Aberrant Toxocara canis in a red fox

During necropsy of a red fox (Vulpes vulpes) heart an adult, male Toxocara canis was found under the pericardium at the junction of the right ventricle and right atrium. The life cycle of T. canis is complex and includes tracheal and somatic migrations of larvae, and they can be found in many tissues throughout the host's body. However, it is rare for adult ascarids to be recovered outside of the small intestine. This is the first report of an adult T. canis inside the pericardial space.     

Fungal ovicidal activity on Toxocara canis eggs

BackgroundVisceral 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.AimsThe 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.MethodsSamples 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 (10³ 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.ResultsAcremonium, 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.ConclusionsThese 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.     

Improved methods for recovering eggs of Toxocara canis from soil

The ingestion of soil in parks and public places containing eggs of Toxocara may constitute a significant health risk, particularly to children. To determine the most efficient method for extracting eggs from experimentally contaminated soil, two consecutive studies were undertaken. Four techniques, including washing, sieving, vacuum, and the one recommended by the World Health Organization, were evaluated. Recovery rates of over 85% were recorded with both washing and sieving methods. Using the washing technique, all combinations of the four pre-treatment solutions, distilled water, acetoacetic solution pH 5, 0.1 n sodium hydroxide and 1% Tween 20, and seven flotation fluids with different specific gravities (S.G.) ranging from 1.20 to 1.35 were assayed. The association of distilled water and saccharose solution with an S.G. of 1.27 showed the best results, with a recovery rate of 99.91%.     

Contamination of soils with eggs of Toxocara in a subtropical city in Argentina

A total of 475 soil samples were collected from five public park playgrounds, 17 kindergarten sandpits and 124 housing estates in Resistencia, a medium-sized subtropical-region city in Argentina, and processed by the centrifugal flotation method. Eggs of Toxocara spp. were present in five (3.4%) of the 146 habitats surveyed and in six (1.3%) of the 475 samples examined. Twenty per cent of public parks, 5.9% of kindergarten sandpits and 2.4% of housing estates were contaminated with Toxocara eggs. Depending on the number of samples examined from the three types of habitat, contamination by Toxocara was 0.7% in public park playgrounds, 1.2% in kindergarten sandpits and 1.6% in the housing estates. High prevalences of Ancylostomidae eggs were also found especially in public park playgrounds with a value of 100%, compared with 19.4% found in housing estates and 11.8% in kindergarten samples. These results suggest that in Resistencia, human infections with Toxocara are likely to occur within the limits of housing estates more so than in public parks or open spaces.     

Fecundity and egg output by Toxocara canis in the red fox, Vulpes vulpes

The role of the red fox Vulpes vulpes in the dissemination of eggs of Toxocara canis into the environment is considered with reference to female worm fecundity and egg output in the faeces of infected foxes collected from four localities in southern England. A significant positive correlation was found between female worm size and the number of eggs in the uterus but there was no significant relationship between T. canis worm numbers and egg output in fox faeces. Reliable estimates of worm burdens in foxes could not, therefore, be determined from faecal egg counts alone. The highest mean egg output of 2145.0 epg recorded from adult foxes indicated that fox cubs are not necessarily the main sources of environmental contamination with T. canis eggs. Saturated magnesium sulphate was found to be a more effective flotation solution than zinc sulphate and sodium chloride for recovering eggs from fox faecal samples.     

Decontamination by anaerobic stabilisation of the environment contaminated with enteronematode eggs Toxocara canis and Ascaris suum

Investigations were carried out under operating conditions of Field Composting Factory in Brezno (Slovak Republic) to determine the effect of anaerobic stabilization of organic wastes from public areas on the survival of model helminth Toxocara canis and Ascaris suum eggs. Due to anaerobic conditions, low temperature, low C:N ratio and changes in physical and chemical properties of organic waste, less than 64% of A. suum eggs remained viable after 150 days of stabilisation. The anaerobic stabilisation had a greater effect on the viability of T. canis eggs than on A. suum eggs. The infectivity of T. canis eggs was confirmed by a follow-up experiment in laboratory mice. A small number of T. canis larvae were found in their brain and muscles on day 28 after infection. The results refer to the risks of dissemination, survival and potential spread of endoparasitic developmental stages in the environment through organic wastes subjected to low temperature stabilisation.     

Loop-mediated isothermal amplification assay for detection and discrimination of Toxocara canis and Toxocara cati eggs directly from sand samples

We developed a novel and simple method, using loop-mediated isothermal amplification (LAMP), for the detection and discrimination of Toxocara canis and Toxocara cati eggs. The new method employs 4 steps: (1) concentration of Toxocara eggs in a small amount of sand; (2) dissolution of the proteinaceous membrane of eggs and simultaneously separation of them from the sand using NaClO treatment; (3) extraction of DNA using NaOH treatment; and (4) detection of T. canis / T. cati DNA using a LAMP assay. All these steps are fast, easy to perform, and do not require expensive equipment or reagents. The novel method was tested both experimentally and in a field study. In the laboratory, we could reliably detect as few as 3 T. canis eggs in artificially contaminated sand, if the experiment was repeated twice. In the field trial, we were able to detect T. cati DNA from 4 natural sandpits having moderate to heavy contamination, although not in a single lightly contaminated sandpit. All of the examined sandpits were found to be contaminated with eggs of T. cati, but none appeared to contain T. canis. Our new method could extract DNA from T. canis and T. cati eggs directly from sand samples as well as detect and distinguish these 2 species in a few easy steps, with markedly reduced time and expense.     

The epidemiology of Toxocara canis

Bred as hunter, companion and pet, the dog has a long and honourable association with man. Yet the domestic dog can host a wide range of parasites - many of which can also infect humans. One of these, the ascarid nematode Toxocara canis (Fig. 1), is of particular interest because of retinal damage that may result from larvae becoming trapped in the eye. At the Hospital for Tropical Diseases in London, about 20-30 patients with toxocariasis are treated annually. Widespread fouling of public parks, playgrounds and pedestrian areas with dog faeces - especially in large cities - is well -recognized as one of the main sources of Toxocara infection. Yet as Stephen Gillespie discusses here, epidemiological indicators vary widely and the risk of infection is often treated too lightly.     

Ultrastructural studies of alterations induced by microwaves in Toxocara canis eggs: prophylactic interest

The ultrastructure of Toxocara canis eggs is described before and after exposure to microwaves. The morphology of normal eggs is compared to that of eggs from other helminths. Following treatment, the complete disorganization of the surface structure of the shell and the loss of much turgidity of the egg are observed. The destruction of the internal structure is most marked in the center of the egg and is associated with the disappearance of some layers of the shell. In addition, there is substantial damage to the synthesis apparatus (ribosomes, endoplasmic reticulum, lipid cisternae). An explanation based on the specific action of microwaves and micro-overheating is proposed, and the prophylactic use of this technique is considered.     

Antagonistic activity of the fungus Pochonia chlamydosporia on mature and immature Toxocara canis eggs

In vitro tests were performed to evaluate the ability of 6 isolates of the nematophagous fungus Pochonia chlamydosporia to infect immature and mature Toxocara canis eggs on cellulose dialysis membrane. There was a direct relationship between the number of eggs colonized and the increase in the days of interaction, as well as between the number of eggs colonized and the increase in the concentration of chlamydospores (P<0.05). Immature eggs were more susceptible to infection than mature eggs. The isolate Pc-04 was the most efficient egg parasite until the 7th day, and showed no difference in capacity to infect mature and immature eggs in comparison to Pc-07 at 14 and 21 days of interaction, respectively. Isolate Pc-04 was the most infective on the two evolutionary phases of the eggs at most concentrations, but its ability to infect immature eggs did not differ from that presented by the isolates Pc-07 and Pc-10 at the inoculum level of 5000 chlamydospores. Colonization of infective larvae inside or outside the egg was observed in treatments with the isolates Pc-03, Pc-04, Pc-07 and Pc-10. The isolate Pc-04 of P. chlamydosporia has great biological capacity to destroy immature and mature T. canis eggs in laboratory conditions.     

Nematophagous fungi of Toxocara canis eggs in a public park of La Plata, Argentina

Fungi have showed a great potential for the biological control of nematodes. However, they have not been evaluated for the control of animal and/or human parasites transmitted by egg contaminated soils. Environmental contamination with Toxocara spp. eggs is a public health problem. Accidental swallowing of Toxocara canis eggs (a nematode of dogs) usually results on a zoonotic infection (toxocarosis). The objectives of this research were: 1) To test the presence of antagonistic fungi against T. canis in the soil in public places of La Plata city, Argentina, infected with eggs of this parasite, 2) To determine the possible association between biotic and abiotic factors of the soil with the presence of fungal parasites of egg nematodes. Soil samples were tested for: textural type, organic matter (%), pH, presence of egg-parasite fungi, of larvae and of nematode eggs, in particular of Toxocara spp. The studied area showed the following characteristics: pH: 6.6-8.0, organic matter: 1.2-70%, with a predominantly loam texture. The following antagonistic fungal genera were identified: Acremonium, Aspergillus, Chrysosporium, Fusarium, Humicola, Mortierella, Paecilomyces and Penicillium. A prevalence of 70% was detected for nematode eggs, of 33% for Toxocara spp. eggs and of 90% for larvae. No association between the presence of egg-parasite fungi and the considered factors was found. More studies are necessary to know the natural antagonism factors to T. canis eggs for its in situ biological control.     

Eosinophilia in chimeric mice infected with Toxocara canis

A marked strain variation in eosinophilia following oral infection with Toxocara canis eggs was observed in mice. Mutual radiation chimeras between high and low responder mice in terms of eosinophilia were made and compared with the respective donor and recipient for eosinophilia after the infection. As a result, the degree and time course of eosinophilia in chimeric mice were similar to those in donors. The result suggested that genes which regulate inheritance of the trait, marked eosinophilia in T. canis-infected mice, might be expressed in bone marrow derived cells.     

The mitochondrial genome of Toxocara canis

Toxocara canis (Ascaridida: Nematoda), which parasitizes (at the adult stage) the small intestine of canids, can be transmitted to a range of other mammals, including humans, and can cause the disease toxocariasis. Despite its significance as a pathogen, the genetics, epidemiology and biology of this parasite remain poorly understood. In addition, the zoonotic potential of related species of Toxocara, such as T. cati and T. malaysiensis, is not well known. Mitochondrial DNA is known to provide genetic markers for investigations in these areas, but complete mitochondrial genomic data have been lacking for T. canis and its congeners. In the present study, the mitochondrial genome of T. canis was amplified by long-range polymerase chain reaction (long PCR) and sequenced using a primer-walking strategy. This circular mitochondrial genome was 14162 bp and contained 12 protein-coding, 22 transfer RNA, and 2 ribosomal RNA genes consistent for secementean nematodes, including Ascaris suum and Anisakis simplex (Ascaridida). The mitochondrial genome of T. canis provides genetic markers for studies into the systematics, population genetics and epidemiology of this zoonotic parasite and its congeners. Such markers can now be used in prospecting for cryptic species and for exploring host specificity and zoonotic potential, thus underpinning the prevention and control of toxocariasis in humans and other hosts.     

Effects of temperature and humidity on the development of eggs of Toxocara canis under laboratory conditions

The influence of temperature and humidity on the survival and development of Toxocara canis eggs in an in vitro model system was investigated. Two soil samples were inoculated with T. canis eggs and maintained at 3% and 50% humidity and temperatures of 19-24 degrees C. Nine soil samples were inoculated with T. canis eggs of which three samples were kept at 4 degrees C with humidities at 3%, 15%, and 30%; three were maintained at 21 degrees C and three more were incubated at 34 degrees C, and at the same three humidity levels. Samples were monitored every 7 days for a total of 2 months, for the presence and development of eggs. With increasing temperature, the number of eggs undergoing development increased (P<0.01); the number of deformed eggs decreased, the number of infective eggs increased (P<0.01), and egg maturation was accelerated. A decrease in the survival of infective eggs occurred at 34 degrees C. An increase in humidity produced a rise in the number of developed eggs at all three temperatures (P<0.01). This study suggests that elevated temperatures accelerated the development as well as the degradation of eggs of T. canis, whereas the range in humidity was directly correlated with egg development.     

Toxocara canis and the allergic process

The protective effect of infectious agents against allergic reactions has beenthoroughly investigated. Current studies have demonstrated the ability of somehelminths to modulate the immune response of infected hosts. The objective of thepresent study was to investigate the relationship between Toxocaracanis infection and the development of an allergic response in miceimmunised with ovalbumin (OVA). We determined the total and differential blood andbronchoalveolar lavage fluid cells using BALB/c mice as a model. To this end, thelevels of interleukin (IL)-4, IL-5 and IL-10 and anti-OVA-IgE were measured using anELISA. The inflammatory process in the lungs was observed using histology slidesstained with haematoxylin and eosin. The results showed an increase in the totalnumber of leukocytes and eosinophils in the blood of infected and immunised animalsat 18 days after infection. We observed a slight lymphocytic inflammatory infiltratein the portal space in all infected mice. Anti-OVA-IgE levels were detected insmaller proportions in the plasma of immunised and infected mice compared with micethat were only infected. Therefore, we concluded that T. canispotentiates inflammation in the lungs in response to OVA, although anti-OVA-IgElevels suggest a potential reduction of the inflammatory process through thismechanism.