Natural-product-inspired design and synthesis of two series of compounds active against Trypanosoma cruzi: Insights into structure-activity relationship, toxicity, and mechanism of action.

Chemical scaffolds of natural products have historically been sources of inspiration for the development of novel molecules of biological relevance, including hit and lead compounds. To identify new compounds active against Trypanosoma cruzi, we designed and synthesized 46 synthetic derivatives based on the structure of two classes of natural products: tetrahydrofuran lignans (Series 1) and oxazole alkaloids (Series 2). Compounds were screened in vitro using a cellular model of T. cruzi infection. In the first series of compounds, 11 derivatives of hit compound 5 (EC50 = 1.1 µM) were found to be active; the most potent (7, 8, and 13) had EC50 values of 5.1-34.2 µM. In the second series, 17 analogs were found active at 50 µM; the most potent compounds (47, 49, 59, and 63) showed EC50 values of 24.2-49.1 µM. Active compounds were assessed for selectivity, hemocompatibility, synergistic potential, effects on mitochondrial membrane potential, and inhibitory effect on trypanothione reductase. All active compounds showed low toxicity against uninfected THP-1 cells and human erythrocytes. The potency of compounds 5 and 8 increased steadily in combination with benznidazole, indicating a synergistic effect. Furthermore, compounds 8, 47, 49, 59, and 63 inhibited parasitic mitochondria in a dose-dependent manner. Although increased reactive oxygen species levels might lead to mitochondrial effects, the results indicate that the mechanism of action of the compounds is not dependent on trypanothione reductase inhibition. In silico calculation of chemical descriptors and principal component analysis showed that the active compounds share common chemical features with other trypanocidal molecules and are predicted to have a good ADMET profile. Overall, the results suggest that the compounds are important candidates to be further studied for their potential against T. cruzi.

Oseltamivir-resistant influenza A(H1N1)pdm09 virus in southern Brazil.

The neuraminidase (NA) genes of A(H1N1)pdm09 influenza virus isolates from 306 infected patients were analysed. The circulation of oseltamivir-resistant viruses in Brazil has not been reported previously. Clinical samples were collected in the state of Rio Grande do Sul (RS) from 2009-2011 and two NA inhibitor-resistant mutants were identified, one in 2009 (H275Y) and the other in 2011 (S247N). This study revealed a low prevalence of resistant viruses (0.8%) with no spread of the resistant mutants throughout RS.

Intracellular persistence of Leishmania tarentolae in primary canine macrophage cells.

Leishmania tarentolae is a non-pathogenic species first isolated from geckoes in the Mediterranean basin. The finding that dogs test positive against both Leishmania infantum and L. tarentolae raises questions regarding the ability of the latter species to persist and adapt to new hosts. This study aimed to evaluate in vitro the capability of L. tarentolae to colonize, survive and persist in canine primary monocyte-derived mononuclear cells. Monocytes were isolated from dog whole blood samples and placed in 24-well plates for differentiation into macrophages and for incubation with L. tarentolae field-isolated strains (RI-325 and SF-178) and laboratory (LEM-124) strain; the parasite burden was assessed at different time points post-infection. The L. infantum laboratory strain (MON-1) was used as control. Infection parameters were evaluated by microscopy, counting the number of amastigotes/200 infected cells, and by duplex real-time PCR from supernatants and detached cells. Similar to L. infantum, L. tarentolae strains developed into round-shaped amastigote-like forms, with higher infection rates detected at 4 h followed by an overall decrease until 48 h. RI-325 presented also a higher infection rate at 72 h. Data showed that L. tarentolae strains infect and persist inside in vitro primary canine mononuclear cells, opening new perspectives for further laboratory studies.

OCCASIONAL DIGESTIVE HEMORRHAGE IN CHILDREN DUE TO STRONGYLOIDIASIS: IMPORTANCE OF PARASITOLOGIC TESTING. / HEMORRAGIA DIGESTIVA OCASIONAL EM CRIANÇA POR ESTRONGILOIDÍASE: IMPORTÂNCIA DO EXAME PARASITOLÓGICO.

OBJECTIVE: To describe an uncommon case of infection by Strongyloides stercoralis (S. stercoralis) in a 4-month-old child and to highlight the importance of early diagnosis. CASE DESCRIPTION: The patient was a male child from the city of Videira, State of Santa Catarina, Southern Brazil, who was born preterm by Cesarean-section, weighing 1,655 g, and stayed in the neonatal intensive care unit for 20 days. At four months of age, the child started presenting blood in stools and the possibility of cow's milk protein allergy was considered, given the symptoms and the use of infant formula in his 1st semester of life, which was then replaced by infant formula with hydrolyzed protein. White blood cell count and a parasitological stool sample were requested. Both tested positive and the stool ova and parasite examination showed a rhabditoid larva of S. stercoralis. The clinician maintained the initial hypothesis and diet, but requested three new stool samples, which tested positive for rhabditoid larvae of S. stercoralis. Since the child presented abdominal pain and vomiting, and there was still blood in stools, treatment with thiabendazole was initiated twice a day for two days. Treatment was repeated after seven days along with a new parasitological examination, which was then negative. COMMENTS: Although strongyloidiasis is usually a mild parasitic infection, it may be severe and disseminated in immunocompromised patients. This agent must be considered in patients who live in endemic areas, and the diagnosis should be established by searching S. stercoralis larvae in tracheal secretions and in stools.

OBJETIVO: Descrever um caso incomum de infecção por Strongyloides stercoralis (S. stercoralis) em paciente de quatro meses de idade e ressaltar a importância do diagnóstico precoce. DESCRIÇÃO DO CASO: Paciente masculino, procedente e residente de Videira, Santa Catarina, Brasil, nasceu pré-termo, parto cesárea, peso de nascimento 1.655 g, e permaneceu na Unidade de Terapia Intensiva neonatal e intermediária por 20 dias. Aos quatro meses de idade, começou a evacuar fezes sanguinolentas e foi feita hipótese de alergia à proteína do leite de vaca, em razão da sintomatologia e do uso da fórmula infantil para o primeiro semestre, para o qual foi indicada a substituição por fórmula infantil com proteína hidrolisada. Foram solicitados a pesquisa de leucócitos e o exame parasitológico das fezes). Ambos se mostraram positivos e o parasitológico revelou a presença de larva rabditoide de S. stercoralis. O clínico manteve a hipótese inicial e a dieta, mas solicitou a coleta de três amostras de fezes, que resultaram em uma amostra para larvas rabditoide, em muda, de S. stercoralis. Como a criança apresentava dor abdominal, vômito e as fezes permaneciam sanguinolentas, foi iniciado o tratamento com tiabendazol - duas vezes/dia por dois dias -, repetido após sete dias, e, em seguida, realizado o exame parasitológico de fezes, tendo sido negativo. COMENTÁRIOS: A estrongiloidíase, apesar de ser uma infecção parasitária frequentemente leve, em pacientes imunocomprometidos pode se apresentar de forma grave e disseminada. Deve-se suspeitar desse agente em pacientes que vivem em áreas endêmicas, sendo o diagnóstico estabelecido por meio da pesquisa das larvas do S. stercoralis na secreção traqueal e nas fezes.

Comparison of Cervicovaginal Cytopathological Samples Collected in Basic Health Units and in Private Clinics in the Midwest of Santa Catarina.

OBJECTIVE: To compare the quality of cervicovaginal samples obtained from basic health units (BHUs) of the Unified Health System (SUS) and those obtained from private clinics to screen precursor lesions of cervical cancer. METHODS: It was an intervention study whose investigated variables were: adequacy of the samples; presence of epithelia in the samples, and cytopathological results. A total of 940 forms containing the analysis of the biological samples were examined: 470 forms of women attended at BHUs of the SUS and 470 forms of women examined in private clinics in January and February of 2016. RESULTS: All the unsatisfactory samples were collected at BHUs and corresponded to 4% of the total in this sector (p < 0.0001). There was a higher percentage of samples containing only squamous cells in the SUS (43.9%). There was squamocolumnar junction (SJC) representativeness in 82.1% of the samples from the private clinics (p < 0.0001). Regarding negative results for intraepithelial lesions and/or malignancies, the percentages obtained were 95.9% and 99.1% (p < 0.0049) in the exams collected in the private system and SUS, respectively. Less serious lesions corresponded to 0.89% of the samples from the SUS and 2.56% of the tests from the private sector; more serious lesions were not represented in the samples obtained from BHUs, whereas the percentage was 1.49% in private institutions. CONCLUSION: Unsatisfactory cervical samples were observed only in exams performed at the SUS. There is a need for guidance and training of professionals who perform this procedure to achieve higher reliability in the results and more safety for women who undergo this preventive test.

OBJETIVO: Comparar a qualidade das amostras cérvico-vaginais colhidas no Sistema Único de Saúde (SUS) e nas clínicas privadas para rastrear lesões precursoras de câncer do colo uterino. MéTODOS: Estudo de intervenção cujas variáveis estudadas foram: adequabilidade da amostra, representação de epitélios na amostra, e resultado do exame citopatológico. Um total de 940 formulários contendo as análises das amostras biológicas foram examinados: 470 formulários de mulheres atendidas nas unidades básicas de saúde do SUS, e 470 formulários de mulheres atendidas em clínicas privadas no período de janeiro a fevereiro de 2016. RESULTADOS: Todas as amostras insatisfatórias foram coletadas nas unidades básicas de saúde do SUS e corresponderam a 4% do total neste setor (p < 0,0001). Observou-se um índice maior de amostras com representatividade somente de células escamosas no SUS (43,9%). Houve representatividade das células da junção escamo-colunar (JEC) em 82,1% das amostras colhidas no setor privado (p < 0,0001). Em relação aos resultados negativos para lesão intraepitelial e/ou malignidade, os percentuais obtidos foram 95,95% e 99,1% (p < 0,0049) para os exames coletados no sistema privado e no SUS, respectivamente. Em relação às lesões menos graves, no SUS obteve-se um resultado de 0,89% e no sistema privado de 2,56%; as lesões mais graves não foram diagnosticadas no SUS, enquanto que no setor privado representaram 1,49% dos exames. CONCLUSãO: As amostras cérvico-vaginais insatisfatórias foram observadas somente em exames realizados no SUS; há necessidade de orientação e capacitação dos profissionais que realizam a coleta do exame citopatológico, possibilitando uma maior confiabilidade nos resultados e mais segurança à mulher que se submete a este exame preventivo.

Performance of direct immunofluorescence assay for the detection of human metapneumovirus under clinical laboratory settings.

INTRODUCTION: Human metapneumovirus (hMPV) is an emergent human respiratory pathogen. This study aimed to evaluate the performance of direct immunofluorescence (DIF) to detect hMPV in a clinical laboratory setting. METHODS: Nasopharyngeal aspirate samples (448) of children and adults with respiratory illness were used to detect hMPV by using DIF and real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assays. RESULTS: In all, 36 (8%) samples were positive by DIF and 94 (21%) were positive by qRT-PCR. Direct immunofluorescence specificity was 99% and sensitivity was 38%. CONCLUSIONS: DIF is not very sensitive under clinical laboratory settings.

Trypanosoma rangeli Transcriptome Project: Generation and analysis of expressed sequence tags.

Trypanosoma rangeli is an important hemoflagellate parasite of several mammalian species in Central and South America, sharing geographical areas, vectors and reservoirs with T. cruzi, the causative agent of Chagas disease. Thus, the occurrence of single and/or mixed infections, including in humans, must be expected and are of great importance for specific diagnosis and epidemiology. In comparison to several Trypanosomatidae species, the T. rangeli biology and genome are little known, reinforcing the needs of a gene discovery initiative. The T. rangeli transcriptome initiative aims to promote gene discovery through the generation of expressed sequence tags (ESTs) and Orestes (ORF ESTs) from both epimastigote and trypomastigote forms of the parasite, allowing further studies of the parasite biology, taxonomy and phylogeny.

Laboratory surveillance of dengue in Rio Grande do Sul, Brazil, from 2007 to 2013.

BACKGROUND: According to official records, dengue was introduced in Brazil in the 80's; since then several epidemics have occurred. Meanwhile, in Rio Grande do Sul (RS, Southern Brazil) the first autochthonous case occurred only in 2007. METHODOLOGY AND PRINCIPAL FINDINGS: In this study we report laboratory surveillance of dengue cases and seasonality of positive cases, describe serotypes and characterize the epidemiological pattern of dengue in RS from 2007 to 2013. A total of 9,779 serum samples from patients with suspected dengue fever were collected and submitted to molecular and/or serological analyses for dengue virus identification and serotyping, based on viral isolation, NS1 antigen detection and qRT-PCR, or Dengue IgM capture ELISA and MAC-ELISA. The first autochthonous dengue case in RS was confirmed in 2007 (DENV-3). While in 2008 and 2009 only imported cases were registered, autochthonous infection waves have been occurring since 2010. The highest number of dengue infections occurred in 2010, with DENV-1 and DENV-2 outbreaks in Northwestern RS. In 2011, another DENV-1 and DENV-2 outbreak occurred in the Northwestern region; moreover, DENV-4 was detected in travelers. In 2012, DENV-1 and DENV-4 co-circulated. DENV-2 circulation was only detected again in 2013, in high frequency (56.7%), co-circulating with DENV-4 (35%). Most infections occur in adults during summer. Differences in prevalence between genders were observed in 2007 (60% females), 2008 (60.8% males) and 2009 (77.5% males). CONCLUSIONS: According to results of dengue surveillance, there was an increase in the number of dengue cases in RS and of cities infested with Aedes aegypti, possibly as a consequence of introduction of new serotypes and the difficulty of health programs to control the vector.

Hemorragia digestiva ocasional em criança por estrongiloidíase: importância do exame parasitológico / Occasional digestive hemorrhage in children due to strongyloidiasis: importance of parasitologic testing

RESUMO Objetivo: Descrever um caso incomum de infecção por Strongyloides stercoralis (S. stercoralis) em paciente de quatro meses de idade e ressaltar a importância do diagnóstico precoce. Descrição do caso: Paciente masculino, procedente e residente de Videira, Santa Catarina, Brasil, nasceu pré-termo, parto cesárea, peso de nascimento 1.655 g, e permaneceu na Unidade de Terapia Intensiva neonatal e intermediária por 20 dias. Aos quatro meses de idade, começou a evacuar fezes sanguinolentas e foi feita hipótese de alergia à proteína do leite de vaca, em razão da sintomatologia e do uso da fórmula infantil para o primeiro semestre, para o qual foi indicada a substituição por fórmula infantil com proteína hidrolisada. Foram solicitados a pesquisa de leucócitos e o exame parasitológico das fezes). Ambos se mostraram positivos e o parasitológico revelou a presença de larva rabditoide de S. stercoralis. O clínico manteve a hipótese inicial e a dieta, mas solicitou a coleta de três amostras de fezes, que resultaram em uma amostra para larvas rabditoide, em muda, de S. stercoralis. Como a criança apresentava dor abdominal, vômito e as fezes permaneciam sanguinolentas, foi iniciado o tratamento com tiabendazol - duas vezes/dia por dois dias -, repetido após sete dias, e, em seguida, realizado o exame parasitológico de fezes, tendo sido negativo. Comentários: A estrongiloidíase, apesar de ser uma infecção parasitária frequentemente leve, em pacientes imunocomprometidos pode se apresentar de forma grave e disseminada. Deve-se suspeitar desse agente em pacientes que vivem em áreas endêmicas, sendo o diagnóstico estabelecido por meio da pesquisa das larvas do S. stercoralis na secreção traqueal e nas fezes.

ABSTRACT Objective: To describe an uncommon case of infection by Strongyloides stercoralis (S. stercoralis) in a 4-month-old child and to highlight the importance of early diagnosis. Case description: The patient was a male child from the city of Videira, State of Santa Catarina, Southern Brazil, who was born preterm by Cesarean-section, weighing 1,655 g, and stayed in the neonatal intensive care unit for 20 days. At four months of age, the child started presenting blood in stools and the possibility of cow's milk protein allergy was considered, given the symptoms and the use of infant formula in his 1st semester of life, which was then replaced by infant formula with hydrolyzed protein. White blood cell count and a parasitological stool sample were requested. Both tested positive and the stool ova and parasite examination showed a rhabditoid larva of S. stercoralis. The clinician maintained the initial hypothesis and diet, but requested three new stool samples, which tested positive for rhabditoid larvae of S. stercoralis. Since the child presented abdominal pain and vomiting, and there was still blood in stools, treatment with thiabendazole was initiated twice a day for two days. Treatment was repeated after seven days along with a new parasitological examination, which was then negative. Comments: Although strongyloidiasis is usually a mild parasitic infection, it may be severe and disseminated in immunocompromised patients. This agent must be considered in patients who live in endemic areas, and the diagnosis should be established by searching S. stercoralis larvae in tracheal secretions and in stools.

Comparison of cervicovaginal cytopathological samples collected in basic health units and in private clinics in the midwest of Santa Catarina / Comparação de amostras citopatológicas cérvicovaginais coletadas nas unidades básicas de saúde e em clínicas privadas no meio-oeste de Santa Catarina

Abstract Objective To compare the quality of cervicovaginal samples obtained from basic health units (BHUs) of the Unified Health System (SUS) and those obtained fromprivate clinics to screen precursor lesions of cervical cancer. Methods It was an intervention study whose investigated variables were: adequacy of the samples; presence of epithelia in the samples, and cytopathological results. A total of 940 forms containing the analysis of the biological samples were examined: 470 forms of women attended at BHUs of the SUS and 470 forms of women examined in private clinics in January and February of 2016. Results All the unsatisfactory samples were collected at BHUs and corresponded to 4% of the total in this sector (p < 0.0001). There was a higher percentage of samples containing only squamous cells in the SUS (43.9%). There was squamocolumnar junction (SJC) representativeness in 82.1% of the samples from the private clinics (p < 0.0001). Regarding negative results for intraepithelial lesions and/or malignancies, the percentages obtained were 95.9% and 99.1% (p < 0.0049) in the exams collected in the private system and SUS, respectively. Less serious lesions corresponded to 0.89% of the samples from the SUS and 2.56% of the tests from the private sector; more serious lesions were not represented in the samples obtained from BHUs, whereas the percentage was 1.49% in private institutions. Conclusion Unsatisfactory cervical samples were observed only in exams performed at the SUS. There is a need for guidance and training of professionals who perform this procedure to achieve higher reliability in the results and more safety for women who undergo this preventive test.

Resumo Objetivo Comparar a qualidade das amostras cérvico-vaginais colhidas no Sistema Único de Saúde (SUS) e nas clínicas privadas para rastrear lesões precursoras de câncer do colo uterino. Métodos Estudo de intervenção cujas variáveis estudadas foram: adequabilidade da amostra, representação de epitélios na amostra, e resultado do exame citopatológico. Um total de 940 formulários contendo as análises das amostras biológicas foram examinados: 470 formulários de mulheres atendidas nas unidades básicas de saúde do SUS, e 470 formulários de mulheres atendidas em clínicas privadas no período de janeiro a fevereiro de 2016. Resultados Todas as amostras insatisfatórias foram coletadas nas unidades básicas de saúde do SUS e corresponderam a 4% do total neste setor (p < 0,0001). Observou-se um índice maior de amostras com representatividade somente de células escamosas no SUS (43,9%). Houve representatividade das células da junção escamo-colunar (JEC) em 82,1% das amostras colhidas no setor privado (p < 0,0001). Em relação aos resultados negativos para lesão intraepitelial e/ou malignidade, os percentuais obtidos foram 95,95% e 99,1% (p < 0,0049) para os exames coletados no sistema privado e no SUS, respectivamente. Em relação às lesões menos graves, no SUS obteve-se um resultado de 0,89% e no sistema privado de 2,56%; as lesões mais graves não foram diagnosticadas no SUS, enquanto que no setor privado representaram 1,49% dos exames. Conclusão As amostras cérvico-vaginais insatisfatórias foram observadas somente em exames realizados no SUS; há necessidade de orientação e capacitação dos profissionais que realizam a coleta do exame citopatológico, possibilitando uma maior confiabilidade nos resultados e mais segurança à mulher que se submete a este exame preventivo.

Capacitação dos profissionais de saúde e seu impacto no rastreamento de lesões precursoras do câncer de colo uterino / Training of health professionals and its impact on the trace of precursoring injuries of the uterine column cancer

Objetivo: Avaliar o impacto da capacitação dos profissionais do SUS envolvidos na coleta de amostras cervicovaginais para rastreamento de lesões precursoras do câncer de colo uterino no município de Videira (SC). Métodos: Estudo de intervenção cujas variáveis estudadas foram: adequabilidade da amostra, epitélios representados e resultado do exame citopatológico. A avaliação da qualificação dos profissionais baseou-se na comparação dos formulários referentes ao período de dezembro de 2015 a junho de 2016, antes da capacitação, com os formulários de junho de 2016 a janeiro de 2017, após a capacitação. Avaliou-se o resultado utilizando-se o teste do c2 de Pearson, com nível de significância de 5%. Resultados: Após a capacitação, houve aumento de amostras satisfatórias, de 97,81% para 99,0%. Observou-se insatisfatoriedade nas amostras coletadas antes e após a capacitação, correspondendo a 2,18% e 1,0%, respectivamente (p<0,0001). Houve índice maior de espécimes com representatividade somente de células escamosas antes da qualificação ­ 44,6 %. Verificou-se aumento da frequência de representação de células endocervicais, 55,39% para 85,03% (p<0,0001). Quanto aos resultados negativos para lesão intraepitelial e/ou malignidade, os percentuais obtidos foram 97,02% e 93,38%, antes e depois da capacitação, respectivamente. Foram constatados 2,41% e 5,14% (p<0,0001) de lesões menos graves, em comparação às lesões mais graves, correspondendo a 0,55% e 1,46% (p=0,0182), nas amostras obtidas antes e após a qualificação, respectivamente. Conclusão: Após a capacitação, houve melhora na adequabilidade da amostra, representação dos epitélios e resultado do exame. Logo, é imprescindível a capacitação dos profissionais coletores do exame citopatológico, proporcionando maior confiabilidade nos resultados.

Leafy Vegetables Free Of Coliforms And Parasites After Washing In Chlorine Treated Water

The production and preparation of vegetables for consumption involve procedures that pose health risks to consumers. In this context the water plays an important role in the quality of minimally processed vegetables. We assessed the hygienic-sanitary quality of leafy vegetables and obtained data of interest to health surveillance agencies and public health. They were tested for the presence of total and faecal coliforms, and also parasitic protozoa and helminths. The vegetable samples were purchased in supermarkets of the Midwest of the State of Santa Catarina, Brazil. We performed bacteriological analyses of wash water samples using chromogenic substrates to obtain the most probable number of the coliform group at 35 °C and Escherichia coli. We used Hoffman's spontaneous sedimentation technique for the parasitological analysis in order to check the occurrence of heavy and light helminths eggs and protozoan cysts. The analyses indicated the presence of coliforms, nematodes and freeliving protozoa, except for the last wash. We concluded that the washing process using running water and no active chlorine was not efficient in reducing the load of microorganisms in the vegetables. Therefore, intensive educational programmes should be implemented by health authorities to encourage hygienic-sanitary practices and risk reduction of food-borne diseases.(AU)

Oseltamivir-resistant influenza A(H1N1)pdm09 virus in southern Brazil

The neuraminidase (NA) genes of A(H1N1)pdm09 influenza virus isolates from 306 infected patients were analysed. The circulation of oseltamivir-resistant viruses in Brazil has not been reported previously. Clinical samples were collected in the state of Rio Grande do Sul (RS) from 2009-2011 and two NA inhibitor-resistant mutants were identified, one in 2009 (H275Y) and the other in 2011 (S247N). This study revealed a low prevalence of resistant viruses (0.8%) with no spread of the resistant mutants throughout RS.