Actinobacteria From Termite Mounds Show Antiviral Activity Against Bovine Viral Diarrhea Virus, A Surrogate Model For Hepatitis C Virus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Extracts from termite-associated bacteria were evaluated for in vitro antiviral activity against bovine viral diarrhea virus (BVDV). Two bacterial strains were identified as active, with percentages of inhibition (IP) equal to 98%. Both strains were subjected to functional analysis via the addition of virus and extract at different time points in cell culture; the results showed that they were effective as posttreatments. Moreover, we performed MTT colorimetric assays to identify the CC50, IC50, and SI values of these strains, and strain CDPA27 was considered the most promising. In parallel, the isolates were identified as Streptomyces through 16S rRNA gene sequencing analysis. Specifically, CDPA27 was identified as S. chartreusis. The CDPA27 extract was fractionated on a C18-E SPE cartridge, and the fractions were reevaluated. A 100% methanol fraction was identified to contain the compound(s) responsible for antiviral activity, which had an SI of 262.41. GC-MS analysis showed that this activity was likely associated with the compound(s) that had a peak retention time of 5 min. Taken together, the results of the present study provide new information for antiviral research using natural sources, demonstrate the antiviral potential of Streptomyces chartreusis compounds isolated from termite mounds against BVDV, and lay the foundation for further studies on the treatment of HCV infection.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2011/50919-5]CNPq [311779/2014-0

Antiviral Activity Of Bacillus Sp. Isolated From The Marine Sponge Petromica Citrina Against Bovine Viral Diarrhea Virus, A Surrogate Model Of The Hepatitis C Virus.

The Hepatitis C virus causes chronic infections in humans, which can develop to liver cirrhosis and hepatocellular carcinoma. The Bovine viral diarrhea virus is used as a surrogate model for antiviral assays for the HCV. From marine invertebrates and microorganisms isolated from them, extracts were prepared for assessment of their possible antiviral activity. Of the 128 tested, 2 were considered active and 1 was considered promising. The best result was obtained from the extracts produced from the Bacillus sp. isolated from the sponge Petromica citrina. The extracts 555 (500 µg/mL, SI>18) and 584 (150 µg/mL, SI 27) showed a percentage of protection of 98% against BVDV, and the extract 616, 90% of protection. All of them showed activity during the viral adsorption. Thus, various substances are active on these studied organisms and may lead to the development of drugs which ensure an alternative therapy for the treatment of hepatitis C.51219-3

Actinobacteria from termite mounds show antiviral activity against bovine viral diarrhea virus, a surrogate model for Hepatitis C virus

Extracts from termite-associated bacteria were evaluated for in vitro antiviral activity against bovine viral diarrhea virus (BVDV). Two bacterial strains were identified as active, with percentages of inhibition (IP) equal to 98%. Both strains were subjected to functional analysis via the addition of virus and extract at different time points in cell culture; the results showed that they were effective as posttreatments. Moreover, we performed MTT colorimetric assays to identify the CC50, IC50, and SI values of these strains, and strain CDPA27 was considered the most promising. In parallel, the isolates were identified as Streptomyces through 16S rRNA gene sequencing analysis. Specifically, CDPA27 was identified as S. chartreusis. The CDPA27 extract was fractionated on a C18-E SPE cartridge, and the fractions were reevaluated. A 100% methanol fraction was identified to contain the compound(s) responsible for antiviral activity, which had an SI of 262.41. GC-MS analysis showed that this activity was likely associated with the compound(s) that had a peak retention time of 5 min. Taken together, the results of the present study provide new information for antiviral research using natural sources, demonstrate the antiviral potential of Streptomyces chartreusis compounds isolated from termite mounds against BVDV, and lay the foundation for further studies on the treatment of HCV infection2015CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP311779/2014-02011/50919-

Actinobacteria from Termite Mounds Show Antiviral Activity against Bovine Viral Diarrhea Virus, a Surrogate Model for Hepatitis C Virus

Extracts from termite-associated bacteria were evaluated for in vitro antiviral activity against bovine viral diarrhea virus (BVDV). Two bacterial strains were identified as active, with percentages of inhibition (IP) equal to 98%. Both strains were subjected to functional analysis via the addition of virus and extract at different time points in cell culture; the results showed that they were effective as posttreatments. Moreover, we performed MTT colorimetric assays to identify the CC50, IC50, and SI values of these strains, and strain CDPA27 was considered the most promising. In parallel, the isolates were identified as Streptomyces through 16S rRNA gene sequencing analysis. Specifically, CDPA27 was identified as S. chartreusis. The CDPA27 extract was fractionated on a C18-E SPE cartridge, and the fractions were reevaluated. A 100% methanol fraction was identified to contain the compound(s) responsible for antiviral activity, which had an SI of 262.41. GC-MS analysis showed that this activity was likely associated with the compound(s) that had a peak retention time of 5 min. Taken together, the results of the present study provide new information for antiviral research using natural sources, demonstrate the antiviral potential of Streptomyces chartreusis compounds isolated from termite mounds against BVDV, and lay the foundation for further studies on the treatment of HCV infection

Antiviral activity of extracts produced by isolated bacteria collected in termite mounds against important human and animal viruses

Resumo: As infecções virais acometem tanto humanos quanto animais. Problemas associados a falta de vacinas e o desenvolvimento de resistência aos medicamentos estimulam a pesquisa por compostos antivirais. As bactérias são responsáveis pela produção de inúmeros compostos com atividades biológicas importantes, portanto, esse trabalho teve como objetivo a busca por compostos antivirais produzidos por bactérias coletadas em cupinzeiros. Extratos provenientes de diferentes isolados de bactérias associadas ao cupim foram avaliados quanto a atividade antiviral in vitro frente a três vírus de importância para a saúde humana e animal: herpes humano tipo 1 (HHV-1), calicivírus felino (FCV)- modelo para o norovírus humano - e o vírus da diarréia viral bovina (BVDV) ¿ modelo para o vírus da hepatite C. Dos 90 extratos analisados, 10% apresentaram porcentagem de inibição (PI%) ? 98% contra pelo menos um dos vírus: cinco foram ativos frente ao HHV-1, três apresentaram atividade contra o FCV e dois foram ativos frente ao BVDV. Das espécies ativas, seis pertencem ao gênero Streptomyces, com uma delas identificada como S. chartreusis e quatro identificadas como sendo do gênero Achromobacter. Três extratos apresentaram índice de seletividade (IS) acima de 3,5 contra o HHV-1 com destaque ao CDPA10 (Streptomyces sp) com IS=17,10. Frente ao FCV e o BVDV dois extratos foram considerados promissores: o LC22 (IS=11,00), identificado como Streptomyces sp e o CDPA27 (S. chartreusis) com IS=3,5, respectivamente. Os extratos que destacaram-se quanto aos valores e IS foram também avaliados em uma triagem de mecanismo de ação. Dos três extratos com IS acima de 3,5 frente ao HHV-1, um apresentou atividade antiviral no pós tratamento. Os outros, incluindo o CDPA10 agem como inativadores virais. O extrato LC22, ativo contra o FCV apresentou atividade no pós tratamento e também como inativador viral. Por fim, o extrato proveniente do S. chartreusis (CDPA27), agiu no pós tratamento frente ao BVDV. Esses três extratos, CDPA10, LC22 e CDPA27 foram selecionados para o fracionamento em coluna C18 fase reversa em ordem crescente de metanol, originando seis frações. Todas as frações foram avaliadas contra seus respectivos vírus e como resultado foi possível observar que a fração 6 (100% metanol) foi a mais promissora dos três extratos. Essa fração do CDPA10 e do CDPA27, ativos contra o HHV-1 e BVDV respectivamente, apresentaram valor de PI=99%, enquanto a fração 6 do extrato LC22 (ativo contra o FCV) apresentou PI=90%. Nas avaliações quanto ao IS dessas frações promissoras, observou-se que os valores aumentaram consideravelmente tanto para o CDPA10 como para o CDPA27, já a fração do LC22 teve o valor de IS reduzido para 3,24. Finalmente, todos os extratos e suas frações promissoras foram analisados por cromatografia gasosa acoplada a espectrometria de massa (CG-MS). Esta análise, quando comparada aos compostos já descritos na biblioteca NIST/2005, não permitiu a identificação dos compostos devido a baixa similaridade. Estudos mais detalhados devem ser realizados a fim de identificar as espécies bacterianas e seus respectivos compostos ativos. Os resultados obtidos revelam novas perspectivas para a pesquisa antiviral a partir de produtos naturais e demonstra o potencial antiviral de compostos produzidos por bacterias associadas ao cupim. ;;Abstract: Viral infections affect both humans and animals. Problems associated with a lack of vaccines and development of drug resistance stimulates the research for antiviral compounds. Bacteria are responsible for producing many compounds with important biological activities, so this study aimed to research for antiviral compounds produced by isolated associated bacteria. Extracts from different isolated termite associated bacteria were evaluated for in vitro antiviral activity against three important viruses to human and animal health: human herpesvirus type 1 (HHV-1), feline calicivirus (FCV) - surrogate model for human norovirus -and bovine viral diarrhea virus (BVDV) - surrogate model for hepatitis C virus. Of 90 extracts analyzed, 10% had percentage of inhibition (PI%) ? 98% against at least one virus: five were active against HHV-1, three showed activity against FCV and two were active against BVDV. Of the active species, six belong to Streptomyces genus, with one of them identified as S. chartreusis and four were identified as Achromobacter genus. Three extracts showed selectivity index (SI) above 3.5 against HHV-1, highlighting CDPA10 (Streptomyces sp) with SI = 17.10. Against the FCV and BVDV only two extracts were considered promising: LC22 (SI = 11.00), identified as Streptomyces sp and CDPA27 (S. chartreusis) with SI = 3.5, respectively. The extracts that stood out about the values of SI were also evaluated in a screening of mechanism of action. Of the three extracts with SI above 3.5 against HHV-1, only one showed antiviral activity in post-treatment, the others, including CDPA10 act as virus inactivation. The LC22, extract active against FCV, showed activity in the post-treatment as well as virus inactivation. Finally, the extract from S. chartreusis (CDPA27), acted in the post-treatment against BVDV. These three extracts CDPA10, LC22 and CDPA27 were then selected for fractionation on column C18 phase reverse with ascending order of methanol, resulting in six fractions. All fractions were evaluated against the respective virus and as a result was observed that the fraction 6 (100% methanol) were the most promising of the three extracts. That fraction of CDPA10 and CDPA27, active against HHV-1 and BVDV respectively, presented value of PI = 99%, while the fraction 6 of LC22 extract (active against FCV) presented PI = 90%. The SI evaluations of these promising fractions showed that the values increased substantially for both CDPA10 and CDPA27; however the fraction of LC22 extract had reduced SI value to 3.24. Finally, all extracts and their promising fractions were analyzed by gas chromatography-mass spectrometry (GC-MS). This analysis, when compared to the compounds already described in the NIST / 2005 library, it was not possible to identify them due to low similarity. More detailed studies should be performed to identify the bacterial species and their active compounds. The result of the present study provide new information for antiviral research using natural sources and demonstrates antiviral potential of compounds produced by termite associated bactéria

Antiviral activity of Brazilian Cerrado plants extracts against Herpesvirus

Orientador: Clarice Weis ArnsDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: Os herpesvírus são responsáveis por enfermidades importantes em humanos e animais. Em animais, estão associados a doenças que causam grandes perdas econômicas. Em humanos, a gravidade da enfermidade é maior quando os pacientes são imunossuprimidos. Além disso, já existem cepas mutantes resistentes aos medicamentos disponíveis. Visto as dificuldades associadas a prevenção e tratamento das infecções por herpesvírus, a utilização de produtos de plantas como antivirais apresenta - se como alternativa. O Cerrado Brasileiro é um bioma que localiza -se praticamente todo no Brasil e apresenta mais de 10.000 espécies de plantas. Essas plantas podem potencialmente servi r de fonte de compostos farmacologicamente ativos. Assim, o presente trabalho teve como objetivos avaliar a atividade antiviral , atividade virucida e o índice de seletividade (SI) de extratos de plantas do Cerrado contra os herpesvírus suíno tipo 1 (SuHV-1), equinotipo 1 (EHV-1) e vírus do herpes simplex tipo 1 (HSV-1) . Inicialmente, os extratos liofilizados foram submetidos aos testes de citotoxicidade em células MDBK e Vero para determinar a concentração máxima não tóxica (CMNT). Dos extratos, quatro apresentaram as mesmas CMNT's em ambas as linhagens mas, em geral , os extratos foram mais citotóxicos para células Vero. A seguir, com base na CMNT, foram realizados os testes de atividade antiviral para os vírus HSV-1 e EHV-1 em células Vero, e SuHV-1 em MDBK. Os resultados demonstraram que 50% dos extratos apresentaram atividade contra pelo menos um dos herpesvírus estudados, com destaque para as espécies Banisteriopsis variabil is , Byrsonima intermedia e Xylopia aromática que foram ativas contra os t rês herpesvírus, e o extrato da Stryphnodendron adstringens , ativo contra o HSV-1 e SuHV-1. Os extratos que apresentaram atividade antiviral foram então testados quanto a atividade virucida e os resultados submetidos ao cálculo do SI. O extrato foi considerado ativo quando o índice de inibição viral ( IIV) foi maior ou igual a 1,5 ou apresentou PI% (porcentagem de inibição) maior ou igual a 97%. Quanto ao SI, foram considerados ativos os extratos que apresentaram valores iguais ou superiores a 4. A atividade virucida foi observada em 75% dos extratos contra pelo menos um dos herpesvírus testados. As espécies que apresentaram os resultados mais promissores foram: B. variabil is, X. aromática, S. adstringens e B. intermedia. Esta última foi então utilizada em testes adicionais com a variação da concentração, e demonstrou atividade antiviral e virucida em concentrações inferiores a CMNT contra os herpesvírus testados. Assim, o presente trabalho demonstra o potencial de plantas do Cerrado como fonte de compostos com atividade antiviral e virucida. Estudos adicionais são necessários para avaliar os mecanismos de ação e os compostos químicos responsáveis pela atividade observadaAbstract: Herpesviruses are responsible for important diseases in humans and animals. In animals, they are associated with economically important diseases worldwide. In humans, they represent serious threats to public health, and the severity of the illness increases in immunocompromised patients. In addition, there are mutant strains that are resistant to available drugs. Because of the difficulties associated with the prevent ion and treatment of herpesvirus infect ions, the use of plant products as antivirals can be an alternative. The Brazilian Cerrado is a biome located almost entirely in Brazil has over 10,000 species of plants. These plants can potentially be used as a source of pharmacologically active compounds. There for , this study aimed to evaluate the antiviral activity, virucidal activity and the selectivity index (SI) of extracts from Cerrado plants against suid herpesvirus type 1 (SuHV-1) equid type 1 (EHV-1) and herpes simplex virus type 1 (HSV-1). Initially, the lyophilized extracts were tested for cytotoxicity in MDBK and Vero cells to identify t he maximum nontoxic concentration (MNTC). Of the extracts, four showed the same MNTC for both cells, but the extracts were generally more toxic to Vero cells. Then, based on the MNTC, antiviral activity tests were performed against HSV-1 and EHV-1 in Vero cells and SuHV-1 in MDBK cells. The results demonstrated that 50% of the extracts showed activity against at least one of the herpesviruses studied. In particular, the extracts from Banisteriopsis variabil is, Byrsonima intermedia and Xylopia aromatica, were active against all of the herpesviruses, and the extract from Stryphnodendron adstringens was active against HSV-1 and SuHV-1. The extracts that showed antiviral activity were also tested for virucidal activity, and the SI was calculated. An extract was considered active when the viral inhibition index (VII) was greater than or equal to 1.5 or showed a PI% (percent inhibition) greater than or equal to 97%. As for the SI, extracts were considered active when the displayed values greater than or equal to four. Virucidal activity was observed in 75% of the extracts against at least one of the herpesviruses tested. The species that showed the most promising results were: B. variabil is , X. aromatica, S. adstringens and B. intermedia. Was used for additional testing with varying concentrations, and demonstrated antiviral and virucidal activities at concentrations lower than the MNTC against the herpesviruses tested. Therefore, this study demonstrates the potential of Cerrado as a source of compounds with antiviral and virucidal activities. Additional studies are necessary to evaluate the mechanisms of act ion and the chemical compounds responsible for the observed activityMestradoCiencias BasicasMestre em Clinica Medic

Antiviral Activity of Bacillus sp. Isolated from the Marine Sponge Petromica citrina against Bovine Viral Diarrhea Virus, a Surrogate Model of the Hepatitis C Virus

The Hepatitis C virus causes chronic infections in humans, which can develop to liver cirrhosis and hepatocellular carcinoma. The Bovine viral diarrhea virus is used as a surrogate model for antiviral assays for the HCV. From marine invertebrates and microorganisms isolated from them, extracts were prepared for assessment of their possible antiviral activity. Of the 128 tested, 2 were considered active and 1 was considered promising. The best result was obtained from the extracts produced from the Bacillus sp. isolated from the sponge Petromica citrina. The extracts 555 (500 µg/mL, SI>18) and 584 (150 µg/mL, SI 27) showed a percentage of protection of 98% against BVDV, and the extract 616, 90% of protection. All of them showed activity during the viral adsorption. Thus, various substances are active on these studied organisms and may lead to the development of drugs which ensure an alternative therapy for the treatment of hepatitis C

A lipidomics approach in the characterization of zika-infected mosquito cells: potential targets for breaking the transmission cycle

Recent outbreaks of Zika virus in Oceania and Latin America, accompanied by unexpected clinical complications, made this infection a global public health concern. This virus has tropism to neural tissue, leading to microcephaly in newborns in a significant proportion of infected mothers. The clinical relevance of this infection, the difficulty to perform accurate diagnosis and the small amount of data in literature indicate the necessity of studies on Zika infection in order to characterize new biomarkers of this infection and to establish new targets for viral control in vertebrates and invertebrate vectors. Thus, this study aims at establishing a lipidomics profile of infected mosquito cells compared to a control group to define potential targets for viral control in mosquitoes. Thirteen lipids were elected as specific markers for Zika virus infection (Brazilian strain), which were identified as putatively linked to the intracellular mechanism of viral replication and/or cell recognition. Our findings bring biochemical information that may translate into useful targets for breaking the transmission cycle1110CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPSem informaçãoSem informação2011/50400-0; 2015/06809-1; 2014/00084-2; 2014/00302-0; 2013/11343-