BACKGROUND: Toxocara canis is a zoonotic disease that commonly infects canids. Mammals and birds are sometimes infected with this disease as paratenic hosts. It can also cause accidental infection in humans. The increase in the number of stray dogs, the expansion of urban gardens, and the proximity of dogs to humans increase the risk of human infection with Toxocara canis.OBJECTIVES: This study aims to determine the prevalence of Toxocara canis infection in dogs and foxes in Zanjan province, Iran.METHODS: A total of 484 fecal samples of stray dogs (n=355), rescue dogs (n=49), guard dogs (n=50), and foxes (n=30) in Zanjan were randomly collected from June 2021 to February 2022. The microscopic examination was done following formalin-ethyl acetate sedimentation procedures. Finally, the PCR method was used to confirm the presence of Toxocara canis in positive samples.RESULTS: Microscopic study revealed that, out of 484 samples, 21 (4.3%) were positive for Toxocara/ Toxascaris eggs. Between these positive samples of dogs and foxes, only 6 samples from dog feces were confirmed as a Toxocara canis infection by the PCR method.CONCLUSIONS: There is an increase in the prevalence of Toxocara canis infection in stray dogs in Zanjan, Iran. Given the presence of dogs in parks and residential areas, there is a risk of human infection with Toxocara canis, emphasizing the importance of adhering to treatment and prevention protocols in dealing with stray dogs.

BACKGROUND: Toxocara canis C-type lectin (T. canis-CTL) is the main protein part of the secretory-excretory product secreted by Toxocara canis infective larvae. T. canis-CTL can stimulate immune response-mediated regulatory T lymphocytes, increase the FOXP3+ cells population, and reduce severe inflammatory responses. T. canis-CTL is a promising candidate for immune modulation in some autoimmune diseases, deserving further investigation.OBJECTIVES: The current research aimed to purify the recombinant T. canis-CTL under denaturation and native condition to increase exploration and maintain biological activity.METHODS: The expression vector, pET32a was constructed with the partial sequence 660bp of T. canis-CTL and expressed in E. coli BL21 (DE3). T. canis-CTL protein expression was induced by IPTG (1 mM) at 37°C after 6 h. In this study, different buffers were used for cell explosion and recombinant protein solubilization, including lysis buffer with urea (8 M, pH=8) and lysozyme enzyme as well as lysis buffer with Imidazole (0.01 M) and lysozyme enzyme, and previous buffers in addition to sonication. The effect of these buffers was evaluated in bacterial cells explosion, using Gram-staining and microscopic examination. Recombinant T. canis-CTL protein was extracted and purified under denaturation and native condition using Ni-NTA affinity chromatography by agarose and sepharose resin. A New Zealand male rabbit was immunized with the recombinant protein to evaluate the bioactivity of the protein. RESULTS: Lysis buffer with urea (8 M, pH=8) and lysozyme enzyme, in addition to sonication, provided acceptable results, and an additional amount of recombinant T. canis-CTL protein was secreted in the buffer. Protein purification under denaturation conditions with Ni-NTA agarose affinity chromatography also provides further recombinant protein. Most of the induction of recombinant T. canis-CTL with 41 KDa molecular weight was collected 6 h after induction at 37°C. Dot-blot results illustrate the brown dot, which showed a 1:500 titer of specific IgG polyclonal antibody has developed in the sera of rabbits immunized with T. canis-CTL recombinant protein.CONCLUSIONS: The denaturation condition did not affect the biological activity of the T. canis-CTL recombinant protein and can recover a further amount of recombinant proteins.

BACKGROUND: Toxocaiasis is an important zoonotic disease caused by the second stage larvae of Toxocara canis and Toxocara cati. Toxocariasis is the most common worm infection in several temperate countries and causes severe complications. C-type lectin is one of the larval stage products of this parasite. It is involved in immune responses, including cellular signals in vertebrate immunity, activation of innate immunity in vertebrates and non-vertebrates, and induction of homeostasis.  OBJECTIVES: The current research aimed to optimize the production of recombinant C-type lectin of Toxocara canis and investigate its antigenic properties. METHODS: Reference nucleotide sequence of lectin type C (CTL) of Toxocara canis (T. canis) was extracted from Genbank. Recombinant Plasmid, pET32a, including the desired sequence was then constructed by GENERAY. The recombinant plasmid was transformed to Escherichia coli strain BL21 (DE3). The expression of the recombinant protein was investigated using SDS-PAGE and dot blot methods and approved with human T. canis positive serum. RESULTS: The findings of the present study showed that optimization and high level production of recombinant protein expression was achieved by selecting Escherichia coli (BL21 (DE3), 37 °C temperature for 4 hours after induction and 1 mM IPTG concentration. After optimization, the recombinant protein was obtained at a concentration of 1160±0.6 µg/mL. The molecular weight of the resulting recombinant protein was 42 kDa. Recombinant plasmid passage in Escherichia coli DH5α strain caused a significant increase in recombinant protein expression. The results of condition optimization evaluation, with SDS-PAGE and Dot blot methods, showed that the highest production of recombinant C type lectin protein of T. canis was obtained under optimized conditions. CONCLUSIONS: Based on these results, to produce reasonable amounts of specific C-type lectin recombinant protein, further studies are needed to evaluate its immunogenicity and protection against Toxocara canis infection.

Eight trials were conducted in dogs to document the efficacy of ivermectin (6 micrograms/kg of body weight) and pyrantel pamoate (5 mg of active pyrantel/kg) in a beef-based chewable formulation against Dirofilaria immitis, Ancylostoma caninum, Uncinaria stenocephala, Toxocara canis, and Toxacaris leonina. Three studies involved induced infection with D immitis, and 5 studies involved induced or natural infection with hookworms and ascarids. In 3 intestinal parasite trials, the efficacy of the combination chewable tablet was compared with each of its components. Results indicated that 1 component did not interfere with the activity of the other. In 1 heartworm and 2 intestinal parasite trials, the efficacy of pyrantel, ivermectin/pyrantel combination, or ivermectin with pyrantel dosage of 10 mg/kg was evaluated. The ivermectin/pyrantel combination was 100% effective in preventing development of D immitis larvae. Efficacy of the combined product against T canis, Toxascaris leonina, A caninum, and U stenocephala was 90.1, 99.2, 98.5, and 98.7%, respectively. In the intestinal parasite trials, each individual component was found not to interfere with the anthelmintic action of the other. Increasing the dosage of pyrantel to 10 mg/kg (2 X that in the combination) did not interfere with the efficacy of ivermectin against heartworm or increase the activity of pyrantel against intestinal parasites.

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.

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.

Toxocara canis and Toxocara cati are roundworms of dogs and cats. The purpose of this study was to investigate the infection caused by these ascarids in cats and dogs, using microscopic and molecular analysis methods. Adult ascarids were gathered from the faeces of dogs and cats in Van province, in 2015–2016. Existing keys and PCR sequencing of the ITS-2 fragment were used to identify the morphological features of the parasite species. It was observed that out of 20 adult ascarids, 17 and 3 were found to be Toxocara canis and Toxocara cati, respectively. The ITS-2 gene region was amplified by PCR to perform molecular analysis. Genotyping indicated that the dogs and cats were infected with T. canis and T. cati, respectively, and none had Toxascaris leonina. To the best of our knowledge, this is the first report on the molecular characteristics of adult ascaridoid nematodes from cats and dogs in Turkey. The molecular approaches established in this study enable molecular identification and genetic structure studies of the ascaridoids.

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.

A male mixed-breed dog of unknown age was presented with a history of bloody diarrhea and cachexia. Toxocara canis in vomitus was identified by a parasitologist. Hematology revealed low hematocrit, eosinophilia, and low albumin. Computed tomography (CT) revealed an enlarged pulmonary artery with an irregular wall, micronodules in the lung, and vicarious excretion of contrast medium to small intestine. CT scan was helpful for identifying lung lesions and the central organs of larval migration and also show vicarious excretion of contrast medium to the small intestine in T. canis infection.

Fourteen cats were inoculated orally with 1 of 2 infective doses of Toxocara canis to induce eosinophilia. Cats were subsequently challenge exposed twice via intraperitoneal injection with 1 of 2 T canis antigen preparations. Peritoneal lavage was performed 2 days after antigenic challenge exposure, and eosinophils in the peritoneal lavage fluid were quantified. None of the cats developed clinical signs of disease after infection. All cats developed peripheral eosinophilia after infection. Significant (P < 0.05) difference in mean eosinophil count from the lavage fluid was observed between lavage 1 (prechallenge exposure) and lavages 2 and 3 (postchallenge exposure) in both groups of cats. Significant difference in eosinophil count was not found between cats given different doses of eggs. After initial challenge exposure, significantly (P < 0.05) more eosinophils were obtained from cats given antigen preparation 2 (prep-2) than from those given antigen prep-1. This difference was no longer observed after the second challenge exposure with higher doses of either antigen prep-1 or prep-2. In cats given antigen prep-2, significant difference was not found between lavages 2 and 3. However, in cats given antigen prep-1, eosinophil count was significantly (P = 0.005) greater in fluid obtained from lavage 3, compared with eosinophil count from lavage 2. Mean +/- SEM percentage of eosinophils in the fluid from lavage 3 in all cats was 70.8 +/- 2.2%. Other cell types included macrophages, neutrophils, lymphocytes, and mast cells. Gross postmortem findings were mild. One- to 3-mm nodular white foci of inflammation were observed on the serosal surfaces of the liver, spleen, kidneys, and omentum. Microscopic examination of tissues revealed pulmonary artery hypertrophy (n = 4), eosinophilic peribronchitis and perivasculitis (n = 10), mild granulomatous interstitial nephritis (n = 6), interstitial pancreatitis (n = 1), focal lymphocytic myocarditis (n = 1), focal eosinophilic granulomatous hepatitis (n = 1), and eosinophilic hyperplasia of bone marrow (n = 14). Large numbers of eosinophils could be harvested from the peritoneal cavity of cats inoculated orally with 500 embryonated T canis eggs and subsequently challenge-exposed intraperitoneally with preparations of parasite antigens. After the second challenge exposure, at least 108 eosinophils could be harvested from each cat, yielding eosinophils in the quantity required to begin isolation of granule constituents.