SUMMARYFoodborne diseases represent operational risks in industrial restaurants. We described an outbreak of nine clustered cases of acute illness resembling acute toxoplasmosis in an industrial plant with 2300 employees. These patients and another 36 similar asymptomatic employees were diagnosed with anti-T. gondii IgG titer and avidity by ELISA. We excluded 14 patients based on high IgG avidity and chronic toxoplasmosis: 13 from controls and one from acute disease other than T. gondii infection. We also identified another three asymptomatic employees with T.gondii acute infection and also anti-T. gondii IgM positive as remaining acute cases. Case control study was conducted by interview in 11 acute infections and 20 negative controls. The ingestion of green vegetables, but not meat or water, was observed to be associated with the incidence of acute disease. These data reinforce the importance of sanitation control in industrial restaurants and also demonstrate the need for improvement in quality control regarding vegetables at risk for T. gondii oocyst contamination. We emphasized the accurate diagnosis of indexed cases and the detection of asymptomatic infections to determine the extent of the toxoplasmosis outbreak.

Introduction: Visceral leishmaniasis is endemic in 88 countries, with a total of 12 million people infected and 350 million at risk. In the search for new leishmanicidal agents, alkaloids and acetogenins isolated from leaves of Annona squamosa and seeds of Annona muricata were tested against promastigote and amastigote forms of Leishmania chagasi. Methods: extracts of A. squamosa leaves and A. muricata seeds were extracted with 10% phosphoric acid and organic solvents to obtain the alkaloid and acetogenin-rich extracts. These extracts were chromatographed on a silica gel column and eluted with a mixture of several solvents in crescent order of polarity. The compounds were identified by spectroscopic analysis. The isolated compounds were tested against Leishmania chagasi, which is responsible for American visceral leishmaniasis, using the MTT test assay. The cytotoxicity assay was evaluated for all isolated compounds, and for this assay, RAW 264.7 cells were used. Results: O-methylarmepavine, a benzylisoquinolinic alkaloid, and a C 37 trihydroxy adjacent bistetrahydrofuran acetogenin were isolated from A. squamosa, while two acetogenins, annonacinone and corossolone, were isolated from A. muricata. Against promastigotes, the alkaloid showed an IC 50 of 23.3 µg/mL, and the acetogenins showed an IC 50 ranging from 25.9 to 37.6 µg/mL; in the amastigote assay, the IC 50 values ranged from 13.5 to 28.7 µg/mL. The cytotoxicity assay showed results ranging from 43.5 to 79.9 µg/mL. Conclusions: These results characterize A. squamosa and A. muricata as potential sources of leishmanicidal agents. Plants from Annonaceae are rich sources of natural compounds and an important tool in the search for new leishmanicidal therapies.

Introduction: Toxoplasmosis is usually a benign infection, except in the event of ocular, central nervous system (CNS), or congenital disease and particularly when the patient is immunocompromised. Treatment consists of drugs that frequently cause adverse effects; thus, newer, more effective drugs are needed. In this study, the possible activity of artesunate, a drug successfully being used for the treatment of malaria, on Toxoplasma gondii growth in cell culture is evaluated and compared with the action of drugs that are already being used against this parasite. Methods: LLC-MK2 cells were cultivated in RPMI medium, kept in disposable plastic bottles, and incubated at 36ºC with 5% CO 2 . Tachyzoites of the RH strain were used. The following drugs were tested: artesunate, cotrimoxazole, pentamidine, pyrimethamine, quinine, and trimethoprim. The effects of these drugs on tachyzoites and LLC-MK2 cells were analyzed using nonlinear regression analysis with Prism 3.0 software. Results: Artesunate showed a mean tachyzoite inhibitory concentration (IC 50 ) of 0.075µM and an LLC MK2 toxicity of 2.003µM. Pyrimethamine was effective at an IC 50 of 0.482µM and a toxicity of 11.178µM. Trimethoprim alone was effective against the in vitro parasite. Cotrimoxazole also was effective against the parasite but at higher concentrations than those observed for artesunate and pyrimethamine. Pentamidine and quinine had no inhibitory effect over tachyzoites. Conclusions: Artesunate is proven in vitro to be a useful alternative for the treatment of toxoplasmosis, implying a subsequent in vivo effect and suggesting the mechanism of this drug against the parasite.

Revista da Sociedade Brasileira de Medicina Tropical 41(6):628-634, nov-dez, 2008 ARTIGO/ARTICLEA toxoplasmose, infecção causada pelo parasita Toxoplasma gondii 11 15 , é especialmente importante na infecção congênita e reativação em imunodeprimidos 26 , sendo geralmente assintomática ou associada a manifestações inespecíficas em indivíduos imunocompetentes 30 .A primo-infecção na gravidez pode afetar o feto, com severas complicações, como nascimento prematuro, dano neurológico permanente e comprometimento visual 20 31 . A freqüência e severidade da infecção congênita dependem do período gestacional 12 e do diagnóstico precoce para intervenção terapêutica específica 34 e redução do risco de infecção fetal. ABSTRACTIn order to investigate the incidence of toxoplasmosis in Cascavel, Paraná, a city near the region where the largest reported epidemic outbreak in the world occurred, 334 serum samples from pregnant women were screened using a commercial IgG immunoenzymatic assay at the Municipal Laboratory in Cascavel and were confirmed at the Institute of Tropical Medicine in São Paulo, by means of IgG immunofluorescence, immunoenzymatic assaying and the in-house IgG avidity test. The IgG seropositivity from the commercial test was 54.2%, from immunofluorescence 54.8% and from the in-house IgG 53.9%, with good agreement between immunofluorescence and the commercial IgG test (kappa = 0.963781; co-positivity = 97.8%; co-negativity = 98,7%) and between immunofluorescence and the in-house IgG (kappa = 0.975857; co-positivity = 97.8%; co-negativity = 100%). The evidence of acute infection among the pregnant women was similar, as estimated both by IgG avidity (2.4%/year) and by statistical trend analysis (χ 2 test) according to age group (2%/year). This suggests that prenatal serological screening and epidemiological surveillance are essential for reducing the risk of toxoplasmosis in the region, although without evidence of an epidemic outbreak. Key-words: Toxoplasma gondii. Immunodiagnosis. IgG avidity.Para o diagnóstico da infecção pelo Toxoplasma gondii, os testes sorológicos de imunofluorescência indireta, aglutinação e imunoenzimáticos são os métodos mais freqüentemente utilizados para pesquisa de anticorpos específicos como IgM e/ou IgA e/ou aumento significante do nível de anticorpos IgG 2 41 .Contudo, o diagnóstico sorológico da toxoplasmose é muito complexo e tem sido discutido extensivamente na literatura, especialmente em gestantes 31 . Os resultados devem ser interpretados com cautela 46 , porque tanto a IgA 18 quanto a IgM podem persistir por anos, levando a erro na determinação da época da infecção materna 24 e falha no diagnóstico da infecção aguda na gestante 22 .

SUMMARYWe detected Toxoplasma gondii oocysts in feces of experimentally infected cats, using a Kato Katz approach with subsequent Kinyoun staining. Animals serologically negative to T. gondii were infected orally with 5x10 2 mice brain cysts of ME49 strain. Feces were collected daily from the 3 rd to the 30 th day after challenge. Oocysts were detected by qualitative sugar flotation and the quantitative modified Kato Katz stained by Kinyoun (KKK). In the experimentally infected cats, oocysts were detected from the 7 th to 15 th day through sugar flotation technique, but oocysts were found in KKK from the 6 th to 16 th day, being sensitive for a larger period, with permanent documentation. The peak of oocysts excretion occurred between the 8 th to 11 th days after challenge, before any serological positive result. KKK could be used in the screening and quantification of oocysts excretion in feces of suspected animals, with reduced handling of infective material, decreasing the possibility of environmental and operator contamination.

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SUMMARYZika virus (ZKV) infection is a huge public health problem in Brazil because of the increased incidence of microcephaly in neonates from infected mothers. Detection of specific IgG antibodies in maternal serum samples constitutes an important approach for diagnosing ZKV infection and evaluating its relationship with neonatal microcephaly. However, as there is no serological test produced in Brazil to detect IgM and IgG antibodies against ZKV, we sought to examine specific IgG in serum samples from patients or suspected mothers to detect previous infection and to test for specificity with regard to flaviviral infections occurring in the same area. Brazilian Zika virus native antigens were obtained from infected Vero cell layers or free virions in the culture medium and then used in ELISA. We tested sera from eight ZKV RNA-diagnosed infected patients (ZKVR), seven neonates with microcephaly and their mothers after delivery (MM), 140 dengue virus IgM-positive (DM) and IgG (DG)-positive patients, and 100 yellow fever (YF)-vaccinated patients. According to the ELISA, ZKVR samples were mostly positive (7/8), and all the MM serum samples were positive for ZKV IgG (7/7). In contrast, cross-reactions for dengue or yellow fever-vaccinated patients were observed, including DM (48/95), DG (10/45) or YF (3/100) serum samples; however, these cross-reactions exhibited low antigen avidity so that 6 M urea largely removed this cross-reactivity, with only a few cross-reacting samples remaining (8/140). ELISA based on extracted virions was much more specific, with all ZKVR (8/8) and MM sera being positive for ZKV IgG (7/7) and only borderline cross-reactivity found for DM (6/95), DG (3/45) or YF (4/100)-vaccinated serum samples. This technique (ELISA) can identify specific IgG in ZKV-infected patients and may be helpful in diagnosing congenital infetions after maternal RNA virus clearance or in epidemiological studies.

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