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.

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.

Evaluation of Toxocara canis Glycosylated TES Produced in Pichia pastoris for Immunodiagnosis of Human Toxocariasis

Abstract Recombinant proteins are a suggested alternative for the diagnosis of toxocariasis. The current Escherichia coli recombinant protein overexpression system usually produces insoluble products. As an alternative, yeast such as Pichia pastoris have secretory mechanisms, which could diminish the cost and time for production. This study aimed to produce recombinant proteins in Pichia pastoris and verify their sensibility and specificity in an indirect ELISA assay. Two sequences (rTES-30 and rTES-120) of Toxocara canis excretory-secretory antigens were cloned in a pPICZαB vector and expressed in P. pastoris KM71H. Sera samples collected from human adults infected by Toxocara spp. were tested by indirect ELISA using rTES-30 and rTES-120 as antigens. Recombinant proteins were detected at 72 hours after induction, in the supernatant, as pure bands between 60~70 kDa with hyperglycosylation. Regarding diagnosis potential, recombinant antigens had high specificity (95.6%); however, sensitivity was 55.6% for rTES-30 and 68.9% for rTES-120. Further deglycosylation of the P. pastoris antigens did not seem to affect ELISA performance (p>0.05). The low sensitivity in the serodiagnosis diminished any advantage that P. pastoris expression could have. Therefore, we do not recommend P. pastoris recombinant TES production as an alternative for the diagnosis of toxocariasis.

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.

Sensitivity and specificity of recombinant proteins in Toxocara spp. for serodiagnosis in humans: Differences in adult and child populations.

Toxocariasis is a neglected zoonosis that affects children and adults. Recombinant proteins have been widely investigated for diagnosis, achieving high sensitivity and specificity in an overall population; however, little is known about age as a factor in its application. This study aims to investigate the diagnostic potential of Toxocara canis TES-30 and TES-120 recombinant proteins in humans, differentiating between its performance in children and adults. Serum samples collected from children and adults seropositive to Toxocara spp. were tested with indirect ELISA using T. canis TES-30 and TES-120 recombinant proteins produced in Escherichia coli. While rTES-30 sensitivity was not affected by age (81.8% in children and 87% in adults), rTES-120 sensitivity severely decreased in children to only 63.6%, down from 95.7% in adults. Furthermore, the sensitivity of rTES-30 increased to 97.8% after Western blotting confirmation. High specificity (>94%) against other geohelminths was reported for both recombinant proteins. Our study favors the use of rTES-30 with total IgG as the primary antibody in an indirect ELISA assay as a tool for epidemiological human studies.

Seroprevalence of Toxoplasma gondii and Toxocara canis in a human rural population of Southern Rio Grande do Sul.

Due to the growing population of pets, especially homeless dogs and cats, zoonoses still represent a significant public health problem. Toxoplasma gondii and Toxocara spp. are epidemiologically important zoonotic agents as they are etiological factors of human toxoplasmosis and toxocariasis, respectively. These parasites remain neglected even though they are substantially prevalent in rural areas. The aim of this study was to investigate T. gondii and T. canis seroprevalence and risk factors of seropositivity in a rural population in Pelotas municipality, Brazil. The study participants (n=344) were patients of a Basic Healthcare Unit (BHU) located in Cerrito Alegre. Blood samples were collected and tested for T. gondii antibodies by indirect immunofluorescence and T. canis antibodies by an indirect ELISA that targets an excreted-secreted antigen (TES). T. gondii seropositivity was 53.2%, with higher titers (1:256 - 1:1,024) in individuals who habitually eat pork, beef, or chicken, while T. canis seropositivity was 71.8% and concomitant T. gondii and T. canis seropositivity was 38.3%. Among the seropositivity risk factors assessed, only habitual undercooked meat consumption was significant (p = 0.046; OR = 3.7) for T. gondii and none of them were associated with T. canis seropositivity. Both parasites have a high prevalence in rural areas, which reinforces the need to invest in rural community education and health.

Detection of Toxocara canis DNA in tissues of experimentally infected mice.

The main etiological agent of toxocariasis is the helminth Toxocara canis. Several difficulties are found in the diagnosis of this disease, because of nonspecific clinical signs and possible cross-reactions that may occur in the available test, the indirect ELISA. Therefore, molecular diagnosis has been indicated as an alternative to conventional diagnosis. The purpose of this study was to evaluate the polymerase chain reaction (PCR) technique for the identification of T. canis in tissues of experimentally infected mice. To this end, nine mice were inoculated with 1500 embryonated eggs and were divided into two groups, the first euthanized 48 h (G1) and the other 30 days post inoculation (G2). Lungs, brain, liver and blood were collected from all the animals for DNA Extraction and tissue digestion, also was collected blood samples for DNA extraction and ELISA test (serum). Toxocara canis DNA was identified in all the inoculated animals using the ITS-2 target gene. The PCR test successfully identified the parasite in the brain, lung and liver of the animals euthanized 48 h PI and 30 days PI. This technique yielded good results in the identification of the parasite in the brain, being more sensitive than the method for the recovery of larvae, in the group with acute infection (48 h PI). The infection was confirmed by PCR within 48 h after infection, while the ELISA indicated serological conversion occurred only 14 days after inoculation. This study demonstrates the ability of PCR to identify T. canis in the liver, lungs and brain during acute and chronic infection.

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.

The role of quorum sensing in Escherichia coli (ETEC) virulence factors.

Quorum sensing (QS) is a signaling system among bacteria mediated by auto-inducer substances (AI). Whenever the concentration of these molecules reaches a threshold corresponding to a high cell density or quorum, the whole population starts a coordinated expression of specific genes. Studies have shown that epinephrine is also responsible for activating specific bacterial genes. This work aimed to investigate the role of conditioned medium (containing AI), epinephrine and their association on growth, motility, F4 fimbriae and heat-labile toxin (LT) expression on enterotoxigenic Escherichia coli (ETEC, E68). A significant increase in motility, F4 and LT expression, was observed in the ETEC culture supplemented with conditioned medium and epinephrine. These findings suggest that ETEC uses some components of conditioned medium (e.g., AI molecules), host molecules (epinephrine), and their association to modulate the expression of important virulence genes.

Recombinant gp19 as a potential antigen for detecting anti-Ehrlichia canis antibodies in dog sera.

The canine monocytic ehrlichiosis, caused by Ehrlichia canis, is endemic in several regions of Brazil. Some serological diagnostic techniques using immunodominant proteins of E. canis as antigens are available, but their specificities and sensitivities are questionable. Based on this, the objective of this study was to test the antigenic potential of the recombinant gp19 protein (rGP19) for subsequent use in diagnostic tests. The rGP19 expressed in the Escherichia coli strain BL21 (DE3) C41 was recognized in the sera from experimentally infected dogs using ELISA and Western blotting. Thus, it was possible to obtain a promising antigen with the ability to differentiate between E. canis-positive and -negative animals, even 1 week after infection.

elevated trans-mammary transmission of Toxocara canis larvae in BALB/c mice.

Toxocariasis is a widespread zoonosis and is considered an important worldwide public health problem. The aim of this study was to investigate the frequency of trans-mammary Toxocara canis infection in newborn BALB/c mice nursed by females experimentally infected with 1,200 eggs after delivery. After 50 days of age, the presence of larvae in different organs of the offspring was investigated. Trans-mammary infection was confirmed in 73.9% of the mice that had been nursed by infected females. These data show a high trans-mammary transmission of T. canis and confirm the significance of this transmission route in paratenic hosts.

Elevated trans-mammary transmission of Toxocara canis larvae in balb/c mice / Elevada transmissão transmamária de larvas de Toxocara canis em camundongos BALB/c

Toxocariasis is a widespread zoonosis and is considered an important worldwide public health problem. The aim of this study was to investigate the frequency of trans-mammary Toxocara canis infection in newborn BALB/c mice nursed by females experimentally infected with 1,200 eggs after delivery. After 50 days of age, the presence of larvae in different organs of the offspring was investigated. Trans-mammary infection was confirmed in 73.9% of the mice that had been nursed by infected females. These data show a high trans-mammary transmission of T. canis and confirm the significance of this transmission route in paratenic hosts.

A toxocaríase é zoonose amplamente difundida e considerada importante problema de saúde pública. O objetivo deste estudo foi avaliar a frequência da transmissão transmamária de Toxocara canis em camundongos BALB/c neonatos amamentados por fêmeas experimentalmente infectadas com 1.200 ovos logo após o parto. Após 50 dias de idade, foi avaliada a presença de larvas em diferentes órgãos dos neonatos. A infecção por via transmamária foi confirmada em 73,9% dos camundongos amamentados por fêmeas infectadas. Estes dados demonstram elevada transmissão transmamária de T. canis e confirmam a importância desta via de transmissão em hospedeiros paratênicos.

Transmammary infection in BALB/c mice with chronic toxocariasis.

Human toxocariasis is a neglected public health problem. Infection of humans generally results from the accidental ingestion of embryonated Toxocara canis eggs, but it is important to broaden knowledge about other forms of transmission. This study aimed to demonstrate the prevalence of transmammary transmission in mice with chronic toxocariasis. BALB/c mice in groups 1 (G1) and 3 (G3) were inoculated with 1200 T. canis eggs 60days before mating, whereas those of group 2 (G2) were not infected. After delivery, the G1 neonates were transferred to G2 females to be nursed, and vice versa. Thus, the mice generated by G2 females and breastfed by G1 females could be infected only during lactation. In the G3 group, offspring were not exchanged. The search for T. canis larvae in the bodies of the lactating females and their offspring was performed after weaning and at 60days old, respectively. The frequency of transmammary infection in the mice generated by G2 uninfected females and breastfed by G1 infected females was 19.8%, which was similar to that observed (19.6%) in the mice bred and fed by G3 females. The frequency of infection in the mice generated by G1 females and breastfed by G2 females was only 4.2%, which was lower than that of G1 (p=0.0064) and G3 (p=0.0062) groups. Transmammary infection by mice with chronic toxocariasis was found to be more prevalent than congenital infection.

Recombinant gp19 as a potential antigen for detecting anti-Ehrlichia canis antibodies in dog sera / gp19 recombinante como um antígeno potencial para detecção de anticorposanti-Ehrlichia canis em soros de cães

The canine monocytic ehrlichiosis, caused by Ehrlichia canis, is endemic in several regions of Brazil. Some serological diagnostic techniques using immunodominant proteins of E. canis as antigens are available, but their specificities and sensitivities are questionable. Based on this, the objective of this study was to test the antigenic potential of the recombinant gp19 protein (rGP19) for subsequent use in diagnostic tests. The rGP19 expressed in the Escherichia coli strain BL21 (DE3) C41 was recognized in the sera from experimentally infected dogs using ELISA and Western blotting. Thus, it was possible to obtain a promising antigen with the ability to differentiate between E. canis-positive and -negative animals, even 1 week after infection.

A erliquiose monocítica canina, causada por Ehrlichia canis, é uma doença endêmica em diversas regiões do Brasil. Algumas técnicas de diagnóstico sorológico, utilizando proteínas imunodominantes de E. canis como antígenos, estão disponíveis, porém suas especificidades e sensibilidades são questionáveis. Com base nesse fato, o objetivo deste trabalho foi testar o potencial antigênico da proteína GP19 recombinante (rGP19) para posterior utilização em testes diagnósticos. A rGP19, expressa em E. coli cepa BL21 (DE3) C41, foi reconhecida por soros de cães experimentalmente infectados pelas técnicas de ELISA e Western blotting. Dessa maneira, conseguiu-se obter um antígeno promissor com a capacidade de diferenciar animais positivos de negativos, até mesmo uma semana após a infecção.

Determination of IgG avidity in BALB/c mice experimentally infected with Toxocara canis.

Toxocariasis is a zoonotic disease in that IgM titers can remain high for long periods making difficult to determine the stage of the disease. The aim of this study is to investigate the applicability of indirect ELISA, associated with urea, to discriminate between the acute and chronic toxocariasis. IgG avidity was evaluated in 25 BALB/c mice experimentally infected with 1000 Toxocara canis eggs. Blood samples were collected, and sera treated with 6 M urea and assayed by ELISA every two weeks. The percent IgG avidity was determined using the mean absorbance of sera treated with urea, divided by the mean absorbance of untreated sera. In the first 15 days post-inoculation, was observed a low percentage, between 7.25 and 27.5%, IgG avidity, characteristic of an acute infection. After 60 days of infection, all the mice showed between 31.4 and 58% IgG avidity, indicating a chronic infection.

Determination of IgG avidity in BALB/c mice experimentally infected with Toxocara canis / Determinação da avidez de IgG em camundongos BALB/c experimentalmente infectados com Toxocara canis

Toxocariasis is a zoonotic disease in that IgM titers can remain high for long periods making difficult to determine the stage of the disease. The aim of this study is to investigate the applicability of indirect ELISA, associated with urea, to discriminate between the acute and chronic toxocariasis. IgG avidity was evaluated in 25 BALB/c mice experimentally infected with 1000 Toxocara canis eggs. Blood samples were collected, and sera treated with 6 M urea and assayed by ELISA every two weeks. The percent IgG avidity was determined using the mean absorbance of sera treated with urea, divided by the mean absorbance of untreated sera. In the first 15 days post-inoculation, was observed a low percentage, between 7.25 and 27.5%, IgG avidity, characteristic of an acute infection. After 60 days of infection, all the mice showed between 31.4 and 58% IgG avidity, indicating a chronic infection.

A toxocaríase é uma zoonose na qual os títulos de IgM podem permanecer elevados por longos períodos, tornando difícil a determinação do estágio em que a doença se encontra. O objetivo deste estudo foi investigar a aplicabilidade de um teste indireto de ELISA, associado com ureia, para fazer a discriminação entre as fases aguda e crônica da toxocaríase. A avidez de IgG foi avaliada em 25 camundongos BALB/c experimentalmente infectados com 1000 ovos embrionados de Toxocara canis. A cada duas semanas, amostras de sangue foram coletadas, o soro tratado com ureia 6M e realizado o ensaio pela técnica de ELISA. O percentual de avidez de IgG foi determinado, usando-se a média das absorbâncias dos soros tratados com ureia dividida pela média das absorbâncias dos soros não tratados. Nos primeiros 15 dias pós-inoculação, foi observado um baixo percentual de avidez de IgG, entre 7,25 e 27,5%, característico da fase aguda da infecção. Após 60 dias de infecção, todos apresentaram avidez de IgG entre 31,4 e 58%, indicando a fase crônica da infecção.

Risk of infection by the consumption of liver of chickens inoculated with low doses of Toxocara canis eggs.

Experimental studies and registries of cases of human toxocariasis have shown that the consumption of raw or undercooked offal of the paratenic host of Toxocara canis may pose a risk of infection. Thus, we evaluated the risk of infection due to the consumption of liver of chickens inoculated with different doses of embryonated T. canis eggs. Doses were 5-100 times smaller than the ones previously employed in this type of study. Groups of five chickens were inoculated with 5000 (control), 1000, 500, 300 or 50 eggs of T. canis, and at 72 h post-inoculation, the liver of each bird was consumed by a BALB/c receptor mouse. Forty-eight hours after consumption, we examined the organs and carcasses of the mice for larvae of T. canis. All mice were positive for larvae, except the group that consumed the chicken liver inoculated with 50 eggs. This group contained only one positive mouse, in which the larva was lodged in the brain. In mice that consumed livers of chickens inoculated with ≥300 eggs, larvae concentration was primarily in the liver and lungs, characterizing the initial phase of infection. We conclude that the consumption of raw poultry liver, under the studied conditions, poses a risk of infection even with a low number of infected T. canis eggs.

Giardia lamblia: diagnóstico com o emprego de métodos microscópicos e Enzyme-linked Immunosorbent Assay (ELISA) / Giardia lamblia: diagnosis using microscopic methods and enzyme-linked immunosorbent assay (ELISA)

O protozoário Giardia lamblia é um dos agentes etiológicos de diarreia em crianças no Brasil.Seu diagnóstico pode tornar-se difícil em consequência da baixa sensibilidade dos métodosusualmente empregados e por causa da eliminação intermitente dos cistos pode produzir resultadosfalso-negativos. Por essa razão, foi desenvolvido um ensaio imunoenzimático (ELISA) parapesquisa de antígenos de G. lamblia em fezes. Este imunoensaio tem sido descrito na literatura comoum método simples, sensível e específico quando aplicado para o diagnóstico de diversas parasitoses.Assim, este estudo objetivou comparar a técnica de ELISA com os métodos coproscópicos decentrífugo-sedimentação (Técnica de Ritchie) e centrífugo-flutuação (Técnica de Faust), visandodemonstrar a importância de um método com maior sensibilidade. Foram examinadas 158 amostrasde fezes de crianças, de 0 a 12 anos, em uma creche municipal pública de Rio Grande, no RioGrande do Sul, Brasil. Os resultados referentes à comparação entre as técnicas mostraram que atécnica de ELISA tem 3,0 vezes mais chances de detectar amostras positivas de G. lamblia que ométodo de centrífugo-flutuação. Quando comparada com o método de centrífugo-sedimentação,a técnica de ELISA demonstrou ter 3,4 vezes mais chances de detectar amostras positivas para G.lamblia.Concluiu-se que a técnica de ELISA desenvolvida mostrou-se mais eficiente que as técnicasadotadas na rotina laboratorial para o diagnóstico desta parasitose, podendo ser aplicada tanto parao diagnóstico individual como em avaliações epidemiológicas, já que permite o processamento devárias amostras simultaneamente.

The protozoan Giardia lamblia is one of the etiological agents of diarrhea in children in Brazil, anddiagnosis may be difficult due to low sensitivity of the methods commonly used, due the intermittentliberation of cysts, which may lead to false-negative results. Thus, an enzyme-linked immunosorbentassay (ELISA) was developed for detection of G. lamblia antigens in fecal samples, consideringthat this immunoassay has been described to be simple, sensitive and specific when applied todiagnose various parasitic diseases. Thus, this study aimed to compare the ELISA technique withsingle stool sample to examination methods using centrifugal sedimentation (Ritchie technique) andcentrifugation-flotation (Faust technique), intending to demonstrate the importance of a method withgreater sensitivity. Fecal samplesfrom a total of 158 children aged 0-12 years, were examined in apublic municipal nursery in the city of Rio Grande, Rio Grande do Sul, Brazil. The comparative resultsof the investigated techniques showed that ELISA has 3.0 times more chance than centrifugationflotation,and 3.4 times more chance than centrifugal sedimentation to detect positive samples ofG. lamblia. Overall, it was concluded that the ELISA was more efficient than the routine laboratorytechniques for the diagnosis of giardiasis, and it may be used for both individual and epidemiologicalassessments, as the technique allows for processing of multiple samples simultaneously.

Vertical transmission of Toxocara canis in successive generations of mice / Transmissão vertical de Toxocara canis em gerações sucessivas de camundongos

Visceral toxocariasis is a neglected zoonosis caused by Toxocara canis larvae in unusual hosts. In dogs, the definitive host, the infection occurs mainly through transplacental and transcolostral transmission. Studies on experimental models have shown that vertical transmission may result from acute infections. Considering that toxocariasis is characterized as a chronic infection, with possible reactivation of larvae present in the brain, this study evaluated the presence of larvae in the brain of female BALB/c mice and their offspring with chronic infection during three successive pregnancies. ELISA-TES was used to evaluate the antibody levels. T. canis larvae were detected in the brain tissue of the mice during the three successive generations evaluated. The offspring's IgG level gradually decreased, and mean absorbance (ABS) above the cutoff point (0.070) was observed only at 30 (0.229) and 50 (0.096) days of age, while IgM was not detected. The infections in the offspring confirmed that vertical transmission of T. canis larvae occurred during chronic toxocariasis in three successive generations of mice.

A toxocaríase visceral é uma zoonose negligenciada causada por larvas de Toxocara canis em hospedeiros não usuais. Em cães, os hospedeiros definitivos, a infecção ocorre normalmente por transmissão transplacentária e através do colostro. Estudos com modelos experimentais têm demonstrado a ocorrência de transmissão vertical durante a infecção aguda. Considerando que a toxocaríase é caracterizada como uma infecção crônica, com uma possível reativação das larvas presentes no cérebro, este estudo avaliou a presença de larvas no cérebro de camundongos Balb/C fêmeas e suas proles com infecção crônica durante três gestações sucessivas. Para avaliar os níveis de anticorpos foi utilizado ELISA-TES. Larvas de T. canis foram detectadas no encéfalo dos animais durante as três gerações sucessivas avaliadas. O nível de IgG das proles foi diminuindo gradualmente e as médias de absorbâncias (ABS) acima do ponto de corte (0,070) foram evidenciadas somente aos 30 (0,229) e 50 dias (0,096) de vida, enquanto que não foi detectada IgM. Infecções das proles confirmam a transmissão vertical de larvas de T. canis durante a toxocaríase crônica em três gerações sucessivas de camundongos.