Chien Wei Liao

Factors affecting disease manifestation of toxocarosis in humans: Genetics and environment

Toxocara canis is regarded as the main cause of human toxocarosis but the relative contribution of T. cati is probably underestimated; serological and other diagnostic methods used in most studies of this zoonotic disease do not distinguish between the two parasites. The definitive hosts for T. canis are caniidae. Pups generally have higher infection rates than adult animals and are a major source of eggs in the environment. Humans usually acquire T. canis infection by accidental ingestion of embryonated eggs or encapsulated larvae from the environment or contaminated food, such infections may lead to visceral larva migrans (VLM), ocular larva migrans (OLM) or covert toxocarosis (CT). Although a mixed Th1- and Th2-mediated immunological response, particularly with high levels of IgE and eosinophilia is observed, the underlying mechanisms of molecular and immunopathogenesis for the development of the symptomatic syndromes of VLM, OLM, or of asymptomatic CT are largely unclear. Studies have indicated that immunological defences against various infectious diseases may be highly influenced by complex interactions of environmental and host genetic factors e.g. MHC class I and II, also known as human leucocyte antigen (HLA). Toxocara spp. infections are associated with a polarized CD4(+) Th2 response with high IgE levels and eosinophilia, mediated mainly by HLA class II molecules. Associations have been made between HLA class II and pathological severity and host genetic effects on exposure to infection. Recent research suggests Foxp3(+) CD4(+)CD25(+)-expressing T regulatory (Treg) cells play a role in regulation of the immunopathology of granulomas in experimental toxocaral granulomatous hepatitis and in enhanced expression of TGF-β1, which is an important factor for the local survival and function of Treg observed during T. canis invasion in the mouse small intestine, liver, muscle, and brain. Since the potential susceptibility loci HLA class II molecules, are considered involved in the regulation of a Th2-dominant immunity which is highly controlled by Foxp3(+) CD4(+)CD25(+) Treg cells by stimulation through TGF-β1, which thus provides a beneficial environment to T. canis larvae but severe injuries to local organs. However, TGF-β1 variant Leu10Pro known to be involved in disease severity warrants further elucidation as this too may have a role in the severity of human toxocarosis. Exploration of TGF-β1 polymorphism, Foxp3(+) CD4(+)CD25(+) Treg cells, and MHC polymorphisms may allow insight into the contribution made by environmental and genetic factors in influencing disease syndrome type and severity in humans with toxocarosis.

Seroepidemiology of Toxocara canis infection among mountain aboriginal schoolchildren living in contaminated districts in eastern Taiwan

We conducted a seroepidemiological study of Toxocara canis infection among mountain aboriginal schoolchildren aged 7–12 years living in contaminated districts in eastern Taiwan. To detect sera IgG (≥1:64) we used a T. canis larval excretory‐secretory antigen‐based enzyme‐linked immunosorbent assay. A short questionnaire elicited information on the practices of raising dogs, playing with soil, eating raw vegetables, or whether the child normally washed his/her hands before eating. The overall seroprevalence was quite high, reaching 76.6% (252/329). Neither age nor gender seemed to be important factors related to a positive serology. Aboriginal schoolchildren who raised dogs (OR = 1.83, 95% CI: 1.04–3.19, P = 0.03), or played with soil (OR = 2.52, 95% CI: 1.49–4.25, P < 0.001) seemed to be more susceptible to T. canis infection than those who did not. Children who habitually washed their hands before eating (OR = 0.57, 95% CI: 0.33–0.97, P = 0.04) had a lower chance of acquiring T. canis infection than those who did not.

Brain injury-associated biomarkers of TGF-beta1, S100B, GFAP, NF-L, tTG, AbetaPP, and tau were concomitantly enhanced and the UPS was impaired during acute brain injury caused by Toxocara canis in mice

BackgroundBecause the outcomes and sequelae after different types of brain injury (BI) are variable and difficult to predict, investigations on whether enhanced expressions of BI-associated biomarkers (BIABs), including transforming growth factor β1 (TGF-β1), S100B, glial fibrillary acidic protein (GFAP), neurofilament light chain (NF-L), tissue transglutaminases (tTGs), β-amyloid precursor proteins (AβPP), and tau are present as well as whether impairment of the ubiquitin-proteasome system (UPS) is present have been widely used to help delineate pathophysiological mechanisms in various BIs. Larvae of Toxocara canis can invade the brain and cause BI in humans and mice, leading to cerebral toxocariasis (CT). Because the parasitic burden is light in CT, it may be too cryptic to be detected in humans, making it difficult to clearly understand the pathogenesis of subtle BI in CT. Since the pathogenesis of murine toxocariasis is very similar to that in humans, it appears appropriate to use a murine model to investigate the pathogenesis of CT.MethodsBIAB expressions and UPS function in the brains of mice inoculated with a single dose of 250 T. canis embryonated eggs was investigated from 3 days (dpi) to 8 weeks post-infection (wpi) by Western blotting and RT-PCR.ResultsResults revealed that at 4 and 8 wpi, T. canis larvae were found to have invaded areas around the choroid plexus but without eliciting leukocyte infiltration in brains of infected mice; nevertheless, astrogliosis, an indicator of BI, with 78.9~142.0-fold increases in GFAP expression was present. Meanwhile, markedly increased levels of other BIAB proteins including TGF-β1, S100B, NF-L, tTG, AβPP, and tau, with increases ranging 2.0~12.0-fold were found, although their corresponding mRNA expressions were not found to be present at 8 wpi. Concomitantly, UPS impairment was evidenced by the overexpression of conjugated ubiquitin and ubiquitin in the brain.ConclusionFurther studies are needed to determine whether there is an increased risk of CT progression into neurodegenerative disease because neurodegeneration-associated AβPP and phosphorylated tau emerged in the brain.

Blood–brain barrier impairment with enhanced SP, NK‐1R, GFAP and Claudin‐5 expressions in experimental cerebral toxocariasis

Infection by Toxocara canis in humans may cause cerebral toxocariasis (CT). Appreciable numbers of T. canis larvae cross the blood–brain barrier (BBB) to invade the brain thus causing CT. In the present studies, we evaluated the BBB permeability and BBB injury as assessed by the cerebral Evans blue (EB) concentration as well as by pathological changes and glial fibrillary acidic protein (GFAP) expression in T. canis‐infected mice monitored from 3 days (dpi) to 8 weeks post‐infection (wpi). The vasodilation neuropeptides, the expressions of substance P (SP) and its preferred binding neurokinin‐1 receptor (NK‐1R) as well as claudin‐5 of tight‐junction proteins associated with BBB impairment were also assessed by Western blotting and reverse‐transcriptase polymerase chain reaction. Results revealed that BBB permeability increased as evidenced by a significantly elevated EB concentration in brains of infected mice. BBB injury appeared due to enhanced GFAP protein and mRNA expressions from 4 to 8 wpi. Leukocytes might have been unrelated to BBB impairment because there was no inflammatory cell infiltration despite T. canis larvae having invaded the brain; whereas markedly elevated SP protein and NK‐1R mRNA expressions concomitant with enhanced claudin‐5 expression seemed to be associated with persistent BBB impairment in this experimental CT model.

Seroprevalence of Toxoplasma gondii infection among children in Swaziland, southern Africa

As there appeared to be no data available on Toxocara canis infection in the children of Swaziland, a serological survey of T. canis infection was recently conducted among 92 children aged 3-12 years from rural slums in the low- and middle-veld. A child was considered seropositive if, in western blots based on the excretory-secretory antigens of larval T. canis, his or her serum gave a positive result when diluted 1 : 64. Forty-one (44.6%) of the children were found seropositive. There were no statistically significant differences in seroprevalence between the 49 boys and 43 girls investigated (46.9% v. 41.8%) or between the eight subjects aged 12 years and the 47 aged < or = 5 years (62.5% v. 38.3%); the corresponding odds ratios were 0.81 (95% confidence interval=0.36-1.86; P=0.62) and 2.69 (95% confidence interval=0.57-12.62; P=0.20), respectively. The 66 subjects from the middleveld were, however, significantly more likely to be seropositive than the 26 subjects from the lowveld (54.5% v. 19.2%; odds ratio=5.04, with a 95% confidence interval of 1.70-14.98; P<0.01). It seems likely that T. canis infection is common among the children who live in slums in Swaziland, particularly in the countrys middleveld, probably as the result of poor hygiene and poor sanitation.

Toxoplasma gondii infection: Relationship between seroprevalence and risk factors among primary schoolchildren in the capital areas of Democratic Republic of São Tomé and Príncipe, West Africa

BackgroundThe status of Toxoplasma gondii infection among primary schoolchildren (PSC) of the Democratic Republic of São Tomé and Príncipe (DRSTP), West Africa, remains unknown to date.MethodsA serologic survey and risk factors associated T. gondii infection among PSC in the DRSTP was assessed by the latex agglutination (LA) test and a questionnaire interview including parents’ occupation, various uncomfortable symptoms, histories of eating raw or undercooked food, drinking unboiled water, and raising pets, was conducted in October 2010. Schoolchildren from 4 primary schools located in the capital areas were selected, in total 255 serum samples were obtained by venipuncture, of which 123 serum samples were obtained from boys (9.8 ± 1.4 yrs) and 132 serum samples were obtained from girls (9.7 ± 1.3 yrs).ResultsThe overall seroprevalence of T. gondii infection was 63.1% (161/255). No significant gender difference in seroprevalence was found between boys (62.6%, 77/123) and girls (63.6%, 84/132) (p = 0.9). The older age group of 10 years had insignificantly higher seroprevalence (69.9%, 58/83) than that of the younger age group of 8 year olds (67.7%, 21/31) (p = 0.8). It was noteworthy that the majority of seropositive PSC (75.8%, 122/161) had high LA titers of ≥1: 1024, indirectly indicating acute or repeated Toxoplasma infection. Parents whose jobs were non-skilled workers (73.1%) showed significantly higher seroprevalence than that of semiskilled- (53.9%) or skilled workers (48.8%) (p < 0.05). Children who had a history of raising cats also showed significantly higher seroprevalence than those who did not (p < 0.001).Children who claimed to have had recent ocular manifestation or headache, i.e. within 1 month, seemed to have insignificantly higher seroprevalence than those who did not (p > 0.05).ConclusionsParents’ educational level and cats kept indoors seemed to be the high risk factors for PSC in acquisition of T. gondii infection. While, ocular manifestation and/or headache of PSC should be checked for the possibility of being T. gondii elicited. Measures such as improving environmental hygiene and intensive educational intervention to both PSC and their parents should be performed immediately so as to reduce T. gondii infection of DRSTP inhabitants including PSC and adults.

Sero-epidemiology of Toxocara canis infection among aboriginal schoolchildren in the mountainous areas of north–eastern Taiwan

Abstract A sero-epidemiological study of Toxocara canis infection was conducted among Atayal schoolchildren (aged 7–12 years) residing in the mountainous areas of north–eastern Taiwan. The 73 children investigated were each checked for anti-Toxocara IgG, in ELISA based on the larval excretory–secretory antigens of T. canis larvae. A short, self-administered questionnaire was then used to collect relevant information from each subject, including data on the keeping of dogs, playing in soil, eating raw vegetables, and whether the subjects normally washed their hands before eating. Once the seropositive children had been identified, odds ratios (OR), with their corresponding 95% confidence intervals (CI) and P-values, were calculated for each potential risk factor. When diluted 1:64, sera from 42 (57.5%) of the children gave a positive result in the ELISA, indicating that these 42 children were seropositive for T. canis infection. Seropositivity did not appear to be associated with the age or gender of the subject, the eating of raw vegetables, or the regular failure to wash hands prior to a meal. Compared with the other subjects, however, those who admitted living in a household where dogs were kept (OR = 3.79; CI = 1.23–11.69; P=0.02) or playing in soil (OR = 3.00; CI = 1.10–8.16; P=0.03) appeared at increased risk of seropositivity.

Prevalence of Enterobius vermicularis Infection among Preschool Children in Kindergartens of Taipei City, Taiwan in 2008

The prevalence of Enterobius vermicularis infection among preschool children was reported to be low based on a 5-year screening program in Taipei City, Taiwan. The Taipei City government intended to terminate the E. vermicularis screening program among preschool children. Thus, we were entrusted with confirming whether pinworm infections among preschool children in Taipei City had truly declined. From each of 12 administrative districts 2-3 kindergartens were randomly selected for investigation. In total, 4,349 children were examined, of which 2,537 were boys and 1,812 were girls. The cellophane tape adhered to a glass slide was used, and all examinations were done by certified medical technologists. Results indicated that the overall prevalence rate of pinworm infections was 0.62% (27/4,349). Although the infection rate was higher among boys (0.67%, 17/2,537) than in girls (0.55%, 10/1,812), no significant difference was found (chi(2) = 0.399, P = 0.62). According to the administrative district, the infection rate ranged from no positive cases of E. vermicularis infection in the Xinyi, Zhongzhen, and Wanhua Districts (0%; 0/299, 0/165, and 0/358, respectively), to 0.26% (1/131) in Songshan District, with the highest rate of 1.88% (7/373) in Wenshan District. Because the overall infection rate (0.62%, 27/4,349) in the present study was unchanged compared to that (0.40%, 197/49,541) previously reported in 2005, we propose that regular pinworm screening and treatment programs should be continued in some parts of Taipei City.

Seroprevalence, disease awareness, and risk factors for Toxocara canis infection among primary schoolchildren in Makoko, an urban slum community in Nigeria

In this study, we investigated the seroprevalence of Toxocara canis infection in southern Nigeria, which previously was unknown, in addition to evaluating disease awareness and potential risk factors for schoolchildren in an urban slum community. In total, 366 primary schoolchildren were investigated for the presence of anti-Toxocara IgG antibodies. Blood was collected and screened by a Western blot analysis based on the excretory-secretory antigens of larval T. canis (TcES), targeting low molecular weight bands of 24-35kDa specific for T. canis. Children were considered seropositive if their serum reacted with TcES when diluted to a titer of 1:32. Questionnaires concerning possible risk factors were given to the schoolchildren to acquire data on this infection. The overall seroprevalence of Toxocara infection was 86.1% (315/366). The logistic regression analysis of risk factors showed that childrens age (odds ratio (OR)=2.88, 95% confidence interval (CI)=1.08-7.66, p=0.03), contact with dogs (OR=0.51, 95% CI=0.28-0.94, p=0.03), the age of the dog (OR=0.34, 95% CI=0.18-0.68, p=0.002), the feeding location of the dog (OR=0.31, 95% CI=0.12-0.79, p=0.01), the consumption of raw vegetables (OR=0.89, 95% CI=0.54-1.48, p=0.004), and the drinking of unboiled water (OR=0.48, 95% CI=0.26-0.90, p=0.02) were risk factors associated with Toxocara infection. Although there was a high awareness of dogs being hosts of some parasites in this study, not much was known about T. canis. This is the first serological investigation of T. canis infection among primary schoolchildren in southern Nigeria. The high seroprevalence recorded is an indication of high transmission with the consequent risk of visceral or ocular larval migrans and neurologic toxocariasis in these children. Our findings suggest the need for prompt interventional measures, particularly health education on personal hygiene.

Enhanced expression of transforming growth factor-β1 in inflammatory cells, α-smooth muscle actin in stellate cells, and collagen accumulation in experimental granulomatous hepatitis caused by Toxocara canis in mice

Although toxocaral granulomatous hepatitis (TGH) characterized with a dominant-Th2 type immune response is a self-limiting disease, little is known concerning the role of fibrosis-related cytokine transforming growth factor-beta 1 (TGF-beta 1) in pathogenesis of TGH. A detailed histological and quantitatively immunohistochemical analysis of TGF-beta 1, alpha-smooth muscle actins (alpha-SMA), and collagen was performed on the liver tissues from mice infected with Toxocara canis as assessed between day 1 and 42 weeks post-infection (DPI or WPI). TGF-beta1 was detected mainly in infiltrating leukocytes in lesions with strong expressions from 4 to 16 WPI. Larvae per se also exhibited strong TGF-beta 1-like molecule expressions in the trial. Alpha-SMA was detected predominantly in hepatic stellate cells (HSC) which surrounded the lesions with moderate expressions largely throughout the period of the entire experiment. Collagen was observed to accumulate in inflammatory lesions and biliary basement with moderate to strong expressions from 1 WPI onwards in the trial. Since many evidences have indicated that leukocytes have the potential to influence HSC by producing TGF-beta 1 which can affect HSC to increase collagen synthesis in various liver diseases, we may propose that persistently elevated TGF-beta 1 expression in infiltrating leukocytes and active HSC with marked alpha-SMA expressions may contribute to healing of injured sites through up-stimulation of collagen deposition; in contrast, abnormally persistent collagen accumulation may cause irreversible fibrotic injury in the TGH.