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.     

Cross-reactions of sera from Toxocara canis-infected mice with Toxascaris leonina and Ascaris suum antigens.

The ELISA method using larval excretory-secretory (E/S) products and homogenized Toxocara canis, Toxascaris leonina and Ascaris suum adult worm extract were used to determine possible cross-reactions in BALB/c and C57BL/10 mice, inoculated with embryonated eggs or adult worm extract of T. canis in single and multiple doses. When we used sera of mice infected with embryonated eggs of T. canis against different heterologous antigens, we observed no cross-reactions in BALB/c mice against A. suum E/S and adult worm extract antigens with a single dose. In multiple doses this was absent too against T. leonina adult worm extract in BALB/c mice, and in both strains against A. suum E/S and adult worm extract. In BALB/c mice inoculated with adult worm extract of T. canis we did not observe cross-reactions with A. suum E/S antigen with both inoculation doses. In the remainder of the experiments, we observed cross-reactions of different intensities.     

Prevalence of Toxocara canis, Toxascaris leonina and Dirofilaria immitis in dogs in Chuncheon, Korea (2004)

The intestines and hearts of dogs were examined for Toxocara canis, Toxascaris leonina, and Dirofilaria immitis, after necropsy between June 26 and September 29, 2004 in Chuncheon, Korea. Of the 662 dogs examined, 6 were infected with T. canis (0.9%), 86 with T. leonina (13.0%). Fifty dogs were infected with D. immitis among 500 dogs examined (10.0%). Five were co-infected with T. canis and T. leonina, and three were co-infected with T. leonina and D. immitis. The cumulative positive infection rate for three species was 134/662 (20.2%). Considering previously reported seropositive rates of T. canis excretory-secretory antigen, i.e., 5% in the adult population in Korea, the possibility of toxocariasis caused by T. leonina should be reevaluated.     

Three years incidence of dermatophytes in a hospital in Porto (Portugal)

We evaluated the incidence of dermatophytes isolated at our hospital in the years of 1997 to 2000 and correlated it with anatomical site and age. Trichophyton rubrum was the predominant species in all anatomical sites, excluding scalp, followed by Microsporum canis, the leading agent of tinea capitis. All dermatophytosis, except tinea capitis by M. canis and Trichophyton schoenleinnii appeared mainly in adult patients. Our results revealed no substantial differences to other portuguese studies regarding the major agents. We found a relatively high incidence of T. schoenleinnii as second tinea capitis agent.     

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.     

Detection of circulating antigens and antibodies in Toxocara canis infection among children in Chengdu, China

In order to determine the seroprevalence of Toxocara spp. infection in children from Chengdu, we performed an enzyme-linked immunosorbent assay (ELISA) and sandwich ELISA (S-ELISA) with excretory-secretory antigens isolated from second-stage larvae of Toxocara canis (TES-Ag ELISA). The seroprevalences of T. canis antibodies in the children from rural areas, urban districts, and urban districts with recent Ascaris lumbricoides infection were 17.7% (59/333), 2.1% (4/186), and 2.6% (1/38), respectively. Among 63 suspected patients with symptoms of T. canis infection, 31 had positive antibodies. The inhibition assay showed an apparent inhibiting capacity of TES-Ag for the antibody against T. canis larvae. The result of S-ELISA demonstrated that circulating antigens of T. canis larvae could be detected in part of the serum with positive antibodies and that the detection rate for circulating antigens in the sera could be improved by polyethylene glycol-acid treatment. This is the first epidemiological study to confirm the existence of T. canis infection and Toxocara-larvae migrans in Chengdu by the combination of TES-Ag ELISA and S-ELISA.     

Intestinal nematode infections in Turkish military dogs with special reference to Toxocara canis

The prevalence and potential zoonotic risk factors of intestinal nematodes of military working dogs, which are used for different military purposes, were assessed. Faecal samples from 352 defined-breed Turkish military dogs were investigated and 107 (30.4%) dogs were found to be infected with one or two nematode species. The following nematodes, with their respective prevalences, were diagnosed in the faecal samples: Toxascaris leonina (21.8%), Toxocara canis (13.3%), Trichuris vulpis (2.9%) and Uncinaria stenocephala (1.2%). Toxocara canis infections were more frequently seen in puppies (0-6 months old). The prevalence of T. canis was significantly higher in male than in female dogs and also higher in dogs which were exercised daily than in those without exercise. The highest prevalence was found in Belgian malinois breed dogs. Toxocara canis infections were not influenced by the floor type of the kennels (i.e. concrete or soil floor). There was no difference in the occurrence of T. canis infection when the last anthelmintic treatment was carried out less or more than 3 months prior to sampling. It is suggested that T. canis infected military dogs would be a threat not only for dog trainers but also for military personnel, notably during national and international operations.     

Dermatophytes isolated from patients in University Hospital,Kuala Lumpur,Malaysia

A total of 576 dermatophytes were isolated from patients with a variety of skin infections from January 1993 to May 2000. Ten species of dermatophytes were identified: Epidermophyton floccosum (0.7%), Microsporum audouinii (1.1%), M. canis (3.1%), M. gypseum (0.3%), Trichophyton concentricum (3.5%), T. equinum (0.2%), T. mentagrophytes (36.1%), T. rubrum (53.8%), T. verrucosum (0.2) and T. violaceum (1.0%). The body sites most frequently affected by dermatophytes were the buttocks, nails and trunk. Anthropophilic dermatophytes made up 60.1% of the isolates; the most common species was T. rubrum, T. mentagrophytes and M. canis were the two main zoophilic dermatophytes. T. mentagrophytes was isolated from all body sites except the scalp. M. canis was found to be associated with domestic dogs and was not isolated from ethnic Malays. The only geophilic dermatophyte was M. gypseum, an uncommon dermatophyte associated with tinea pedis.     

Cross-reactivity between sera from dogs experimentally infected with Dirofilaria immitis and crude extract of Toxocara canis

This study was performed to investigate whethere there is cross-reactivity between Dirofilaria immitis and three intestinal nematodes of dogs. In ELISA, D. immitis-infected dog sera obtained at the 4th molting stage (9-11 weeks) and microfilaremic stage (25-30 weeks) were shown to be highly reactive with crude extract of T. canis. In immunoblotting, some antigenic fractions, 44, 57, 88, 100 kDa of crude extract of T. canis, were found to be positive reaction with sera of dogs infected with D. immitis. However, little or no cross-reaction were observed between sera of D. immitis-infected dogs and crude extract antigen of T. vulpis or A. caninum. These result suggest that there are partial cross reaction between sera of D. immitis-infected dogs and the antigen of T. canis.     

Prevalence and intensity of Toxocara canis (Werner, 1782) in dogs and its potential public health significance in Ile-Ife, Nigeria

A study on the prevalence and intensity of Toxocara canis (Werner, 1782) in dogs was carried out in Ile-Ife, Nigeria. Faecal samples were collected from 269 dogs between January and December 2004, processed by the Kato-Katz technique and then examined for T. canis eggs. The prevalence of T. canis obtained was 33.8%. The intensity of infection, measured as mean egg count per gram of faeces ( +/- SEM) was 393.8 +/- 83.4. The prevalence and intensity of T. canis in dogs aged 0-6 months were significantly higher (P < 0.05) than older age groups. The prevalence and intensity of T. canis infection were significantly higher in males than in female dogs (P < 0.05). Since T. canis is known to cause visceral larva migrans (VLM) in young children, there is the possibility that the high prevalence of T. canis infection obtained in this study might constitute an important risk factor for transmission to humans. Therefore, there is the need to educate the residents of Ile-Ife on the danger of close association of their children with household pets.     

Transmission of hepatozoon canis to dogs by naturally-fed or percutaneously-injected Rhipicephalus sanguineus ticks.

Hepatozoon canis is an apicomplexan protozoan parasite of dogs, prevalent in Asia, Africa, and southern Europe. Experimental transmission of H. canis to dogs was performed with laboratory-reared Rhipicephalus sanguineus nymphs that fed on a naturally infected dog or were percutaneously injected with canine blood containing H. canis gamonts. Dogs were inoculated by oral ingestion of adult ticks containing H. canis oocysts. Transstadial transmission of H. canis was recorded, whereas transovarial transmission could not be demonstrated. Oocysts were detected in 85% of the adult ticks that had engorged as nymphs on an infected dog and in 61% of the adult ticks resulting from nymphs injected percutaneously with blood from the same dog. Nine of 12 dogs (75%) inoculated with naturally fed or percutaneously injected ticks became parasitologically positive, and all showed seroconversion. Meronts were initially detected in the bone marrow 13 days postinoculation and gamonts 28 days after infection. The variation in the time of initial detection of parasitemia among infected dogs and the rapid appearance of gamonts in dogs immunosuppressed with corticosteroids suggest that immune mechanisms play an important role in controlling H. canis parasitism. The artificial acquisition of Hepatozoon parasites by percutaneous injection of ticks, demonstrated here for the first time, may serve as a useful tool for studies on transmission, vector-host relationships, and the immunology of infection with Hepatozoon species.     

Characterization of a Toxocara canis species-specific excretory-secretory antigen (TcES-57) and development of a double sandwich ELISA for diagnosis of visceral larva migrans

This study describes the isolation of a Toxocara canis species-specific excretory-secretory (ES) antigen and the development of an enzyme-linked immunosorbent assay (ELISA) based on this antigen. Analysis of the ES antigens of T. canis, Toxocara vitulorum, Ascaris lumbricoides and Necator americanus larval antigen was performed by SDS-PAGE followed by western blotting. A 57 kDa T. canis-specific antibody fraction (TcES-57) was identified by western blotting and labelling with anti-Toxocara antibodies (from experimental rabbits and human patients) and tracing with anti-human or anti-rabbit peroxidase conjugate. No protein fraction of 57 kDa was detected in ES or larval antigens collected from T. canis, T. vitulorum, A. lumbricoides and N. americanus. Using TcES-57, a specific antiserum was produced in rabbits and a double sandwich ELISA was developed. This test was validated using known seropositive sera from toxocariasis patients, sera from A. lumbricoides or N. americanus patients, and 50 serum samples from cats. These tests revealed that TcES-57 antigen is specific to T. canis infection and does not cross react with sera of other related infections. Thus, ELISA based on TcES-57 antigen was proven to be an effective tool in the diagnosis of toxocariasis and studies on the role of T. canis in the epidemiology of human toxocariasis.     

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.     

Cross-reactivity induced by Anisakis simplex and Toxocara canis in mice

The aim of this study was to verify whether cross-reactivity appeared between Toxocara canis and Anisakis simplex in an experimental rodent model. No cross-reactions were detected using sera from mice infected with T. canis eggs. When responses obtained against T. canis ES antigen using sera from BALB/c and C57BL/10 mice infected with T. canis eggs were compared with those obtained by testing sera from mice infected with one A. simplex L3, an increase in cross-reactions was observed using the C57BL/10 strain.     

Toxocara infection in pigs. The use of indirect fluorescent antibody tests and an in vitro larval precipitate test for detecting specific antibodies.

An in vitro larval precipitate test using second-stage Toxocara canis larvae and an indirect fluorescent antibody (IFA) test employing cuticles of T. canis larvae as antigen were evaluated using antisera produced in pigs experimentally infected with T. canis, T. cati, Ascaris suum, Toxascaris leonina and Parascaris equorum. The former test was both specific and sensitive and is suggested as a reliable and simple method of detecting Toxocara antibodies in pigs. The latter test was considered unsuitable because of cross-reactions that occurred when sera from pigs infected with other ascarids were tested. An IFA test for Ascaris antibodies, employing cuticles of A. suum larvae as antigen, is described. The degree of specificity of this test suggests that it may be of value in the detection of antibodies to Ascaris in pigs under natural conditions.     

Human and dogs Toxocara canis infection in a poor neighborhood in Bogota

Prevalence of Toxocara canis antibodies was studied in a poor community of Bogotá, Colombia. Two-hundred-seven patients, from both sexes and all age groups, were studied. Positive ELISA titers were found in 47.5% of the population, a high prevalence compared with reports from developed countries. T. canis ova were positive in 43.6% of fecal samples from dog puppies. An endemic pattern of the disease is described: socioeconomic status, weather, pollution, poor hygiene and a significant population of infected dogs. Neither the physical examination nor ELISA titers could detect any case of T. canis disease.     

Evidence for the involvement of the optic nerve as a migration route for larvae in ocular toxocariasis of Mongolian gerbils

Although Toxocara canis, an important pathogen of ocular disease, tends to migrate to the eye, the precise migratory route has yet to be determined experimentally. Mongolian gerbils, Meriones unguiculatus, known as a useful animal model for human toxocariasis, were used to investigate the migration route toward the eyes. Infective larvae of T. canis were directly inoculated into the intracranial region. Haemorrhagic lesions or larvae were observed in 56.3% of cases. Histopathologically, a larva was observed in the optic nerve of gerbils 6 days after inoculation, and two larvae were found in the optic chiasma in the gerbils having a haemorrhage in the retina 9 days after inoculation. These results indicate that T. canis migrates from the brain to the eye through the optic nerve. Considering these data and previous studies showing that the ocular changes appear as early as 3 days of infection in the oral-administrated gerbils, there are two phases in the migration to the retina: a haematogenous early phase and an optic nerve route late phase.     

Toxocariasis associated with chronic cough in childhood: a longitudinal study in Hungary

Chronic cough lasting 8 weeks or more often seems to be an intractable problem in childhood. Toxocara infection is associated with an increased prevalence of airway symptoms and may be the possible aetiological agent of chronic cough. Of 425 children aged 2-17 years with chronic cough who were investigated for toxocariasis and the distribution of bronchial asthma (BA), cough variant asthma (CVA) and non-asthmatic eosinophilic bronchitis (NAEB), 136 (32%) were seropositive for Toxocara canis antigens. Ninety-three of the 136 were adequately assessed, diagnosed and followed up during 1 year. BA was diagnosed in 40%, CVA in 27% and NAEB in 33% of the children. The eosinophil cell count, serum T. canis IgG levels and symptoms are predictors of the improvement or the decline of the condition. Presuming the aetiopathogenetic role of T. canis in the inflammatory process of chronic cough, we treated the children not only with inhaled corticosteroid (ICS), but also with a 1-week course of anthelminthics. We could significantly decrease the dose of ICS in 23 (62%) of the 37 with BA. The administration of anthelminthics and the avoidance of sensitizers were sufficient for those with NAEB; none needed ICS. ICS therapy could be stopped 2-3 months later in 17 (68%) of the 25 with CVA. We found that 8 of the 25 with CVA (32%) presented asthmatic symptoms at the end of the 1-year period. In Hungary, T. canis may be a potential sensitizer for chronic cough in seropositive children. Deworming therapy will then alleviate the airway symptoms without exacerbation in patients with BA, and have a positive effect on those with NAEB and the majority of those with CVA.     

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.     

Regulation of toxocariasis in mice selectively reared for high and low immune responses to Nematospiroides dubius

Test mice have been selectively reared for high (H) or low (L) immune responses to Nematospiroides dubius. After secondary infection with N. dubius, the L mice voided ten times as many eggs in their faeces as the H mice, and at necropsy, 71% versus 20% of the inoculum of N. dubius were recovered as adult worms from the L and H mice respectively. Furthermore, N. dubius were more fecund in the L than in H mice. High or low immune responsiveness was not restricted to N. dubius infection in these mice but was also observed during Toxocara canis infection. The migration of T. canis larvae from gut via the liver to skeletal muscle and CNS was inhibited in H versus L mice. Many more larvae were recovered from the livers of H compared with L mice which was indicative of greater immunity in the H mice. The protective immune response in H compared with L mice to both N. dubius and T. canis included pronounced eosinophilia and elevated antiparasite antibody titres.