Monitoramento da inserção do patrimônio genético de Biomphalaria tenagophila do Taim (RS), linhagem resistente ao Schistosoma mansoni, após a sua introdução em uma área endêmica para esquistossomose no Município de Bananal/SP, com transmissão mantida por B. tenagophila

Submitted by Nuzia Santos ([email protected]) on 2012-09-11T16:58:58Z No. of bitstreams: 1 Dissertação FINAL 08 09 12.pdf: 2298162 bytes, checksum: 6e2f996852589f87fd2d6180f0571f3e (MD5)Made available in DSpace on 2012-09-11T16:58:58Z (GMT). No. of bitstreams: 1 Dissertação FINAL 08 09 12.pdf: 2298162 bytes, checksum: 6e2f996852589f87fd2d6180f0571f3e (MD5)FIOCRUZFUNDEPCNPqCAPESFundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.O córrego Herivelton Martins foi escolhido para dar continuidade aos estudos com a introdução da linhagem Biomphalaria tenagophila do Taim/RS, resistente ao Schistosoma mansoni, em coleções hídricas do Município de Bananal/SP, iniciados em 2008. Oitocentos exemplares B. tenagophila adultos da linhagem Taim/RS foram marcados e introduzidos. Pelo método de marcação da concha foi estimada a densidade populacional da linhagem introduzida em relação à local. Após 15 dias da introdução 37,5% dos caramujos coletados estavam marcados. Após 4, 11 e 14 meses da introdução, caramujos jovens (≤ 5 mm de diâmetro) foram coletados e submetidos a técnica de PCR_RFLP para identificação do marcador molecular de 350pb, típico da linhagem do Taim e com caráter dominante. A proporção do marcador molecular nestes caramujos foi 37,1%, 35,7 e 60% respectivamente. Foram realizados três testes de infecção com a cepa SJ de S. mansoni com os descendentes F1 de caramujos coletados nos diferentes períodos após a introdução. Os sobreviventes foram submetidos à técnica de PCR_RFLP para verificar se havia alguma correlação entre a resistência e a presença do marcador de 350pb. No primeiro desafio com o S. mansoni 38,6% dos exemplares provenientes dos caramujos locais, coletados antes da introdução, estavam positivos, bem como 14,9% dos descendentes coletados após 4 meses da introdução. A análise molecular realizada com os caramujos positivos para S. mansoni, oriundos de exemplares pós-introdução, demonstraram que apenas 10% apresentaram o marcador de 350 pb, sendo que este marcador estava presente em 60,8% dos caramujos negativos. No segundo desafio, os descendentes de caramujos coletados antes, após 4 e 11 meses da introdução apresentaram taxas de infecção de 25%, 4,1%, 17%, respectivamente. Os caramujos positivos para S. mansoni não apresentaram o marcador molecular. A percentagem do marcador molecular nos caramujos negativos, coletados após 4 e 11 meses da introdução, foi de 66,6% e 56,2%, respectivamente. No terceiro desafio, as taxas de infecção dos descendentes foram de 26,5% antes da introdução, 6,8% após 11 meses e 2,1% após 14 meses. A análise molecular dos descendentes neste desafio está em andamento. Os dados apontam para uma significativa diminuição da suscetibilidade dos caramujos coletados após a intervenção. O marcador molecular de 350pb típico da linhagem do Taim foi detectado em proporções muito maiores nos caramujos negativos. Os resultados parciais obtidos permitem a inferência que o patrimônio genético da linhagem resistente B. tenagophila do Taim está sendo transmitido com êxito aos descendentes após intervenção tendo como consequência uma maior resistência à infecção por S. mansoni.The stream Herivelton Martins was chosen to continue the studies on introduction of Biomphalaria tenagophila from Taim/RS, resistant to Schistosoma mansoni, in hydric collections of Bananal/SP, initiated in 2008. Eight hundred adult specimens of B. tenagophila belonging to the lineage of Taim/RS were labeled and introduced. The populational density of the introduced lineage in relation to the local one was estimated using the labeling method of the shell. Fifteen days post-introduction, 37.5% of the collected snails were labeled. After 4, 11 and 14 months of the introduction, juvenile snails (≤ 5 mm diameter) were collected and submitted to PCR-RFLP technique for identification of the molecular marker 350 pb, typical of the Taim lineage and with dominant character. The proportions of the molecular marker in these snails were 37.1%, 35.7% and 60%, respectively. Three infection tests were carried out with the SJ strain of S. mansoni, using the descendants F1 of the snails collected on different periods after introduction. The surviving snails were submitted to the PCR-RFLP technique, in order to verify if there was any correlation between resistance and the presence of the marker 350 pb. In the first challenge with S. mansoni, 38.6% of the specimens belonging to the local population, and collected before introduction, were found to be positive, as well as 14.9% of the offsprings collected 4 months post-introduction. The molecular analysis performed with positive snails to S. mansoni, related to specimens after introduction, demonstrated that only 10% presented the molecular marker 350 pb, and this marker was present in 60.8% of the negative snails. In the second challenge, the offsprings of the snails collected before or 4 and 11 months post-introduction presented infection rates of 25%, 4.1% and 17%, respectively. The positive snails for S. mansoni did not present the molecular marker. The proportions of the molecular marker in the negative snails, collected 4 and 11 months after introduction, were 66.6% and 56.2%, respectively. In the third challenge, the infection rates of the descendants were 26.5% before introduction, 6.8% after 11 months and 2.1% after 14 months. The molecular analysis of the descendants related to this challenge is in progress. Data point out to a significant decrease in the susceptibility of the snails collected after intervention. The molecular marker 350 pb, typical of the Taim lineage, was detected in higher proportions in the negative snails. The partial results obtained allow to infer that the genetic heritage of the resistant B. tenagophila strain from Taim is being successfully transmitted to the descendants after intervention, showing as a result a higher resistance to S. mansoni infection

Reduced Susceptibility of a Biomphalaria tenagophila Population to Schistosoma mansoni after Introducing the Resistant Taim/RS Strain of B. tenagophila into Herivelton Martins Stream

Submitted by Nuzia Santos ([email protected]) on 2015-02-23T15:54:00Z No. of bitstreams: 1 2014_097.pdf: 480405 bytes, checksum: bebd815dcc4b7c94099cad10fea5543d (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2015-02-23T15:54:07Z (GMT) No. of bitstreams: 1 2014_097.pdf: 480405 bytes, checksum: bebd815dcc4b7c94099cad10fea5543d (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2015-02-23T15:58:38Z (GMT) No. of bitstreams: 1 2014_097.pdf: 480405 bytes, checksum: bebd815dcc4b7c94099cad10fea5543d (MD5)Made available in DSpace on 2015-02-23T15:58:38Z (GMT). No. of bitstreams: 1 2014_097.pdf: 480405 bytes, checksum: bebd815dcc4b7c94099cad10fea5543d (MD5) Previous issue date: 2014Fundação Oswaldo Cruz. Centro de Pesquisa Rene Rachou. Laboratório de Esquistossomose. Belo Horizonte, MG, BrazilUniversidade Federal de Juiz de Fora. Instituto de Ciencias Biologicas. Laboratorio de Parasitologia. Sao Pedro, MG, BrazilChacara Santa Ines. Bananal, SP, BrazilUniversidade Federal de Minas Gerais. Instituto de Ciencias Biologicas. Laboratório de Esquistossomose. Belo Horizonte, MG, BrazilSuperintendencia de Controle de Endemias. Sao Paulo, SP, Brazil,Fundação Oswaldo Cruz. Centro de Pesquisa Rene Rachou. Laboratório de Helmintologia e Malacologia Medica. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa Rene Rachou. Moluscário Dr. Lobato Paraense. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa Rene Rachou. Laboratório de Esquistossomose. Belo Horizonte, MG, BrazilStudies performed in the last 30 years demonstrated that a strain of B. tenagophila from the Taim Biological Reserve is completely resistant to Schistosoma mansoni infection. This resistance to parasite infection is a dominant characteristic during crossbreeding with susceptible B. tenagophila strains. These experiments also identified a 350 bp molecular marker that is exclusive to the Taim strain and does not occur in other geographic strains of this snail species. The Taim strain (Taim/RS) of Biomphalaria tenagophila was bred on a large scale, physically marked and introduced into a stream in which previous malacological analyses had revealed the presence of only parasite-susceptible B. tenagophila. Samples of offspring captured 4, 11 and 14 months after the introduction of the Taim strain were examined, and the susceptibility of the snails to S. mansoni infection dropped from 38.6–26.5% to 2.1% during the 14 months after the introduction of the Taim snail strain. A significant correlation was also observed between the absence of infection and the identification of the Taim molecular marker. These results demonstrate that the genetic marker from the Taim strain was successfully introduced into the wild snail population. In addition, a significant relationship exists between the marker and resistance to infection

Silver-stained 6% polyacrylamide gel showing RFLP profiles.

<p>Lane 1: molecular size markers (PhiX 174); Lane 3: negative control (i.e., no template DNA); Lane 13: pool of <i>S. mansoni</i> cercariae; Lane 14: <i>B. tenagophila</i> Taim; Lanes 6 and 9: profile of the local strain of <i>B. tenagophila</i>; Lanes 2, 4–5, 7–8 and 10–12: profiles of <i>B. tenagophila</i> collected 14 months after Taim introduction (i.e., containing the 350 bp marker).</p

Susceptibility of <i>B. tenagophila</i> collected before and after the introduction of the resistant lineage to infection after exposure to the SJ strain of <i>S. mansoni</i>.

<p>PI: post-introduction.</p><p>*Significant differences between <i>B. tenagophila</i> collected before and after the introduction of the resistant lineage (<i>p</i><0.05). Pearson’s chi-square test which is suitable for comparisons between proportions, and Fisher’s exact test were used at frequencies lower than 5.</p

Protease inhibitors from Theobroma cacao impair SARS-CoV-2 replication in vitro

SARS-CoV-2 is a newly emerging virus from the Coronaviridae family that has already infected over 700 million people worldwide and killed over 6 million. This virus uses protease molecules to replicate and infect the host, which makes these molecules targets for therapeutic substances to eliminate the virus and treat infected people. Through the protein-protein molecular docking approach, we detected two cystatins from Theobroma cacao, TcCYS3 and TcCYS4, described as papain-like protease inhibitors. These inhibitors decreased SARS-CoV-2 genomic copies without toxicity to Vero cells. There is a need to perform comprehensive studies in relevant animal models and to investigate the action mechanisms of protease inhibitors from Theobroma cacao that control the replication of SARS-CoV-2 in human cells