Maria Elisabeth Aires Berne

Presence of Toxocara canis eggs on the hair of dogs: A risk factor for Visceral Larva Migrans

The close contact between dogs and humans poses a high risk of exposure to Toxocara canis eggs, which can lead to Visceral Larva Migrans (VLM) syndrome. The objective of this study was to investigate the existence of these nematode eggs on the hair of the perianal region in stray and owned dogs. Samples of hair from 104 dogs of different ages were collected: 25 (24%) were positive for T. canis eggs, with mean of 614.8 eggs per gram of hair. Puppies were responsible for 99% of the total number of eggs. The ages of the animal and hair length were factors that influenced the intensity of the observed eggs. This study showed that dog hair contaminated by T. canis eggs in different developmental stages represents a potential source of VLM infection for humans.

Seroprevalence of Toxocara Infection in Children from Southern Brazil

Abstract: The seroprevalence of Toxocara canis antibodies in children aged from 1 to 12 yr old was evaluated in Pelotas City, Rio Grande do Sul, Brazil. Human toxocariasis or visceral larva migrans (VLM) was diagnosed with the use of an ELISA based on the T. canis excretory–secretory (TES) antigens; Western blotting was used to confirm the ELISA-positive results. From 427 samples, 50.6% were positive for the presence of anti-TES antibodies. A confirmatory test (Western blot) was carried out on a sample of the ELISA-positive sera (n = 70), and all were positive. The Western blots had specific banding pattern characteristics, where the 30-kDa fraction demonstrated the highest reactivity. This fraction could be important for the specific diagnosis of toxocariasis.

Isolation and detection of Fasciola hepatica DNA in Lymnaea viatrix from formalin-fixed and paraffin-embedded tissues through multiplex-PCR

Detection of Fasciola hepatica infection in Lymnaea viatrix through analysis of histological cuts is based upon morphological characters of the parasite during the intra-mollusk phase of parasitism. At this stage, trematode forms are very similar and, thus, very difficult to differentiate. Specific detection may also be impaired by the presence of other helminthes in the mollusk. Histological samples are usually fixed in formalin, embedded in paraffin, sectioned and HE stained. In the current study, a method for the extraction of DNA from formalin-fixed, paraffin-embedded tissues was standardized by means of deparaffinizing with xylol and digesting with proteinase K. Extracted DNA was amplified in a multiplex-PCR, by using simultaneous primers in a single reaction under high stringency conditions. Results showed specific amplification of DNA from the trematode and the snails. The technique was sensitive enough to detect F. hepatica infections in L. viatrix, in histological sections in which the presence of larval stages could not be observed through brightfield microscopy. The profiles generated were: stair bands referring to F. hepatica DNAmt amplification; a band of 1200 bp referring to L. viatrix ITS and another of 1300 bp referring to F. hepatica ITS and other trematodes. Multiplex-PCR has shown to be a fast, safe, highly sensitive and specific method, which is able to amplify DNA from fixed tissues, despite a low DNA quantity and its degradation caused by fixation processes. Such methodology may be useful in studies on fascioliasis epidemiology, enabling the use of material from histological collections.

Saccharomyces boulardii reduces infection intensity of mice with toxocariasis.

Several studies have shown the benefit of the use of probiotics in the prevention and treatment of diseases; however, few of them have investigated the effect of probiotics on parasitosis. In this study, the effect of Saccharomyces boulardii on the intensity of infection of mice with toxocariasis was evaluated. The animals were fed with a diet supplemented with S. boulardii for 15 days before inoculation with Toxocara canis eggs and for 2 or 60 days post-inoculation. S. boulardii promoted a reduction of approximately 36% in the average number of recovered T. canis larvae, suggesting that it can be used as an alternative to help control toxocariasis.

Association between microsatellite markers of sheep chromosome 5 and faecal egg counts

Abstract This study analyses the associations between alleles at seven microsatellite markers located at the sheep chromosome 5—where the interleukin IL-3, IL-4 and IL-5 genes were mapped—and faecal egg counts (FEC) in two different flocks. The associations were analysed by estimating the average excess of alleles at each microsatellite marker on FEC. Three of the studied markers (CSRD2138, OarAE129 and TGLA176) showed significant associations with FEC in the Corriedale flock. Seven alleles at these microsatellite loci were associated with FEC and their effects ranged from −28 to +20%, relative to the FEC population mean. On the other hand, four alleles at CSRD2138 and TGLA176 microsatellite markers had significant associations with FEC in the Polwarth mob, with effects varying from −22 to −5%, relative to the FEC population mean. All microsatellites investigated presented high diversity in both flocks. Some alleles were breed specific. The CSRD2138 ∗ A allele was the only marker associated with consistent reductions in FEC for both breeds. These results indicate that markers or genes nearby the interleukin genes, such as the CSRD2138, might enhance host resistance to internal parasites in sheep by probably increasing effector cells and antibody responses.

Development of an indirect ELISA-NcSRS2 for detection of Neospora caninum antibodies in cattle

Neosporosis is of alarming economic concern in the cattle industry. The effectiveness of diagnostic tests for detecting specific antibodies against Neospora caninum is hampered by potential cross-reaction with other coccidia. Use of a single specific antigen might improve test specificity. An indirect enzyme-linked immunosorbent assay (ELISA) was developed using the truncated protein NcSRS2 expressed in Escherichia coli. The ELISA results were compared with those of the indirect fluorescence antibody test (IFAT). Receiver Operating Characteristic (ROC) and Tests in the Absence of a Gold Standard (TAGS) analysis revealed an assay having 96% specificity and 95% sensitivity when applied to 145 positive and 352 negative sera from two distinct cattle populations. Using OD ≤ 0.095 as the cut-off point, the assays negative and positive predictive values ranged from 98.8% to 50.8% and from 58.8% to 99.1%, respectively, depending on neosporosis prevalence in a given area. The novel ELISA-NcSRS2 format described in the present report constitutes a specific and sensitive method for detecting N. caninum in cattle.

Presence of eggs of Toxocara spp. and hookworms in a student environment in Rio Grande do Sul, Brazil

Environmental contamination by parasite forms was studied on a university campus in the municipality of Pelotas, Rio Grande do Sul. Soil samples were analyzed using the modified Caldwell & Caldwell technique to identify parasite forms. Nematode eggs were detected in 62% of the samples. Among the parasites detected, eggs of Toxocara spp. and Ancylostomatidae were the most prevalent parasites in the studied area throughout the study period. The results demonstrated that there is significant environmental contamination, thereby representing a risk of zoonotic infection for humans frequenting the study area.

Fungal ovicidal activity on Toxocara canis eggs

BACKGROUND Visceral toxocariasis is a parasitic zoonosis caused by Toxocara canis. The prevalence of this parasite in dogs, soil contamination and the resistance of eggs increase human exposure to the disease. Moreover, the difficulties of the control measures justify the need for alternative ones. AIMS The objective of this study was to evaluate the in vitro ovicidal activity of fungi isolated from soils from public places in the city of Pelotas, Rio Grande do Sul, Brazil, on Toxocara canis. METHODS Samples of soil from ten localities were inoculated onto Petri dishes with 2% water-agar (WA) that contained antibiotics, and incubated at 25°C/21 days. Isolated fungi were tested in vitro for ovicidal activity, with five replicates. One mL of an embryonated Toxocara canis egg suspension (10(3) eggs) was poured over the fungal cultures after 10 days of growth. At intervals of 7, 14 and 21 days, 100 eggs were removed from each plaque and evaluated by optical microscopy. RESULTS Acremonium, Aspergillus, Bipolaris, Fusarium, Gliocladium, Mucor and Trichoderma were isolated from the soil. A significant ovicidal type 3 effect was observed in Trichoderma, Fusarium solani complex and Acremonium. Those isolates from the genus Trichoderma showed their ovicidal effect on the 14th day of fungus-egg interaction. The other fungal genera tested showed a type 2 effect. CONCLUSIONS These results suggest that the use of Trichoderma and Fusarium solani complex in biological control of T. canis is promising; however, further studies should be performed.

Expression of Neospora caninum NcSRS2 surface protein in Pichia pastoris and its application for serodiagnosis of Neospora infection

Abstract Neospora caninum is considerd a major cause of abortion in cattle worldwide. The antigenic domain of NcSRS2 in N. caninum is an important surface antigen present in the membrane of this parasite. In the present study, the Pichia pastoris expression system proved to be a useful tool for the production of recombinant protein. The truncated NcSRS2 gene (by removal of the N-terminal hydrophobic sequence), was cloned in the vector pPICZalphaB, and integrated on the genome of the methylotrophic yeast P. pastoris. Subsequently, the NcSRS2 protein was expressed, purified, and characterized using naturally infected cattle sera and Mab 6xhistag. The recombinant protein NcSRS2 was present in the supernatant of the culture, where later it was concentrated and purified using ammonium sulfate (∼100 mg/ml). An indirect immunoenzymatic assay (ELISA) was performed using cattle sera from endemic N. caninum area.

Prevalence of Chagas disease among pregnant women in the southern region of Rio Grande do Sul

Anti-Trypanosoma cruzi antibodies in the umbilical cord of 351 parturients in the city of Pelotas, Rio Grande do Sul were investigated to determine the prevalence of Chagas disease among pregnant women. One case was identified (0.3%), without detection of congenital transmission. This highlights the importance of investigating Chagas disease among pregnant women living in or originating from endemic areas.