Hepatoprotective properties from the seaweed Bryothamnion triquetrum (S.G.Gmelin) M.A.Howe against CCl4-induced oxidative damage in rats

Context: Seaweeds are seen as a traditional food and folk medicine by different coastal countries. The red seaweed Bryothamnion triquetrum is a widely distributed species that grows in shallow waters, and different authors have demonstrated a possible application of the seaweeds as a source of natural antioxidants and relative diseases. Aims: To evaluate the hepatoprotective properties on CCl4-induced oxidative stress in rats that were associated with the antioxidant activity from the polyphenol-rich fractions of the red seaweed Bryothamnion triquetrum. Methods: Polyphenols were determined by Folin-Cioacalteu. Antioxidant activity from phenolic compounds-rich fractions was measured by different assays (DPPH, Reducing power, (beta-Carotene/linoleic acid assay and Inhibition of lipoperoxidation). Aqueous extract from B. triquetrum was administered during 20 days to rats and submitted CCl4-Induced oxidative damage. The peroxidation and hepatic damage (TBARS, ASAT and ALAT), antioxidant metabolite and enzymes (glutathione, catalase and superoxide dismutase) were evaluated. Also, it was evaluated the expression of antioxidant enzymes by RT-PCR. Results: The antioxidant activity determined by different assays with polyphenolic fractions. Free Phenolic Acid was more active: DPPH, 20 mu g 87%; Reducing power OD = 0.490, 20 g; beta-carotene/linoleic acid 1 mu g 53%, and inhibition of lipid peroxidation 0.250 mu g 100%. Rats treated displayed lower liver TBARS, ASAT and ALAT than CCl4-treated group and catalase activity was increased. It was demonstrated expression of catalase. Conclusions: Data suggest that Bryothamnion triquetrum protects the liver against oxidative stress by modulating its antioxidant enzymes and oxidative status with potential use as phytodrug or functional food.

Presence of respiratory viruses in equines in Brazil.

Equines are susceptible to respiratory viruses such as influenza and parainfluenza. Respiratory diseases have adversely impacted economies all over the world. This study was intended to determine the presence of influenza and parainfluenza viruses in unvaccinated horses from some regions of the state of São Paulo, Brazil. Blood serum collected from 72 equines of different towns in this state was tested by hemagglutination inhibition test to detect antibodies for both viruses using the corresponding antigens. About 98.6% (71) and 97.2% (70) of the equines responded with antibody protective titers (≥ 80 HIU/25µL) H7N7 and H3N8 subtypes of influenza A viruses, respectively. All horses (72) also responded with protective titers (≥ 80) HIU/25µL against the parainfluenza virus. The difference between mean antibody titers to H7N7 and H3N8 subtypes of influenza A viruses was not statistically significant (p > 0.05). The mean titers for influenza and parainfluenza viruses, on the other hand, showed a statistically significant difference (p < 0.001). These results indicate a better antibody response from equines to parainfluenza 3 virus than to the equine influenza viruses. No statistically significant differences in the responses against H7N7 and H3N8 subtypes of influenza A and parainfluenza 3 viruses were observed according to the gender (female, male) or the age (≤ 2 to 20 years-old) groups. This study provides evidence of the concomitant presence of two subtypes of the equine influenza A (H7N7 and H3N8) viruses and the parainfluenza 3 virus in equines in Brazil. Thus, it is advisable to vaccinate equines against these respiratory viruses.

Presence of resiiratory viruses in equines in Brazil / Presença de vírus respiratórios em equinos do Brasil

Equines are susceptible to respiratory viruses such as influenza and parainfluenza. Respiratory diseases have adversely impacted economies all over the world. This study was intended to determine the presence of influenza and parainfluenza viruses in unvaccinated horses from some regions of the state of São Paulo, Brazil. Blood serum collected from 72 equines of different towns in this state was tested by hemagglutination inhibition test to detect antibodies for both viruses using the corresponding antigens. About 98.6% (71) and 97.2% (70) of the equines responded with antibody protective titers (≥ 80 HIU/25µL) H7N7 and H3N8 subtypes of influenza A viruses, respectively. All horses (72) also responded with protective titers (≥ 80) HIU/25µL against the parainfluenza virus. The difference between mean antibody titers to H7N7 and H3N8 subtypes of influenza A viruses was not statistically significant (p > 0.05). The mean titers for influenza and parainfluenza viruses, on the other hand, showed a statistically significant difference (p < 0.001). These results indicate a better antibody response from equines to parainfluenza 3 virus than to the equine influenza viruses. No statistically significant differences in the responses against H7N7 and H3N8 subtypes of influenza A and parainfluenza 3 viruses were observed according to the gender (female, male) or the age (≤ 2 to 20 years-old) groups. This study provides evidence of the concomitant presence of two subtypes of the equine influenza A (H7N7 and H3N8) viruses and the parainfluenza 3 virus in equines in Brazil. Thus, it is advisable to vaccinate equines against these respiratory viruses.

Os equinos são susceptíveis aos vírus respiratórios, como o vírus influenza, e também tem sido citado o vírus parainfluenza. Doenças respiratórias têm impactado a economia em todo mundo. Este estudo intencionou determinar a presença dos vírus influenza e parainfluenza em equinos não vacinados de certas regiões do Estado de São Paulo, Brasil. Os soros coletados de 72 equinos, de diferentes cidades deste Estado, foram submetidos ao teste de Inibição da Hemaglutinação (IH) com objetivo de detectar anticorpos contra os referidos vírus, usando antígenos correspondentes. Cerca de 98,8% (72) e 97,2% (70) desses equinos responderam com títulos protetores (≥ 80 UIH/25µL) para os subtipos H7N7 e H3N8 de vírus influenza, respectivamente. Todos equinos (72) responderam com títulos protetores (≥ 80 UIH/25µL) contra o vírus parainfluenza 3. A diferença entre as médias de anticorpos contra o vírus influenza A não foi estatisticamente significante (p > 0,05). As médias de títulos dos vírus influenza e parainfluenza, por outro lado, demonstraram diferença estatisticamente significante (p < 0,001). Esses resultados indicam melhor resposta de anticorpos pelos equinos ao vírus parainfluenza 3 do que ao vírus da influenza equina. Nenhuma diferença estatística foi observada nas respostas contra os vírus da influenza equina A (H7N7 e H3N8) e parainfluenza 3, com relação ao gênero (fêmeas e machos) e grupo etário (≤ 2 até 20 anos) nos equinos avaliados. Este estudo fornece evidência da presença concomitante dos dois subtipos vírus influenza A (H7N7 e H3N8) e do parainfluenza 3 em cavalos no Brasil. Portanto, é aconselhável a vacinação dos cavalos contra esses vírus respiratórios.

Influenza viruses in adult dogs raised in rural and urban areas in the state of São Paulo, Brazil / Incidência do vírus influenza em cães adultos criados em áreas rural e urbana do estado de São Paulo, Brasil

In 1970, searching for the interspecies transmission of influenza viruses led to the first study on influenza viruses in domestic animals. Birds and mammals, including human beings, are their natural hosts; however, other animals may also play a role in the virus epidemiology. The objective was to investigate the incidence of influenza viruses in adult dogs raised in rural (9, 19.56%) and urban (37, 80.43%) areas in the state of São Paulo, Brazil. Dog serum samples were examined for antibodies to influenza viruses by the hemagglutination inhibition (HI) test using the corresponding antigens from the circulating viruses in Brazil. Dogs from rural areas presented antibodies to influenza A H3N2, and influenza A H7N7 and H3N8. In rural areas, dog sera displayed mean titers as 94.37, 227.88, 168.14, 189.62 HIU/25 µL for subtypes H1N1, H3N2, H7N7, H3N8, respectively. About 84% and 92% of dogs from urban areas exhibited antibodies to human influenza A H1N1 and H3N2, respectively, with statistical difference at p < 0.05 between the mean titers of antibodies to H1N1 and H3N2. About 92% and 100% were positive for H7N7 and H3N8, respectively. In dogs from urban areas, the mean titers of antibodies against influenza A H1N1, H3N2, H7N7 and H3N8, were 213.96, 179.42, 231.76, 231.35 HIU/25 µL respectively. The difference among them was not statistically significant at p > 0.05. In conclusion, these dogs were positive for both human and equine influenza viruses. The present study suggests the first evidence that influenza viruses circulate among dogs in Brazil.

A transmissão interespecífica do vírus influenza é relatada em estudo sobre influenza com animais domésticos desde 1970. Pássaros e mamíferos, incluindo o homem, são seus hospedeiros naturais, porém outros animais podem participar da sua epidemiologia. Foi investigada a incidência do vírus influenza em cães adultos criados em zonas rural (9, 19,56%) e urbana (37, 80,43%), do Estado de São Paulo. Os soros dos cães foram examinados pelo teste de inibição da hemaglutinação (IH), usando antígeno dos vírus influenza circulantes no Brasil. Nos cães rurais foram detectados títulos médios de 94,37, 227,88, 168,14 e 189,62 UIH/25 mL (unidades inibidoras de hemaglutinação/25 mL) para os subtipos H1N1, H3N2, H7N7, H3N8 de vírus influenza A, respectivamente, com diferenças estatisticamente significativas (p<0,05) entre as médias de títulos de anticorpos contra H1N1 e H3N2. Cerca de 84% e 92% dos cães urbanos responderam aos vírus influenza A humano H1N1 e H3N2, respectivamente e destes 92% e 100% foram positivos para os vírus eqüinos H7N7 e H3N8, respectivamente. Para esses cães as médias de títulos de anticorpos para os vírus influenza A H1N1, H3N2, H7N7 e H3N8 foram 213,96, 179,42, 231,76 e 231,35UIH/25 mL, respectivamente. As diferenças entre as médias não foram estatisticamente significativas (p>0,05). Conclui-se que os cães apresentaram positividade para ambos vírus influenza humano e equino. O presente estudo sugere, pela primeira vez, evidências de que há circulação do vírus influenza em cães, no Brasil.

Influenza viruses in adult dogs raised in rural and urban areas in the state of São Paulo, Brazil.

In 1970, searching for the interspecies transmission of influenza viruses led to the first study on influenza viruses in domestic animals. Birds and mammals, including human beings, are their natural hosts; however, other animals may also play a role in the virus epidemiology. The objective was to investigate the incidence of influenza viruses in adult dogs raised in rural (9, 19.56%) and urban (37, 80.43%) areas in the state of São Paulo, Brazil. Dog serum samples were examined for antibodies to influenza viruses by the hemagglutination inhibition (HI) test using the corresponding antigens from the circulating viruses in Brazil. Dogs from rural areas presented antibodies to influenza A H3N2, and influenza A H7N7 and H3N8. In rural areas, dog sera displayed mean titers as 94.37, 227.88, 168.14, 189.62 HIU/25 µL for subtypes H1N1, H3N2, H7N7, H3N8, respectively. About 84% and 92% of dogs from urban areas exhibited antibodies to human influenza A H1N1 and H3N2, respectively, with statistical difference at p < 0.05 between the mean titers of antibodies to H1N1 and H3N2. About 92% and 100% were positive for H7N7 and H3N8, respectively. In dogs from urban areas, the mean titers of antibodies against influenza A H1N1, H3N2, H7N7 and H3N8, were 213.96, 179.42, 231.76, 231.35 HIU/25 µL respectively. The difference among them was not statistically significant at p > 0.05. In conclusion, these dogs were positive for both human and equine influenza viruses. The present study suggests the first evidence that influenza viruses circulate among dogs in Brazil.

VaxcineTM: an oil-based adjuvant for influenza vaccines

Vaccination is the method of choice for the prevention of influenza infection. However, the quantity of the antigen available, especially in the case of pandemics, often fails to meet the global demand. However, improved adjuvants can overcome this problem. Preliminary results obtained in this study revealed that one year after a single subcutaneous immunisation with influenza A H3N2 virus in an oil-based carrier, VaxcineTM, outbreed mice produced a high immunoglobulin G response that lasted for up to one year and exhibited less variation in titre compared with the response of the control group treated with alum. The haemagglutination-inhibition titres induced by VaxcineTM were also higher than those generated by alum. These data indicate that VaxcineTM is a good adjuvant candidate for seasonal influenza vaccines.

Analysis of in vitro and in vivo antioxidant properties of hydrophilic fractions from the Seaweed Halimeda Monile L

In this work, in vitro and in vivo antioxidant properties of the marine algae Halimeda monile were assessed and the levels of some of its compounds likely to be responsible for such properties were determined. The estimated contents of total polyphenols, chlorophylls a and b and carotenoids were 179.5, 356.3, 452.8 and 42.2 µg/g dry weight seaweed, respectively. The presence of terpenoids and flavonoids was also observed. The antioxidant activity of two polar fractions from H. monile (lyophilized aqueous extract and free phenolic acid fraction) was evaluated using three antioxidant assays: ferric reducing antioxidant power, 1,1-diphenyl-2-picrylhydrazyl and lipid peroxidation. Treatment of CCl4-induced liver damage in rats with extracts resulted in lower serum thiobarbituric acid-reactive substances levels and higher hepatic glutathione concentrations compared to those observed in the CCl4-treated group. Also, a significant increase in catalase activity was detected after treatment with the extracts. These results suggest that the seaweed H. monile could be a potential source for natural antioxidants.

Vaxcine(TM): an oil-based adjuvant for influenza vaccines.

Vaccination is the method of choice for the prevention of influenza infection. However, the quantity of the antigen available, especially in the case of pandemics, often fails to meet the global demand. However, improved adjuvants can overcome this problem. Preliminary results obtained in this study revealed that one year after a single subcutaneous immunisation with influenza A H3N2 virus in an oil-based carrier, Vaxcine(TM), outbreed mice produced a high immunoglobulin G response that lasted for up to one year and exhibited less variation in titre compared with the response of the control group treated with alum. The haemagglutination-inhibition titres induced by Vaxcine(TM) were also higher than those generated by alum. These data indicate that Vaxcine(TM) is a good adjuvant candidate for seasonal influenza vaccines.

Compostos fenólicos da polpa e sementes de romã ( Púnica granatum, L)atividade antioxidante e protetrora em células MDCK

Os radicais livres, dentre os quais as EROs e as ERNs, são espécies reativas envolvidas no início de reaçõescomo a peroxidação, que no sistema biológico pode dar início ao desenvolvimento de diversos processos patológicos. Para conter o avanço da cadeia oxidativa iniciada pelos radicais livres e minimizar os seus efeitos deletérios ao organismo, algumas substâncias antioxidantes como compostos fenólicos, dentre os quais os ácidos fenólicos, podem ser encontrados em diversas fontes vegetais. Asfrutas são uma boa fonte de compostos antioxidantes, e a romã (Punica granatum, L.) apresenta expressiva quantidade de compostos fenólicos. Os objetivos deste trabalho foram identificar os principais ácidos fenólicos, presentes na fruta (polpa e sementes), através de cromatografi a gasosa, a avaliação da capacidade antioxidante, utilizando dois métodos: o de co-oxidação de substratos (β- caroteno e ácido linoléico) e o da redução do radical 1,1- diphenyl- 2- picryl- hydrazyl (DPPH•). Ainda, foi verifi cado o efeito da fração de ácidos fenólicos livres da polpa da romã (AFLp) sobre a proliferação e viabilidade em cultura de células MDCK. Pôde-se verifi car que a fração AFLp apresentou o maior conteúdo de compostos redutores (1.399,02μg/mL) enquanto que a de ácidos fenólicos ligados a compostos solúveis apresentou o maior conteúdo (770,58 μg/mL) nas sementes(AFESs). Todas as frações de ácidos fenólicos apresentaram resultados signifi cativamente (p<0,05) superiores aos do antioxidante sintético butil-hidroxitolueno, utilizado como padrão. Os principais ácidos fenólicos encontrados foram o salicílico, o protocatequínico e o gálico. As célulastratadas com baixas concentrações da fração AFLp apresentaram aumento da proliferação celular e a viabilidade manteve-se alta.

Free phenolic acids from the seaweed Halimeda monile with antioxidant effect protecting against liver injury.

Phenolic compounds are found in seaweed species together with other substances presenting antioxidant activity. The objective of this work was to evaluate the antioxidant activity of the free phenolic acids (FPA) fraction from the seaweed Halimeda monile, and its activity to protect the expression of hepatic enzymes in rats, under experimental CCl4 injury. The antioxidant activity was measured by the DPPH method. The FPA fraction (80 mg/kg, p.o.) was administered during 20 consecutive days to rats. The peroxidation was performed by thiobarbituric acid reactive substances (TBARS). The SOD and CAT enzymatic expressions were measured by RT/PCR. The histology technique was used to evaluate liver injuries. The expression of both, CAT and SOD genes, was more preserved by FPA. Only partial injury could be observed by histology in the liver of rats receiving FPA as compared with the control group; and CCl4 administration induced 60% more peroxidation as compared with the rats receiving FPA. These data suggest that FPA could modulate the antioxidant enzymes and oxidative status in the liver through protection against adverse effects induced by chemical agents.

Inhibition of DNA virus: herpes-1 (HSV-1) in cellular culture replication, through an antioxidant treatment extracted from rosemary spice / A inibição do vírus DNA: herpes-1 (HSV-1) na replicação de cultura celular, através de um tratamento antioxidante extraídos de especiarias alecrim

This work aimed to evaluate antiviral properties in antioxidants from spices. Phenolic compounds extracted from rosemary (Rosmarinus officinallis, L) by hot water, had their antioxidant activity determined by spectrophotometry using β carotene/linoleic acid system. The rosemary extract was evaluated by antiviral assay of Herpes Virus type-1 (HSV-1) replication in VERO cells, in the presence or absence of the spice. 10,000 TCID50/mL of the HSV-1 was kept for 3 h at 4º C, with 300 ppm of rosemary extract, and 100 ppm of butyl hydroxyl toluene (BHT). Then, these viruses were inoculated in VERO cells incubated at 37º C in CO2-5 percent, for seven days. Daily, they were examined and the end point was based on 100 percent of CPE in virus control (without antioxidants). The HSV-1 replication inhibition percentage (IP) measured the antiviral action from antioxidants, showing viral reductions of the 82.0, 82.5 percent, in the presence of rosemary and rosemary + BHT, respectively. As an extension, cell test corresponded to the similar viral decrease (IP = 85.0 and 86.3 percent) in both aforementioned situations. Results lead to conclude that phenolic compounds from rosemary revealed an antiviral action on herpesvirus-1.

Neste estudo foi avaliada a ação antiviral de antioxidantes de especiaria. Extrato aquoso de alecrim (Rosmarinus officinalis, L), que apresentou atividade antioxidante através de espectrofotometria usando o sistema β caroteno/ácido linoléico, foi avaliado em ensaios com vírus herpes-1 na replicação em células VERO. Nestes ensaios foram utilizados 10.000 TCID50 por cento/mL do vírus HSV-1, mantidos em contato com 300 ppm do extrato de alecrim e com 100 ppm de butil hidroxi tolueno (BHT), durante 3h a 4ºC. Esses vírus, em seguida, foram inoculados em células VERO incubadas a 37 ºC/5 por cento de CO2 por sete dias. Pelo efeito citopático (ECP) e o "end point" de ECP do controle de vírus (sem antioxidante), foi possível observar que houve reduções na replicação viral de 82 e 82,5 por cento na presença do alecrim e do alecrim + BHT, respectivamente. Nessa situação,avaliou-se ainda a redução da adsorção viral às células, que apresentou índices similares de 85,0 e 86,3 por cento de redução na capacidade da adsorção. Estas reduções no desempenho do HSV foram medidas pela fórmula de porcentagem de inibição da replicação viral (PI). Os resultados levam a concluir que os compostos fenólicos do alecrim apresentam ação antiviral sobre o HSV-1.

Influenza virus and proteolytic bacteria co-infection in respiratory tract from individuals presenting respiratory manifestations.

A role for proteolytic bacteria in the exacerbation of influenza virus has been shown in natural hosts such as pigs and humans. Four hundred seven samples were collected from the respiratory tract of individuals presenting clinical manifestations, during influenza season (2003-2005) in São Paulo City. The aim of this study was to evaluate the incidence of determined bacteria co-infecting virus in human respiratory tract. Tests, such as bacteriological, immunofluorescence (IF), RT/PCR and hemagglutination (HA) were used for bacterial and viral investigation. Thirty seven (9.09%) positive for influenza virus were screened by IF. The RT/PCR confirmed the presence of influenza virus in these samples. Bacterial and agar casein tests demonstrated that 18 (48.64%) individuals were infected with proteolytic bacteria such as Staphylococcus spp., Streptococcus spp. and Pseudomonas spp. Among these samples, 13 (35.13%) were co-infected with influenza A virus. Influenza type B, co-infecting bacteria were found in five (13.51%) samples. In vitro the S. aureus protease increased the influenza HA titer after contact for 30 min at 25 masculineC. Results revealed the occurrence of co-infection with proteolytic bacteria and influenza in the evaluated individuals. This finding corroborates that virus versus bacteria synergism could be able to potentiate respiratory infection, increasing damage to hosts.

Influenza virus and proteolytic bacteria co-infection in respiratory tract from individuals presenting respiratory manifestations / Vírus influenza e bactéria proteolítica co-infectantes em trato respiratório de indivíduos com manifestações respiratórias

A role for proteolytic bacteria in the exacerbation of influenza virus has been shown in natural hosts such as pigs and humans. Four hundred seven samples were collected from the respiratory tract of individuals presenting clinical manifestations, during influenza season (2003-2005) in São Paulo City. The aim of this study was to evaluate the incidence of determined bacteria co-infecting virus in human respiratory tract. Tests, such as bacteriological, immunofluorescence (IF), RT/PCR and hemagglutination (HA) were used for bacterial and viral investigation. Thirty seven (9.09 percent) positive for influenza virus were screened by IF. The RT/PCR confirmed the presence of influenza virus in these samples. Bacterial and agar casein tests demonstrated that 18 (48.64 percent) individuals were infected with proteolytic bacteria such as Staphylococcus spp., Streptococcus spp. and Pseudomonas spp. Among these samples, 13 (35.13 percent) were co-infected with influenza A virus. Influenza type B, co-infecting bacteria were found in five (13.51 percent) samples. In vitro the S. aureus protease increased the influenza HA titer after contact for 30 min at 25 ºC. Results revealed the occurrence of co-infection with proteolytic bacteria and influenza in the evaluated individuals. This finding corroborates that virus versus bacteria synergism could be able to potentiate respiratory infection, increasing damage to hosts.

O papel da bactéria proteolítica na exacerbação do vírus influenza tem sido demonstrado em hospedeiros naturais como porcos e humanos. Foram coletadas 407 amostras do trato respiratório de indivíduos apresentando manifestações clínicas, durante a estação da influenza (2003-2005) na cidade de São Paulo. Este trabalho teve como objetivo avaliar a incidência de determinadas bactérias que junto com vírus co-infectarem o trato respiratório humano. Testes bacteriológicos, e virológicos como imunofluorescência (IF), RT/PCR e hemaglutinação (HA) foram usados nas investigações viral e bacteriana. Pelo teste de IF foram selecionadas trinta e sete (9,09 por cento) amostras positivas para o vírus influenza. A presença do vírus influenza foi confirmada pela técnica de RT/PCR. Pelos testes bacteriológicos e do agar caseina, verificou-se que 18 (48,64 por cento) dos indivíduos foram infectados com bactérias proteolíticas tais como Staphylococcus spp., Streptococcus spp. e Pseudomonas spp. Destas amostras, 13 (35,13 por cento) foram co-infectadas com vírus influenza tipo A, e 5 (13,51 por cento) com influenza tipo B. No experimento in vitro com influenza e S. aureus, detectou-se aumento do título hemaglutinante deste vírus, após contacto de 30 min a 25 ºC. Os resultados obtidos revelaram a ocorrência de co-infecção com bactéria proteolítica e vírus influenza nos indivíduos avaliados. Estes achados corroboram com a investigação do sinergismo, entre bactéria e vírus, que poderia ser capaz de potencializar infecção respiratória, aumentando os riscos aos hospedeiros.

Co-infection between influenza virus and flagellated bacteria.

Trypsin is required in the hemagglutinin (HA) cleavage to in vitro influenza viruses activation. This HA cleavage is necessary for virus cell entry by receptor-mediated endocytosis. Bacteria in the respiratory tract are potential sources of proteases that could contribute to the cleavage of influenza virus in vivo. From 47 samples collected from horses, pigs, and from humans, influenza presence was confirmed in 13 and these samples demonstrated co-infection of influenza with flagellated bacteria, Stenotrophomonas maltophilia from the beginning of the experiments. Despite treatment with antibiotics, the bacteria remained resistant in several of the co-infected samples (48.39%). These bacteria, considered opportunistic invaders from environmental sources, are associated with viral infections in upper respiratory tract of hosts. The protease (elastase), secreted by Stenotrophomonas maltophilia plays a role in the potentiation of influenza virus infection. Proteolytic activity was detected by casein agar test. Positive samples from animals and humans had either a potentiated influenza infectivity or cytopathic effect (CPE) in MDCK and NCI H292 cells, Stenotrophomonas maltophilia were always present. Virus and bacteria were observed ultrastructurally. These in vitro findings show that microbial proteases could contribute to respiratory complications by host protease activity increasing inflammation or destroying endogenous cell protease inhibitors.

Co-infection between influenza virus and flagellated bacteria

Tripsina é necessária na ativação da clivagem do vírus influenza A in vitro. Esta clivagem é importante para entrada do vírus na célula por endocitose mediada pelo receptor celular. Bactérias presentes no trato respiratório são fontes de proteases que podem contribuir na replicação do vírus influenza in vivo. Entre 47 amostras coletadas de cavalos, suínos e humanos, a influenza foi isolada e confirmada em 13 que estavam co-infectadas com bactéria flagelada: Stenotrophomonas maltophilia desde o início destes experimentos. Apesar do tratamento das amostras com antibióticos, as bactérias resistiram em diversas delas (48.39%). A protease (elastase), secretada pela Stenotrophomonas maltophilia, desenvolveu papel decisivo na potencialização da infecção pelo vírus influenza. Essa atividade proteolítica foi detectada pelo teste de ágar-caseína. Amostras positivas para o vírus influenza isolado em animais, bem como em humanos tiveram potencialização da infectividade (ECP) em células MDCK e NCI-H292, sempre que a Stenotrophomonas maltophilia esteve presente. Os referidos microorganismos, bactéria e vírus foram observados ultra-estruturalmente. Esses achados in vitro demonstram como complicações respiratórias podem ocorrer in vivo, através da contribuição de protease microbiana, provocando aumento da inflamação ou destruição dos inibidores celulares de proteases endógenas, nos hospedeiros susceptíveis à influenza.

Investigation of influenza in migrating birds, the primordial reservoir and transmitters of influenza in Brazil

Os mais importantes reservatórios do vírus influenza são os pássaros. A manutenção do vírus influenza em hospedeiros naturais, inclusive o homem, permite que esse vírus realize rearranjos entre as suas cepas. O recente relato de uma cepa influenza aviária A(H5N1), em humanos, se deu em uma criança com doença respiratória fatal, na China em 1977. O presente estudo foi conduzido para elucidar o transporte da influenza por pássaros que migram, anualmente, através de ambos hemisférios o do Norte e do Sul, com especial atenção voltada à espécies Vireo olivaceo [Juruviara(BR) e Red-eyed vireo(USA)] que viaja do USA para o Brasil, e vice-versa, e a espécie Elaenia mesoleuca [Tuque(BR) e (USA)] que voa por todo o Hemisfério Sul. Essas espécies de pássaros, que residem e migram em São Paulo, e que demonstram transportar o vírus influenza, foram selecionadas. As partículas virais isoladas foram observadas por microscópio eletrônico. O vírus influenza foi detectado pelos testes: House Duplex/PCR e Gloria. Os resultados revelam que os pássaros das espécies: Elaenia mesoleuca e Vireo olivaceus são transportes do vírus influenza enquanto cruzam ambos Hemisférios. Para o conhecimento da função que os pássaros migratórios podem desempenhar na epidemia de influenza, no Brasil, caracterização dos subtipos deste vírus estão sendo realizados.

Anti-human influenza protector antibody detected in horses as a zoonotic viruses / Anticorpo protetor anti influenza humana detectado em cavalos, como virose zoonótica

This study aimed to investigate the incidence of the influenza virus, and its interspecies transmission cycle among horses. A comparative serological survey was performed using horse sera following challenge with both specific (equine) and non-specific (human) influenza virus strains. Bleedings of the 22 horses were performed during the years 1999 and 2000. Following treatment with kaolin (20%), added in rooster erythrocytes suspension (50%), for removal of non-specific antibodies, sera were titered against both Human and Equine Influenza virus, by the Hemagglutination Inhibition Assay(HI), recommended by WHO. The HI results of horse serological responses demonstrate cross reaction between both the specific strain, A/Eq1 (H7N7) ( 62.75%) and A/Eq2 (H3N8) (60.65%), and the non- specific strains, type A (H1N1) (79.5%) and A (H3N2) (94.45%) and type B (77.75%). It was noteworthy the high percentage of protection responses in equine sera aginst the non-specific strains, as compared with the specific strains. This finding suggests direct interspecies transmission of influenza virus as zoonotic viruses, particulary for the type B strain which is considered restricted to humans. It was the first report, in Brazil. Further studies are required to achieve a complete understanding of the incidence of influenza in our environment. (AU)

O objetivo deste estudo foi investigar em cavalos, a incidência do vírus influenza e seu ciclo de transmissão interespécies. Portanto, levantamento sorológico foi realizado em soro de cavalos, confrontados com ambas cepas, as específicas (eqüino) e não específicas (humana) deste vírus. Sangrias de cavalos realizadas nos anos de 1999 e de 2000, forneceram soros que, após tratamento com Caolim (20%) e hemácias de galo(50%) para remoção dos anticorpos inespecíficos, foram titulados contra ambas referidas cepas, através do teste de Inibição da Hemaglutinação (recomendado pela OMS). Os resultados, demonstraram que as respostas sorológicas dos cavalos apresentaram reação cruzada entre as cepas específicas e as não específicas. As porcentagens de títulos IH obtidos foram de 62,75% e de 60,65% para as cepas específicas A/Eq1 (H7N7) e A/Eq2 (H3N8), respectivamente. E às cepas não específicas essas porcentagens foram de: 79,05% para A (H1N1), de 94,45% para A (H3N2) e de 77,75% ao tipo B. O mais relevante nestes dados comparativos com vírus influenza, foi a alta porcentagem de resposta protetora à cepa não específica comparada àquela específica, detectada nos soros eqüinos. Considerando o fato de que o tipo B, deste vírus, ser restrito à espécie humana, portanto a resposta de proteção nos cavalos sugere uma direta transmissão interspécies, como em viroses zoonóticas. Os autores relatam pela primeira vez este tipo de evento no Brasil. (AU)

Anti-human influenza protector antibody detected in horses as a zoonotic viruses

O objetivo deste estudo foi investigar em cavalos, a incidência do vírus influenza e seu ciclo de transmissão interespécies. Portanto, levantamento sorológico foi realizado em soro de cavalos, confrontados com ambas cepas, as específicas (eqüino) e não específicas (humana) deste vírus. Sangrias de cavalos realizadas nos anos de 1999 e de 2000, forneceram soros que, após tratamento com Caolim (20%) e hemácias de galo(50%) para remoção dos anticorpos inespecíficos, foram titulados contra ambas referidas cepas, através do teste de Inibição da Hemaglutinação (recomendado pela OMS). Os resultados, demonstraram que as respostas sorológicas dos cavalos apresentaram reação cruzada entre as cepas específicas e as não específicas. As porcentagens de títulos IH obtidos foram de 62,75% e de 60,65% para as cepas específicas A/Eq1 (H7N7) e A/Eq2 (H3N8), respectivamente. E às cepas não específicas essas porcentagens foram de: 79,05% para A (H1N1), de 94,45% para A (H3N2) e de 77,75% ao tipo B. O mais relevante nestes dados comparativos com vírus influenza, foi a alta porcentagem de resposta protetora à cepa não específica comparada àquela específica, detectada nos soros eqüinos. Considerando o fato de que o tipo B, deste vírus, ser restrito à espécie humana, portanto a resposta de proteção nos cavalos sugere uma direta transmissão interspécies, como em viroses zoonóticas. Os autores relatam pela primeira vez este tipo de evento no Brasil.

[Influenza in heterothermic animals]. / Influenza em animais heterotérmicos.

The objective was to study Orthomyxovirus in heterothermic animals. Blood samples from snakes (genus Bothrops and Crotalus) and from toads and frogs (genus Bufo and Rana) were collected to evaluate the red cell receptors and antibodies specific to influenza virus by the hemagglutination and hemagglutination inhibition tests, respectively. Both snakes and toads kept in captivity presented receptors in their red cells and antibodies specific to either influenza virus type A (human and equine origin) or influenza type B. The same was observed with recently captured snakes. Concerning the influenza hemagglutination inhibition antibodies protective levels were observed in the reptiles' serum, against influenza type A and type B. Unlike the toads, 83.3% of the frogs presented mean levels of Ab 40HIU for some influenza strains. It was concluded that heterothermic animals could offer host conditions to the influenza virus and also susceptibility to the infection.

Influenza em animais heterotérmicos / Influenza in heterothermics

O objetivo foi pesquisar Ortomyxovirus em animais heterotérmicos. Coletou-se sangue de serpentes dos gêneros Bothrops e Crotalus e de sapo e rãs dos gêneros Bufo e Rana, para a detecção dos receptores de hemácias e anticorpos específicos, ao vírus influenza, pelos testes de hemaglutinação e inibição da hemaglutinação, respectivamente. Pelo teste de hemaglutinação, verificou-se que serpentes e sapos em cativeiro apresentaram receptores em suas hemácias para o vírus influenza, humano e eqüino do tipo A e tipo B. O mesmo ocorreu com serpentes recém chegadas. Quanto ao teste de inibição da hemaglutinação dos soros dos répteis observou-se títulos protetores de anticorpos aos vírus influenza tipo A (origens humana e eqüina) e tipo B. Com soro de sapo não se observou reação de inibição da hemaglutinação porém, 83,3 por cento das rãs obtiveram médias de 40UIH para algumas cepas. Conclui-se que animais heterotérmicos podem oferecer condições de hospedeiros aos vírus influenza, assim como susceptibilidade à infecção.