Prevalencia de Toxoplasma gondii en mujeres embarazadas asintomáticas en Quito, Ecuador 2020 / Prevalence of Toxoplasma gondii in asymptomatic pregnant women in Quito, Ecuador 2020

La toxoplasmosis es causada por un protozoo intracelular, Toxoplasma gondii, que tiene una amplia distribución geográfica. La forma congénita resulta en una forma gestacional que puede presentar una parasitemia temporal que infectará al feto. Es adquirido por consumo de carne cruda o poco cocida, alimentos o agua contaminada e infección congénita a través de la placenta. El objetivo de este estudio fue determinar la prevalencia de infección por toxoplasmosis y sus posibles factores de riesgo asociados con mujeres embarazadas que asistieron al servicio de control prenatal en el Hospital Gineco Obstétrico "Isidro Ayora", Ecuador. Las muestras de sangre se analizaron para detectar la presencia de anticuerpos IgG e IgM contra T. gondii utilizando una prueba de hemoaglutinación. Se probó la significación de los factores de riesgo mediante análisis bivariado. Se consideró un valor de p <0,05 como estadísticamente significativo. La prevalencia de anticuerpos IgG e IgM fue de 16,32%. Se observó una asociación significativa entre la seroprevalencia y antecedentes de aborto (p=0,00804), contacto con gatos domésticos (p<0,0001) y hábitos higiénicos incorrectos (p<0.0001). Los hallazgos demostraron que el contacto con gatos, antecedentes de abortos y hábitos higiénicos incorrectos se identificaron como factores de riesgo de infección por T. gondii(AU)

Toxoplasmosis is caused by an intracellular protozoan, Toxoplasma gondii, which has a wide geographical distribution. The congenital form results in a gestational form that can present a temporary parasiteamia that will infect the fetus. It is acquired by consumption of raw or undercooked meat, food or water contaminated and congenital infection through the placenta. Tthe objective of this study was to determine the prevalence of toxoplasmosis infection and its possible risk factors associated with pregnant women who attended the prenatal control service at the Gyneco-Obstetric Hospital "Isidro Ayora", Ecuador. Blood samples were tested for presence of IgG and IgM antibodies against T. gondii using a hemagglutination test. The risk factors were tested for significance using bivariate analysis. P-value <0.05 was considered statistically significant. The prevalence of IgG and IgM antibodies was 16.32%. A significant association was observed between seroprevalence and history of abortion (p=0,00804), contact with domestic cats (p<0,0001) and incorrect hygiene habits (p<0,0001). The findings demonstrated that contact with cats, a history of miscarriages, and poor hygiene habits were identified as risk factors for T. gondii infection(AU)

Seroprevalence of Brucella ovis-epididymitis, smooth-Brucella, leptospirosis, toxoplasmosis, and Maedi-Visna in sheep slaughtered in Minas Gerais State, Brazil / Soroprevalência de Brucella ovis, Brucella lisa, leptospirose, toxoplasmose e Maedi-Visna em ovinos abatidos em Minas Gerais, Brasil

The present study aimed to estimate the prevalence of antibodies against Brucella ovis-epididymitis, smooth-Brucella, leptospirosis, toxoplasmosis and Maedi-visna in sheep slaughtered in Minas Gerais, Brazil and to study their simultaneous occurrence, including caseous lymphadenitis, at sheep and flock levels. The study was conducted at a sheep slaughterhouse with Federal Inspection Service. Sera from 594 animals from 21 flocks were collected, in 2007. The agar gel immunodiffusion (AGID) was employed to detect anti-B. ovis and anti-Maedi Visna antibodies, whereas Rose Bengal (RB) and the 2-mercaptoethanol test (2ME) were used to test anti-smooth Brucella antibodies. For the detection of anti-Leptospiraantibodies, sera were examined by microscopic agglutination test (MAT), while for the detection of IgG antibodies to Toxoplasma gondii ELISA was used. Prevalence of antibodies against smooth Brucella, B. ovis-epididimitis, Leptospiraspp., toxoplasmosis and Maedi-Visna found in sheep from Minas Gerais was 0.00%, 24.04%, 25.96%, 10.46% and 3.08%, respectively; whereas the seroprevalence in flocks was 0.00%, 80.95%, 90.48%, 71.43% and 23.81%, respectively. Moreover, when data on antibodies anti-Corynebacterium pseudotuberculosis, previously obtained, were included, about 60% of the flocks showed animals that were exposed to four or more of the studied agents. However, only 25.47% of the sheep exhibited simultaneously antibodies against more than one pathogen. Thus, data from the present study on sheep slaughtered in Minas Gerais, Brazil, showed no antibodies to smooth-Brucella and a low frequency of antibodies anti-Maedi Visna lentivirus, and a high and widespread seroprevalence of B. ovis, Leptospira spp., and T. gondii among animals and flocks.(AU)

O presente estudo teve como objetivo estimar a prevalência de anticorpos contra Brucella ovis (epididimite ovina), Brucellalisa, leptospirose, toxoplasmose e Maedi-visna em ovinos abatidos em Minas Gerais, Brasil, e estudar sua ocorrência simultânea, incluindo linfadenite caseosa, nos ovinos e nos rebanhos. O estudo foi realizado em um abatedouro de ovinos com Serviço de Inspeção Federal. Soros de 594 animais de 21 rebanhos foram coletados, em 2007. A imunodifusão em gel de ágar (IDGA) foi empregada para detectar anticorpos anti-B. ovis e anticorpos anti-Maedi Visna, enquanto o teste do antígeno acidificado tamponado (AAT) e o teste de 2-mercaptoetanol (2ME) foram utilizados para testar anticorpos anti-Brucella lisa. Para a detecção de anticorpos anti-Leptospira, os soros foram examinados pelo teste de aglutinação microscópica (MAT), enquanto que para a detecção de anticorpos IgG para Toxoplasma gondii, foi usado o ELISA. A prevalência de anticorpos anti-Brucella lisa, B. ovis, Leptospira spp., toxoplasmose e Maedi-Visna encontrados em ovinos de Minas Gerais foi de 0,00%, 24,04%, 25,96%, 10,46% e 3,08%, respectivamente; enquanto a soroprevalência em rebanhos foi de 0,00%, 80,95%, 90,48%, 71,43% e 23,81%, respectivamente. Além disso, quando dados de anticorpos anti-Corynebacterium pseudotuberculosis, previamente obtidos, foram incluídos, cerca de 60% dos rebanhos apresentaram animais expostos a quatro ou mais dos agentes estudados. No entanto, apenas 25,47% dos ovinos exibiram simultaneamente anticorpos contra mais de um patógeno. Assim, os dados do presente estudo sobre ovinos abatidos em Minas Gerais, Brasil, mostram que ausência de anticorpos anti-Brucella lisa e baixa frequência de anticorpos anti-Maedi Visna, e uma soroprevalência alta e generalizada de B. ovis, Leptospira spp. e T. gondii entre animais e rebanhos.(AU)

Cross-sectional survey for toxoplasmosis in free-range chickens (Gallus gallus domesticus) from the Atlantic Forest area in Alagoas state, Northeastern Brazil / Estudo transversal para toxoplasmose em galinhas caipiras (Gallus gallus domesticus) de área de Mata Atlântica do estado de Alagoas, Nordeste do Brasil

Abstract Toxoplasmosis is a parasitic disease, which is prevalent across the world and has a strong environmental component in its transmission chain. The present study determined the seroprevalence and factors related to Toxoplasma gondii infection among free-range chickens from an Atlantic Forest area in Northeastern Brazil. A total of 550 free-range chickens were tested for T. gondii antibodies usingthe indirect fluorescent antibody test (IFAT, cut-off point 1:16), and 269 (48.9%; 95% CI = 44.7-53.1%) were positive. Among the 61 properties 57 (93.4%) had at least one seropositive animal. The robust Poisson regression model showed that the variables/categories associated with the seroprevalence of T. gondii were: region of origin of the animals/urban area (prevalence ratio [PR] = 2.346; P<0.001), management system/use of cages (PR = 1.591; P = 0.019), presence of rodents/yes (PR = 1.295; P = 0.035), and the type of food/use of food scraps (PR = 1.603; P = 0.009). The high prevalence found suggest that this scenario demands careful management, mainly regarding the use of scraps of undercooked or raw food, adequate environmental hygiene and frequent rodentcontrol.

Resumo A toxoplasmose é uma doença parasitária mundial com um forte componente ambiental em sua cadeia de transmissão. Nesta pesquisa, foram determinados a soroprevalência e os fatores associados em galinhas caipiras de uma área de Mata Atlântica no Nordeste do Brasil. Um total de 550 galinhas caipiras foi testado para anticorpos anti-Toxoplasma gondii pela reação de imunfluorescência indireta (RIFI) com ponto de corte de 1:16, e 269 (48,9%; IC95% = 44,7- 53,1%) foram positivas. Das 61 propriedades visitadas, 57 (93,4%) apresentaram pelo menos um animal positivo. O modelo de regressão de Poisson robusta mostrou que as variáveis/categorias associadas à soroprevalência de T. gondii foram: origem dos animais/área urbana (razão de prevalência - RP = 2,346; P<0,001), sistema de manejo/uso de gaiolas (RP = 1,591; P = 0,019), presença de roedores/sim (RP = 1,295; P = 0,035), e tipo de alimento/uso de sobras de alimentos (RP = 1,603; P = 0,009). A alta prevalência sugereque este cenário demanda cuidados de manejo, principalmente no que se refere ao fornecimento de sobras de alimentos mal cozidos ou crus, higiene adequada do meio ambiente e controle frequente de roedores.

Condução da toxoplasmose gestacional / Management of gestational toxoplasmosis

A toxoplasmose é uma doença proveniente do Toxoplasma gondii, um protozoário que tem os felinos como seu hospedeiro definitivo e os mamíferos e aves como seu hospedeiro intermediário. Tem um curso benigno e autolimitado quando acomete um indivíduo imunocompetente, no entanto a infecção durante a gestação acarreta até 50% de chance de toxoplasmose congênita, podendo causar danos severos ao feto. A virulência dos genótipos encontrados nas Américas Central e do Sul é a mais alta, comparada a Europa e América do Norte, tendo a doença um comportamento mais agressivo. Os estudos relatam a diminuição da infecção fetal em até 60% com o uso da espiramicina, usada ainda na profilaxia. Este artigo discute sobre a triagem materna pré-natal e sua necessidade, a profilaxia e o tratamento da infecção fetal ainda intraútero, com o objetivo de diminuir a transmissão vertical e as sequelas neonatais com suas implicações ao longo da vida.(AU)

Toxoplasmosis it is a disease originating from Toxoplasma gondii, a protozoan that has felines at as ultimate host and mammals and birds at as intermediate host. Has a benign and self-limiting course when affects immunocompetent individual, however, infection during pregnancy leads 50% chance of congenital toxoplasmosis and can cause severe damage to the fetus. The virulence of genotypes found in Central and South America is the highest compared to Europe and North America, having the disease a more aggressive behavior. Studies report a reduction in fetal infection 60% with the use spiramycin still used for prophylaxis. This article discusses prenatal maternal screening, prophylaxis and treatment of fetal infection still in utero with the objective of decreasing vertical transmission and neonatal sequelae with their lifelong implications.(AU)

Immunological and Molecular Diagnosis of T. gondii Infection among Aborted Women in Sana’a Capital and Capital Trusteeship, Yemen

This study was conducted to evaluate polymerase chain reaction (PCR) assay to determine recent infections with Toxoplasma gondii in pregnant women. T. gondii DNA was detected by using the B1 gene as a target for amplification which is highly specific for T. gondii. The overall prevalence of Toxoplasma gondii infection was: PCR was positive in 32 subjects (8.0%) and 2 of controls (2.0%), 44 (11%) had positive IgM and 1.0 (1.0%) of controls had positive for IgM, whereas IgG antibody was positive in 176 (44%) compared to 29 (29%) was positive in the control group. There was no significant association between positivity PCR and abortion whereas there was a significant association between IgM positive and abortion. There was a significant association between eating raw or undercooked meat, contact with cats, contact with soil, and having domestic animals and positive IgM ELISA and PCR positive. There was no significant relationship between age, residency, educational level and either ELISA or PCR. This study showed that the use of PCR for a confirmatory test to detect primary acute toxoplasmosis in pregnant women is very important. It demonstrates the possibility of defining and selecting the high-risk cases for mother-to-child transmission of infection by combining specific serology and PCR tests to formulate a specific approach.

Therapeutic Targets on Toxoplasma gondii Parasite in Combatting Toxoplasmosis

The status of toxoplasmosis management is challenged by drug intolerance, compromised efficacy and potential development of drug resistance. However, currently, there are very limited targets on T. gondii that serve as the site of action of current medications. This review aimed to explore more potential targets that are essential to parasite survival and are absent in humans. Many unexplored targets on T. gondii exist and their specificities to the parasite make them ideal drug targets. The present review had searched relevant English databases such as PubMed, Scopus, Google scholar and Science Direct, for relevant literature on the therapeutic target of T. gondii. Many of the enzyme systems in several pathways are essentially palatable as drug targets. Establishing compounds that can target these enzymes on T. gondii will greatly be beneficial in treating acute and chronic toxoplasmosis in humans.

Dipenyleneiodonium Induces Growth Inhibition of Toxoplasma gondii through ROS Induction in ARPE-19 Cells

Based on the reactive oxygen species (ROS) regulatory properties of diphenyleneiodonium (DPI), we investigated the effects of DPI on host-infected T. gondii proliferation and determined specific concentration that inhibit the intracellular parasite growth but without severe toxic effect on human retinal pigment epithelial (ARPE-19) cells. As a result, it is observed that host superoxide, mitochondria superoxide and H2O2 levels can be increased by DPI, significantly, followed by suppression of T. gondii infection and proliferation. The involvement of ROS in anti-parasitic effect of DPI was confirmed by finding that DPI effect on T. gondii can be reversed by ROS scavengers, N-acetyl-L-cysteine and ascorbic acid. These results suggest that, in ARPE-19 cell, DPI can enhance host ROS generation to prevent T. gondii growth. Our study showed DPI is capable of suppressing T. gondii growth in host cells while minimizing the un-favorite side-effect to host cell. These results imply that DPI as a promising candidate material for novel drug development that can ameliorate toxoplasmosis based on ROS regulation.

Virulence of VEG strain Toxoplasma gondii oocysts against Kunming mice / 中国人兽共患病学报

To explore the pathogenicity of VEG strain Toxoplasma gondii oocysts on China Kunming mice,T.gondii oocysts of ＜ 1 and 102-108 were chosen to feed the mice orally.And the modified agglutination test (MAT),H&E and IHC were used to check the infection of mice.The infection rate,the survival rate of mice,and number of cysts in brain were analyzed.Results showed that 100％ of the mice fed with ≥102 oocysts were infected,the minimum lethal dose was 102 oocysts and the 100％ lethal dose was 108 oocysts.The time of death in acute infection was 7 DPI to 14 DPI.T.gondii cysts formation rate was 32.14％ (9/28),and the number of cysts was 9 to 857 per mouse.The survival rate of infected mice was 67.44％ (29/43),and the longest survival time was more than 390 DPI.Accordingly,the virulence of T.gondii VEG strain is medium,and has a higher cysts formation rate.

Toxoplasmic encephalitis: role of Human Leucocyte Antigens/alleles associated with rapid progression to Acquired Immunodeficiency Syndrome

Abstract Background/aims The frequency of Human Leucocyte Antigens/alleles associated with rapid progression from Human Immunodeficiency Virus infection to Acquired Immunodeficiency Syndrome was evaluated in Brazilian patients with Acquired Immunodeficiency Syndrome with and without Toxoplasmic Encephalitis. Methods 114 patients with Acquired Immunodeficiency Syndrome (41 with Toxoplasmic Encephalitis, 43 with anti-Toxoplasma gondii antibodies, without Toxoplasmic Eencephalitis, and 30 without anti-Toxoplasma gondii antibodies circulating and without Toxoplasmic Encephalitis) were studied. Results Human Leucocyte Antigens/alleles associated with rapid progression to Acquired Immunodeficiency Syndrome, particularly HLA-B35, -DR3, and -DR1 allele group, were significantly less represented in patients with Toxoplasmic Encephalitis and Acquired Immunodeficiency Syndrome. Conclusion The presence of these Human Leucocyte Antigens/Alleles that predispose to Acquired Immunodeficiency Syndrome progression was associated with resistance to Toxoplasmic Encephalitis among Human Immunodeficiency Virus-1 carriers.

Occurrence of antibodies to Toxoplasma gondii in opossums caught in urban areas of Bauru, state of São Paulo, Brazil / Ocorrência de anticorpos para Toxoplasma gondii em gambás capturados na área urbana de Bauru, Estado de São Paulo, Brasil

Toxoplasmosis is one of the most common zoonoses worldwide. It was initially described in rodents and rabbits. There are few data on the morbidity and mortality of this disease among Brazilian marsupial fauna, such as opossums. These animals are of great importance regarding the epidemiology of this disease, given that they are prey for felids and other carnivores. With the aim of ascertaining the serological response to Toxoplasma gondii among marsupials (Didelphis spp.), 38 animals that had been caught in 14 districts of the urban area of the municipality of Bauru, state of São Paulo, were evaluated. The modified agglutination test (MAT) showed that 26.3% (10/38) of the samples analyzed were seropositive. It can be suggested that the opossums' behavior and persistent proximity to human housing results in contact with cats and T. gondii infection, based on the frequency found in this study. This was the first study on the seroprevalence of T. gondii in opossums caught in the urban area of the municipality of Bauru, SP, and it highlights the need for environmental and health authorities of the municipality to monitor this zoonosis.(AU)

A toxoplasmose é uma das zoonoses mais comuns no mundo, tendo sido descrita inicialmente em roedores e em coelhos. Todavia, poucos são os dados sobre morbidade e mortalidade da toxoplasmose nos marsupiais da fauna brasileira, como os gambás, sendo de grande importância na epidemiologia da doença, como presas para felídeos e outros carnívoros. Com o objetivo de verificar a resposta sorológica para Toxoplasma gondii em marsupiais (Didelphis spp.), foram avaliados 38 animais capturados em 14 regiões da área urbana do município de Bauru-SP. Foi encontrada uma frequência, de acordo com o teste de aglutinação modificada (MAT), de 26,3% (10/38) nas amostras analisadas. Pode-se sugerir que o comportamento dos gambás e sua permanência próxima a habitações humanas resultam em contato com gatos e infecção por T. gondii, tendo em vista a frequência encontrada neste estudo. Este é o primeiro estudo de soroprevalência de T. gondii em gambás capturados na área urbana do município de Bauru-SP, alertando-se para a necessidade do monitoramento desta zoonose pelas autoridades de vigilância ambiental e sanitária do município.(AU)

Occurrence of anti-Toxoplasma gondii antibodies in meat and dairy goat herds in Rio Grande do Norte, Brazil / Ocorrência de anticorpos anti-Toxoplasma gondii em rebanhos caprinos de corte e leiteiros do Rio Grande do Norte, Brasil

Toxoplasmosis is caused by Toxoplasma gondii, which is the main causative agent of abortion in small ruminants. Goats are among the animals that are most susceptible to this protozoon, and the disease that it causes leads to significant economic losses and has implications for public health, since presence of the parasite in products of goat origin is one of the main sources of human infection. Because of the significant economic impact, there is an urgent need to study the prevalence of T. gondii infection among goats in Sertão do Cabugi, which is the largest goat-producing region in Rio Grande do Norte. In the present study, the ELISA assay was used to test 244 serum samples from nine farms, located in four different municipalities in the Sertão do Cabugi region, which is an important goat-rearing region. The results showed that the prevalence of anti-T. gondii antibodies was 47.1% and that there was a significant association between positivity and the variables of age (≥ 34 months), location (Lajes, Angicos and Afonso Bezerra) and farm (all the farms). The avidity test was applied to all the 115 ELISA-positive samples to distinguish between acute and chronic infection. One hundred and three samples (89.6%) displayed high-avidity antibodies, thus indicating that most of the animals presented chronic infection, with a consequent great impact on the development of the goat production system and a risk to human health.

A toxoplasmose é causada pelo Toxoplasma gondii, principal agente causador de aborto em pequenos ruminantes. Os caprinos são uns dos animais mais suscetíveis a esse protozoário, levando a perdas econômicas significativas e implicações para a saúde pública, uma vez que a presença do parasito em produtos de origem caprina é uma das principais fontes de infecção humana. Devido ao impacto econômico significativo torna-se urgente estudar a prevalência da infecção, pelo T. gondii, entre caprinos do Sertão do Cabugi, a maior região produtora de caprinos no Rio Grande do Norte. O presente estudo utilizou o ELISA para testar 244 amostras de soro de 9 fazendas, situadas em 4 diferentes cidades na região do Sertão do Cabugi; uma importante região de criação de cabras. Os resultados mostraram uma prevalência de 47,1% para anticorpos anti- T. gondii e uma significativa associação entre a positividade e as variáveis idade (≥ 34 meses), localização (Lajes, Angicos e Afonso Bezerra e propriedade (todas as fazendas). O teste de avidez foi aplicado a todas as 115 amostras positivas pelo ELISA para discriminar entre infecção aguda e crônica. Cento e três amostras (89,6%) apresentaram anticorpos de alta avidez; indicando que a maioria dos animais estavam em infecção crônica, gerando um grande impacto sobre o desenvolvimento do sistema de produção em cabras e um risco para a saúde humana.

Formas alternasdde transmisión de toxoplasma gondii / Alternative ays of toxoplasma gondii transmission

El Toxoplasma gondii, es un protozoario descubierto hace poco más de un siglo, es un parásito de gran importancia epidemiológica, que puede ocasionar en mujeres embarazadas desde complicaciones neurológicas en el producto, hasta muerte fetal, incluso abortos y encefalitis en pacientes con SIDA, convirtiéndose en Colombia en un grave problema de salud pública. En la revisión de la evidencia disponible, se ha llegado a concluir que las formas en que el parásito llega a infectar al ser humano son muy variadas, incluyendo las formas conocidas desde tiempo atrás como: alimentos contaminados, contacto con heces felinas; además de otras formas no tan conocidas como son: la vía transfusional, por trasplantes de órganos, la vía acuática, la mecánica, la inhalación de ooquistes y la vía sexual; involucrando en el ciclo de vida del parásito, a especies animales diferentes a los felinos, tales como perros e insectos. Así mismo, se resalta la importancia de factores antrópicos y medio ambientales, como moduladores de las dinámicas de transmisión y del ciclo de vida del parasito. Esta revisión, proporciona bases sólidas para alertar al sistema de salud pública colombiano, acerca de la necesidad de adoptar medidas higiénicas y sanitarias para evitar la difusión de esta patología entre poblaciones de alto riesgo.

Toxoplasma gondii is a protozoan discovered more than a century ago. It is a parasite of great epidemiologic importance which can cause from neurologic complications in the product to fetal death in pregnant women, abortion and encephalitis in patients with AIDS, thus becoming a public health problem in Colombia. In this review of the available evidence, it was concluded that the forms in which the parasite infects human beings are very varied, including the forms known long time ago such as contaminated food, contact with feline feces, as well as other not so well known ways like transfusions, organ transplants, waterborne, and mechanic way, ooquist inhalation and sexual transmission, involving animal species different from felines in the parasite lifecycle, such as dogs and insects. Likewise, the importance of anthropic and environmental factors such as modulators of the transmission dynamics and lifecycle of the parasite are highlighted. This review, provides solid bases for alerting the public health systems about the need to adopt hygienic and sanitary measures to avoid the diffusion of this pathology among high risk populations.

Laboratory diagnosis of Toxoplasma gondii infection with direct and indirect diagnostic techniques.

Background: Toxoplasma gondii infection, common parasitic zoonoses, is an important cause of spontaneous abortion, mental retardation, encephalitis, ocular disease and death worldwide. Today the major diagnostic techniques for the toxoplasmosis are serological assays, but its have many limitations. Aim : The goal in this study is to improve the diagnostic accuracy of T. gondii infection, using direct (Real Time PCR) and indirect (IgM, IgA, IgG and IgG avidity) diagnostic techniques. Materials and Methods: In the period between 2007 and 2008, 96 non consecutive different clinical samples (38 blood, 40 amniotic fluids, 8 cerebrospinal fluids, 10 vitreous humors) and 96 sera have been studied simultaneously through molecular biology and serological techniques. Results: Direct and indirect diagnostic techniques used in this study for laboratory diagnosis of T. gondii infection were always concordant. Conclusions : The high correlation between direct and indirect diagnostic techniques exhibit that serologic techniques are accurate diagnostic assays as screening test in laboratory diagnosis of toxoplasmosis.

Primary culture of intestinal epithelial cells as a potential model for Toxoplasma gondii enteric cycle studies

The primary culture of intestinal epithelial cells from domestic cats is an efficient cellular model to study the enteric cycle of Toxoplasma gondii in a definitive host. The parasite-host cell ratio can be pointed out as a decisive factor that determines the intracellular fate of bradyzoites forms. The development of the syncytial-like forms of T. gondii was observed using the 1:20 bradyzoite-host cell ratio, resulting in similar forms described in in vivo systems. This alternative study potentially opens up the field for investigation into the molecular aspects of this interaction. This can contribute to the development of new strategies for intervention of a main route by which toxoplasmosis spreads.

Spontaneous stage differentiation of mouse-virulent Toxoplasma gondii RH parasites in skeletal muscle cells: an ultrastructural evaluation

Although the predilection for Toxoplasma gondii to form cysts in the nervous system and skeletal and heart muscles has been described for more than fifty years, skeletal muscle cells (SkMCs) have not been explored as a host cell type to study the Toxoplasma-host cell interaction and investigate the intracellular development of the parasite. Morphological aspects of the initial events in the Toxoplasma-SkMC interaction were analysed and suggest that there are different processes of protozoan adhesion and invasion and of the subsequent fate of the parasite inside the parasitophorous vacuole (PV). Using scanning electron microscopy,Toxoplasma tachyzoites from the mouse-virulent RH strain were found to be attached to SkMCs by the anterior or posterior region of the body, with or without expansion of the SkMC membrane. This suggests that different types of parasite internalization occurred. Asynchronous multiplication and differentiation of T. gondii were observed. Importantly, intracellular parasites were seen to display high amounts of amylopectin granules in their cytoplasm, indicating that tachyzoites of the RH strain were able to differentiate spontaneously into bradyzoites in SkMCs. This stage conversion occurred in approximately 3 percent of the PVs. This is particularly intriguing as tachyzoites of virulent Toxoplasma strains are not thought to be prone to cyst formation. We discuss whether biological differences in host cells are crucial to Toxoplasma stage conversion and suggest that important questions concerning the host cell type and its relevance in Toxoplasma differentiation are still unanswered.

Expression and identification of the multiple gene ROP2-P30 of Toxoplasma gondii in E.coli BL21 / 中国人兽共患病学报

To obtain the functional fusion protein of rhoptry protein 2, compound rhoptry protein2 and surface antigen 1 of Toxoplasma gondii. the ROP2 and P30 genes from genomic DNA of T.gondii RH strain were amplified by PCR, and were inserted into pMD18-T cloning vector. Then the ROP2 fragment was subcloned to pET-30a(+) plasmid digested by EcoRⅠand Hind Ⅲ to construct plasmid pET-ROP2. Furthermore,the P30 fragment was subcloned into pET-ROP2 digested by BglⅡand EcoRⅠto create plasmid pET-ROP2-P30, the resulting recombinant plasmids , transformed into E.coli BL21 (DE3), were induced with IPTG. and the proteins identified by SDS-PAGE were further purified and refolded. The biological activity was analyzed by Western blot with specific antibody. It was found that the sizes of ROP2 and ROP2-P30 were 1212 and 1896bp with corresponding molecular weight 50- kDa and 75-kDa, respectively. The recombinant protein ROP2 (50-kDa) could specifically react with rabbit-polyclonal antiserum, and complex fusion protein ROP2-P30 (75- kDa) could react with P30 monoclonal antibody.

Over-expression and purification of the recombinant p30 antigen of Toxoplasma gondii / 中国人兽共患病学报

To provide the basis for preparation of diagnostic kits and vaccines in Toxoplasma gondii infection, the gene coding for the qualified recombinant p30 protein (SAG1) of this parasite was amplified by PCR, and the amplified gene was cloned into prokaryotic expression vector pET-30a(+) to construct the recombinant plasmid, and then transformed to E.coli DH5α. The positive recombinant plasmid was screened by PCR and double enzymes digestion, and the nucleotide sequence of p30 gene was determined by automated DNA sequencing. Meanwhile, the identified recombinant plasmid was transformed to E.coli BL21(DE3) with the expression of p30 on bacteria induced by IPTG and the expressed protein was identified by SDS-PAGE. The protein obtained was then further purified and refolded, and its biological activity was checked by Western blotting. It was shown that the size of the amplified gene was 750 bp with molecular weight of 30 ku, and this protein could specifically react with monoclonal antibody against p30 protein.

AMPLIFICATION, CLONING AND PROKARYOTIC EXPRESSION OF GENE FOR MAJOR SURFACE ANTIGEN OF TOXOPLASMA GONDII / 中国人兽共患病学报

Aim To amplify P30 gene and express P30 fusion with GST Methods P30 gene was smplified from T. gondii chromosomal DNA and ligated to pGEM-T and pGEX-4T-1. Screening-positive recombinants were induced for expression, which was subsequently detected by WB Results P30 gene was amplified and GST-fusion was confirmed by rabbit antiT. gondii serum. Conclusions The construction of pGEM-T-P30 and pGEX-4T-1-P30, together with the recombinant protein would lay a base for further investigation of P30 at a molecule-level and application to diagnosis and vaccination