Probiotic Bacillus amyloliquefaciens Strain H57 Improves the Performance of Pregnant and Lactating Ewes Fed a Diet Based on Palm Kernel Meal

Probiotic supplements are single or mixed strain cultures of live microorganisms that benefit the host by improving the properties of the indigenous microflora (Seo et al 2010). In a pilot study at the University of Queensland, Norton et al (2008) found that Bacillus amyloliquefaciens Strain H57 (H57), primarily investigated as an inoculum to make high-quality hay, improved feed intake and nitrogen utilisation over several weeks in pregnant ewes. The purpose of the following study was to further challenge the potential of H57 -to show it survives the steam-pelleting process, and that it improves the performance of ewes fed pellets based on an agro-industrial by-product with a reputation for poor palatability, palm kernel meal (PKM), (McNeill 2013). Thirty-two first-parity White Dorper ewes (day 37 of pregnancy, mean liveweight = 47.3 kg, mean age = 15 months) were inducted into individual pens in the animal house at the University of Queensland, Gatton. They were adjusted onto PKM-based pellets (g/kg drymatter (DM): PKM, 408; sorghum, 430; chick pea hulls, 103; minerals and vitamins; Crude protein, 128; ME: 11.1MJ/kg DM) until day 89 of pregnancy and thereafter fed a predominately pelleted diet incorporating with or without H57 spores (10 9 colony forming units (cfu)/kg pellet, as fed), plus 100g/ewe/day oaten chaff, until day 7 of lactation. From day 7 to 20 of lactation the pelleted component of the diet was steadily reduced to be replaced by a 50:50 mix of lucerne: oaten chaff, fed ad libitum, plus 100g/ewe/day of ground sorghum grain with or without H57 (10 9 cfu/ewe/day). The period of adjustment in pregnancy (day 37-89) extended beyond expectations due to some evidence of mild ruminal acidosis after some initially high intakes that were followed by low intakes. During that time the diet was modified, in an attempt to improve palatability, by the addition of oaten chaff and the removal of an acidifying agent (NH4Cl) that was added initially to reduce the risk of urinary calculi. Eight ewes were removed due to inappetence, leaving 24 ewes to start the trial at day 90 of pregnancy. From day 90 of pregnancy until day 63 of lactation, liveweights of the ewes and their lambs were determined weekly and at parturition. Feed intakes of the ewes were determined weekly. Once lambing began, 1 ewe was removed as it gave birth to twin lambs (whereas the rest gave birth to a single lamb), 4 due to the loss of their lambs (2 to dystocia), and 1 due to copper toxicity. The PKM pellets were suspected to be the cause of the copper toxicity and so were removed in early lactation. Hence, the final statistical analysis using STATISTICA 8 (Repeated measures ANOVA for feed intake, One-way ANOVA for liveweight change and birth weight) was completed on 23 ewes for the pregnancy period (n = 11 fed H57; n = 12 control), and 18 ewes or lambs for the lactation period (n = 8 fed H57; n = 10 control). From day 90 of pregnancy until parturition the H57 supplemented ewes ate 17 more DM (g/day: 1041 vs 889, sed = 42.4, P = 0.04) and gained more liveweight (g/day: 193 vs 24.0, sed = 25.4, P = 0.0002), but produced lambs with a similar birthweight (kg: 4.18 vs 3.99, sed = 0.19, P = 0.54). Over the 63 days of lactation the H57 ewes ate similar amounts of DM but grew slower than the control ewes (g/day: 1.5 vs 97.0, sed = 21.7, P = 0.012). The lambs of the H57 ewes grew faster than those of the control ewes for the first 21 days of lactation (g/day: 356 vs 265, sed = 16.5, P = 0.006). These data support the findings of Norton et al (2008) and Kritas et al (2006) that certain Bacillus spp. supplements can improve the performance of pregnant and lactating ewes. In the current study we particularly highlighted the capacity of H57 to stimulate immature ewes to continue to grow maternal tissue through pregnancy, possibly through an enhanced appetite, which appeared then to stimulate a greater capacity to partition nutrients to their lambs through milk, at least for the first few weeks of lactation, a critical time for optimising lamb survival. To conclude, H57 can survive the steam pelleting process to improve feed intake and maternal liveweight gain in late pregnancy, and performance in early lactation, of first-parity ewes fed a diet based on PKM.

Caracterização de um peptídeo antimicrobiano produzido por linhagem de Bacillus amyloliquefaciens isolada de solo

Em anos recentes, surgiram numerosos casos de intoxicação alimentar envolvendo patógenos emergentes. Estes casos levaram a um aumento da preocupação com a preservação dos alimentos minimamente processados e com a segurança alimentar. Este fato está induzindo a pesquisa por inibidores para estes patógenos e fatores para prolongar a vida de prateleira de produtos alimentícios. Entre as novas alternativas na preservação está a utilização de peptídeos antimicrobianos produzidos por bactérias. No presente trabalho uma bactéria identificada como Bacillus amyloliquefaciens LBM 5006 isolada de solos de mata Atlântica de Santa Catarina foi selecionada dentre outros microrganismos e sua capacidade de produzir antimicrobianos foi avaliada. O extrato bruto da cultura do isolado LBM 5006 foi caracterizado, sendo ativo contra importantes bactérias patogênicas e deteriorantes como Listeria monocytogenes, Bacillus cereus, Erwinia carotovora, Escherichia coli, dentre outras. Houve maior produção do antimicrobiano quando a bactéria foi propagada em caldo infusão de cérebro e coração (BHI) a 37o C durante 48 h. Após concentração, a atividade antimicrobiana resistiu ao tratamento com enzimas proteolíticas. A atividade antimicrobiana foi verificada em pHs ácidos, sendo inibida em pH 9 e 10. O extrato foi purificado por meio de cromatografia de gel filtração e extração com butanol. O teste qualitativo de ninidrina, juntamente com a espectroscopia de infravermelho e ultravioleta, feitos com a substância purificada revelou que o antimicrobiano possui natureza protéica. O antimicrobiano apresentou um efeito bacteriostático contra 106 UFC/mL de Listeria monocytogenes na concentração de 25 AU/ml.

Keratinase production by new strain of Bacillus Amyloliquefaciens: Application of statistical experimental design for optimization of keratinase production

Feathers are rich in amino acids and can be employed as a dietary protein supplement for animal feed. Microbial degradation is an alternative technology for improving the nutritional value of feathers. Other potential applications of keratinase include use in the leather industry, detergents and medicine as well as the pharmaceutical for the treatment of acne, psoriasis and calluses. A new keratinolytic enzyme production bacterium was isolated from a poultry processing plant. To improve keratinase yield, statistically based experimental designs were applied to optimize three significant variables: temperature, substrate concentration (feathers) and agitation speed. Response surface methodology demonstrated an increase in keratinolytic activity at temperature, agitation speed and substrate concentration of 26.6°C, 150 rpm and 2%, respectively. Liquid chromatography revealed the release of amino acids in the Bacillus amyloliquefaciens culture broth, thereby demonstrating the potential of feather meal in the animal feed industry. © Global Science Publications.

Otimização da produção de queratinase por Bacillus amyloliquefaciens

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Efeito probiótico do Bacillus amyloliquefaciens no desempenho produtivo e nos parâmetros hematológicos, morfométricos e ultraestruturais do intestino de tilápias-do-Nilo cultivadas em tanque-rede

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Ultra high pressure homogenization (UHPH) inactivation of Bacillus amyloliquefaciens spores in phosphate buffered saline (PBS) and milk

Ultra high pressure homogenization (UHPH) opens up new areas for dynamic high pressure assisted thermal sterilization of liquids. Bacillus amyloliquefaciens spores are resistant to high isostatic pressure and temperature and were suggested as potential surrogate for high pressure thermal sterilization validation. B. amyloliquefaciens spores suspended in PBS buffer (0.01 M, pH 7.0), low fat milk (1.5%, pH 6.7), and whole milk (3.5%, pH 6.7) at initial concentration of similar to 10(6) CFU/mL were subjected to UHPH treatments at 200, 300, and 350 MPa with an inlet temperature at similar to 80 degrees C. Thermal inactivation kinetics of B. amyloliquefaciens spores in PBS and milk were assessed with thin wall glass capillaries and modeled using first-order and Weibull models. The residence time during UHPH treatments was estimated to determine the contribution of temperature to spore inactivation by UHPH. No sublethal injury was detected after UHPH treatments using sodium chloride as selective component in the nutrient agar medium. The inactivation profiles of spores in PBS buffer and milk were compared and fat provided no clear protective effect for spores against treatments. Treatment at 200 MPa with valve temperatures lower than 125 degrees C caused no reduction of spores. A reduction of 3.5 log(10)CFU/mL of B. amyloliquefaciens spores was achieved by treatment at 350 MPa with a valve temperature higher than 150 degrees C. The modeled thermal inactivation and observed inactivation during UHPH treatments suggest that temperature could be the main lethal effect driving inactivation.

Production responses of reproducing ewes to a by-product-based diet inoculated with the probiotic Bacillus amyloliquefaciens strain H57

The potential application of the spore-forming probiotic Bacillus amyloliquefaciens strain H57 (H57) as a novel probiotic for ruminants was evaluated in reproducing ewes. Performance responses were determined by delivering H57 in a pelleted diet based mainly on palm kernel meal (PKM) and sorghum grain. PKM is an agro-industrial by-product with a reputation for poor palatability and the availability of the starch in sorghum grain can be limited in ruminants. The hypothesis was that H57 improves the feeding value of a relatively low quality concentrate diet. Twenty-four first-parity white Dorper ewes were fed PKM-based pellets manufactured with or without H57 (109 cfu/kg pellet) in late pregnancy. During this phase of late pregnancy, the H57 ewes ate 17% more dry matter (1019 vs 874 g/day, P = 0.03), gained more weight (194 vs 30 g/day, P = 0.008) and retained more nitrogen (6.13 vs 3.34 g/day, P = 0.01), but produced lambs with a similar birthweight (4.1 vs 4.2 kg, P = 0.73). Rumen fluid collected from H57 ewes in late pregnancy had higher pH (7.1 vs 6.8, P = 0.07), acetate : propionate ratio (3.4 vs 2.7, P = 0.04), lower ammonia (69 vs 147 mmol/L, P = 0.001) and total volatile fatty acid concentrations (40 vs 61 mg/L, P = 0.02). The digestibility of dry matter, organic matter and fibre were similar between the two groups. The lambs of the H57 ewes grew faster than those of the Control ewes for the first 21 days of lactation (349 vs 272 g/day, P = 0.03), but not thereafter. H57 can improve feed intake and maternal liveweight gain in late pregnancy of first-parity ewes fed a diet based on PKM.

Near complete genome sequence of the animal feed probiotic, Bacillus amyloliquefaciens H57

Bacillus amyloliquefaciens H57 is a bacterium isolated from lucerne for its ability to prevent feed spoilage. Further interest developed when ruminants fed with H57-inoculated hay showed increased weight gain and nitrogen retention relative to controls, suggesting a probiotic effect. The near complete genome of H57 is ~3.96 Mb comprising 16 contigs. Within the genome there are 3,836 protein coding genes, an estimated sixteen rRNA genes and 69 tRNA genes. H57 has the potential to synthesise four different lipopeptides and four polyketide compounds, which are known antimicrobials. This antimicrobial capacity may facilitate the observed probiotic effect.

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Situação atual do marco regulatório sobre o acesso ao patrimônio genético frente às atividades de pesquisa e desenvolvimento de produtos biológicos. Principais atividades da coordenação de fertilizantes, inoculantes e corretivos no biênio 2012-2014. Resultados das análises da fiscalização de produtos inoculantes para leguminosas nos anos de 2012-2013 e estabelecimento de um ensaio interlaboratorial. Perspectivas de uso de inoculantes microbianos no Brasil: a visão da ANPII. A rede ?FBN_ABC?: compromisso com a promoção dos benefícios da Fixação Biológica do Nitrogênio ( FBN) através dos inoculantes. Método alternativo para contagem e viabilidade celular para inoculantes: novo horizonte baseado em citometria de fluxo. Alternativa metodológica para contagem de Azospirillum em inoculantes. Sugestão para alterações nos meios de cultura e análises de Azospirillum nas instruções normativas Nº 30 e Nº 13 do MAPA. Análise de nodulação de soja e uso de um inoculante padrão em casa de vegetação para determinar a qualidade de produtos comerciais. Eficiência simbiótica de estirpes de Bradyrhizobium para a produção de mudas de Centrolobium paraense. Avaliação da eficiência agronômica de novas estirpes de rizóbio para a cultura do feijoeiro. Caracterização e avaliação agronômica de novos isolados de rizóbio obtidos de nódulos de genótipos silvestes de feijoeiro-comum. Resposta do feijoeiro irrigado à inoculação em fazendas do GTEC-feijão em Unaí-MG: safras 2012 e 2013. Recomendação de nova estirpe de Rhizobium para feijão-comum. Sobrevida en semillas de soja de esporas de Penicillium bilaiae. Uso y sistema de aplicación de promotores del crecimiento em semilla de soja. Novas tecnologias na cultura da soja: métodos de inoculação, densidades de plantas e novas estirpes de Bradyrhizobium. Relato dos resultados de ensaios de adubação nitrogenada na cultura da soja pelo CESB (Comitê Estratégico Soja Brasil). Resposta do amendoinzeiro à inoculação. Efeito de biofertilizante na promoção de crescimento e aumento de produtividade de arroz. Alterações de germinação e vigor de sementes da cultivar de arroz BRS pampa tratadas com indutores de crescimento. Promoção do crescimento de milho por novas estirpes de bactérias associativas: resultados de ensaios em rede conduzidos pelo instituto nacional de ciência e tecnologia da Fixação Biológica do Nitrogênio (INCT-FBN) Metodologias de inoculação de Azospirillum brasilense na cultura de milho. O conteúdo de exopolissacarídeos e polihidroxibutirato influenciam a sobrevivência de Azospirillum brasilense e o desenvolvimento de raízes de plântulas de milho. Reclassificação de espécies de estirpes autorizadas para as culturas da soja e do feijoeiro e revelações obtidas no sequenciamento dos seus genomas. Coinoculaçâo da soja e do feijoeiro com rizóbios e Azospirillum brasilense. Validação de estirpes de rizóbio para a inoculação de espécies arbóreas, adubos verdes e forrageiras visando metas do programa ABC e do novo código florestal. Evaluación de un inoculante formulado con cepas de Bacillus amyloliquefaciens para el control biológico de hongos fitopatógenos. Relação de membros credenciados na Relare. Ata técnica da assembleia geral ordinária da XVII Relare. Ata da eleição da diretoria da Relare para o biênio 2014 ? 2016. Relação dos participantes da XVII Relare.

Trypsin Inhibitor from Edible Mushroom Pleurotus floridanus Active against Proteases of Microbial Origin

Protease inhibitors can be versatile tools mainly in the fields of medicine, agriculture and food preservative applications. Fungi have been recognized as sources of protease inhibitors, although there are only few such reports on mushrooms. This work reports the purification and characterization of a trypsin inhibitor from the fruiting body of edible mushroom Pleurotus floridanus (PfTI) and its effect on the activity of microbial proteases. The protease inhibitor was purified up to 35-fold by DEAE-Sepharose ion exchange column, trypsin-Sepharose column and Sephadex G100 column. The isoelectric point of the inhibitor was 4.4, and its molecular mass was calculated as 37 kDa by SDS-PAGE and 38.3 kDa by MALDI-TOF. Inhibitory activity confirmation was by dot-blot analysis and zymographic activity staining. The specificity of the inhibitor toward trypsin was with Ki of 1.043×10−10 M. The inhibitor was thermostable up to 90 °C with maximal stability at 30 °C, active over a pH range of 4–10 against proteases from Aspergillus oryzae, Bacillus licheniformis, Bacillus sp. and Bacillus amyloliquefaciens. Results indicate the possibility of utilization of protease inhibitor from P. floridanus against serine proteases

Anti-diabetic or anti-hypertensive dietary supplement

An anti-diabetic or anti-hypertensive fish protein hydrolysate is provided, in which the fish is of the genus Salmo or Oncorhynchus, and wherein the fish protein is hydrolysed by a metalloendopeptidase obtainable from Bacillus amyloliquefaciens. Methods of making and methods for using such fish protein hydrolysates are also provided.

Estimation of accumulated lethality under pressure-assisted thermal processing

A study was conducted to develop an integrated process lethality model for pressure-assisted thermal processing (PATP) taking into consideration the lethal contribution of both pressure and heat on spore inactivation. Assuming that the momentary inactivation rate was dependent on the survival ratio and momentary pressure-thermal history, a differential equation was formulated and numerically solved using the Runge-Kutta method. Published data on combined pressure-heat inactivation of Bacillus amyloliquefaciens spores were used to obtain model kinetic parameters that considered both pressure and thermal effects. The model was experimentally validated under several process scenarios using a pilot-scale high-pressure food processor. Using first-order kinetics in the model resulted in the overestimation of log reduction compared to the experimental values. When the n th-order kinetics was used, the computed accumulated lethality and the log reduction values were found to be in reasonable agreement with the experimental data. Within the experimental conditions studied, spatial variation in process temperature resulted up to 3.5 log variation in survivors between the top and bottom of the carrier basket. The predicted log reduction of B. amyloliquefaciens spores in deionized water and carrot purée had satisfactory accuracy (1.07-1.12) and regression coefficients (0.83-0.92). The model was also able to predict log reductions obtained during a double-pulse treatment conducted using a pilot-scale high-pressure processor. The developed model can be a useful tool to examine the effect of combined pressure-thermal treatment on bacterial spore lethality and assess PATP microbial safety. © 2013 Springer Science+Business Media New York.

Potencial in vitro de rizobactérias para o controle de nematóides-chave da cana-de-açúcar, dinâmica populacional e manejo de Pratylenchus zeae

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Isolamento de microrganismos produtores de queratinase: estudo da biodegradação da queratina oriunda de penas de abatedouro de frangos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Produção de biossurfactantes utilizando resíduos agroindustriais como substrato

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)