samples were confirmed by SNT. Samples of 89 animals showed a cytotoxic effect in the SNT and were excluded from seroprevalence calculation, resulting in an overall seroprevalence of 1.5% (60/3,996). Seroprevalence was higher among wild animals (wild boar: 2.9% [34/1,190], roe deer: 2.7% [4/149], red deer: 1.7% [1/60], fallow deer: 5.6% [1/18]) than among domestic animals (dogs: 1.1% [15/1,317], horses: 0.8% [4/505], sheep: 0.4% [1/266]). No anti-TBEV-antibodies were detected in the other wild animal species as well as goats and cats. A notable clustering of positive samples was observed in districts where TBEV transmission foci have been described. Further clusters in other districts suggest the existence of so far undetected transmission foci, underlining the fact that both wild and domestic animals are useful sentinels for monitoring the spread of TBEV.

Changing climatic conditions and other anthropogenic influences have altered tick distribution, abundance and seasonal activity over the last decades. In Germany, the two most important tick species are Ixodes ricinus and Dermacentor reticulatus, the latter of which has expanded its range across the country during the past three decades. While I. ricinus was rarely found during the colder months in the past, D. reticulatus is known to be active at lower temperatures. To quantify tick appearance during winter, specimens were monitored in quasi-natural tick plots three times a week. Additionally, the questing activities of these two tick species were observed throughout the year at nine field collection sites that were regularly sampled by the flagging method from April 2020 to April 2022. Furthermore, tick winter activity in terms of host infestation was analysed as part of a nationwide submission study from March 2020 to October 2021, in which veterinarians sent in ticks mainly collected from dogs and cats. All three study approaches showed a year-round activity of I. ricinus and D. reticulatus in Germany. During the winter months (December to February), on average 1.1% of the inserted I. ricinus specimens were observed at the tops of rods in the tick plots. The average questing activity of I. ricinus amounted to 2 ticks/100 m² (range: 1-17) in the flagging study, and 32.4% (211/651) of ticks found infesting dogs and cats during winter 2020/21 were I. ricinus. On average 14.7-20.0% of the inserted D. reticulatus specimens were observed at the tops of rods in the tick plots, while the average winter questing activity in the field study amounted to 23 specimens/100 m² (range: 0-62), and 49.8% (324/651) of all ticks collected from dogs and cats during winter 2020/21 were D. reticulatus. Additionally, the hedgehog tick Ixodes hexagonus was found to infest dogs and cats quite frequently during the winter months, accounting for 13.2% (86/651) of the collected ticks. A generalized linear mixed model identified significant correlations of D. reticulatus winter activity in quasi-natural plots with climatic variables. The combined study approaches confirmed a complementary main activity pattern of I. ricinus and D. reticulatus with climate change-driven winter activity of both species. Milder winters and a decrease of snowfall, and consequently high winter activity of D. reticulatus, among other factors, may have contributed to the rapid spread of this tick species throughout the country. Therefore, an effective year-round tick control is strongly recommended to not only efficiently protect dogs and cats with outdoor access from ticks and tick-borne pathogens (TBPs), but also to limit the further geographical spread of ticks and TBPs to so far non-endemic regions. Further measures, including information of the public, are necessary to protect both, humans and animals, in a One Health approach.

Abstract Background Ticks and tick-borne diseases play a major role in companion animal health. Additionally, the European tick fauna is changing, for instance due to the spread of Dermacentor reticulatus, displaying a higher likelihood of winter activity than Ixodes ricinus. Therefore, we investigated current tick infestations in dogs and cats in Germany and in parts of Austria and the seasonal infestation risk. Methods Overall, 219 veterinary practices were invited to collect ticks from cats and dogs on a monthly basis. Ticks were morphologically identified and female I. ricinus specimens were measured to estimate attachment duration. Results In total, 19,514 ticks, 17,789 (91.2%) from Germany and 1506 (7.7%) from Austria, were received between March 2020 and October 2021, with 10,287 specimens (52.7%) detached from dogs, 8005 from cats (41.0%) and 1222 from other species (6.3%). In Germany, the most common tick species collected from dogs were I. ricinus (78.0%) and D. reticulatus (18.8%), while cats mainly harboured I. ricinus (91.3%) and I. hexagonus (5.5%) and only few D. reticulatus (0.6%). In Austria, collected I. ricinus reached similar proportions in dogs (90.4%) and cats (95.3%), followed by D. reticulatus in both dogs (5.2%) and cats (1.5%), with I. hexagonus (0.9%) collected only marginally from cats. The average infestation intensity amounted to 1.62 ticks/dog and 1.88 ticks/cat. The single to multiple infestation ratio was 79.1% to 20.9% in dogs and 69.0% to 31.0% in cats, with cats being significantly more often multiple infested than dogs, while the proportion of mixed-species infestations was 2.0% for both dogs and cats. The average attachment duration of female I. ricinus specimens amounted to 78.76 h for dogs and 82.73 h for cats. Furthermore, year-round tick exposure was confirmed, with 108 D. reticulatus and 70 I. ricinus received on average per month during December 2020 to February 2021. Conclusions The study shows a year-round tick infestation risk, with activity of both D. reticulatus and I. ricinus during winter, and confirms the widespread occurrence of D. reticulatus in Germany. Additionally, long average attachment durations and frequent multiple infestations underline the need for adequate year-round tick control, even during the winter months. Graphical Abstract

Abstract Background In the last years, research on feline cardio-pulmonary parasites has considerably increased in Europe. Not only domestic cats (Felis catus), but also European wildcats (Felis silvestris) can serve as definitive hosts for these nematodes. The F. silvestris population in Germany has been growing rapidly within the last decades; therefore, the assessment of its cardio-pulmonary parasite status is of importance to unravel whether the wildcat population serves as a substantial reservoir for these nematodes and might pose a health threat to domestic cats. Methods As part of a nature conservation project for European wildcats in the German federal state Rhineland-Palatinate, lungs (n = 128) and hearts (n = 111) of 128 F. silvestris found dead were examined for cardio-pulmonary parasites. All isolated parasites were identified morphologically, and results were confirmed by molecular analysis of a total of 3–11 specimens of each worm species. Results A total of 70.3% (90/128) wildcats were positive for at least one lungworm species. Angiostrongylus chabaudi was most common (53.1% [68/128]), followed by Aelurostrongylus abstrusus (42.2% [54/128]), Troglostrongylus brevior (31.3% [40/128]) and Capillaria spp. (3.1% [4/128]). Of note, about two-thirds of the infected wildcats harboured coinfections. Infection intensities ranged from 1 to 167 nematodes per wildcat. Generalised linear models revealed a strong correlation between A. chabaudi and A. abstrusus infection, and prevalences were higher in adult than in younger wildcats, except for T. brevior. Moreover, the T. brevior prevalence varied significantly with nutritional status. Conclusions This study shows that feline cardio-pulmonary nematodes are common parasites in European wildcats in Germany but do not appear to have a serious impact on the overall health of the population. Due to presumed spillover events via prey, cardio-pulmonary nematodes may circulate between the wildcat population and domestic cats and might therefore pose a health risk to individual domestic cats. Graphical abstract

Abstract Background The porcine roundworm Ascaris suum impairs feed conversion and weight gain, but its effects on intestinal microbiota remain largely unexplored. Methods Modulation of the intestinal microbiota was assessed in pigs that were infected once with 10,000 A. suum eggs and pigs that received a trickle infection (1000 eggs/day over 10 days), compared with a non-infected control group. Six pigs each were sacrificed per group at days 21, 35 and 49 post-infection (p.i.). Faecal samples taken weekly until slaughter and ingesta samples from different intestinal compartments were subjected to next-generation sequencing of the bacterial 16S rRNA gene. Results The results revealed marked differences between the single- and the trickle-infected group. Single infection caused a remarkable but transient decrease in microbial diversity in the caecum, which was not observed in the trickle-infected group. However, an increase in short-chain fatty acid-producing genera in the caecum on day 21 p.i., which shifted to a decrease on day 35 p.i., was common to both groups, possibly related to changes in excretory–secretory products following the parasite’s final moult. Faecal microbial interaction networks were more similar between the single-infected and control group than the trickle-infected group. In addition, a lower degree of similarity over time indicated that A. suum trickle infection prevented microbiota stabilization. Conclusions These different patterns may have important implications regarding the comparability of experimental infections with natural scenarios characterized by continuous exposure, and should be confirmed by further studies. Graphical Abstract

Sarcoptic mange was detected in five free-ranging raccoon dogs (Nyctereutes procyonoides) in the federal state of Schleswig-Holstein, Germany, during a health assessment study of invasive species, including raccoon dogs, carried out between 2021 and 2022. Four raccoon dogs showed severe lesions, including extensive alopecia with thickening and hyperpigmentation of the skin (lichenification). The fifth animal was less affected, showing only thinning of the hair coat in multiple body locations. Skin scrapings were performed and confirmed the presence of Sarcoptes scabiei. Histopathology of the skin revealed diffuse epidermal hyperplasia and hyperkeratosis, mild eosinophilic dermatitis, and varying amounts of intralesional mites. Staphylococcus pseudintermedius and Corynebacterium auriscanis were detected in the skin samples of the affected animals, indicating a secondary bacterial infection. The source of sarcoptic mange remains unclear; interspecies transmission via direct or indirect contact seems likely. Raccoon dogs are therefore a potential vector for sarcoptic mange, and their behaviour could contribute to disease spread and persistence.

In the northernmost German federal state Schleswig-Holstein, populations of European brown hares (Lepus europaeus Pallas, 1778) show diverse densities and varying courses over the years. To examine differences in pathomorphological findings and infectious diseases as possible reasons for varying population dynamics, we assessed 155 hunted hares from three locations in Schleswig-Holstein from 2016 to 2020. We investigated the association of location, year, age, and sex of animals to certain pathomorphological findings and infectious diseases. Frequent pathomorphological findings were intestinal parasites (63.9%), hepatitis (55.5%), nephritis (31.0%), steatitis (23.2%), enteritis (13.5%), and pneumonia (5.2%). Body condition differed significantly between locations, and the prevalence of pneumonia was significantly higher in females. Enteritis was not detected in 2019, when much more juveniles were sampled. Hepatitis and nephritis occurred significantly more often in 2016 and among adults. Additionally, more adults showed hepatitis with concurrent serotitre for European brown hare syndrome virus (EBHSV), while intestinal parasitosis as well as high excretion rates of coccidia were more common in juveniles. Sampled animals showed high infection rates with Eimeria spp. (96.1%), Trichostrongylus spp. (52.0%), Graphidium strigosum (41.2%), and a high seroprevalence (90.9%) for EBHSV, without severe symptoms. This study revealed a low prevalence of infectious pathogens, but a high prevalence of chronic inflammations of unknown origin in the tested brown hare populations. Overall, our results indicate a rather minor importance of infectious diseases for observed population dynamics of analysed hare populations in Schleswig-Holstein.

Abstract Background Parasitic bronchopneumonia in domestic cats in Europe, which can manifest with moderate to severe clinical signs, is frequently caused by Troglostrongylus brevior. Data on epizootiological and clinical relevance of cat troglostrongylosis have been published in the last decade but treatment options are still limited. Promising effectiveness data have been generated from clinical cases and field trials for a spot-on formulation containing 1% w/v moxidectin and 10% w/v imidacloprid (Advocate®, Elanco Animal Health). Therefore, two studies have been conducted to confirm under experimental conditions the efficacy of moxidectin 1% contained in Advocate® for the treatment of cat troglostrongylosis. Methods Sixteen and 20 cats experimentally infected with T. brevior were included in two separate studies, i.e., Study 1 and 2, respectively. Cats were infected with T. brevior third-stage larvae via gastric tube. In both studies cats were randomized to untreated (control, Group 1) and treatment (Group 2) groups. In Study 1 and Study 2, the two groups comprised eight and 10 cats each. Treated cats received Advocate® spot-on twice at a 4-week interval. The primary efficacy criterion was the number of viable adult T. brevior counted at necropsy. Throughout the trial, the fecal shedding of first-stage larvae (L1) was assessed in treated and untreated control cats. Results The experimental model was successful in both studies, as all cats started shedding T. brevior L1 within 25 days post-infection. At necropsy, T. brevior adults were found in 4/8 and 4/10 cats of the control groups in Study 1 and 2, respectively, while none of the treated cats harbored adult worms. The necropsy worm counts in controls did not meet relevant guideline requirements for adequacy of infection, with fewer than six infected cats in the control groups, thus limiting conclusions on treatment efficacy. The fact that 6/8 and 8/10 control cats in Study 1 and 2, respectively, shed L1 up to necropsy while larval shedding ceased in all treated animals after the first treatment provides supporting evidence on the level of efficacy. No remarkable adverse events were recorded in the two studies. Conclusion These results indicate that Advocate® spot-on is a safe and effective option for treating cats infected by T. brevior. Graphical Abstract

Abstract Background Nature conservation with reduced drainage of pastures has been increasingly promoted in agriculture in recent years. However, moisture on pastures is a crucial factor for the development of free-living stages of many parasite species in ruminants. Hence, for the first time, we conducted a field study between 2015 and 2017 at the German North Sea coast to investigate the long-term effect of pasture rewetting (since 2004) on endoparasite infections in sheep and cattle. Methods We examined faecal samples of 474 sheep and 646 cattle from five farms in spring, summer and autumn each year for the presence of endoparasite infections. Animals were kept on conventionally drained, undrained and rewetted pastures. The association between pasture rewetting and endoparasite infection probability was analysed in generalized linear mixed models and including further potential confounders. Results Infection frequencies for gastrointestinal strongyles, Eimeria spp. and Strongyloides papillosus were significantly higher in sheep (62.9%, 31.7% and 16.7%) than in cattle (39.0%, 19.7% and 2.6%). Fasciola hepatica was detected with a frequency of 13.3% in sheep and 9.8% in cattle, while rumen fluke frequency was significantly higher in cattle (12.7%) than in sheep (3.8%). Nematodirus spp., lungworms (protostrongylids, Dictyocaulus viviparus), Moniezia spp., Trichuris spp. and Dicrocoelium dendriticum were identified in less than 7% of samples. Co-infection with more than three endoparasite taxa was present significantly more often in sheep than in cattle. We identified significant positive correlations above 0.2 for excretion intensities between S. papillosus with strongyles, Eimeria spp. and Nematodirus spp. in sheep and between strongyles and Nematodirus spp. in cattle. Pasture rewetting had no long-term effect on endoparasite infections, neither in sheep nor in cattle. Interestingly, F. hepatica infections decreased significantly in sheep and cattle from 2015 (10.9% and 13.9%) to 2017 (1.4% and 2.1%). Conclusions Pasture rewetting for nature conservation did not increase endoparasite infection probability in ruminants in the long term. This finding should be confirmed in ongoing studies aimed at further animal welfare parameters. The rapid decrease in F. hepatica infections over 3 years may suggest climatic impact or competition with rumen flukes in addition to potential anthelmintic treatment after feedback of the results to the farmers. Graphical Abstract

Lyme borreliosis, caused by Borrelia burgdorferi sensu lato (s.l.) spirochaetes, is the most common tick-borne disease (TBD) in the Northern Hemisphere. Rising incidences indicate that its epidemiology may be affected by global changes. Therefore, the current study aimed to assess changes in tick infection rates with Borrelia spp. over a 15-year monitoring period in the city of Hanover, Germany, as a follow-up to previous prevalence studies (years 2005, 2010 and 2015). To assess the epidemiological risk, ticks of the Ixodes ricinus/inopinatus-complex were sampled from April to October 2020 by the flagging method at 10 frequently visited recreation areas in Hanover. Analysis by quantitative real-time PCR of 2100 individual ticks revealed an overall Borrelia prevalence of 25.5% (535/2100). Regarding different tick developmental stages, nymphs showed a significantly lower Borrelia prevalence (18.4% [193/1050]) than adult ticks (32.6% [342/1050]). Comparison with previous years revealed a stable total Borrelia prevalence along with consistent infection rates in the different developmental stages over the 15-year monitoring period. Borrelia species differentiation by Reverse Line Blot was successful in 67.3% of positive ticks collected in 2020, with B. afzelii being the dominating species (59.2% of the differentiated infections), besides B. burgdorferi sensu stricto (s.s.), B. garinii, B. valaisiana, B. spielmanii, B. bavariensis and B. bissettiae and the relapsing fever spirochaete B. miyamotoi. Additionally, the proportion of infections attributed to B. afzelii showed a significant increase in 2020 compared to 2005 and 2015 (59.2% vs. 37.6% and 32.0% of successfully differentiated infections, respectively). Coinfections with Anaplasma phagocytophilum and Rickettsia spp. stayed stable comparing 2020 with previous years. Therefore, although changes in the Borrelia prevalence in questing ticks were not observed throughout the 15-year monitoring period, shifts in Borrelia species distribution may alter the epidemiological risk.

Fasciola hepatica is one of the economically most important endoparasites in cattle production. The aim of the present work was to evaluate the relevance of production level on the associations of on-farm presence of F. hepatica with farm-level milk yield, milk fat, and milk protein in Holstein cows, a specialised dairy breed, and in Simmental cows, a dual purpose breed. Furthermore, we investigated whether differential associations were present depending on breed. Data from 560 dairy farms across Germany housing 93,672 cows were analysed. The presence of F. hepatica antibodies was determined via ELISA on bulk tank milk samples. Quantile regression was applied to model the median difference in milk yield, milk fat, and milk protein depending on the interaction of breed and fluke occurrence. Whereas a reduction in milk yield (-1,206 kg, p < 0.001), milk fat (-22.9 kg, p = 0.001), and milk protein (-41.6 kg, p

Leucocytozoonosis is a vector-borne infection of birds, caused by members of the haemosporidian genus Leucocytozoon. The clinical presentation may range from asymptomatic to severe disease. Consequences of Leucocytozoon infection on blood profiles remain to be described, especially for different host species in the wild. In the current study, the prevalence of Leucocytozoon infection was determined in wild nestlings of three European raptor species, the common buzzard (Buteo buteo, n = 464), red kite (Milvus milvus, n = 46) and northern goshawk (Accipiter gentilis, n = 18). Among 528 nestlings, 51.9% (n = 274) were infected with Leucocytozoon spp., whereby the highest prevalence was found in common buzzards (54.9%), followed by red kites (32.6%) and northern goshawks (22.2%). For a subset of 87 individuals (50 common buzzards, 29 red kites, 8 northern goshawks), a detailed analysis of differential leukocyte counts and several blood chemistry parameters in response to infection was conducted: AP (alkaline phosphatase), AST (aspartate aminotransferase), GLDH (glutamate dehydrogenase), LDH (lactate dehydrogenase), GGT (gamma glutamyl transferase), CK (creatine kinase), BuChE (butyrylcholinesterase), BA (bile acids), ALB (albumin) and TP (total protein). Even though in the physiological range, infected nestlings displayed significantly increased levels of heterophils, aspartate aminotransferase, lactate dehydrogenase, bile acids and butyrylcholinesterase, but decreased lymphocyte and monocyte values compared to uninfected ones. Furthermore, significant species differences with regard to blood parameters, but no sex differences were found. Overall, obtained results show a high prevalence, but a low pathogenicity of Leucocytozoon spp. in wild raptor chicks, presumably resulting from coevolutionary adaptation, but show signatures of infection in the haematological and blood chemistry profiles.

Abstract Background The roundworm Ascaris suum is one of the parasites with the greatest economic impact on pig farming. In this context, lower weight gain is hypothesized to be due to decreased nutrient absorption. This study aims at characterizing the effects of A. suum infection on intestinal nutrient transport processes and potential molecular mechanisms. Methods Three groups of six piglets each were infected orally (10,000 embryonated A. suum eggs) in a single dose (“single infection”). Another three groups were infected orally (1000 embryonated eggs) for 10 consecutive days (“trickle infection”). Animals were necropsied 21, 35 and 49 days post-infection (dpi). Three groups served as respective controls. The Ussing chamber technique was applied for the functional characterization of small intestinal tissues [short-circuit currents (I sc) as induced by glucose, alanine and peptides; 3H-glucose net flux rates; tissue conductance (G t)]. Transcription and expression levels of relevant cytokines and nutrient transporters were evaluated (qPCR/western blot). Results Peptide- and alanine-induced changes in I sc were significantly decreased in the jejunum and ileum of the trickle-infected group at 49 dpi and in the ileum of the single-infected group at 49 dpi. No significant differences regarding glucose transport were observed between the Ascaris-infected groups and the control group in Ussing chamber experiments. Transcription levels of the glucose and peptide transporters as well as of selected transcription factors (transcription of signal transducer and activator of transcription 6 [STAT6] and hypoxia-inducible factor 1-alpha [Hif-1α]) were significantly increased in response to both infection types after some periods. The transcription of interleukins 4 and 13 varied between decrease and increase regarding the respective time points, as did the protein expression of glucose transporters. The expression of the peptide transporter PepT1 was significantly decreased in the ileal single-infected group at 35 dpi. Hif-1α was significantly increased in the ileal tissue from the single-infected group at 21 dpi and in the trickle-infected group at 35 dpi. The expression levels of Na+/K+-ATPase and ASCT1 remained unaffected. Conclusions In contrast to the current hypothesis, these results indicate that the nutrient deprivation induced by A. suum cannot be explained by transcriptional or expression changes alone and requires further studies. Graphical abstract

A considerable range expansion of Dermacentor reticulatus has been observed in several European countries, which is concerning in the light of its vector function for several pathogens, including Babesia canis and tick-borne encephalitis virus (TBEV). The present study provides an update on the distribution of Dermacentor ticks in Germany, using a citizen science approach. Ticks were collected by citizens from March 2020 to May 2021, and submitted along with information on the date and location of collection, potential hosts and details about the circumstances of discovery. In total, 3,292 Dermacentor specimens were received, of which 76.4% (2,515/3,292) were identified as D. reticulatus and 23.0% (758/3,292) as D. marginatus, while 0.6% (19/3,292) were too damaged for species-level identification. Dermacentor reticulatus was received from all federal states of Germany. Maxent species distribution models predicted suitable environmental conditions for D. reticulatus throughout Germany. Findings on the vegetation or on pastured animals without travel history confirmed the occurrence of this tick species as far north as the most northern German federal state Schleswig-Holstein. In contrast, the distribution of D. marginatus still appears to be limited to southwestern Germany, although the northward shift of the distribution limit observed in the preceding citizen science study, as compared with previous published distributions, was confirmed. This shift was also predicted by Maxent species distribution models, reflecting the broader distribution of the tick occurrence data contributed by citizens. Most D. reticulatus ticks were found on dogs (1,311/1,960, 66.9%), while D. marginatus was mainly discovered on hoofed animals (197/621, 31.7%) and humans (182/621, 29.3%). Human tick bites were reported in 0.7% (14/1,960) of host-assigned D. reticulatus and 3.4% (21/621) of host-assigned D. marginatus. Further studies to investigate an increasing endemisation of Babesia canis in Germany as well as the relevance of D. reticulatus for TBEV spread throughout the country, e.g., by traveling dogs, are urgently needed. In view of the activity of D. reticulatus during winter or the colder months, which complements that of Ixodes ricinus, a year-round tick protection of at least dogs is strongly recommended.

Ascarid infections constitute a major concern for both human and animal health risk assessment. Although being effectively transmitted by soil, water and contaminated food, reliable detection of ascarid eggs in environmental media often remains challenging. However, contamination of the environment with ascarid ova has gained more attention as a decisive part of proper risk assessment in recent years. Due to various factors, such as sample matrices, dissociation detergents and flotation solutions, defined and standardised protocols for the isolation of eggs from complex environmental matrices are difficult to establish and therefore limited. Thus, this study reviews common techniques used for the recovery of ascarid eggs from environmental media with special emphasis on sampling strategies, purification procedures and microscopic as well as molecular detection of egg contamination. Despite various advancements, mainly in the field of molecular methods leading to more reliable and sensitive detection, it can be concluded that there is still a need for unified guidelines for sampling and recovery of ascarid eggs derived from complex environmental matrices.

Abstract Background Toxoplasma gondii is an obligate intracellular parasite with a worldwide distribution. Congenital infection in humans and animals may lead to severe symptoms in the offspring, especially in the brain. A suitable animal model for human congenital toxoplasmosis is currently lacking. The aim of this study is to establish and validate the guinea pig as a model for human congenital toxoplasmosis by investigating the impact of the T. gondii infection dose, the duration of infection and the gestational stage at infection on the seroconversion, survival rate of dams, fate of the offspring, T. gondii DNA loads in various offspring tissues and organs and the integrity of the offspring brain. Methods Pregnant guinea pigs were infected with three different doses (10, 100, 500 oocysts) of T. gondii strain ME49 at three different time points during gestation (15, 30, 48 days post-conception). Serum of dams was tested for the presence of T. gondii antibodies using immunoblotting. T. gondii DNA levels in the dam and offspring were determined by qPCR. Offspring brains were examined histologically. Results We found the survival rate of dams and fate of the offspring to be highly dependent on the T. gondii infection dose with an inoculation of 500 oocysts ending lethally for all respective offspring. Moreover, both parameters differ depending on the gestational stage at infection with infection in the first and third trimester of gestation resulting in a high offspring mortality rate. The duration of infection was found to substantially impact the seroconversion rate of dams with the probability of seroconversion exceeding 50% after day 20 post-infection. Furthermore, the infection duration of dams influenced the T. gondii DNA loads in the offspring and the integrity of offspring brain. Highest DNA levels were found in the offspring brain of dams infected for ≥ 34 days. Conclusion This study contributes to establishing the guinea pig as a suitable model for human congenital toxoplasmosis and thus lays the foundation for using the guinea pig as a suitable animal model to study scientific questions of high topicality and clinical significance, which address the pathogenesis, diagnosis, therapy and prognosis of congenital toxoplasmosis. Graphical abstract

Abstract Deleterious effects of habitat loss and fragmentation on biodiversity have been demonstrated in numerous taxa. Although parasites represent a large part of worldwide biodiversity, they are mostly neglected in this context. We investigated the effects of various anthropogenic environmental changes on gastrointestinal parasite infections in four small mammal hosts inhabiting two landscapes of fragmented dry forest in northwestern Madagascar. Coproscopical examinations were performed on 1,418 fecal samples from 903 individuals of two mouse lemur species, Microcebus murinus (n = 199) and M. ravelobensis (n = 421), and two rodent species, the native Eliurus myoxinus (n = 102) and the invasive Rattus rattus (n = 181). Overall, sixteen parasite morphotypes were detected and significant prevalence differences between host species regarding the most common five parasites may be explained by parasite–host specificity or host behavior, diet, and socioecology. Ten host‐ and habitat‐related ecological variables were evaluated by generalized linear mixed modeling for significant impacts on the prevalence of the most abundant gastrointestinal parasites and on gastrointestinal parasite species richness (GPSR). Forest maturation affected homoxenous parasites (direct life cycle) by increasing Lemuricola, but decreasing Enterobiinae gen. sp. prevalence, while habitat fragmentation and vegetation clearance negatively affected the prevalence of parasites with heterogenic environment (i.e., Strongyloides spp.) or heteroxenous (indirect cycle with intermediate host) cycles, and consequently reduced GPSR. Forest edges and forest degradation likely change abiotic conditions which may reduce habitat suitability for soil‐transmitted helminths or required intermediate hosts. The fragility of complex parasite life cycles suggests understudied and potentially severe effects of decreasing habitat quality by fragmentation and degradation on hidden ecological networks that involve parasites. Since parasites can provide indispensable ecological services and ensure stability of ecosystems by modulating animal population dynamics and nutrient pathways, our study underlines the importance of habitat quality and integrity as key aspects of conservation.

Pseudaliid lungworm (Metastrongyloidea) infections and associated secondary bacterial infections may severely affect the health status of harbour porpoises (Phocoena phocoena) in German waters. The presented retrospective analysis including data from 259 harbour porpoises stranded between 2006 and 2018 on the German federal state of Schleswig-Holstein's North Sea coast showed that 118 (46%) of these stranded individuals harboured a lungworm infection. During this 13-year period, a significant difference in annual lungworm prevalence was only observed between the years 2006 and 2016. Lungworm coinfections of bronchi and pulmonary blood vessels were observed in 85.6% of positive cases. Mild infection levels were detected in 22.9% of infected animals and were most common in the age class of immature individuals (74.1%). Moderate and severe infections were present in 38.1% and 39.0% of the lungworm positive animals, respectively. Their distribution in immatures (51.1% and 54.3%) and adults (48.9% and 43.4%) did not show significant differences. In stranded animals, lungworm diagnosis can be easily obtained via necropsy, while reliable lungworm diagnosis in living porpoises requires invasive bronchoscopy or faecal examination, which is difficult to obtain in cetaceans. To overcome this issue, an enzyme-linked immunosorbent assay (ELISA) and immunoblot based on recombinant major sperm protein (MSP) of the cattle lungworm were evaluated as potential diagnostic tools in harbour porpoises. However, in contrast to hitherto other investigated host species, no reliable antibody response pattern was detectable in harbour porpoise serum/plasma or whole blood samples. Thus, MSP-based serological tests are considered unsuitable for lungworm diagnosis in harbour porpoises.

Abstract Background The feline lungworm Aelurostrongylus abstrusus affects the lower respiratory tract in cats worldwide. As infections may lead to chronic respiratory changes or even death, preventive treatment in cats with outdoor access is warranted. Methods The preventive efficacy of a spot-on solution (Bravecto® Plus spot-on solution for cats, MSD) against cat aelurostrongylosis was evaluated using three different preventive treatment regimes in a negative controlled, randomized and partially blinded laboratory efficacy study with 31 purposed-bred cats. The minimum recommended dose of 2.0 mg moxidectin + 40 mg fluralaner/kg bodyweight was applied once 12 (Group [G]1), 8 (G2) or 4 (G3) weeks before experimental infection with 300 third-stage larvae (L3) of A. abstrusus. Another group served as untreated control (G4). Individual faecal samples were analysed as of day 30 post infection (pi) to monitor larvae excretion. Necropsy was performed at days 47–50 pi. The lungs were examined macroscopically for pathological findings and (pre-)adult worms were counted to assess preventive efficacy. Results Beginning at day 32–40 pi, all cats of the control group were constantly shedding larvae of A. abstrusus, whereas only one animal of G1 excreted larvae at several consecutive days. In addition, two cats of G1 and G3 and three of G2 were positive on a single occasion. The geometric mean (GM) of the maximum number of excreted larvae was 7574.29 in the control group compared to 1.10 (G1), 1.19 (G2) and 0.53 (G3), resulting in a GM reduction of > 99.9% in all treatment groups. All lungs of the control animals showed severe or very severe alterations at necropsy, while in 94.44% of the treated cats lung pathology was rated as absent or mild. The GM number of (pre-)adult A. abstrusus retrieved from the lungs was 26.57 in the control group, 0.09 in G1 and 0.00 in G2 and G3. Thus, GM worm count reduction was 99.66% in G1 and 100% in G2 and G3. Conclusions A single application of Bravecto® Plus spot-on solution at a dose of 2.0 mg moxidectin + 40 mg fluralaner/kg bodyweight reliably prevents cat aelurostrongylosis for at least 12 weeks.

Abstract Background Ixodes ricinus constitutes the main European vector tick for the Lyme borreliosis pathogen Borrelia burgdorferi (sensu lato), the relapsing fever borrelia Borrelia miyamotoi, as well as Anaplasma phagocytophilum and several Rickettsia species. Under laboratory conditions, a transovarial transmission to the next tick generation is described for Rickettsia spp. and Borrelia spp., especially regarding B. miyamotoi, whereas the efficiency of transovarial transfer under field conditions is largely unstudied. Methods In order to better estimate the potential infection risk by tick larvae for humans and animals, 1500 I. ricinus larvae from 50 collected “nests” (larvae adhering to the flag in a clumped manner) were individually examined for Borrelia, Rickettsia and A. phagocytophilum DNA using quantitative real-time PCR (qPCR). Results Thirty-nine of 50 nests each (78.0%, 95% CI: 64.0–88.5%) were positive for Borrelia spp. and Rickettsia spp. DNA, and in three nests (6.0%, 95% CI: 1.3–16.5%) A. phagocytophilum DNA was detected. Overall, DNA from at least one pathogen could be detected in 90.0% (45/50, 95% CI: 78.2–96.7%) of the nests. Of the 1500 larvae, 137 were positive for Borrelia spp. DNA (9.1%, 95% CI: 7.7–10.7%), 341 for Rickettsia spp. DNA (22.7%, 95% CI: 20.6–24.9%) and three for A. phagocytophilum DNA (0.2%, 95% CI: 0–0.6%). Quantity of Borrelia spp. and Anaplasma spp. DNA in positive larvae was low, with 2.7 × 100 Borrelia 5S-23S gene copies and 2.4 × 101 A. phagocytophilum msp2/p44 gene copies detected on average, while Rickettsia-positive samples contained on average 5.4 × 102 gltA gene copies. Coinfections were found in 66.0% (33/50, 95% CI: 51.2–78.8%) of the nests and 8.6% (38/443, 95% CI: 6.1–11.6%) of positive larvae. In fact, larvae had a significantly higher probability of being infected with Borrelia spp. or Rickettsia spp. when both pathogens were present in the nest. Conclusions This study provides evidence for transovarial transmission of Rickettsia spp. and Borrelia spp. in I. ricinus under field conditions, possibly facilitating pathogen persistence in the ecosystem and reducing the dependence on the presence of suitable reservoir hosts. Further studies are needed to prove transovarial transmission and to explain the surprisingly high proportion of nests containing Rickettsia and/or Borrelia DNA-positive larvae compared to infection rates in adult ticks commonly reported in other studies.

Abstract Background In three randomized, controlled laboratory efficacy studies, the efficacy in the prevention of patent infections of a topical combination of imidacloprid 10%/moxidectin 1% (Advocate® spot-on formulation for cats, Bayer Animal Health GmbH) against larval stages and immature adults of Aelurostrongylus abstrusus, as well as the treatment efficacy of a single or three monthly treatments against adult A. abstrusus, were evaluated. Methods Cats were experimentally inoculated with 300–800 third-stage larvae (L3). Each group comprised 8 animals and the treatment dose was 10 mg/kg bodyweight (bw) imidacloprid and 1 mg/kg bw moxidectin in each study. Prevention of the establishment of patent infections was evaluated by two treatments at a monthly interval at three different time points before and after challenge infection. Curative efficacy was tested by one or three treatments after the onset of patency. Worm counts at necropsy were used for efficacy calculations. Results In Study 1, the control group had a geometric mean (GM) of 28.8 adult nematodes and the single treatment group had a GM of 3.4 (efficacy 88.3%). In Study 2, the control group had a GM of 14.3, the prevention group had a GM of 0 (efficacy 100%), while the treatment group had a GM of 0.1 (efficacy 99.4%). In Study 3, the GM worm burden in the control group was 32.6 compared to 0 in all three prevention groups (efficacy 100% for all of those groups). Conclusions The monthly administration of Advocate® reliably eliminated early larval stages and thereby prevented lung damage from and patent infections with A. abstrusus in cats. Regarding treatment, a single application of Advocate® reduced the worm burden, but it did not sufficiently clear the infection. In contrast, three monthly treatments were safe and highly efficacious against A. abstrusus.

Fasciola hepatica and Ostertagia ostertagi are internal parasites of cattle compromising physiology, productivity, and well-being. Parasites are complex in their effect on hosts, sometimes making it difficult to identify clear directions of associations between infection and production parameters. Therefore, unsupervised approaches not assuming a structure reduce the risk of introducing bias to the analysis. They may provide insights which cannot be obtained with conventional, supervised methodology. An unsupervised, exploratory cluster analysis approach using the k-mode algorithm and partitioning around medoids detected two distinct clusters in a cross-sectional data set of milk yield, milk fat content, milk protein content as well as F. hepatica or O. ostertagi bulk tank milk antibody status from 606 dairy farms in three structurally different dairying regions in Germany. Parasite-positive farms grouped together with their respective production parameters to form separate clusters. A random forests algorithm characterised clusters with regard to external variables. Across all study regions, co-infections with F. hepatica or O. ostertagi, respectively, farming type, and pasture access appeared to be the most important factors discriminating clusters (i.e. farms). Furthermore, farm level lameness prevalence, herd size, BCS, stage of lactation, and somatic cell count were relevant criteria distinguishing clusters. This study is among the first to apply a cluster analysis approach in this context and potentially the first to implement a k-medoids algorithm and partitioning around medoids in the veterinary field. The results demonstrated that biologically relevant patterns of parasite status and milk parameters exist between farms positive for F. hepatica or O. ostertagi, respectively, and negative farms. Moreover, the machine learning approach confirmed results of previous work and shed further light on the complex setting of associations a between parasitic diseases, milk yield and milk constituents, and management practices.

Feline troglostrongylosis caused by Troglostrongylus brevior is increasingly reported in European countries. Although the disease can be severe and potentially life-threatening, especially in kittens and young cats, effective treatment options are still limited. Two administrations of emodepside 2 weeks apart have shown promising results for the treatment of T. brevior infection in single cases and in a field trial. Therefore, the present study has been conducted to evaluate the efficacy of two spot-on combinations containing emodepside (i.e. 2.14% w/v emodepside and 8.58% w/v praziquantel - Profender®, and 2.04% w/v emodepside, 8.14% w/v praziquantel and 9.79% w/v tigolaner - Felpreva®) in the treatment of troglostrongylosis under experimental conditions. Twenty-four cats were experimentally infected with T. brevior and randomly assigned to one of three groups of eight cats each, i.e. (i) Group 1 (G1) left untreated, (ii) Group 2 (G2) receiving Profender® on Days 28 and 44, and (iii) Group 3 (G3) receiving Felpreva® on Day 28 and Profender® on Day 44. Doses corresponded to the minimum effective dose of 0.140 and 0.148 ml/kg body weight, for Profender® and Felpreva®, respectively. The primary efficacy criterion was the number of viable adult T. brevior counted at necropsy conducted between Days 69 and 72. The fecal shedding of first-stage larvae (L1) was also assessed. L1 of T. brevior were detected in samples from all cats within 20 days post-infection. At necropsy, 4 of 8 G1 cats harbored adult T. brevior, while no adult T. brevior worms or other development stages were recovered from any of the G2 and G3 cats. The primary efficacy criterion was not evaluated as the worm counts in G1 did not meet VICH guideline requirements. After the first treatment (Day 28), most G2 and G3 cats were negative at the Baermann examination. After the second treatment (Day 44), L1 were found in two cats from G2 on Day 49 and in one G3 cat on Day 51. No adverse events occurred in G2 and G3 cats. These results indicate that two applications of emodepside spot-on given 2 weeks apart represent a safe and efficacious treatment regime against troglostrongylosis.

The newly discovered group of Jingmenviruses has been shown to infect a wide range of hosts and has been associated with febrile illness in humans. During a survey for Jingmenviruses in ticks from Lower Saxony, Germany, Alongshan virus (ALSV) was identified in Ixodes spp. ticks. Additional virus screenings revealed the presence of ALSV in the bodies and saliva of ticks collected at several locations in Lower Saxony. Vector competence studies that included Ixodes ricinus and Dermacentor reticulatus validated the replication of ALSV within those tick species. In vitro feeding experiments with ALSV-injected Ixodes ricinus demonstrated effective viral transmission during blood feeding. To evaluate the potential viral transmission during a natural blood meal, sera from wild game and domestic animals were investigated. One serum sample from a red deer was found to be positive for ALSV RNA, while serological screenings in game and domestic animals revealed the presence of ALSV-specific antibodies at different locations in Lower Saxony. Overall, those results demonstrate the broad distribution of ALSV in ticks in Lower Saxony and hypothesize frequent exposure to animals based on serological investigations. Hence, its potential risk to human and animal health requires further investigation.

The European brown hare (Lepus europaeus) is a quite adaptable species, but populations have been decreasing for several decades in different countries, including Germany. To investigate infectious diseases as possible influences on observed population decline in the German federal state Schleswig-Holstein, 118 deceased free-ranging European brown hares were collected between 2017 and 2020 and underwent detailed postmortem examination with extended sampling. Infectious diseases were a major cause of death (34.7%). The number of juveniles found exceeded the adult ones. The main pathomorphological findings were hepatitis (32.8%), pneumonia (22.2%), nephritis (19.1%), liver necrosis (12.9%), and enteritis (40.7%). An unusual main finding was steatitis (20.9%) of unknown origin. Animals were mainly emaciated and showed high infection rates with Eimeria spp. (91.3%) and Trichostrongylus spp. (36.2%). European Brown Hare Syndrome Virus reached an epidemic status with few fatal infections (4.2%) and high seroprevalence (64.9%), whereas the prevalence of Rabbit Haemorrhagic Disease Virus 2 was very low (0.8%) in hares in Schleswig-Holstein. Pathogens such as Yersinia pseudotuberculosis (5.9%), Pasteurella multocida (0.8%), and Staphylococcus aureus (3.4%) only caused sporadic deaths. This study illustrates the wide distribution of various infectious pathogens with high mortality and even zoonotic potential. Infectious diseases need to be considered as an important influence on population dynamics in Schleswig-Holstein.

Abstract Background Parafilaria bovicola (Nematoda: Filariidae) causes cutaneous bleedings in bovine species. Flies serve as intermediate hosts. In recent years, reports on bovine parafilariosis have become more frequent, corroborating the necessity of reliable diagnostic interventions especially since no molecular or serological test has been available. We aimed to establish a polymerase chain reaction assay to detect DNA of P. bovicola in flies, skin biopsies and serohemorraghic exudates of bleeding spots. Methods PCRs targeting the cytochrome c oxidase subunit 1 (cox1) gene and the internal transcribed spacer region (ITS) of the ribosomal RNA gene cluster were evaluated for their diagnostic sensitivity as well as performance and specificity on biopsy and serohemorrhagic exudate samples from P. bovicola-infected cattle. Results Using serohemorrhagic exudates (n = 6), biopsies (n = 2) and flies (n = 1), the PCR targeting the cox1 gene resulted in a gel band of almost 700 bp. Cloning, sequencing, and removal of primer sequences yielded a 649-bp fragment of the P. bovicola cox1 gene. The PCR targeting the ITS region showed a band of about 1100 bp. Cloning, sequencing, and removal of primer sequences resulted in a 1083 bp stretch of the P. bovicola ITS region. Testing samples from presumably affected animals, the cox1-PCR resulted in bands with the expected size and they were all confirmed as P. bovicola by sequencing. In contrast, the ITS-PCR proved to be less sensitive and less specific and additionally amplified the ITS region of Musca domestica or buttercup DNA. When analysing for sensitivity, the cox1-PCR yielded visible bands up to 2 ng of genomic DNA, whereas the ITS-PCR produced bands up to 3 ng. In a plasmid dilution series, the minimum number of target DNA copies was 102 for the cox1-PCR and 101 in the ITS-PCR. Conclusions The evaluated cox1-PCR enables reliable detection of P. bovicola DNA in skin biopsies and serohemorrhagic exudates. This PCR and, to a limited extent, the ITS-PCR, may help evaluate different therapeutic approaches. Furthermore, the cox1-PCR may be useful for epidemiological studies on the geographical distribution of P. bovicola. Further understanding of the epidemiology of this parasite will help develop and implement effective control strategies.

Small rodents serve as intermediate or paratenic hosts for a variety of parasites and may participate in the transmission of these parasites into synanthropic cycles. Parasites with neuroinvasive stages, such as Toxoplasma gondii or Toxocara canis, can cause detrimental damage in the brain of intermediate or paratenic hosts. Therefore, the occurrence of neuroinvasive parasite stages was evaluated in brains of wild rodents captured in the city of Leipzig, Germany. In addition, a few specimens from the cities of Hanover, Germany, and Vienna, Austria were included, resulting in a total of 716 rodents collected between 2011 and 2016. Brains were investigated for parasitic stages by microscopic examination of native tissue, artificially digested tissue as well as Giemsa-stained digestion solution to verify positive results. Infective stages of zoonotic ascarids or other helminths were not detected in any sample, while coccidian cysts were found in 10.1% (95% CI: 7.9–12.5%; 72/716) of examined brains. The most abundant rodent species in the study was the bank vole (Myodes glareolus; Arvicolinae), showing an infection rate with cerebral cysts of 13.9% (95% CI: 11.0–17.8%; 62/445), while 2.7% (95% CI: 1.0–5.8%; 6/222) of yellow-necked mice (Apodemus flavicollis; Murinae) were infected. Generalized linear modelling revealed a statistically significant difference in prevalence between M. glareolus and A. flavicollis, significant local differences as well as an effect of increasing body mass on cyst prevalence. Coccidian cysts were differentiated by amplification of the 18S rRNA gene and subsequent sequencing. The majority of identifiable cysts (97.9%) were determined as Frenkelia glareoli, a coccidian species mainly circulating between M. glareolus as intermediate and buzzards (Buteo spp.) as definitive hosts. The zoonotic pathogen Toxoplasma gondii was confirmed in one M. glareolus originating from the city of Leipzig. Overall, it can be concluded that neuroinvasion of zoonotic parasites seems to be rare in M. glareolus and A. flavicollis. Keywords: Toxocara, Toxoplasma, Frenkelia glareoli, Zoonosis, Myodes glareolus, Apodemus flavicollis

The dynamic relationship of vector-borne parasites, arthropod vectors and their hosts is prone to change under the influence of climate change, global integration, shifting demographics and deforestation. It is therefore essential to better understand parasitism in wildlife populations, including parasites transmitted by blood-feeding vectors, and explore host range and heterogeneity of parasitic infections. We investigated Giemsa stained blood smears of two sympatric Malagasy primate species (Microcebus murinus: 184 samples from 69 individuals and M. ravelobensis: 264 samples from 91 individuals) for blood-stage parasites and tested for a potential influence of host species, sex, body mass and sampling month on blood-stage parasite prevalence and infection intensity. No protozoan parasites were detected in either host species. A host-specific difference was observed in filarial nematode infections, with higher risk of infection in M. murinus (prevalence 30.43%), than in M. ravelobensis (prevalence 6.59%), which may be explained by differences in host behavior and/or immune competence, linked to the period of host-parasite coevolution. Neither sex nor sampling month influenced infection prevalence or intensity significantly. We did not observe a negative effect of microfilarial infections on host fitness when taking body mass as a proxy. Our results support the hypothesis of a long-term evolutionary adaptation of hosts and parasites, leading to persistent infection with low morbidity. Morphological and molecular analyses indicate the finding of a new species, “Lemurfilaria lemuris”. Genetic analysis furthermore showed >99% sequence identity with microfilariae described from a sympatric, larger-bodied lemur species of a different genus, suggesting low host-specificity of the detected filariae and pathogen transmission across genus boundaries. Findings contribute to a more comprehensive picture of vector-borne diseases of Malagasy lemurs. Keywords: Microcebus spp., Onchocercidae, Microfilaria, Evolution, Ecology, Madagascar

Life history theory predicts that species with shorter lifespan should show higher investments into growth and reproduction at the expense of immune defenses. Labord's chameleon (Furcifer labordi) is the tetrapod with the shortest known life span. To investigate to which extent immunosenescence influences the die-off of these chameleons when they are only about 6 months old, we examined the gastrointestinal-, blood- and ectoparasite burden in F. labordi in Kirindy Forest (western Madagascar) and compared them with sympatric and longer living F. cf. nicosiai. Moreover, we included data from wild F. labordi that were singly housed under ambient conditions with daily food and water supply. Gastrointestinal parasite prevalence of wild F. labordi increased dramatically during the last 3 months of their lives, which include the reproductive period. Furcifer cf. nicosiai was found to have a belated increase in gastrointestinal parasites compared to F. labordi. In F. cf. nicosiai higher prevalence of blood parasites were found, which probably result from the longer exposure to the arthropod intermediate host. Both species showed infestations with ectoparasites, which peaked in the rainy season but disappeared towards the dry season. Male F. labordi showed a significantly higher prevalence of gastrointestinal - and ectoparasites and higher intensities of coccidians and ectoparasites than females. Males of F. cf. nicosiai exhibited higher prevalence of blood- and ectoparasites, as well as higher intensities in ectoparasites. Caged individuals of both sexes showed delayed senescence, reduced parasite burden and lived longer than their wild conspecifics. Overall, the increase in the prevalence in gastrointestinal - and blood parasites towards the disappearance of the wild population of F. labordi indicates that this species invests comparatively less energy in efficient immune system function, supporting the prediction of life history theory. Keywords: Parasites, Chameleons, Immunosenescence, Aging, Furcifer