Environmental impact plays a pivotal role in forming the welfare of shelter dogs exposed to chronic stress. Standard methods of animal health monitoring, such as psychological evaluation or cortisol measurements, do not fully reflect modulation of the immune system. Functional cellular changes may be subtle and observed only at the molecular level. Therefore, the aim of this study was to characterize the immune function of shelter dogs kept on different timetables in comparison with client-owned dogs. We focused on potential alterations of antigen processing by neutrophils and monocytes in animals undergoing different durations of stress. Hematological and biochemical parameters were evaluated, and changes in TLR4 and MHC Class II expression on neutrophils and monocytes isolated from peripheral blood were determined. Additionally, we measured the percentage of apoptotic cells within these leukocyte populations. Our study revealed that stressful conditions can alter the molecular pattern of surface receptors on neutrophils and monocytes, as well as the leukocytes apoptosis rate. The obtained data also indicated that the dogs’ duration of stay in the shelter plays an important role in immunomodulation and triggering their adaptation mechanisms. These results bring a new perspective and will be crucial in developing improved guidelines for monitoring and promoting the welfare of shelter dogs.

Introduction:Dirofilaria repens is a zoonotic parasitic filarial nematode that infects carnivores and occasionally humans. Knowledge of the host-parasite molecular interactions enabling the parasite's avoidance of the host immune response in subcutaneous dirofilariasis remains limited. Parasitic orthologues of host macrophage migration inhibitory factor (MIF) are molecules potentially involved in this process. Material and Methods:Complementary DNA encoding two D. repens MIF orthologues (rDre-MIF-1 and rDre-MIF-2) was cloned into a pET-28a expression vector. The recombinant proteins were produced in Escherichia coli and purified using affinity nickel chromatography. The reactivity of both recombinant proteins was analysed with infected dog and immunised mouse sera. Results:Stronger antibody production was induced by rDre-MIF-1 in mice, as evidenced by significantly higher levels of anti-rDre-MIF-1 total IgG, IgG2 and IgE antibodies than of anti-rDre-MIF-2 immunoglobulins. Additionally, a significantly different level of antibodies specific to both proteins was noted between the sera of infected dogs and those of uninfected dogs. Conclusion:This study is the first attempt to characterise MIF orthologues from the filarial parasite D. repens, which may affect the immune response during infection.

American mink (Neovison vison) is an invasive species in the sylvatic environment of Poland. Mink are exposed to different parasite infections as their preys serve as intermediate and/or paratenic hosts. The study aimed to discriminate the pattern of intestinal parasite infections in mink inhabiting Biebrza (BNP) and Narew (NNP) national parks. Gastrointestinal tract examinations revealed Coccidia, Echinostomatidae, Taenidae, and Capillariidae parasites. There was no significant difference in the parasite burden of mink, but patterns of infections varied between both localizations. Coccidia were found in 3.8% of BNP vs. 6.7% of NNP mink. Fluke prevalence was significantly higher in NNP 27.5% compared to 7.7% in BNP mink. Tapeworms were only found in 3.4% of NNP mink. Significantly more Aonchotheca eggs were found in BNP 34.6% vs. 11.4% in NNP mink. The intensity of coccidiosis and aonchothecosis was low in both parks. Fluke intensity varied between low to moderate (ranging from 1 to 16) in BNP and low to massive (ranging from 1 to 117) in NNP mink. Coinfections of various parasite species were noted in both areas. Morphological and DNA analysis revealed that flukes belonged to Isthiomorpha melis and tapeworms to Versteria mustelae. It was the first isolation of V. mustelae in mink of those localizations. In conclusion, our study showed that mink indwelling Biebrza and Narew national parks are moderately infested with parasites. Results suggest that mink play an important role as a reservoir for parasites endangering endemic mustelids, becoming also a potential risk factor in case of accidental transmissions to farm mink. That is why, more strict biosecurity measures are required to protect farm mink.

Introduction:The aim of this study was to investigate changes in haematological parameters and the antioxidant system in the early and late pregnancy of red deer (Cervus elaphus). Material and Methods:Blood samples were collected from 30 red deer females 50 days after impregnation and 40 days before calving. Complete blood counts and stained blood smears were assessed. Paraoxonase 1 (PON1), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR) activities, glutathione disulphide (GSSG), total glutathione, total bilirubin, total protein, albumin, uric acid, malondialdehyde (MDA), beta-hydroxybutyrate (BHB), low-density lipoprotein (LDL), high-density lipoprotein (HDL), triglycerides, cholesterol levels and total antioxidant status (TAS) were measured. Results:The haematological characteristics of blood collected in the third trimester showed higher haemoglobin levels, haematocrit, mean corpuscular volume (P < 0.05) and a lower mean cell haemoglobin concentration (P < 0.05) in comparison to samples collected on the 50th day after mating. Activity of CAT and MDA, LDL, and triglyceride levels were lower while GR, GSSG, total glutathione, total protein and BHB levels were increased in blood samples drawn in the third trimester compared with the first trimester blood samples. There were no changes in SOD, PON1 activities, TAS, bilirubin, albumin, HDL, cholesterol or uric acid concentrations. Conclusion:Red deer's compensatory mechanisms facilitate its optimal adaptation to seasonal changes evidenced by mild pronounced haematological disturbances and an effective antioxidant system during pregnancy.

Dirofilaria repens is a parasitic nematode causing vector-borne disease (dirofilariasis), considered an emerging problem in veterinary and human medicine. Although main hosts are carnivores, particularly dogs, D. repens shows high zoonotic potential. The disease spreads uncontrollably, affecting new areas. Since there is no vaccine against dirofilariasis, the only way to limit disease transmission is an early diagnosis. Currently, diagnosis depends on the detection of microfilariae in the host bloodstream using modified Knott's test or multiplex PCR. However, the efficacy of tests relying on microfilariae detection is limited by microfilariae periodic occurrence. Therefore, a new reliable diagnostic test is required. Our study aimed to select new diagnostic markers for dirofilariasis with potential application in diagnostics. We focused on single epitopes to ensure high specificity of diagnosis and avoid cross-reactivity with the other parasite infections common in dogs. Using phage display technology and 12-mer peptides library, we selected epitopes highly reactive with IgG from sera of infected dogs. Additionally, our study presents the possibility of detecting D. repens specific cell-free DNA in dogs with no microfilaria but high IgG and IgM antibody levels against parasite somatic antigen.

Dirofilaria repens is the causative agent of the zoonotic canine skin condition called subcutaneous dirofilariosis. Despite being endemic throughout much of Europe, little is known about host humoral responses to D. repens. To address this, we analyzed serum immunoglobulin isotypes recognizing D. repens somatic antigens (DrSA) in naturally infected dogs. Titers of anti-DrSA IgG and IgE, but not IgM were significantly higher in dogs infected with D. repens compared to those that were negative. Moreover, microfilaremic infections were associated with higher levels of IgG1 than IgG2, while occult infections occurred with significantly higher levels of IgG2 than IgG1. Finally, the measurement of anti-DrSA IgG antibodies allowed the detection of occult subcutaneous dirofilariosis in dogs.

Dirofilariarepens is a parasitic nematode causing a vector-borne zoonotic infection (dirofilariosis), considered an emerging problem in human and veterinary medicine. Currently, diagnosis is based on the detection of the adult parasite and microfilariae in the host tissues. However, the efficacy of tests relying on microfilariae detection is limited by microfilariae periodic occurrence. Therefore, a new reliable and affordable serological diagnostic method is needed. Better characteristic of the parasite biology and its interaction with host immune system should help to achieve this goal. This study analyzes adult and microfilariae proteomes, and the use of one-dimensional electrophoresis (1-DE) and two-dimensional electrophoresis (2-DE) proteomics, immunoproteomics, and LC-MS/MS mass spectrometry allowed us to identify 316 potentially immunogenic proteins (75 belong to adult stage, 183 to microfilariae, and 58 are common for both). Classified by their ontology, the proteins showed important similarities and differences between both parasite stages. The most frequently identified proteins are structural, metabolic, and heat shock proteins. Additionally, real-time PCR analysis of some immunogenic targets revealed significant differences between microfilariae and adult life stages. We indicated molecules involved in parasite-host interactions and discussed their importance in parasite biology, which may help to reveal potential diagnostic antigens or select drug and vaccine targets.

Subcutaneous dirofilariosis is a zoonotic disease emerging throughout Europe caused by the filarial nematodeDirofilaria repens. Despite its increasing prevalence, there is a large gap in knowledge of the impact of the parasite on the host. Currently classified as being non-pathogenic, recent evidence suggests that skin dirofilariosis is associated with dermatological conditions including concomitant pruritus, neoplastic processes, inflammation, and even blindness in dogs and humans. The aim of this study was to determine if natural canineD. repensinfection leads to biological changes in the canine host. In a real-life veterinary clinic setting, animals are often presented to clinicians for unrelated issues, andD. repensis incidentally identified during inspection. As such, we compared hematological and biochemical parameters of 218 uninfected and 197 dogs naturally infected withD. repens. Interestingly, animals infected withD. repenshad lower numbers of lymphocytes (p< 0.001), red blood cells (p< 0.001), and thrombocytes (p= 0.025), decreased hematocrit (p< 0.001), and increased alkaline phosphatase (p= 0.016) and creatinine activity (p= 0.023) compared to uninfected dogs. We further selected a subpopulation of 214 dogs havingprima faciehematological and biochemical results within normal reference ranges to evaluate the effect ofD. repensinfections in seemingly healthy dogs. Among these patients, 93 dogs infected withD. repenshad lower numbers of lymphocytes (p= 0.031), red blood cells (p= 0.025), and hematocrit (p= 0.002), higher glucose levels (p= 0.023), and border line elevated alkaline phosphatase levels (p= 0.054) compared to 121 uninfected animals. Despite being categorized as asymptomatic, we have observed hematological and biochemical changes associated withD. repensinfections in dogs, and our data suggest that dirofilariosis may induce a state of chronic stress. These results link the presence of skin dirofilariosis to biological changes in the canine host, suggesting a mechanism for pathogenicity and shedding new light on the host-parasite relationship.

Subcutaneous dirofilariosis in dogs, caused by Dirofilaria repens, is an underdiagnosed disease, now recognized for its zoonotic potential, and growing distribution and prevalence across Europe and Asia. Our understanding of the pathogenicity in human and canine host remains unclear, but case reports suggest that microfilariae (Mf) as well as adult D. repens may directly cause internal organs damage or may be a factor complicating the course of other ailments. The purpose of the study was to report high Mf in dogs and to discuss potential relevance with co-morbidity. Our data from a modified Knott's test performed on 62 infected dogs indicate that the median Mf count in D. repens infections is 675 Mf/ml and we consider microfilaremia above 10,000 Mf/ml as high intensity. This collection of case reports discusses 4 cases of high intensity D. repens microfilaremia in companion dogs; one presenting pathology from a very high intensity of adult D. repens with post-treatment complications, and 3 dogs in which high microfilaremia was detected incidentally during the management of other primary illnesses. To our knowledge this report describes the highest D. repens microfilaremia ever detected in a dog, at 178,000 Mf/ml. The issue of high microfilaremic infections in dogs is poorly studied and there is growing need to identify the presentation and understand the mechanisms of associated pathogenesis in the host-parasite relationship.

INTRODUCTION AND OBJECTIVE:The aim was to review available data concerning the occurrence of protozoan parasites affecting carnivorous fur farm animals, such as: American mink, blue and silver foxes, and raccoon dogs. Although, many protozoan pathogens have been isolated in wild fur animals, some are still not recognised as relevant to particular fur farm animal species. Protozoans that have been isolated as causative agents of clinical cases and reported in these animals are: Cryptosporidium spp., Eimeria spp., Isospora spp., Leishmania infantum, Neospora caninum, Sarcocystis spp., and Toxoplasma gondii. Unfortunately, in most reports, neither clinical symptoms nor detailed pathogenicity and clinical pathology are satisfactory discriminated. This data is essential for preparing treatment and preventive measure protocols. Additionally, attempts to better understand the infections might be useful in the case of outbreaks.RESULTS:The results of the literature research revealed that some detected infections may have zoonotic potential, and are quite likely to be transmitted in both directions. The role of vectors, e.g. small rodents, birds, insects (sand-flies), or pets (cats and dogs), which might be involved in the transmission of various parasitic infections, is also discussed. The summarized list of protozoans involved can be used for further studies on the health and welfare aspects of fur farm animal breeding and public health issues.

Introduction: Canine roundworm T. canis and T. leonina infections were investigated in experimentally infected farm mink (Neovison vison) to describe the pattern of pathological lesions in this paratenic host. Material and Methods: Infections in mink developed following ingestion of embryonated eggs of either parasite or mice tissue infected with both parasite species. Results: Comparative analysis of haematoxylin- and eosin-stained slides showed essential differences among the experimental groups. The lesions observed included eosinophil and mononuclear inflammatory infiltrates of the intestinal wall and local lymph nodes, inflammation and haemorrhages in liver tissues, and interstitial inflammation and mineralisation of the kidneys and lungs. Larvae migrating through the minks' bodies also caused particularly salient enlargement of lymphoid follicles in the spleen and inflammatory infiltrates of mononuclear cells in skeletal and heart muscles. Conclusions: It is assumed that histopathological lesions appeared as a local and general host response to invasive L3 T. canis and T. leonina larvae migrating through the tissues of infected farm mink Interestingly, mink infected with embryonated eggs had more pronounced lesions than animals infected with tissue larvae. Detailed histopathological examinations of parenchymal organs and striated muscles revealed lesions resembling those observed in other paratenic host species due to toxocarosis.

INTRODUCTION:Farm mink (Neovison vison) can be naturally exposed to T. canis and T. leonina pathogens on the farm. If mink were hosts, it would imply some veterinary public health as well as animal welfare issues. For this reason, the aim of the study was to determine whether mink might be definitive or paratenic hosts of these parasites.MATERIAL AND METHODS:Four groups of mink were infected with both parasite species using larvated eggs or feed containing mouse tissue previously infected with the parasites. Following inoculation, the infections were monitored in vivo by faecal examination for 14 weeks p.i., and then western blotting and ELISA were performed.RESULTS:Coprology did not reveal any canine roundworm eggs, neither were nematodes found in mink intestines during post mortem examination. The specific IgG antibodies recognising excretory/secretory (ES) antigens of both parasite species were identified in mink sera. Single T. leonina tissue larvae were found in digested organs.CONCLUSIONS:Our results confirm that farm mink may contribute both T. canis and T. leonina infections. It was proved that farm mink were not their definitive hosts, and therefore mink faeces need not be considered a source of canine roundworm eggs in any soil it fertilises. Nonetheless, as farm mink may be a paratenic host for both parasite species, this may have some impact on the health and welfare of infected animals.

The aim of the study was to compare the intestinal parasites in different groups of pigs (sows, weaners, fatteners) before and after the application of a deworming programme. The experiment was carried out twice, in February and in June 2011 on a farm keeping pigs in a deep litter system. Samples of faeces for analyses were collected twice from 15 sows, 7 weaners and 18 fatteners. In total, 80 samples of faeces were examined using flotation and the McMaster technique. Three parasitic species belonging to the phylum Nematoda (Strongyloides ransomi, Ascaris suum, Oesophagostomum dentatum) and one protozoan species from the phylum Apicomplexa (Eimeria sp.) were detected. The highest prevalence of parasite infection was recorded in sows in the period before deworming (86.7%). Differences in the intensity and prevalence of intestinal parasite invasion were observed before and after the prevention program was applied. The intensity of Ascaria suum infestation in sows before and after deworming differed significantly (P≤0.05). The highest intensity of parasitic invasion was observed after deworming in the group of fatteners (26 800 oocysts of Coccidia per 1 gram of faeces). Multi-species invasions were also observed in some infected pigs. These results indicate the necessity to implement parasitological screening protocols in pig herds to control and implement prevention and/or control programmes.

Angiostrongylus chabaudi (arrows) on the endothelium of the right ventricle of a wildcat in Northern Greece

The effect of Toxocara larval antigens on cytokine secretion by mouse splenocytes was studied in vitro. Recombinant mucins were produced in Pichia pastoris yeast, and Toxocara excretory-secretory (TES) antigens were collected from in vitro culture of L2 larvae. Tc-MUC-2, Tc-MUC-3, Tc-MUC-4, and Tc-MUC-5 were expressed as glycoproteins and were specifically recognized by Toxocara canis-infected dog serum antibodies. Mouse splenocytes stimulated with recombinant mucins produced IL-5, IL-6, and TGF-β. Cell stimulation with whole TES products was more effective and resulted in secretion of IL-4, IL-5, IL-6, IL-10, and TGF-β and downregulation of TNF-α production. IFN-γ and IL-17 secretion was noted only after ConA treatment. Cells originating from infected animals produced significantly smaller amounts of these two cytokines compared to control cells, which suggests that Th1 and Th17 response in infected mice is strongly inhibited. However, splenocyte stimulation with both TES and ConA upregulated the production of IFN-γ and IL-17. This shows that TES antigens have strong immunomodulatory properties and are able to induce a broad range of effects on murine immune cells.