Antibacterial activity of phytopathogenic Streptomyces strains against bacteria associated to clinical diseases / Atividade antimicrobiana de Streptomyces fitopatogênicas contra bactérias associadas a doenças de importância clínica

The genus Streptomyces is associated with the ability to produce and excrete a variety of bioactive compounds, such as antibiotic, antifungal and antiviral. Biological active polyketide and peptide compounds with applications in medicine, agriculture and biochemical research are synthesized by PKS-I and NRPS genes. The evaluation of the presence of these genes associated with the biosynthesis of secondary metabolites in different phytopathogenic Streptomyces strains were performed using degenerated primers. The positive signal was observed in 58/63 Streptomyces strains for NRPS gene, 43/63 for PKS-I, and for PKS-II all the 63 strains showed positive signal of amplification. These strains also were tested with double layer agar-well technique against bacterial with clinical importance, and it was possible to observe the Streptomyces spp. strains were able to inhibit the growth of 14, 20, 13 and 3 isolates Gram-positive and Gram-negative bacteria, Staphylococcus aureus (ATCC 25923), Bacillus cereus (ATCC 14579), Pseudomonas aeruginosa (ATCC 27853) and Escherichia coli (ATCC 11775) respectively. The Streptomyces sp. strains IBSBF 2019 and IBSBF 2397 showed antibacterial activity against all four bacteria-target tested.(AU)

O gênero Streptomyces apresenta alta capacidade de produzir e excretar uma grande variedade de compostos biologicamente ativos, como antibióticos, antifúngicos e antivirais. Compostos biologicamente ativos de policetídeos e peptídeos com aplicações na medicina, agricultura e pesquisas bioquímicas são sintetizados pelos genes PKS-I e NRPS. A avaliação da presença desses genes associados à biossíntese de metabólitos secundários em diferentes linhagens de Streptomyces fitopatogênicas foi realizada através do uso de primers degenerados. O sinal positivo foi observado em 58/63 linhagens de Streptomyces para o gene NRPS, 43/63 para o gene PKS-I e, para o gene PKS-II, todas as 63 linhagens apesentaram o sinal positivo de amplificação. Essas linhagens também foram testadas através da técnica de dupla camada contra bactérias de importância clínica e foi possível observar que as linhagens de Streptomyces spp. foram capazes de inibir o crescimento de 14, 20, 13 e 3 isolados de bactérias Gram-positivas e Gram-negativas, Staphylococcus aureus (ATCC 25923), Bacillus cereus (ATCC 14579), Pseudomonas aeruginosa (ATCC 27853) e Escherichia coli (ATCC 11775), respectivamente. As linhagens de Streptomyces sp. ISBSF 2019 e 2397 apresentaram atividade antibacteriana contra todas as bactérias-alvo testadas.(AU)

Antibacterial activity of phytopathogenic Streptomyces strains against bacteria associated to clinical diseases / Atividade antimicrobiana de Streptomyces fitopatogênicas contra bactérias associadas a doenças de importância clínica

The genus Streptomyces is associated with the ability to produce and excrete a variety of bioactive compounds, such as antibiotic, antifungal and antiviral. Biological active polyketide and peptide compounds with applications in medicine, agriculture and biochemical research are synthesized by PKS-I and NRPS genes. The evaluation of the presence of these genes associated with the biosynthesis of secondary metabolites in different phytopathogenic Streptomyces strains were performed using degenerated primers. The positive signal was observed in 58/63 Streptomyces strains for NRPS gene, 43/63 for PKS-I, and for PKS-II all the 63 strains showed positive signal of amplification. These strains also were tested with double layer agar-well technique against bacterial with clinical importance, and it was possible to observe the Streptomyces spp. strains were able to inhibit the growth of 14, 20, 13 and 3 isolates Gram-positive and Gram-negative bacteria, Staphylococcus aureus (ATCC 25923), Bacillus cereus (ATCC 14579), Pseudomonas aeruginosa (ATCC 27853) and Escherichia coli (ATCC 11775) respectively. The Streptomyces sp. strains IBSBF 2019 and IBSBF 2397 showed antibacterial activity against all four bacteria-target tested.(AU)

O gênero Streptomyces apresenta alta capacidade de produzir e excretar uma grande variedade de compostos biologicamente ativos, como antibióticos, antifúngicos e antivirais. Compostos biologicamente ativos de policetídeos e peptídeos com aplicações na medicina, agricultura e pesquisas bioquímicas são sintetizados pelos genes PKS-I e NRPS. A avaliação da presença desses genes associados à biossíntese de metabólitos secundários em diferentes linhagens de Streptomyces fitopatogênicas foi realizada através do uso de primers degenerados. O sinal positivo foi observado em 58/63 linhagens de Streptomyces para o gene NRPS, 43/63 para o gene PKS-I e, para o gene PKS-II, todas as 63 linhagens apesentaram o sinal positivo de amplificação. Essas linhagens também foram testadas através da técnica de dupla camada contra bactérias de importância clínica e foi possível observar que as linhagens de Streptomyces spp. foram capazes de inibir o crescimento de 14, 20, 13 e 3 isolados de bactérias Gram-positivas e Gram-negativas, Staphylococcus aureus (ATCC 25923), Bacillus cereus (ATCC 14579), Pseudomonas aeruginosa (ATCC 27853) e Escherichia coli (ATCC 11775), respectivamente. As linhagens de Streptomyces sp. ISBSF 2019 e 2397 apresentaram atividade antibacteriana contra todas as bactérias-alvo testadas.(AU)

Antimicrobial activity of enterocin obtained from Enterococcus durans on Shiga-like toxin-producing Escherichia coli / Atividade antimicrobiana de enterocina produzida por Enterococcus durans sobre Escherichia coli produtora de toxina Shiga-like

Shiga-like toxin-producing Escherichia coli (STEC) is an important source of food contamination that presents risks to human health. Several industrial food processes eliminate this microorganism; however, these processes can alter the characteristics of the product. Alternative methods of preservation have been identified as an option to control these foodborne pathogens. The purpose of this study was to evaluate the action of bacteriocins produced by Enterococcus durans MF5 in STEC cells. Cell-free supernatant (CFS) containing enterocins from the MF5 isolate was tested over different time points (6, 18, and 24 h). Enterocins present in the crude CFS showed inhibition against STEC at all time points. In the investigation of cell integrity, using propidium iodide and fluorescence microscopy, considerable cell death was observed within 6 h of the cells being in contact with the enterocins, which was also observed at the 18 and 24 h time points. These results showed that the enterocins produced by the MF5 isolate have potential use in the control of STEC.(AU)

Escherichia coli, produtora de toxina Shiga-like (STEC), apresenta riscos à saúde humana, constituindo uma importante fonte de contaminação na indústria de alimentos. Diversos processos industriais eliminam esse microrganismo, contudo podem alterar as características do produto. Métodos alternativos de conservação tem sido uma opção para controlar esse microrganismo de alimentos. O objetivo desta pesquisa foi avaliar a ação de bacteriocinas produzidas por Enterococcus durans MF5 em células de E. coli STEC. Foram utilizados sobrenadante livre de células (CFS) contendo enterocina, nos tempos 6, 18 e 24 horas de incubação. A enterocina presente no CFS bruto apresentou inibição contra E. coli STEC em todos os tempos testados. Na observação da integridade celular utilizando iodeto de propídio e observação em microscópio de fluorescência, observou-se que em 6h da célula em contato com a enterocina, já havia considerável morte celular, estendendo até os tempos de 18 e 24 horas. Os resultados obtidos mostraram que a enterocina produzida pelo isolado MF5 apresenta uso potencial no controle de E. coli STEC.(AU)

Co-supplementation of dietary seaweed powder and antibacterial peptides improves broiler growth performance and immune function

The feasibility of Laminaria japonica powder (LJP) combined with cecropin as a dietary supplement to enhance broiler growth performance and immune function was evaluated in this study. In total, 648 one-day-old Arbor Acres broiler chicks were randomly distributed into nine numerically-equal treatment groups: T1 (control group; fed a basal diet); T2 (fed the basal diet supplemented with 1% LJP);T3 (fed the basal diet supplemented with 300mg cecropin/kg); and T4,T5,T6,T7,T8 and T9, individually fed with the dietary supplemented with varying levels of LJP and cecropin). Compared with the control, dietary of LJP or cecropin supplementation slightly improved feed conversion ratio (FCR). However, the dietary supplementation of LJP combined with cecropin significantly improved broiler growth performance during the periods of 1-21,21-42, and 1-42 days (p<0.05).The dietary supplementation of 3% LJP combined with 300 mg/kg cecropin significantly increased FCR, and serum Newcastle disease antibody titers and lymphocyte numbers during the periods of 1-21, 21-42, and 1-42 days (p<0.05). Cecal microorganisms were cultivated and the number of Escherichia coli and Lactobacillus colonies were counted. The dietary supplementation of LJP combined with cecropin remarkably inhibited E. coli growth and increased Lactobacillus growth. The results of this study demonstrate the feasibility of using LJP and cecropin as feed supplement for improving the growth performance and enhancing the immune function of broilers.(AU)

Co-supplementation of dietary seaweed powder and antibacterial peptides improves broiler growth performance and immune function

The feasibility of Laminaria japonica powder (LJP) combined with cecropin as a dietary supplement to enhance broiler growth performance and immune function was evaluated in this study. In total, 648 one-day-old Arbor Acres broiler chicks were randomly distributed into nine numerically-equal treatment groups: T1 (control group; fed a basal diet); T2 (fed the basal diet supplemented with 1% LJP);T3 (fed the basal diet supplemented with 300mg cecropin/kg); and T4,T5,T6,T7,T8 and T9, individually fed with the dietary supplemented with varying levels of LJP and cecropin). Compared with the control, dietary of LJP or cecropin supplementation slightly improved feed conversion ratio (FCR). However, the dietary supplementation of LJP combined with cecropin significantly improved broiler growth performance during the periods of 1-21,21-42, and 1-42 days (p<0.05).The dietary supplementation of 3% LJP combined with 300 mg/kg cecropin significantly increased FCR, and serum Newcastle disease antibody titers and lymphocyte numbers during the periods of 1-21, 21-42, and 1-42 days (p<0.05). Cecal microorganisms were cultivated and the number of Escherichia coli and Lactobacillus colonies were counted. The dietary supplementation of LJP combined with cecropin remarkably inhibited E. coli growth and increased Lactobacillus growth. The results of this study demonstrate the feasibility of using LJP and cecropin as feed supplement for improving the growth performance and enhancing the immune function of broilers.

Effect of ultrasound on survival and growth of Escherichia coli in cactus pear juice during storage

The aim of this study was to investigate the effectiveness of ultrasound as a conservation method for the inactivation of Escherichia coli inoculated into cactus pear juices (green and purple). Total soluble solids, pH, titratable acidity, and the kinetics of E. coli in cactus pear juices treated by ultrasound (60%, 70%, 80% and 90% amplitude levels for 1, 3 and 5 min) were evaluated over 5 days. Total inactivation was observed in both fruit juices after 5 min of ultrasound treatment at most amplitude levels (with the exception of 60% and 80%). After one and two days of storage, the recovery of bacteria counts was observed in all cactus pear juices. Ultrasound treatment at 90% amplitude for 5 min resulted in non-detectable levels of E. coli in cactus pear juice for 2 days. The parameters of pH, titratable acidity and soluble solids were unaffected.(AU)

The importance of protozoan bacterivory in a subtropical environment (Lobo-Broa Reservoir, SP, Brazil) / A importância da bacterivoria por protozoários em um ambiente subtropical (Reservatório do Lobo-Broa, SP, Brasil)

This study evaluated the importance of heterotrophic nanoflagellates (HNF) and ciliates bacterivory in a mesotrophic subtropical environment (Lobo-Broa Reservoir, Brazil) by the quantification of their ingestion rates. The in situ experiments using fluorescently labelled bacteria (FLB) were carried out bimonthly over one year (three surveys in the dry season and three in the rainy one) at the sub-surface of two sampling points that have different trophic degrees. The ingestion rates for both ciliates and HNF were higher in the meso-eutrophic region (point 2) due to the higher water temperatures, which accelerate the metabolism of protozoans and the higher bacteria densities. Concerning total protozoan bacterivory, the HNF had the greatest grazing impact on bacterial community, especially the HNF <5µm. The data showed that HNF grazing, in addition to regulating the bacteria abundance, also induced changes to the bacterial community structure, such as increasing size and numbers of bacterial filaments. The ciliates were also important to the system bacterivory, especially in point 2, where there were high densities and ingestion rates. The protozoan bottom-up control was more important in the dry season and the top-down control was more important in the rainy season, so, these two forces are equally important to the bacterial abundance regulation in this reservoir in an annual basis.(AU)

Este estudo avaliou a importância da bacterivoria por nanoflagelados heterotróficos (NFH) e ciliados em um ambiente subtropical mesotrófico (Reservatório do Lobo-Broa, Brasil) pela quantificação de suas taxas de ingestão. Os experimentos in situ utilizando bactérias marcadas por fluorescência (FLB) foram realizados bimestralmente durante um ano (três estudos no período seco e três no período chuvoso) na superfície de dois pontos de coleta que apresentam graus de trofia diferentes. As taxas de ingestão, tanto para os ciliados quanto para os NHF, foram maiores no ponto 2 (P2) devido provavelmente as maiores temperaturas da água, que aceleram o metabolismo dos protozoários, e as maiores densidades de bactérias. Em relação à bacterivoria total por protozoários, as populações de NFH causaram maior impacto de predação sobre a comunidade bacteriana do reservatório, principalmente os NFH menores que 5µm. Os dados indicaram que a predação por NFH, além de regular a abundância de bactérias, provavelmente também induziu modificações na estrutura da comunidade bacteriana, como o aumento do tamanho e da quantidade de filamentos bacterianos. Os ciliados também contribuíram significativamente para a bacterivoria no sistema, principalmente no P2, onde apresentaram altas densidades e taxas de ingestão. O controle bottom-up teve maior importância no período seco e o controle top-down maior importância no período chuvoso, assim, estas duas forças são igualmente importantes na regulação da abundância bacteriana neste reservatório em uma base anual.(AU)

Regulation of gene expression: Cryptic beta-glucoside (bgl) operon of Escherichia coli as a paradigm

Bacteria have evolved various mechanisms to extract utilizable substrates from available resources and consequently acquire fitness advantage over competitors. One of the strategies is the exploitation of cryptic cellular functions encoded by genetic systems that are silent under laboratory conditions, such as the bgl (β-glucoside) operon of E. coli. The bgl operon of Escherichia coli, involved in the uptake and utilization of aromatic β-glucosides salicin and arbutin, is maintained in a silent state in the wild type organism by the presence of structural elements in the regulatory region. This operon can be activated by mutations that disrupt these negative elements. The fact that the silent bgl operon is retained without accumulating deleterious mutations seems paradoxical from an evolutionary view point. Although this operon appears to be silent, specific physiological conditions might be able to regulate its expression and/or the operon might be carrying out function(s) apart from the utilization of aromatic β-glucosides. This is consistent with the observations that the activated operon confers a Growth Advantage in Stationary Phase (GASP) phenotype to Bgl+ cells and exerts its regulation on at least twelve downstream target genes.

Toxicity of herbicides on Escherichia coli growth / Toxicidade de herbicidas sobre o crescimento de Escherichia coli

Agriculture uses a huge variety and quantity of chemicals. If, on one hand, the goal is to increase productivity, on the other hand these products contaminate aquatic environments. Among these products, herbicides deserve greater attention in relation to contamination of aquatic environments due to their extensive use to weed control. This study was carried out because the effects of these molecules on aquatic microorganisms such as Escherichia coli, is still unclear. Using microdilution plate assays, Escherichia coli were exposed to various commercial formulations of herbicides widely used in Brazil. The herbicide paraquat was the only one able to prevent the growth of Escherichia coli and is characterized as bacteriostatic.

A agricultura utiliza uma enorme variedade de produtos químicos, e se por um lado a meta é aumentar a produtividade, por outro lado estes produtos contaminam ambientes aquáticos. Entre estes produtos, herbicidas merecem maior atenção em relação à contaminação de ecossistemas aquáticos, devido a seu amplo emprego para controlar plantas daninhas. Este estudo foi realizado porque os efeitos destas moléculas em micro-organismos aquáticos, tais como Escherichia coli, ainda são pouco estudados. Através de ensaios de microdiluição, E. coli foi exposta a várias formulações comerciais de herbicidas amplamente utilizados no Brasil e foi verificado que o herbicida paraquat foi o único capaz de impedir o crescimento de E. coli, caracterizando-se assim como bacteriostático.

Dissemination of bacteria labeled with technetium-99m after laparotomy and abdominal insufflation with different CO2 pressures on rats / Disseminação de bactérias marcadas com tecnécio-99m após laparotomia e insuflação com diferentes pressões de CO2 em ratos

PURPOSE: To asses the dissemination of bacteria labeled with technetium-99m (99mTc) from peritoneal cavity after different surgical procedures. METHODS: Bacteria of the Escherichia coli species labeled with 99mTc were used in a concentration of 10(8) units of colony-makers for ml (UFC/ml) and 1ml was inoculated through intra-peritoneal via. Forty-eight rats were divided into four groups: control, laparotomy, pneumoperitoneum with 10mmHg and pneumoperitoneum with 20mmHg of CO2. Procedures were performed 20 min after injection of the inoculum and lasted 30 min. Animals were sacrificed after six hours (Group 1) and 24 hours (Group 2). Samples of blood, liver and spleen were collected for radioactivity counting. RESULTS: After six hours, indirect detection of the bacteria in different organs was uniform in all groups. After 24 hours, a larger detection of technetium was observed in the livers of animals of the group insufflated with 20mmHg of CO2, when compared with those of control group (p<0.01). The other groups did not present statistically significant variations. CONCLUSIONS: The use of a higher intra-abdominal pressure was associated with a higher bacterial dissemination to the liver. The application of lower intra-abdominal pressures may be associated with a lower dissemination of the infectious status during laparoscopic approach of peritonitis status.(AU)

OBJETIVO: Avaliar a disseminação de bactérias marcadas com tecnécio-99m (99mTc) a partir da cavidade peritoneal após diferentes procedimentos cirúrgicos. MÉTODOS: Foram utilizadas bactérias da espécie Escherichia coli marcadas com 99mTc em uma concentração de10(8) unidades formadoras de colônia por ml (UFC/ml) sendo inoculado 1ml por via intra-peritoneal. Quarenta e oito ratos foram divididos em quatro grupos: controle, laparotomia, pneumoperitôneo com 10 mmHg e pneumoperitôneo com 20 mmHg de CO2. Os procedimentos foram realizados 20 minutos após a injeção do inóculo e duraram 30 minutos. Os animais foram sacrificados após seis horas (grupo 1) e 24 horas (grupo 2). Foram coletadas amostras de sangue, fígado e baço para contagem radioativa. RESULTADOS: Após seis horas, a detecção indireta das bactérias nos diferentes órgãos foi uniforme em todos os grupos. Após 24 horas, observou-se uma maior detecção de tecnécio nos fígados dos animais do grupo insuflado com 20 mmHg de CO2, quando comparados aos do grupo controle (p<0,01). Os outros grupos não apresentaram variações estatisticamente significativas. CONCLUSÕES: A utilização de pressões intra-abdominais mais elevadas associou-se a uma maior disseminação bacteriana para o fígado. A utilização de pressões intra-abdominais menos elevadas na abordagem da peritonite pode estar associada a uma menor disseminação do quadro infeccioso.(AU)

Inbred F344 rats as a biologic model of intra-abdominal sepsis / Ratos isogênicos F344 como modelo biológico de sepsis intra-abdominal

Aiming to design a biologic model of acute intra-abdominal sepsis for experimental studies, conventional, isogenic rats F344 were infected with the bacteria Escherichia coli (E.coli), strain 11775, serotype H7:01:K1. Six hours after inoculation with E. coli, the animals - males and females - showed the following symptoms: piloerection, hyperpnea and decreased motor activity. The lethal dosis (LD50), was 6 x 10(5) CFU/ml, analyzed in 32 males and 32 females. The highest mortality rate was observed on the first 24 hours. Liver dysfunction, common in intra-abdominal sepsis, was evaluated at 0, 24, 48 and 168 hours after inoculation, by means of serum enzyme activities. Study of the migration of polymorphonuclear-neutrophil cells (PMN) and mononuclear-macrophage ones (MN) showed a significant increase of PMN in E. coli inoculated males (z > 4.7; P<0.003) and females (z > 6.2; P < 0.0003), when compared to control groups. As for MN cells, there were no differences between inoculated and control groups, in males (z = 2.3; P = 0.0107) and in females (z = 1.8; P = 0.0359) as well. In conclusion, these results show that inbred F344 rats are adequate biologic models for studies of acute, intra-abdominal sepsis.(AU)

Com o objetivo de estudar um modelo biológico de sepsis intra-abdominal aguda para estudos experimentais, foram infectados ratos isogênicos F344, convencionais, com a bactéria Escherichia coli (E.coli), cepa ATCC 11775, sorotipo H7:O1:K1. Os animais inoculados, machos e fêmeas, apresentaram 6 horas após a inoculação por E.coli os seguintes sintomas: arqueamento do dorso, piloereção, hiperpnéia e diminuição das atividades motoras. A dose que produziu 50% de mortalidade (DL50) após 7 dias, determinada pelo método Reed & Muench, foi de 6 x 105 CFU/ml (analisado em 32 machos e 32 fêmeas). A maior concentração de mortalidade foi observada nas primeiras 24 horas. A disfunção hepática, comum em sepsis intra-abdominal, foi avaliada por provas enzimáticas, em 0, 24, 48 e 168 horas após a inoculação. O estudo da migração de células polimorfonucleares-neutrófilos (PMN) e mononucleares-macrófagos (MN) apontou um aumento significante de PMN entre o grupos de machos (z ³ 4,7; p < 0,003) e de fêmeas (z ³ 6,2; p < 0,0003) inoculados E.coli, quando comparados ao grupos controles. Quanto às células MN, não houve diferença entre os grupos inoculados e os controles, tanto para os machos (z=2,3; p = 0,0107), como para as fêmeas (z=1,8; p =0,0359). Em conclusão, estes resultados demonstram que os ratos isogênicos F344 são modelos biológicos adequados para estudos de sepsis intra-abdominal aguda.(AU)