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1.
Ecotoxicol Environ Saf ; 213: 112048, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33610941

RESUMO

We conducted a large-scale epidemiological investigation to detect the prevalence of Toxoplasma gondii in four marine bivalve shellfish species collected from six representative coastal regions of Weihai, eastern China. Between January 2018 and December 2018, 14,535 marine bivalve shellfish pooled into 2907 samples were randomly collected and examined for T. gondii DNA by a nested PCR assay targeting B1 gene. The results showed that 2.8% (82) of the 2907 pooled samples were tested positive for T. gondii DNA. Two T. gondii genotype (ToxoDB Genotype #9 and ToxoDB Genotype #1) were identified PCR-restriction fragment length polymorphism analysis. Factors that were found significantly associated with the presence of T. gondii DNA in marine bivalve shellfish included the source of samples (being wild) (odds ratio [OR], 3.34; 95% confidence interval [CI], 2.00-5.84; p < 0.01), surface runoff near the sampling site (OR, 2.64; 95% CI, 1.47-4.72; p < 0.01), and presence of cats near the sampling site (OR, 1.77; 95% CI, 1.02-3.07; p = 0.04). Moreover, the prevalence of T. gondii DNA in marine bivalve shellfish correlated with temperature (Pearson's correlation: R = 0.75, p = 0.0049) and precipitation (R = 0.87, p = 0.00021). These findings provide new insights into the presence of T. gondii DNA in marine bivalve shellfish and highlight the impact of human activity on marine pollution by such an important terrestrial pathogen pollutant.


Assuntos
Bivalves/parasitologia , Toxoplasmose Animal/epidemiologia , Animais , Bivalves/genética , China/epidemiologia , Genótipo , Humanos , Reação em Cadeia da Polimerase/métodos , Prevalência , Fatores de Risco , Frutos do Mar , Toxoplasma/genética
2.
Rev Bras Parasitol Vet ; 30(1): e028520, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33605391

RESUMO

This study aimed to identify members of the Sarcocystidae family in naturally infected wild birds at a rescue center in the state of Minas Gerais, southeastern Brazil. The heart and brain of 44 wild birds were evaluated by bioassay in mice to detect T. gondii, and extracted DNA was used for nested PCR of the 18S ribosomal DNA gene to detect members of the Sarcocystidae family. The positive samples were sequenced, assembled, edited and compared with sequences deposited in GenBank. Toxoplasma gondii was isolated from six (13.6%) out of 44 birds. Toxoplasma gondii DNA was identified in 10/44 (22.7%) of the birds. The amplified sequences exhibited 100% similarity with the DNA of the ME49 strain of T. gondii. Sarcocystis DNA (99% similarity) was identified in 5/44 (11.4%) of the birds. T. gondii and Sarcocystis spp. are common in wild birds in Minas Gerais, Brazil.


Assuntos
Doenças das Aves , Coccidiose , Sarcocystidae , Animais , Bioensaio , Doenças das Aves/epidemiologia , Doenças das Aves/parasitologia , Aves , Brasil , Coccidiose/epidemiologia , DNA de Protozoário , Camundongos , Reação em Cadeia da Polimerase , RNA Ribossômico 18S/genética , Sarcocystidae/genética , Sarcocystis/genética , Toxoplasma/genética , Toxoplasmose Animal/epidemiologia
3.
Nat Commun ; 12(1): 120, 2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33402698

RESUMO

Mitochondrial ATP synthase plays a key role in inducing membrane curvature to establish cristae. In Apicomplexa causing diseases such as malaria and toxoplasmosis, an unusual cristae morphology has been observed, but its structural basis is unknown. Here, we report that the apicomplexan ATP synthase assembles into cyclic hexamers, essential to shape their distinct cristae. Cryo-EM was used to determine the structure of the hexamer, which is held together by interactions between parasite-specific subunits in the lumenal region. Overall, we identified 17 apicomplexan-specific subunits, and a minimal and nuclear-encoded subunit-a. The hexamer consists of three dimers with an extensive dimer interface that includes bound cardiolipins and the inhibitor IF1. Cryo-ET and subtomogram averaging revealed that hexamers arrange into ~20-megadalton pentagonal pyramids in the curved apical membrane regions. Knockout of the linker protein ATPTG11 resulted in the loss of pentagonal pyramids with concomitant aberrantly shaped cristae. Together, this demonstrates that the unique macromolecular arrangement is critical for the maintenance of cristae morphology in Apicomplexa.


Assuntos
Mitocôndrias/ultraestrutura , Membranas Mitocondriais/ultraestrutura , ATPases Mitocondriais Próton-Translocadoras/química , Subunidades Proteicas/química , Proteínas de Protozoários/química , Toxoplasma/ultraestrutura , Sítios de Ligação , Cardiolipinas/química , Cardiolipinas/metabolismo , Microscopia Crioeletrônica , Expressão Gênica , Mitocôndrias/genética , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , ATPases Mitocondriais Próton-Translocadoras/genética , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Modelos Moleculares , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Proteínas/química , Proteínas/genética , Proteínas/metabolismo , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Especificidade por Substrato , Termodinâmica , Toxoplasma/genética , Toxoplasma/metabolismo
4.
Nat Commun ; 12(1): 116, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33414462

RESUMO

Apicomplexan parasites have evolved efficient and distinctive strategies for intracellular replication where the timing of emergence of the daughter cells (budding) is a decisive element. However, the molecular mechanisms that provide the proper timing of parasite budding remain unknown. Using Toxoplasma gondii as a model Apicomplexan, we identified a master regulator that controls the timing of the budding process. We show that an ApiAP2 transcription factor, TgAP2IX-5, controls cell cycle events downstream of centrosome duplication. TgAP2IX-5 binds to the promoter of hundreds of genes and controls the activation of the budding-specific cell cycle expression program. TgAP2IX-5 regulates the expression of specific transcription factors that are necessary for the completion of the budding cycle. Moreover, TgAP2IX-5 acts as a limiting factor that ensures that asexual proliferation continues by promoting the inhibition of the differentiation pathway. Therefore, TgAP2IX-5 is a master regulator that controls both cell cycle and developmental pathways.


Assuntos
Ciclo Celular/fisiologia , Divisão Celular/fisiologia , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Toxoplasma/genética , Toxoplasma/fisiologia , Proliferação de Células , Centrossomo , Replicação do DNA , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Organismos Geneticamente Modificados , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
5.
J Parasitol ; 106(6): 772-788, 2020 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-33326588

RESUMO

Toxoplasma gondii infections are common in humans and animals worldwide. The ingestion of food or water contaminated with oocysts excreted by infected cats or ingesting uncooked or undercooked meat containing tissue cysts of T. gondii are the 2 major modes of transmission of T. gondii. Viable T. gondii is more prevalent in pork and lamb than in beef. In the past decade, there have been many articles on the high seroprevalence in cattle, particularly from China. There is a report of an outbreak of acute toxoplasmosis in humans suspected to be linked to the ingestion of Artisan fresh cheese from cow's milk. There are conflicting reports concerning the rate of congenital transmission of T. gondii in cattle, especially from Brazil. In a report from Brazil, viable T. gondii was isolated from the blood of 1 of 60 pregnant cows slaughtered at an abattoir and from 1 fetus. The role of beef in the epidemiology of T. gondii infections is still not clear. Here, we review prevalence, persistence of infection, clinical disease, epidemiology, and public health risks of T. gondii infections in cattle from beef and cow's milk worldwide for the past decade.


Assuntos
Doenças dos Bovinos/parasitologia , Saúde Pública , Toxoplasmose Animal/parasitologia , Animais , Anticorpos Antiprotozoários/sangue , Brasil/epidemiologia , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/transmissão , Variação Genética , Genótipo , Saúde Global , Humanos , Carne/parasitologia , Leite/parasitologia , Estudos Soroepidemiológicos , Toxoplasma/classificação , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasmose Animal/diagnóstico , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/transmissão , Toxoplasmose Congênita/transmissão
6.
PLoS Pathog ; 16(12): e1009067, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33383579

RESUMO

Inorganic ions such as phosphate, are essential nutrients required for a broad spectrum of cellular functions and regulation. During infection, pathogens must obtain inorganic phosphate (Pi) from the host. Despite the essentiality of phosphate for all forms of life, how the intracellular parasite Toxoplasma gondii acquires Pi from the host cell is still unknown. In this study, we demonstrated that Toxoplasma actively internalizes exogenous Pi by exploiting a gradient of Na+ ions to drive Pi uptake across the plasma membrane. The Na+-dependent phosphate transport mechanism is electrogenic and functionally coupled to a cipargarmin sensitive Na+-H+-ATPase. Toxoplasma expresses one transmembrane Pi transporter harboring PHO4 binding domains that typify the PiT Family. This transporter named TgPiT, localizes to the plasma membrane, the inward buds of the endosomal organelles termed VAC, and many cytoplasmic vesicles. Upon Pi limitation in the medium, TgPiT is more abundant at the plasma membrane. We genetically ablated the PiT gene, and ΔTgPiT parasites are impaired in importing Pi and synthesizing polyphosphates. Interestingly, ΔTgPiT parasites accumulate 4-times more acidocalcisomes, storage organelles for phosphate molecules, as compared to parental parasites. In addition, these mutants have a reduced cell volume, enlarged VAC organelles, defects in calcium storage and a slightly alkaline pH. Overall, these mutants exhibit severe growth defects and have reduced acute virulence in mice. In survival mode, ΔTgPiT parasites upregulate several genes, including those encoding enzymes that cleave or transfer phosphate groups from phosphometabolites, transporters and ions exchangers localized to VAC or acidocalcisomes. Taken together, these findings point to a critical role of TgPiT for Pi supply for Toxoplasma and also for protection against osmotic stresses.


Assuntos
Osmorregulação/genética , Fosfatos/metabolismo , Proteínas Cotransportadoras de Sódio-Fosfato/fisiologia , Toxoplasma , Animais , Animais Geneticamente Modificados , Transporte Biológico/genética , Células Cultivadas , Humanos , Camundongos , Proteínas Cotransportadoras de Sódio-Fosfato/genética , Toxoplasma/genética , Toxoplasma/metabolismo
7.
Nat Commun ; 11(1): 5258, 2020 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-33067458

RESUMO

Macrophages play an essential role in the early immune response against Toxoplasma and are the cell type preferentially infected by the parasite in vivo. Interferon gamma (IFNγ) elicits a variety of anti-Toxoplasma activities in macrophages. Using a genome-wide CRISPR screen we identify 353 Toxoplasma genes that determine parasite fitness in naїve or IFNγ-activated murine macrophages, seven of which are further confirmed. We show that one of these genes encodes dense granule protein GRA45, which has a chaperone-like domain, is critical for correct localization of GRAs into the PVM and secretion of GRA effectors into the host cytoplasm. Parasites lacking GRA45 are more susceptible to IFNγ-mediated growth inhibition and have reduced virulence in mice. Together, we identify and characterize an important chaperone-like GRA in Toxoplasma and provide a resource for the community to further explore the function of Toxoplasma genes that determine fitness in IFNγ-activated macrophages.


Assuntos
Interferon gama/imunologia , Macrófagos/imunologia , Toxoplasma/genética , Toxoplasmose/imunologia , Animais , Feminino , Genoma de Protozoário , Interações Hospedeiro-Parasita , Humanos , Interferon gama/genética , Macrófagos/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Toxoplasma/crescimento & desenvolvimento , Toxoplasma/metabolismo , Toxoplasma/patogenicidade , Toxoplasmose/genética , Toxoplasmose/parasitologia , Virulência
8.
Nat Commun ; 11(1): 4813, 2020 09 23.
Artigo em Inglês | MEDLINE | ID: mdl-32968076

RESUMO

Artemisinins have revolutionized the treatment of Plasmodium falciparum malaria; however, resistance threatens to undermine global control efforts. To broadly explore artemisinin susceptibility in apicomplexan parasites, we employ genome-scale CRISPR screens recently developed for Toxoplasma gondii to discover sensitizing and desensitizing mutations. Using a sublethal concentration of dihydroartemisinin (DHA), we uncover the putative transporter Tmem14c whose disruption increases DHA susceptibility. Screens performed under high doses of DHA provide evidence that mitochondrial metabolism can modulate resistance. We show that disrupting a top candidate from the screens, the mitochondrial protease DegP2, lowers porphyrin levels and decreases DHA susceptibility, without significantly altering parasite fitness in culture. Deleting the homologous gene in P. falciparum, PfDegP, similarly lowers heme levels and DHA susceptibility. These results expose the vulnerability of heme metabolism to genetic perturbations that can lead to increased survival in the presence of DHA.


Assuntos
Antimaláricos/farmacologia , Artemisininas/farmacologia , Resistência a Medicamentos/genética , Testes Genéticos/métodos , Heme/genética , Heme/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Técnicas de Inativação de Genes , Humanos , Malária Falciparum/tratamento farmacológico , Proteínas de Membrana Transportadoras/metabolismo , Mutação , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/genética , Proteínas de Protozoários/genética , Toxoplasma/efeitos dos fármacos , Toxoplasma/genética
9.
Exp Parasitol ; 218: 108006, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32991867

RESUMO

This study aimed to elucidate the cellular immune response against Toxoplasma gondii in chronically infected mice reinfected with different strains of the parasite to elucidate the immunological basis for chronicity or virulence and to uncover the involvement of genes that encode virulence proteins and modulate the immune response. BALB/c mice were infected by oral gavage with non-virulent D8 strain and challenged 45 days post-infection with virulent EGS or CH3 strains. Cytokine measurement was performed 2 days post-challenge in cell extracts of the small intestine and 2, 7, and 14 days post-challenge in serum. Virulence gene allele type of these strains was analyzed. Challenged mice survived by avoiding exacerbated inflammation and inhibiting the overproduction of cytokines. Local and systemic cytokine response in challenged mice was similar to chronic controls and quite distinct in mice acutely infected with the EGS or CH3 strains. Allelic combinations of the virulence genes ROP5/ROP18 was predictive of virulence in mice when tested in these T. gondii strains. Other allelic combinations of rhoptries and dense granules genes showed discrepancies.


Assuntos
Citocinas/biossíntese , Toxoplasma/imunologia , Toxoplasmose Animal/imunologia , Alelos , Animais , Doença Crônica , Citocinas/sangue , Citocinas/genética , Cães , Feminino , Íleo/imunologia , Imunidade Celular , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Proteínas de Protozoários/genética , Toxoplasma/classificação , Toxoplasma/genética , Toxoplasma/patogenicidade , Toxoplasmose Animal/parasitologia , Virulência
10.
Parasitol Res ; 119(11): 3771-3776, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32914221

RESUMO

Toxoplasma gondii causes serious clinical toxoplasmosis in humans mostly due to its asexual life cycles, which can be artificially divided into five tightly coterminous stages. Any radical or delay for the stage will result in tremendous changes immediately behind. We previously demonstrated that TgERK7 is associated with the intracellular proliferation of T. gondii, but during the process, other stages before were not meanwhile determined. To further clarify the function of ERK7 gene in T. gondii, the complemental strain of ΔTgERK7 tachyzoites created previously was engineered via electric transfection with the recombinant pUC/Tgerk7 plasmid, named pUC/TgERK7 strain in this study, and was used together with ΔTgERK7 and wild-type GT1 strains to retrospect the phenotypic changes including invasion and attachment. The results showed that TgERK7 protein can be re-expressed in the ΔTgERK7 tachyzoites and eradication of this protein leads to significantly lower invasion of T. gondii at 1 h and 2 h post-infection (P < 0.05), which is the key factor causing the following slow intracellular proliferation, in comparison with wild-type GT1 and pUC/TgERK7 parasites; noteworthily, at other early time points including 15 min for attachment assay was no statistical difference (P > 0.05). The data suggested that ERK7 protein in T. gondii is an important virulence factor that participates in the invasion of this parasite.


Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas de Protozoários/metabolismo , Toxoplasma/patogenicidade , Animais , MAP Quinases Reguladas por Sinal Extracelular/genética , Teste de Complementação Genética , Humanos , Estágios do Ciclo de Vida/genética , Mutação , Proteínas de Protozoários/genética , Toxoplasma/genética , Toxoplasma/crescimento & desenvolvimento , Toxoplasmose/parasitologia , Virulência/genética
11.
Artigo em Inglês | MEDLINE | ID: mdl-32667393

RESUMO

Infection by the protozoan Toxoplasma gondii during pregnancy demands greater attention from the health authorities due to the risk of placental transmission, which can have devastating consequences to the foetus and newborn. This study was conducted in a high-risk prenatal care outpatient clinic of a university teaching hospital. Pregnant women screened for specific IgM and IgG anti -T. gondii, attended from January 2009 to August 2018 were included. From 530 suspected patients, 218 were followed up and they presented positive IgM and IgG anti- T. gondii. From these patients, 83 (38.0%) had low IgG avidity, 39 (18%) seroconverted in the second or third trimester of pregnancy, 19 (8.7%) had no avidity test, 69 (31.6%) had high IgG avidity after 16 weeks of gestation, five had recurrent chorioretinitis (2.2%) and three (1.3%) were seropositive to HIV. Complementary diagnoses were made in 30/48 (62.5%) of the patients revealing the presence of specific IgA antibodies raised to T. gondii; 3/63 (4.8%) peripheral blood samples and 1/57 (1.8%) amniotic fluid sample. There were eight foetal deaths, one case of neonatal hepatomegaly and one case of T. gondii DNA detected in a peripheral blood sample. Of the 139 newborn deliveries at the teaching hospital, there was a 38% loss of follow-up. The prevalence of congenital toxoplasmosis was 1.2 cases/1,000 live births in this study area, according to the retrospective survey of cases. Prenatal treatment may have helped to reduce the risk of vertical transmission.


Assuntos
Complicações Parasitárias na Gravidez/diagnóstico , Toxoplasma/isolamento & purificação , Toxoplasmose/diagnóstico , Instituições de Assistência Ambulatorial , Anticorpos Antiprotozoários/sangue , Brasil , Feminino , Hospitais de Ensino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Recém-Nascido , Gravidez , Complicações Parasitárias na Gravidez/tratamento farmacológico , Cuidado Pré-Natal , Estudos Prospectivos , Toxoplasma/genética , Toxoplasmose/sangue , Toxoplasmose/tratamento farmacológico
12.
Korean J Parasitol ; 58(3): 257-265, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32615739

RESUMO

The outbreak of human toxoplasmosis can be attributed to ingestion of food contaminated with Toxoplasma gondii. Toxoplasmosis recently increased in domestic and stray dogs and cats. It prompted studies on the zoonotic infectious diseases transmitted via these animals. Sero- and antigen prevalences of T. gondii in dogs and cats were surveyed using ELISA and PCR, and B1 gene phylogeny was analyzed in this study. Toxoplasmosis antibodies were measured on sera of 403 stray cats, 947 stray dogs, 909 domestic cats, and 2,412 domestic dogs collected at nationwide regions, Korea from 2017 to 2019. In addition, whole blood, feces, and tissue samples were also collected from stray cats (1,392), stray dogs (686), domestic cats (3,040), and domestic dogs (1,974), and T. gondii-specific B1 gene PCR was performed. Antibody prevalence of stray cats, stray dogs, domestic cats, and domestic dogs were 14.1%, 5.6%, 2.3%, and 0.04%, respectively. Antigen prevalence of these animals was 0.5%, 0.2%, 0.1%, and 0.4%, respectively. Stray cats revealed the highest infection rate of toxoplasmosis, followed by stray dogs, domestic cats, and domestic dogs. B1 gene positives were 5 of stray cats, and identified to high/moderate pathogenic Type I/III group. These findings enforce that preventive hygienic measure should be strengthened at One Health level in dogs and cats, domestic and stray, to minimize human toxoplasmosis infections.


Assuntos
Doenças do Gato/epidemiologia , Doenças do Gato/parasitologia , Gatos/parasitologia , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Cães/parasitologia , Genes de Protozoários/genética , Toxoplasma/genética , Toxoplasmose Animal/parasitologia , Animais , Filogenia , Reação em Cadeia da Polimerase , República da Coreia/epidemiologia , Estudos Soroepidemiológicos , Toxoplasmose/parasitologia , Toxoplasmose/prevenção & controle
13.
Korean J Parasitol ; 58(3): 327-331, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32615747

RESUMO

Toxoplasma gondii are intracellular protozoa that can cause neurological disease or death in fetuses and even in immunocompromised human adults. Ticks are recognized as vectors of many microorganisms including viruses, bacteria, and protozoa. Recent studies detected T. gondii in various tick species in many countries. In this study, we performed PCR detection of the T. gondii B1 gene from Haemaphysalis ticks collected from vegetation in 4 localities, Wonju, Gunsan, Miryang, and Yangsan, in Korea. We analyzed DNA from 314 ticks (268 Haemaphysalis longicornis and 46 Haemaphysalis flava) and the B1 gene of T. gondii was detected in 13 of these. The detection of T. gondii in ticks differed significantly by region (P=0.021). T. gondii was detected in the following percentages of collected ticks: 3.7% (7 of 189) in Gunsan, 10% (5 of 50) in Wonju, 16.7% (1 of 6) in Yangsan, and 0% (0 of 69) in Miryang. The detection of T. gondii in ticks was not associated with tick species or development stage. This is the first report of T. gondii detection in ticks in Korea. Our results provide important information necessary to understand toxoplasmosis transmission.


Assuntos
Carrapatos/parasitologia , Toxoplasma/genética , Toxoplasma/isolamento & purificação , Animais , Vetores Aracnídeos , Reação em Cadeia da Polimerase , República da Coreia , Toxoplasmose/transmissão
14.
Arch Microbiol ; 202(10): 2689-2695, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32725599

RESUMO

Depression disorder is one of the most common psychological recognitions that characterized by sadness, low self-confidence, and disinterest in every activity. Considering evidence showing the effects of toxoplasmosis on the psychological disease, this study conducted to investigate the serological and molecular aspects of Toxoplasma gondii infection among patients with depression. In this study, after selecting the patients with depression and control groups under the supervision of a psychologist, the blood samples were collected and the serum samples and buffy coat were separated. The specific anti-Toxoplasma IgG antibodies in serum samples were evaluated using the commercial ELISA kit. Then the desired region of the Toxoplasma B1 gene was amplified using the specific primers. To confirm the specificity of primers to amplify the B1 gene of Toxoplasma, the extracted PCR product was sequenced. The overall prevalence of toxoplasmosis in patients with depression was 59.8 and 60.19% by ELISA and PCR, respectively. In the control group, the prevalence of Toxoplasma was 56.3 and 40.2% by serology and PCR. There was a significant correlation between the prevalence of toxoplasmosis and depression. Moreover, a significant difference was found between the variables of age, sex, kind of nutrition, level of education and toxoplasmosis among the two cases and control groups. The higher prevalence of Toxoplasma infection among patients with depression compared with the control group indicates the probable impact of this parasite on depression and exacerbates its symptoms, which requires special attention of specialist physicians and patient's relatives.


Assuntos
Anticorpos Antiprotozoários/sangue , Depressão/complicações , Depressão/parasitologia , Toxoplasma , Toxoplasmose/complicações , Toxoplasmose/epidemiologia , Anticorpos Antiprotozoários/genética , Estudos de Casos e Controles , Ensaio de Imunoadsorção Enzimática , Feminino , Genes de Protozoários/genética , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Reação em Cadeia da Polimerase , Prevalência , Fatores de Risco , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasmose/imunologia
15.
PLoS Pathog ; 16(7): e1008650, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32628723

RESUMO

Toxoplasma gondii is an obligate intracellular parasite that can invade any nucleated cell of any warm-blooded animal. In a previous screen to identify virulence determinants, disruption of gene TgME49_305140 generated a T. gondii mutant that could not establish a chronic infection in mice. The protein product of TgME49_305140, here named TgPL3, is a 277 kDa protein with a patatin-like phospholipase (PLP) domain and a microtubule binding domain. Antibodies generated against TgPL3 show that it is localized to the apical cap. Using a rapid selection FACS-based CRISPR/Cas-9 method, a TgPL3 deletion strain (ΔTgPL3) was generated. ΔTgPL3 parasites have defects in host cell invasion, which may be caused by reduced rhoptry secretion. We generated complementation clones with either wild type TgPL3 or an active site mutation in the PLP domain by converting the catalytic serine to an alanine, ΔTgPL3::TgPL3S1409A (S1409A). Complementation of ΔTgPL3 with wild type TgPL3 restored all phenotypes, while S1409A did not, suggesting that phospholipase activity is necessary for these phenotypes. ΔTgPL3 and S1409A parasites are also virtually avirulent in vivo but induce a robust antibody response. Vaccination with ΔTgPL3 and S1409A parasites protected mice against subsequent challenge with a lethal dose of Type I T. gondii parasites, making ΔTgPL3 a compelling vaccine candidate. These results demonstrate that TgPL3 has a role in rhoptry secretion, host cell invasion and survival of T. gondii during acute mouse infection.


Assuntos
Proteínas de Protozoários/metabolismo , Toxoplasma/patogenicidade , Toxoplasmose/metabolismo , Fatores de Virulência/metabolismo , Animais , Camundongos , Fosfolipases/genética , Fosfolipases/metabolismo , Proteínas de Protozoários/genética , Toxoplasma/genética , Toxoplasmose/enzimologia , Virulência
16.
Parasitol Res ; 119(9): 2907-2916, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32686022

RESUMO

The protozoan parasite Toxoplasma gondii secretes a number of dense granule proteins (GRAs) from the dense granule organelle to manipulate the host cell. Two of these effector proteins (GRA17 and GRA23) are involved in the trafficking of molecules between the parasitophorous vacuole (PV) and the host cell cytoplasm. However, their roles in establishing chronic infection remain obscured. In this study, CRISPR-Cas9 was used to delete gra17 or gra23 gene in T. gondii Pru strain (type II). The growth, the virulence, the ability to establish chronic infection, and the immunogenicity of the constructed mutant strains were investigated in Kunming mice. Pru:Δgra17 and Pru:Δgra23 mutants developed PVs with abnormal morphology and exhibited reduced growth rate, compared with the wild-type Pru strain. Deletion of gra17 abrogated acute infection and blocked cyst formation. Although the deletion of gra23 caused slight attenuation of the parasite virulence in mice, it caused a significant reduction in cyst formation. Immunization with Pru:Δgra17 induced high levels of IgG (IgG1 and IgG2a) antibodies and cytokines (interleukin-2 [IL-2], IL-10, IL-12, and interferon gamma [IFN-γ]), which conferred significant protection in mice challenged with virulent type I (RH), ToxoDB#9 (PYS) strains, or less virulent type II (Pru) strain of T. gondii. These findings show that GRA17 and GRA23 play important roles in T. gondii chronic infection and that irreversible deletion of gra17 in T. gondii type II Pru strain can be a viable option for stimulating protective immunity to T. gondii infection.


Assuntos
Antígenos de Protozoários/imunologia , Citocinas/metabolismo , Proteínas de Protozoários/genética , Toxoplasma , Fatores de Virulência/genética , Animais , Anticorpos Antiprotozoários/sangue , Anticorpos Antiprotozoários/imunologia , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Feminino , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Camundongos , Toxoplasma/genética , Toxoplasma/crescimento & desenvolvimento , Toxoplasma/imunologia , Toxoplasma/patogenicidade , Toxoplasmose Animal/parasitologia , Virulência/genética
17.
Adv Clin Exp Med ; 29(6): 669-675, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32573993

RESUMO

BACKGROUND: Toxoplasma gondii (T. gondii) is a ubiquitous protozoan parasite which causes a serious disease called toxoplasmosis. The high prevalence of T. gondii infection has attracted a great deal of interest in its diagnosis and treatment. The use of pure antigens shows high sensitivity and specificity, but challenges such as cross-reactivity remain diagnostic difficulties. OBJECTIVES: The aim of this study was to use 3 surface antigens (SAGs) of T. gondii to design gene-encoding a multi-epitope and immunogenic protein as a serodiagnostic marker. MATERIAL AND METHODS: The multi-epitope antigen was expressed using Escherichia coli BL21 (DE3) cells and purified using affinity chromatography. To evaluate acute toxoplasmosis, 95 human sera with anti-T. gondii IgG, 25 human sera without anti-T. gondii IgG and 6 serum samples with nosocomial infections were collected and submitted to an enzyme-linked immunosorbent assay (ELISA) analysis. The potential of purified protein as a diagnostic marker of T. gondii infection was also investigated using ELISA analysis. RESULTS: The western blot analysis for both protein expression and purification confirmed that the protein was expressed and purified successfully. The results of validation showed a sensitivity of 72.6% and a specificity of 90.3% for recombinant ELISA. CONCLUSIONS: Although this protein showed potential for detecting T. gondii, the sensitivity and specificity were lower than in tests that use the whole body of the parasite.


Assuntos
Antígenos de Protozoários , Toxoplasma , Toxoplasmose , Antígenos de Protozoários/análise , Ensaio de Imunoadsorção Enzimática , Epitopos , Humanos , Proteínas Recombinantes , Sensibilidade e Especificidade , Testes Sorológicos , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasmose/diagnóstico , Toxoplasmose/genética
18.
J Parasitol ; 106(3): 395-399, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32556163

RESUMO

The objective of the present study was to determine the characterization of Toxoplasma gondii in cats, rats, and chickens in the border areas of Yunnan Province. A total of 259 samples was collected from 10 border areas in Yunnan Province including 94 cats, 58 rats, and 107 chickens. Samples were screened by a nested polymerase chain reaction (PCR) assay and the positive products were analyzed by multilocus PCR-restriction fragment length polymorphism (RFLP) to determine the genotypes. Toxoplasma gondii deoxyribonucleic acid (DNA) was detected from 15.96% of 94 cats, 15.52% of 58 rats, and 6.54% of 107 chickens, respectively, and the average infection rate is 11.97%. Using the multilocus PCR-RFLP, we found that the genotype of T. gondii in cats and rats was ToxoDB#9. Because of low DNA concentration, no genotype was determined from chickens. These results fill the gaps of knowledge in the prevalence and genotype of T. gondii in the border areas of Yunnan Province and have implications for the better control of T. gondii infection in humans and animals.


Assuntos
Doenças do Gato/epidemiologia , Galinhas/parasitologia , Doenças das Aves Domésticas/epidemiologia , Ratos/parasitologia , Doenças dos Roedores/epidemiologia , Toxoplasmose Animal/epidemiologia , Animais , Encéfalo/parasitologia , Doenças do Gato/parasitologia , Gatos , China/epidemiologia , DNA de Protozoário/química , DNA de Protozoário/isolamento & purificação , Marcadores Genéticos , Técnicas de Genotipagem/veterinária , Tipagem de Sequências Multilocus/veterinária , Reação em Cadeia da Polimerase/veterinária , Polimorfismo de Fragmento de Restrição , Doenças das Aves Domésticas/parasitologia , Prevalência , Doenças dos Roedores/parasitologia , Toxoplasma/classificação , Toxoplasma/genética , Toxoplasmose Animal/parasitologia
19.
Parasitol Res ; 119(8): 2727-2731, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32518965

RESUMO

We report a case of severe congenital toxoplasmosis that involved an atypical T. gondii genotype in a newborn baby from Alagoas state in Northeastern Brazil. A pregnant woman presented IgM and IgG anti-T. gondii antibodies, as detected by the chemiluminescence immunoassay on the second trimester of pregnancy. A mouse bioassay was performed using umbilical cord blood and one isolate was obtained. The isolate was designated TgCTBrAL1 and genetic characterization revealed genotype ToxoDB #162. Genotype results of the rhoptry genes, ROP5 and ROP18, could predict the high virulence of the isolate in mice, which was confirmed by an in vivo virulence assay. This is the first report of generating a T. gondii isolate from a newborn baby with congenital toxoplasmosis in Northeastern Brazil.


Assuntos
Toxoplasma/isolamento & purificação , Toxoplasmose Congênita/parasitologia , Animais , Brasil , Genótipo , Humanos , Recém-Nascido , Masculino , Camundongos , Proteínas de Protozoários/genética , Toxoplasma/genética , Toxoplasma/patogenicidade , Virulência/genética
20.
Parasitol Res ; 119(7): 2299-2307, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32476060

RESUMO

In the intermediate hosts, tachyzoites of T. gondii predominate in the acute stage while bradyzoites persist inside tissue cysts with the potential for reactivation. The two stages exhibit different metabolic and antigenic characters. The present study aimed to investigate temporal expression of Toxoplasma SAG1 and BAG1 genes in the brain tissue and the coincident parasitological and histopathological findings in mice models of toxoplasmosis. The study included group A: mice infected with RH strain and sacrificed 7 days post-infection (p.i.); group B: mice infected with RH strain and treated with sulfamethoxazole-trimethoprim (30 mg/kg/day and 150 mg/kg/day respectively) 24 h p.i. until sacrificed at days 5, 10, or 20 post-treatment; group C: mice infected with ME-49 strain and sacrificed at days 7, 27, 47, or 67 p.i; and group D: mice infected with ME-49 strain and received dexamethasone daily starting at day 68 p.i. and scarified at days 6 or 10 post-treatment. All mice were inspected daily for abnormal physical signs. Peritoneal exudate and brain homogenate were examined for detection of Toxoplasma stages. Brain sections were examined histopathologically. SAG1 and BAG1 gene expression was evaluated using reverse transcription real-time polymerase chain reaction and the ΔΔCt method. Results revealed that marked BAG1 upregulation is consistent with detection of Toxoplasma cysts and degenerative changes while predominance of tachyzoites and inflammatory infiltrate is compatible with SAG1 upregulation. The study sheds light on the potential for using stage-specific gene expression pattern as markers for evaluation of toxoplasmosis disease progression in clinical settings.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Estágios do Ciclo de Vida/genética , Toxoplasma/genética , Toxoplasmose Animal/patologia , Toxoplasmose Animal/parasitologia , Animais , Encéfalo/parasitologia , Encéfalo/patologia , Feminino , Genes de Protozoários/genética , Camundongos , Encistamento de Parasitas/genética , Toxoplasma/crescimento & desenvolvimento
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