Ovicidal in vitro activity of the fixed oil of Helianthus annus L. and the essential oil of Cuminum cyminum L. against Fasciola hepatica (Linnaeus, 1758).

The fascioliasis is a parasitic disease of importance in veterinary medicine and public health. For this parasitosis, the treatment by synthetic fasciolicides is used and due to their intense use although they have been shown less effective because of the establishment of resistant Fasciola hepatica population to these drugs, with a global concern. The use of derived products of plants with biological activity has been shown promising in the control of parasites. In this context, we evaluated the chemical composition and action of ovicidal in vitro fixed oil of Helianthus annuus L. (FOH) and essential oil of Cuminum cyminum L. (EOC), as well as their combination (FOH + EOC) of F. hepatica. In the assay in vitro of F. hepatica were submitted to different concentrations of oils, such as FOH (2.3 mg/mL + 0,017 mg/mL); EOC (2.07 mg/mL + 0,004 mg/mL) and the combination of (1.15 mg/mL + 1.03 mg/mL to 0,0085 mg/mL + 0,008 mg/mL) as well as a positive control of thiabendazole (0.025 mg/mL) and a negative control with distilled water and tween. The identification of the majority chemical compounds was performed by gas chromatography. The -cell viability of the oils was tested in MDBK cellular line by the MTT method. The majority compounds in the FOH were the linoleic (53.6%) and oleic (35.85%) unsaturated fatty acids, and the majority phytochemicals compounds in the EOC were the Cumaldehyde (26.8%) and the 2-Caren 10-al (22.17%). The EOC and the combination presented effectiveness of 99% (±1) and of 94% (±1) in the concentration of 0.03 mg/mL and 0.035 mg/mL+0.03 mg/mL, respectively, and the FOH was insufficiently active as ovicidal. The cell viability at this concentration of EOC was 93%. From the results above we could infer that the EOC is promising as a new alternative for the fascioliasis control.

Essential oil from Brazilian Red Propolis exhibits anthelmintic activity against larvae of Toxocara cati.

Toxocara spp. are responsible for causing toxocariasis, a zoonotic disease of global importance, which is difficult to treat as the available drugs have moderate efficacy in the clinical resolution of the disease. A promising alternative to the existing drugs is Propolis, which is known for having biological and pharmacological properties such as antiparasitic, antioxidant, and antitumor activities. In this study, we report the in vitro anthelmintic activity of essential oil from Brazilian Red Propolis (EOP) against larvae of Toxocara cati. Approximately 100 larvae per well were cultivated in microplates containing RPMI-1640 medium and incubated in the presence of EOP (18.75, 37.5, 75, 150, 300 and 600 µg/mL) to determine the Minimum Inhibitory Concentration (MIC) and IC50 (concentration required to inhibit 50% of the population) values. Then, T. cati larvae treated with the MIC of EOP were inoculated in mice to evaluate their progression in vivo. A concentration of 600 µg/mL of EOP showed 100% larvicidal activity after exposure for 48 h, while 300 µg/mL represented the IC50 and CC50. The anthelmintic activity of EOP was confirmed by the inability of the treated T. cati larvae to infect the mice. Our findings demonstrate the potential of EOP as an anthelmintic.

Reactivity of recombinant Toxocara canis TES-30/120 in experimentally infected mice.

AIM: While the use of recombinant antigens is being widely investigated in the diagnosis of human toxocariasis, relatively little attention has been given to animal diagnostic models. For this reason, this study aimed to investigate the diagnosis potential of Toxocara canis TES-30 and TES-120 recombinant antigens in mice, the animal model for toxocariasis studies. METHODS AND RESULTS: Serum samples obtained from mice infected with T. canis or Toxocara cati were tested by indirect ELISA using T. canis TES-30 and TES-120 recombinant antigens produced in Escherichia coli. 90% of the samples reacted with rTES-30, whereas there was almost no reactivity with rTES-120. CONCLUSION: Despite rTES-120 being a good antigen for diagnosis in humans, it could not reproduce its reactivity in this animal model. As rTES-30 has good reactivity in mice, it is a valuable tool for diagnosis.

Ovicidal in vitro activity of 2-aryl-3-(2-morpholinoethyl)thiazolidin-4-ones and 2-aryl-3-(3-morpholinopropyl)thiazolidin-4-ones against Fasciola hepatica (Linnaeus, 1758).

Although there is a variety of biological activity reports regarding compounds derived from thiazolidin-4-ones, no data related to ovicidal activity against trematodes, particularly Fasciola hepatica are available. Since there are reports about anthelmintic resistance in F. hepatica, new drugs are required. Thus, this study evaluated ovicidal action in vitro against F. hepatica eggs in two systematic series of thiazolidin-4-ones: 2-aryl-3-(2-morpholinoethyl)thiazolidin-4-ones (1a-h) and 2-aryl-3-(3-morpholinopropyl)thiazolidin-4-ones (2a-h) at different concentrations (20, 2, 0.2, 0.02 and 0.002 µg/ml). The egg hatch assay (EHA) was used to evaluate the ovicidal action property of such compounds. In addition, potential negative effects of the compounds on metabolic activity of bovine kidney (MDBK) cells were evaluated by determining mitochondrial dehydrogenase activity. The eggs used in the EHA were obtained from parasites removed from the liver of cattle, which were discarded by slaugh after sanitary inspection. The results of EHA showed that compounds 2a-h exhibited ovicidal activity, especially compounds 2b which showed 90% ovicidal activity and viability of 93% MDBK cells at the concentration of 2 µg/ml; and 2e with 96-99% ovicidal activity at 0.2 µg/ml, 0.02 µg/ml and 0.002 µg/ml. The results show the potential of compound 2b to continue the studies in the production of new compounds with anthelmintic action.

Lactobacillus rhamnosus reduces parasite load on Toxocara canis experimental infection in mice, but has no effect on the parasite in vitro.

Human toxocariasis is a neglected global parasitic zoonosis. The efficacy of drug treatment for this disease has been hindered by the biological complexity of the main etiological agent, the nematode Toxocara canis. Experimental studies have shown the potential of probiotics to promote a reduction in the parasite load of T. canis larvae. This study aimed to evaluate the effect of probiotic Lactobacillus rhamnosus ATCC 7469 on the parasite load of BALB/c mice with acute toxocariasis and evaluate the direct effect of this probiotic on T. canis larvae in vitro. In vivo administration of probiotics reduced the parasite load of T. canis larvae by 53.3% (p = 0.0018) during the early stage of infection in mice. However, when analyzed in vitro, it was observed that the probiotic did not present a deleterious effect on the larvae, as approximately 90% of these remained viable. These results demonstrate the potential of the probiotic L. rhamnosus in the reduction of T. canis larvae in BALB/c mice and suggest it could be used as an alternative means for the controlling of visceral toxocariasis. However, further studies are required to elucidate the mechanisms of action promoted by this probiotic.

Toxocara canis: Larvicidal activity of fatty acid amides.

Considering the therapeutic potential of fatty acid amides, the present study aimed to evaluate their in vitro activity against Toxocara canis larvae and their cytotoxicity for the first time. Linoleylpyrrolidilamide was the most potent, with a minimal larvicidal concentration (MLC) of 0.05 mg/mL and 27% cytotoxicity against murine peritoneal macrophages C57BL/6 mice, as assessed by the MTT assay.

Larvicidal activity of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus.

Effective control of gastrointestinal parasites is necessary in sheep production. The development of anthelmintics resistance is causing the available chemically based anthelmintics to become less effective. Biological control strategies present an alternative to this problem. In the current study, we tested the larvicidal effects of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus larvae. Bacterial suspensions [2 × 108 colony-forming units (CFU) g-1 of the feces] of B. thuringiensis var. israelensis and recombinant Escherichia coli expressing Cry11Aa toxin were added to naturally H. contortus egg-contaminated feces. The larvae were quantified, and significant reductions of 62 and 81% (P < 0·001) were, respectively observed, compared with the control group. A 30 mL bacterial suspension (1 × 108 CFU mL-1) of B. thuringiensis var. israelensis and recombinant E. coli expressing Cry11Aa toxin were then orally administered to lambs naturally infected with H. contortus. Twelve hours after administration, feces were collected and submitted to coprocultures. Significant larvae reductions (P < 0·001) of 79 and 90% were observed respectively compared with the control group. The results suggest that the Cry11Aa toxin of B. thuringiensis var. israelensis is a promising new class of biological anthelmintics for treating sheep against H. contortus.

Toxocariasis and childhood asthma: A case-control study.

OBJECTIVE: The objective of this study is to investigate the association between anti-Toxocara IgG seropositivity and asthma in children. METHODS: This was a case-control study conducted in a university hospital in south Brazil between May 2012 and June 2013. Were recruited 208 children up to 12 years old of whom 156 had asthma (cases) and 52 did not have asthma (controls), with a case-control ratio of 3:1 matched by age. Children's parents or guardians were interviewed using a structured questionnaire with closed questions. Serology was performed using enzyme-linked immunosorbent assay (ELISA) with excretory-secretory antigen of Toxocara canis (TES). RESULTS: The seroprevalence of IgG anti-T. canis antibodies was 12.8% in the cases and 7.7% in the controls. There was no significant association between seropositivity to T. canis and risk of asthma (adjusted odds ratio [OR]: 1.89, 95% CI: 0.52 to 6.89, p = 0.33). Household income < 2 minimum salaries, paternal school years < 9, allergic rhinitis in children, a positive family history of asthma and rhinitis and contact with cats were significantly associated with asthma, with adjusted ORs (95% CIs) of 3.05 (1.21 to 7.73), 2.83 (1.11 to 7.18), 10.5 (4.32 to 25.6), 2.65 (1.14 to 6.17), 2.49 (1.07 to 5.78) and 2.73 (1.03 to 7.27), respectively. CONCLUSIONS: This study did not find a statistically significant association between seropositivity to Toxocara sp. and risk of asthma in children. Low family income, low paternal education level, concomitant allergic rhinitis, family history of asthma and allergic rhinitis and contact with cats were independent factors associated with childhood asthma.

Diagnostic potential of anti-rNcp-43 polyclonal antibodies for the detection of Neospora caninum.

Neosporosis is a disease caused by the apicomplexan parasite Neospora caninum, which is closely related to Toxoplasma gondii. N. caninum infection represents an important cause of reproductive failure in sheep, goats, horses, and cattle worldwide. The diagnosis of neosporosis is based on the detection of pathogen-specific antibodies in animal sera or the presence of tissue cysts. However, morphological similarities and serological cross-reactivity between N. caninum and T. gondii can result in the misdiagnosis. In this study, the N. caninum tachyzoite surface protein Ncp-43 was expressed in a recombinant form to elicit polyclonal antibodies (pAb) response. The pAb was purified and conjugated to horseradish peroxidase (HRP) or fluorescein isothiocyanate (FITC) to detect the recombinant and native Ncp-43 proteins, respectively. The pAb and pAb/HRP were able to recognize rNcp-43 by dot blot and ELISA, and pAb/FITC immunolabeled the apical complex of tachyzoites. A blocking enzyme-linked immunosorbent assay (b-ELISA) was performed to evaluate pAb/HRP as a diagnostic tool. The mean percent inhibition for the positive and negative serum samples from cattle with neosporosis was significantly different (P < 0.0001). These results suggest that the pAb may bind to the same epitopes of Ncp-43 as anti-N. caninum antibodies in the positive samples tested. The b-ELISA using the pAb/HRP can facilitate diagnostic testing for neosporosis, since fewer steps are involved, and cross-reactivity with secondary antibodies is avoided. In summary, this report describes the production of antibodies against N. caninum, and evaluates the potential of these tools for the development of new diagnostic tests for neosporosis.

An evaluation of techniques to diagnose Dioctophyme renale in dogs.

Dioctophyme renale is a nematode with zoonotic potential that affects the kidneys of carnivorous, wild, and domestic mammals. In this study, we sought to evaluate the indirect ELISA method against routine methods used to diagnose dioctophimosis. Hence, 38 dogs parasitized by D. renale, as confirmed by surgery, were selected. The dogs were evaluated by abdominal ultrasound and urinalysis, and their sera were tested by indirect ELISA using D. renale adult secretion and excretion antigen (DES). Five dogs were followed up with serum collections on day 0 (day of surgery) and 30, 60, and 90 days after surgery to evaluate antibody kinetics. Abdominal ultrasound and indirect ELISA successfully diagnosed 37 dogs parasitized by D. renale, while urinalysis diagnosed 29 animals. The positive animals were parasitized with 1-7 parasites; 17 dogs were infected by male and female parasites, 15 only by female parasites, and six were parasitized only by male parasites. When assessing specificity and sensitivity, all techniques showed 100% specificity and 81.6%, 97.4%, and 97.4% sensitivity for urinalysis, ultrasound, and ELISA, respectively (p < 0.001). The five positive dogs that were followed up after surgery showed a progressive decrease in mean absorbances in indirect ELISA (0.644, 0.516, 0.511, and 0.440, respectively). This study demonstrated that the indirect ELISA using the DE antigen could diagnose dioctophimosis regardless of the number, sex, and location of the parasites, with the potential to be used in epidemiological research and implementing immunological and molecular studies, opening new lines of research on D. renale.

Dioctophyme renale é um nematódeo que possui potencial zoonótico e acomete o rim de mamíferos carnívoros, silvestres e domésticos. Neste estudo busca-se avaliar a técnica de ELISA indireto frente metodologias de rotina utilizadas no diagnóstico da dioctofimose. Para isto, 38 cães participaram do estudo, sendo todos parasitados por D. renale, confirmados por cirurgia. Esses cães foram avaliados por ultrassom abdominal, urinálise e seus soros testado por ELISA indireto utilizando antígeno de excreção e secreção (DES) de adultos de D. renale. Destes, cinco cães foram acompanhados com coletas de soro, no dia zero (dia da cirurgia) e após 30, 60 e 90 dias da cirurgia para avaliar a cinética dos anticorpos. O ultrassom abdominal e ELISA indireto apresentaram sucesso no diagnóstico de 37 cães parasitados por D. renale, enquanto que a urinálise diagnosticou 29 animais. Os animais positivos possuíam de um a sete parasitos, 17 cães apresentaram infecções por macho e fêmeas, 15 somente por fêmeas e seis foram parasitados apenas por machos. Na avaliação da especificidade e sensibilidade, todas as técnicas apresentaram 100% de especificidade e 81,6%, 97,4%, 97,4% de sensibilidade para urinálise, ultrassom e ELISA, respectivamente (p < 0,001). Os cinco cães positivos que foram acompanhados após a cirurgia apresentaram diminuição progressiva nas médias de absorbâncias no ELISA indireto (0,644, 0,516, 0,511 e 0,440, respectivamente). O estudo demonstrou que o ELISA indireto utilizando o antígeno DES poderia diagnosticar dioctofimose, independentemente do número, sexo e localização dos parasitos, com potencial para ser utilizada em estudos epidemiológicos e na implementação de estudos imunológicos e moleculares, abrindo novas linhas de pesquisa sobre Dioctophyme renale.

Evaluation of the initial and chronic phases of toxocariasis after consumption of liver treated by freezing or cooling.

Human toxocariasis is a neglected parasitic zoonosis of worldwide distribution. The consumption of raw or undercooked meat and offal from paratenic hosts of the Toxocara canis nematode can cause infection in humans, but there have been a lack of studies examining specific prophylactic measures to combat this mode of transmission. The aim of this study was to evaluate the establishment of infection by T. canis larvae at the initial and chronic phases of visceral toxocariasis after the consumption of mouse liver subjected to cold treatment. This study was divided into two stages using groups (G) of five donor mice inoculated with 2,000 eggs of T. canis. Two days post-inoculation, the livers of donor mice in G1 and G2 were kept at -20 °C and between 0 and 4 °C, respectively, for 10 days. In the first stage of the study, the livers of mice from G1, G2, and G3 (control) were subjected to a tissue digestion technique and found to be positive for infection. In the second stage, which evaluated infection in mice that had consumed livers from donor mice, receiver mice of G4 and G7 were fed with livers of donor mice from G1 (freezing), receiver mice of G5 and G8 were fed with livers of donor mice from G2 (cooling), and receiver mice of G6 and G9 with livers from G3 (control). Then, the tissue digestion technique was performed for recovering larvae from organs and carcasses of mice, at 2 days (G4, G5, and G6) and 60 days after liver consumption (G7, G8, and G9). It was observed that freezing inhibited the viability of 100 % of the larvae, while cooling promoted 87.7 and 95.7 % reductions in the intensity of infection at 2 and 60 days after liver consumption, respectively. Under the studied conditions, cold treatment shows great potential to help control this parasitosis, both in the initial and chronic phases of toxocariasis.

Toxocara spp. seroprevalence in sheep from southern Brazil.

Visceral toxocariasis is a neglected parasitic zoonosis that occurs through the ingestion of embryonated Toxocara spp. eggs. A wide range of animal species can act as paratenic hosts for this ascarid. The main risk factor for humans is the ingestion of the eggs from contaminated soil; however, infection can also occur through the ingestion of contaminated raw or undercooked infected meat from paratenic hosts. The aim of this study was to verify the presence of Toxocara spp.-specific antibodies in sheep and to determine the risk factors associated with the infection of sheep in Rio Grande do Sul (a major sheep-producing and sheep-consuming state) in southern Brazil. Serum samples collected from 1,642 sheep were tested using an IgG enzyme-linked immunosorbent assay based on the excretory-secretory Toxocara canis antigen. Seroprevalence was 29.0% (477/1,642), and every farm included in the study contained at least one seropositive animal. These results indicate that T. canis infection is widely distributed among sheep herds in Rio Grande do Sul and that it represents a potential risk to human health.

Immunomodulation in the intestinal mucosa of mice supplemented with Lactobacillus rhamnosus (ATCC 7469) and infected with Toxocara canis.

Toxocariasis is an anthropozoonosis caused by the helminth Toxocara canis that shows different clinical manifestations as visceral, ocular, or neurological toxocariasis forms. Probiotics have been studied as alternatives to prevent and treat this parasitosis. Lactobacillus rhamnosus is a prospect that presents immunomodulatory activity that acts to strengthen the intestinal barrier. In this context, the main objective of this study was to evaluate the protective capacity and immunomodulatory action of the probiotic Lactobacillus rhamnosus at the level of the intestinal mucosa in different stages of T. canis infection (acute and chronic). Mice were supplemented by oral gavage with 1 × 107 UFC/mL L. rhamnosus for 15 days before inoculation with 100 embryonated eggs of T. canis. Euthanasia of mice was conducted at three different time points: 2, 15 and 30 days post-inoculation (PI). The brain, lungs and liver were collected to evaluate the intensity of infection. The small intestines were removed, and mucosal cells of the duodenum were collected to perform gene analysis of IFN-γ, IL-10, IL-4 and IL-13 by real-time polymerase chain reaction (qPCR). Jejunum and ileum segments were analysed by histological techniques. A reduction of 51% in infection intensity was observed in the tissue of supplemented animals evaluated 2 days PI; however, analysis of groups 15 and 30 days PI did not show a protective effect. The intestinal mucosa of supplemented animals presented an inflammatory process that initiated at 2 days PI, persisted at 15 days PI and had regressed at 30 days PI. IL-13 transcription was increased in the probiotic group 2 days after supplementation ended; however, the same increase was not observed in the group that was supplemented and infected. Toxocara canis modulated the local immune system, with suppression of IFN-γ at 2 days PI and increased levels of IL-4 and IL-10 at 15 days PI. These results indicate that, under the studied conditions, the protective effect of Lactobacillus rhamnosus against infection caused by T. canis is not related to IL-4, IL-10 or IFN-γ but could be influenced by IL-13 action at 2 days PI. The probiotic stimulated immune cell recruitment to the intestinal mucosa, which can be involved in the diminished capacity of larval penetration in the mucosa, resulting in the reduced infection intensity observed during acute infection.

Evaluation of the immunosuppressive effect of cyclophosphamide and dexamethasone in mice with visceral toxocariasis.

Visceral toxocariasis is a serious public health problem with a cosmopolitan distribution. Children are susceptible due to their immature immune system and high risks of infection. Nevertheless, the few completed studies about immunosuppression have had controversial results. To evaluate the effect of two immunosuppressive drugs on the larval burden of Toxocara canis, four groups of ten Swiss strain mice each were inoculated on day 0 with 1,200 embryonated T. canis eggs. Fifteen days before the experimental infection, group 1 (control) was treated via intraperitoneal injection (IP) with sterile distilled water and groups 2 and 3 were treated with dexamethasone (DEX) at 1 and 5 mg/kg/day, respectively. Additionally, group 4 was treated IP with cyclophosphamide (CY) at 50 mg/kg at two times per week for 2 weeks. Sixty days following infection, the mice were euthanised to recover the larvae by means of the tissue digestion technique. The levels of antibodies detected by indirect ELISA were not associated with the larval burden. Administration of CY (50 mg/kg) and DEX (5 mg/kg) resulted in an increase of the larval burden of 162.1% and 50.8%, respectively, in relation to the control group. These two treatments, especially CY (50 mg/kg), promoted immunosuppression and the establishment of a significant larval burden, supporting its further utilisation in studies related to immunosuppression in visceral toxocariasis.

Dioctophyme renale: in vitro culture, production of excretory and secretory antigens, and their use in immunodiagnosis.

The nematode Dioctophyme renale affects mainly the right kidney of domestic and wild mammals, in addition to having zoonotic potential. Therefore, this study aimed to produce excretion and secretion antigens of adult D. renale (DES) to diagnose canine dioctophimosis. To obtain the excretion and secretion antigens (DES), five D. renale adults were maintained for three weeks in supplemented RPMI medium with weekly supernatant collections. After the DES concentration, 2200 mL of the collected supernatant was used, which was centrifuged, followed by two filtrations and dialyzed, resulting in 12.5 mL of DES with a protein concentration of 0.59 mg/mL. The polyacrylamide gel electrophoresis (SDS-PAGE) of the DES antigen showed fractions with molecular weights ranging from 30 to 250 Kda. In the indirect ELISA with the DES antigen, the mean absorbance of the positive sera (38) was 0.839 ± 0.267, and the mean of the negative control sera (7) was 0.208 ± 0.083; the specificity and sensitivity of the ELISA were 100 and 97.4 %, respectively, being as effective as the surgical method in the presence of viable parasites. Thus, for the first time, this study describes the production of excretion and secretion antigens of adult D. renale and standardizes an indirect ELISA to diagnose dioctophimosis.

Antiparasitic treatment using herbs and spices: A review of the literature of the phytotherapy.

This study sought to make a literature review of the medicinal plants Origanum majorana, Origanum vulgare L., Thymus vulgaris L., Cuminum cynimum L., and Rosmarinus officinalis L. with antiparasitic potential. Articles and theses were selected from the LILACS, PubMed, and Google Scholar databases, which comprised the period from 2000 to 2021 (22 years). In all, 49 studies were selected, and the majority were with the plant Origanum vulgare L. (oregano), followed by Thymus vulgaris L. (thyme). Twenty-five genera of parasites were detected, which were described being tested with phytotherapic. The nematode Haemonchus spp. was the most evaluated in these studies, followed by the parasite genera Leishmania, Trichostrongylus, and Toxocara. All plants showed antiparasitic effects, with more or less action, therefore with the potential to continue research in the search for biomolecules to control these parasites.

O presente trabalho faz uma revisão bibliográfica das plantas medicinais Origanum majorana, Origanum vulgare L., Thymus vulgaris L., Cuminum cynimum L. e Rosmarinus officinalis L. com potencial antiparasitário. Foram selecionados artigos e teses nos bancos de dados LILACS, PubMed e Google Acadêmico que compreendiam o período de publicação de 2000 a 2021 (22 anos). Ao todo, foram selecionados 49 estudos, sendo que na maioria constava a planta Origanum vulgare L. (orégano), seguido de Thymus vulgaris L. (tomilho). Foram detectados 25 gêneros de parasitos, os quais foram descritos sendo testados frente a algum fitoterápico. O nematoda Haemonchus spp. foi o mais avaliado nestes estudos, seguido dos gêneros dos parasitos Leishmania, Trichostrongylus e Toxocara. Todas as plantas apresentaram efeitos antiparasitários, com maior ou menos ação, portanto com potencial para dar continuidade aos estudos em buscas de biomoléculas para controle destes parasitos.

Protective effect of the probiotic Lactobacillus acidophilus ATCC 4356 in BALB/c mice infected with Toxocara canis.

Human toxocariasis consists of chronic tissue parasitosis that is difficult to treat and control. This study aimed to evaluate the action of the probiotic Lactobacillus acidophilus ATCC 4356 on larvae of Toxocara canis and the effect of IFN-γ cytokine on parasite-host in vivo (1.109 CFU) and in vitro (1.106, 1.107, 1.108, 1.109 CFU) interactions. Four groups of six BALB/c mice were formed: G1 - L. acidophilus supplementation and T. canis infection; G2 - T. canis infection; G3 - L. acidophilus supplementation; and G4 - PBS administration. Mice were intragastrically suplemented with probiotics for 15 days before inoculation and 48 h after inoculation with 100 T. canis eggs. The inoculation of T. canis was also perfomed intragastrically. The recovery of larvae took place through digestion of liver and lung tissues; the evaluation of IFN-γ gene transcription in leukocytes was performed by qPCR. The in vitro test consisted of incubating the probiotic with T. canis larvae. The supplementation of probiotics produced a reduction of 57.7% (p = 0.025) in the intensity of infection of T. canis larvae in mice, whereas in the in vitro test, there was no larvicidal effect. In addition, a decrease in the IFN-γ gene transcription was observed in both, T. canis-infected and uninfected mice, regardless of whether or not they received supplementation. The probiotic L. acidophilus ATCC 4356 reduced T. canis infection intensity in mice, however, the probiotic did not have a direct effect on larvae, demonstrating the need of interaction with the host for the beneficial effect of the probiotic to occur. Yet, the proinflammatory cytokine IFN-γ did not apparently contributed to the observed beneficial effect of probiotics.

Quantification of Toxocara canis DNA by qPCR in mice inoculated with different infective doses.

The nematode Toxocara canis is of public health importance and is the main causative agent of toxocariasis in humans. This disease is difficult to diagnose due to several factors, including the possibility of cross-reactions with other nematodes in the ELISA. To overcome this problem, molecular tests have been recommended as an alternative to identify the parasite. The quantitative real-time polymerase chain reaction (qPCR) technique was used in this study to identify and quantify the parasite load of T. canis in the mouse brain. To this end, 24 mice were divided into six groups, five of which were challenged with different infective doses of T. canis larvae (L3) (1000, 500, 250, 100 and 50 larvae), while the sixth group, uninfected, acted as negative control. Forty-five days after infection, the animals were euthanized to collect the brain, from which two portions of 20 mg of tissue were taken for DNA extraction, while the rest of the brain tissue was digested to quantify the number of larvae by microscopy. The number of DNA copies was calculated from the standard DNA quantification curve, showing values of E = 93.4%, R2 = 0.9655 and Y = -3.415. A strong positive correlation (R = 0, 81; p < .001) was found between the number of copies and the recovery of larvae from brain. However, the parasite's DNA was also identified even in animals from whose brain no larvae were recovered after tissue digestion. The results of this study therefore confirm that the qPCR technique can be a valuable tool for the detection and quantification of T. canis DNA in murine hosts, even in animals whose with tissues contain very few parasites.

Biomphalaria molluscs (Gastropoda: Planorbidae) in Rio Grande do Sul, Brazil.

The present study was aimed at characterising Biomphalaria species using both morphological and molecular (PCR-RFLP) approaches. The specimens were collected in 15 localities in 12 municipalities of the southern region of the state of Rio Grande do Sul, Brazil. The following species were found and identified: Biomphalaria tenagophila guaibensis, Biomphalaria oligoza and Biomphalaria peregrina. Specimens of the latter species were experimentally challenged with the LE Schistosoma mansoni strain, which showed to be refractory to infection.

The serodiagnostic potential of recombinant proteins TES-30 and TES-120 in an indirect ELISA in the diagnosis of toxocariasis in cattle, horses, and sheep.

Toxocariasis is a zoonotic disease that affects humans and animals alike. Although recombinant proteins are widely used for its diagnosis in humans, their performance in companion and production animals remains unknown. This study aimed to investigate the serodiagnostic potential of the recombinant proteins rTES-30 and rTES-120 from Toxocara canis in an indirect ELISA for cattle, horses, and sheep. Serum samples collected from the animals were tested with indirect ELISA and Western Blotting using T. canis TES-30 and TES-120 recombinant proteins produced in Escherichia coli, as well as native-TES. In the ELISA, rTES-30 showed high serodiagnostic potential in sheep and horses (92.6% and 85.2%, respectively), while the sensitivity of rTES-120 was higher in cattle and horses (97.2% and 92.6%, respectively). Furthermore, a highly positive association was observed between native and recombinant proteins in seropositive samples, while a moderately positive association was observed in seronegative samples, probably due to the lower specificity of native TES. In conclusion, our study indicates that the use of recombinant proteins in an indirect ELISA is an effective tool for the serodiagnosis of toxocariasis in animals, with the choice of protein being species-dependent.