RESUMO
Gluconacetobacter diazotrophicus is a species of great agronomic potential due to its growth-promotion traits. Its colonization process in different plants has been reported. However, there have been no studies regarding its structural colonization in elephant grass. This is a fast-growing C4-Poaceae plant, and its application in Brazil is mainly aimed at feeding dairy cattle, due to its high nutritional value. Also, in the last decade, this grass has been applied in the production of biofuels. The present study aimed to monitor the colonization process of strain LP343 of G. diazotrophicus inoculated in elephant grass seedlings of PCEA genotype, by using a mCherry-tagged bacterium. Samples of roots and shoots collected at different periods were visualized by confocal laser-scanning microscopy. The colony-counting assay was used to compare the number of cells recovered in different niches and a qPCR was performed for the quantification of endophytic cells in root and shoot tissues. Results suggested that the strain LP343 quickly recognized the PCEA roots as host, attached to the elephant grass roots at 6 h, and 7 days after inoculation were able to colonize the xylem vessels of roots and shoots of elephant grass. This study advances our knowledge about the colonization process of G. diazotrophicus species in elephant grass, contributing to future studies involving the plant-bacteria interaction cultivated under gnotobiotic conditions.
Assuntos
Gluconacetobacter/crescimento & desenvolvimento , Pennisetum/microbiologia , Raízes de Plantas/microbiologia , Brotos de Planta/microbiologia , Brasil , Vida Livre de Germes , Gluconacetobacter/genética , Gluconacetobacter/isolamento & purificação , Pennisetum/crescimento & desenvolvimento , Raízes de Plantas/crescimento & desenvolvimento , Brotos de Planta/crescimento & desenvolvimento , Plântula/crescimento & desenvolvimento , Plântula/microbiologiaRESUMO
Bacterial endophytes are well known inhabitants of living plant system and perform important assignments in maintaining plant growth and health. Currently, limited reports are available on the endophytes of pearl millet (Pennisetum glaucum) reflecting antagonistic and plant growth promoting (PGP) attributes. Therefore, the major objectives of current investigation were to identify antagonistic strains of endophytic Bacillus from pearl millet and further illustrate their PGP capabilities. In this study, 19 endophytic Bacillus strains (EPP5, EPP21, EPP30, EPP32, EPP35, EPP42, EPP49, EPP55, EPP62, EPP65, EPP70, EPP71, EPP74, EPP78, EPP83, EPP86, EPP93, EPP100, and EPP102) displaying antagonistic activity towards Rhizoctonia solani (RS), Sclerotium rolfsii (SR), and Fusarium solani (FS) were isolated from different sections (root, leaf, stem, and root) of pearl millet. Phenotypic (shape, colony, gram staining reaction, endospore formation, and motility) and biochemical features (catalase, oxidase, citrate, gelatinase, urease, Voges Proskauer's, methyl red, indole, and nitrate reduction), along with the similarly comparison of 16S rRNA gene sequence with type strains identified eight antagonistic endophyhtes as B. amyloliquefaciens (EPP35, EPP 42, EPP62, and EPP 102), Bacillus subtilis subsp. subtilis (EPP65), and Bacillus cereus (EPP5, EPP71, and EPP74). The production of indole acetic acid and siderophores was varied among the isolated endophytes. Besides displaying enzymatic activities, these isolates varied in solubilizing capabilities of phosphate, potassium, and zinc. The presence of three antimicrobial peptide genes (ituD, bmyC, and srfA) also confirmed their antifungal nature. Further, single treatment of three promising strains (EPP5, EPP62, and EPP65) offered protection ranging from 35.68 to 45.74% under greenhouse conditions. However, microbial consortium (EPP5+ EPP62 + EPP65) provided the highest protection (71.96%) against root rot and wilt infection with significant increase in plant biomass. Overall, the current study indicated that pearl millet plant harbors various species of endophytic Bacillus that possess excellent biocontrol and growth promotion activities.
Assuntos
Antifúngicos/isolamento & purificação , Endófitos/isolamento & purificação , Pennisetum , Doenças das Plantas , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Bacillus/genética , Bacillus/isolamento & purificação , Bacillus/metabolismo , Bacillus amyloliquefaciens/genética , Bacillus amyloliquefaciens/isolamento & purificação , Bacillus amyloliquefaciens/metabolismo , Bacillus cereus/genética , Bacillus cereus/isolamento & purificação , Bacillus cereus/metabolismo , Bacillus subtilis/genética , Bacillus subtilis/isolamento & purificação , Bacillus subtilis/metabolismo , Basidiomycota/efeitos dos fármacos , Agentes de Controle Biológico , Endófitos/genética , Endófitos/metabolismo , Fusarium/efeitos dos fármacos , Genes Bacterianos , Ácidos Indolacéticos/metabolismo , Consórcios Microbianos , Interações Microbianas , Pennisetum/crescimento & desenvolvimento , Pennisetum/microbiologia , Fosfatos/metabolismo , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Raízes de Plantas/microbiologia , Potássio/metabolismo , Rhizoctonia/efeitos dos fármacos , Sideróforos/metabolismo , Microbiologia do Solo , Zinco/metabolismoRESUMO
Bacterial endophytes are considered to have a beneficial effect on host plants, improving their growth by different mechanisms. The objective of this study was to investigate the capacity of four endophytic Bacillus strains to solubilize iron phosphate (Fe-P), produce siderophores and indole-acetic acid (IAA) in vitro, and to evaluate their plant growth promotion ability in greenhouse conditions by inoculation into pearl millet cultivated in a P-deficient soils without P fertilization, with Araxá rock phosphate or soluble triple superphosphate. All strains solubilized Fe-P and three of them produced carboxylate-type siderophores and high levels of IAA in the presence of tryptophan. Positive effect of inoculation of some of these strains on shoot and root dry weight and the N P K content of plants cultivated in soil with no P fertilization might result from the synergistic combination of multiple plant growth promoting (PGP) traits. Specifically, while B1923 enhanced shoot and root dry weight and root N P content of plants cultivated with no P added, B2084 and B2088 strains showed positive performance on biomass production and accumulation of N P K in the shoot, indicating that they have higher potential to be microbial biofertilizer candidates for commercial applications in the absence of fertilization.
Assuntos
Bacillus/metabolismo , Endófitos/metabolismo , Nutrientes/metabolismo , Pennisetum/crescimento & desenvolvimento , Pennisetum/microbiologia , Fosfatos/metabolismo , Bacillus/genética , Endófitos/genética , Ácidos Indolacéticos/metabolismo , Ferro/metabolismo , Pennisetum/metabolismo , Fosfatos/análise , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Sideróforos/metabolismoRESUMO
Abstract Bacterial endophytes are considered to have a beneficial effect on host plants, improving their growth by different mechanisms. The objective of this study was to investigate the capacity of four endophytic Bacillus strains to solubilize iron phosphate (Fe-P), produce siderophores and indole-acetic acid (IAA) in vitro, and to evaluate their plant growth promotion ability in greenhouse conditions by inoculation into pearl millet cultivated in a P-deficient soils without P fertilization, with Araxá rock phosphate or soluble triple superphosphate. All strains solubilized Fe-P and three of them produced carboxylate-type siderophores and high levels of IAA in the presence of tryptophan. Positive effect of inoculation of some of these strains on shoot and root dry weight and the N P K content of plants cultivated in soil with no P fertilization might result from the synergistic combination of multiple plant growth promoting (PGP) traits. Specifically, while B1923 enhanced shoot and root dry weight and root N P content of plants cultivated with no P added, B2084 and B2088 strains showed positive performance on biomass production and accumulation of N P K in the shoot, indicating that they have higher potential to be microbial biofertilizer candidates for commercial applications in the absence of fertilization.
Assuntos
Bacillus/metabolismo , Alimentos/metabolismo , Pennisetum/crescimento & desenvolvimento , Pennisetum/microbiologia , Endófitos/metabolismo , Ácidos Indolacéticos/metabolismo , Fosfatos/análise , Fosfatos/metabolismo , Bacillus/genética , Sideróforos/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Pennisetum/metabolismo , Endófitos/genética , Ferro/metabolismoRESUMO
Saccharum officinarum bagasse (common name: sugarcane bagasse) and Pennisetum purpureum (also known as Napier grass) are among the most promising feedstocks for bioethanol production in Argentina and Brazil. In this study, both biomasses were assessed before and after acid pretreatment and following hydrolysis with Nasutitermes aquilinus and Cortaritermes fulviceps termite gut digestome. The chemical composition analysis of the biomasses after diluted acid pretreatment showed that the hemicellulose fraction was partially removed. The (hemi) cellulolytic activities were evaluated in bacterial culture supernatants of termite gut homogenates grown in treated and untreated biomasses. In all cases, we detected significantly higher endoglucanase and xylanase activities using pretreated biomasses compared to untreated biomasses, carboxymethylcellulose and xylan. Several protein bands with (hemi) cellulolytic activity were detected in zymograms and two-dimensional gel electrophoresis. Some proteins of these bands or spots were identified as xylanolytic peptides by mass spectrometry. Finally, the diversity of cultured cellulolytic bacterial endosymbionts associated to both Argentinean native termite species was analyzed. This study describes, for the first time, bacterial endosymbionts and endogenous (hemi) cellulases of two Argentinean native termites as well as their potential application in degradation of lignocellulosic biomass for bioethanol production.
Assuntos
Ácidos/química , Biomassa , Celulases/metabolismo , Isópteros/enzimologia , Pennisetum/enzimologia , Polissacarídeos/metabolismo , Saccharum/enzimologia , Animais , Argentina , Celulases/química , Sistema Digestório/enzimologia , Sistema Digestório/microbiologia , Proteínas de Insetos/metabolismo , Isópteros/classificação , Isópteros/microbiologia , Microscopia Eletrônica de Varredura , Pennisetum/microbiologia , Saccharum/microbiologia , SimbioseRESUMO
To test the accuracy of creatinine as a marker for estimating urinary volume and its use as a nutritional index, the possible interference of forage intake and forage quality over creatinine excretion was evaluated. For this, sheep were fed different levels of pearl millet (Pennisetum americanum(L) Leeke) or Italian ryegrass (Lolium multiflorum Lam). The experiment consisted of a compilation of digestibility trials (n=6) with pearl millet or Italian ryegrass in completely randomized designs with four replications and four forage levels: 1.5, 2.0, 2.5% (kg dry matter (DM)/ 100 kg of live weight (LW)). The trials were repeated at different periods to evaluate how stable the average metabolic excretion of creatinine is. In each trial, total urine collection was performed individually during a period of 24 hours for five consecutive days and subsequently analyzed by colorimetry for creatinine and purine derivatives. The creatinine excretion was not affected (P>0.05) by forage offer or forage type, but there were period effects (P=0.0001). The average creatinine excretion for both forages was 0.21mmol/kg PV0,75. Linear regressions between the purine derivatives:creatinine index with total excretion of purine derivatives were detected for pearl millet (P<0.0001, R2= 0.64) and Italian ryegrass (P=0.02, R2=0.20). These results demonstrate that creatinine excretion is independent of the type and availability of forage and can be a marker for urinary volume prediction and nutritional measures under grazing systems.
Para testar a precisão da creatinina como marcador para estimativas de volume urinário e índice nutricional, foram avaliadas a possível influência do consumo e a qualidade da forragem sobre esse marcador. Para isso, ovinos foram alimentados com diferentes níveis de milheto (Pennisetum americanum (L) Leeke) ou azevém (Lolium multiflorum Lam). O experimento consistiu de uma compilação de ensaios de digestibilidade (n=6) com milheto ou azevém, em um desenho experimental de blocos completamente ao acaso, com quatro repetições e quatro níveis de forragem: 1,5; 2,0; 2,5% (kg de matéria seca (MS)/ 100kg de peso vivo (PV)). Os ensaios foram repetidos em diferentes períodos, com ambas as forragens, se para avaliar a estabilidade da excreção média de creatinina metabólica. Em cada ensaio, foi coletado o volume total de urina individualmente, durante períodos de 24 horas, por cinco dias consecutivos. Posteriormente, esses ensaios foram analisados por colorimetria para creatinina e derivados de purina. A excreção de creatinina não foi afetada (P>0,05) pelo consumo de forragem ou pelo tipo de forragem, mas foi influenciada pelo período (P=0,0001). A excreção média de creatinina para ambas as forragens foi 0,21mmol/kg PV0,75. Regressões lineares entre os índices derivados de purina:creatinina com a excreção total de derivados de purina foram detectadas para milheto (P<0,0001; R2=0,64) e azevém (P=0,02; R2=0,20). Os resultados demonstraram que a excreção de creatinina é independente do tipo e do consumo de forragem e pode ser usada como marcador preditivo do volume urinário e do status nutricional em sistemas de pastejo.
Assuntos
Animais , Biomarcadores/urina , Creatinina/análise , Pennisetum/microbiologia , Ovinos , Matéria Orgânica/análise , Sistema Urinário , UrinaRESUMO
To test the accuracy of creatinine as a marker for estimating urinary volume and its use as a nutritional index, the possible interference of forage intake and forage quality over creatinine excretion was evaluated. For this, sheep were fed different levels of pearl millet (Pennisetum americanum(L) Leeke) or Italian ryegrass (Lolium multiflorum Lam). The experiment consisted of a compilation of digestibility trials (n=6) with pearl millet or Italian ryegrass in completely randomized designs with four replications and four forage levels: 1.5, 2.0, 2.5% (kg dry matter (DM)/ 100 kg of live weight (LW)). The trials were repeated at different periods to evaluate how stable the average metabolic excretion of creatinine is. In each trial, total urine collection was performed individually during a period of 24 hours for five consecutive days and subsequently analyzed by colorimetry for creatinine and purine derivatives. The creatinine excretion was not affected (P>0.05) by forage offer or forage type, but there were period effects (P=0.0001). The average creatinine excretion for both forages was 0.21mmol/kg PV0,75. Linear regressions between the purine derivatives:creatinine index with total excretion of purine derivatives were detected for pearl millet (P<0.0001, R2= 0.64) and Italian ryegrass (P=0.02, R2=0.20). These results demonstrate that creatinine excretion is independent of the type and availability of forage and can be a marker for urinary volume prediction and nutritional measures under grazing systems.(AU)
Para testar a precisão da creatinina como marcador para estimativas de volume urinário e índice nutricional, foram avaliadas a possível influência do consumo e a qualidade da forragem sobre esse marcador. Para isso, ovinos foram alimentados com diferentes níveis de milheto (Pennisetum americanum (L) Leeke) ou azevém (Lolium multiflorum Lam). O experimento consistiu de uma compilação de ensaios de digestibilidade (n=6) com milheto ou azevém, em um desenho experimental de blocos completamente ao acaso, com quatro repetições e quatro níveis de forragem: 1,5; 2,0; 2,5% (kg de matéria seca (MS)/ 100kg de peso vivo (PV)). Os ensaios foram repetidos em diferentes períodos, com ambas as forragens, se para avaliar a estabilidade da excreção média de creatinina metabólica. Em cada ensaio, foi coletado o volume total de urina individualmente, durante períodos de 24 horas, por cinco dias consecutivos. Posteriormente, esses ensaios foram analisados por colorimetria para creatinina e derivados de purina. A excreção de creatinina não foi afetada (P>0,05) pelo consumo de forragem ou pelo tipo de forragem, mas foi influenciada pelo período (P=0,0001). A excreção média de creatinina para ambas as forragens foi 0,21mmol/kg PV0,75. Regressões lineares entre os índices derivados de purina:creatinina com a excreção total de derivados de purina foram detectadas para milheto (P<0,0001; R2=0,64) e azevém (P=0,02; R2=0,20). Os resultados demonstraram que a excreção de creatinina é independente do tipo e do consumo de forragem e pode ser usada como marcador preditivo do volume urinário e do status nutricional em sistemas de pastejo.(AU)
Assuntos
Animais , Creatinina/análise , Ovinos , Pennisetum/microbiologia , Biomarcadores/urina , Matéria Orgânica/análise , Sistema Urinário , UrinaRESUMO
Foram realizados dois experimentos para avaliação de aditivos microbianos na ensilagem de forragens tropicais. No primeiro experimento avaliou-se o efeito de doses crescentes de Lactobacillus buchneri, em diferentes tempos de armazenamento, sobre o valor nutritivo, perfil fermentativo e estabilidade aeróbia de silagens de milho. No segundo experimento avaliou-se o efeito de L. buchneri, de forma exclusiva, ou em combinação com Lactobacillus plantarum, em silagens de milho, cana-de-açúcar e capim elefante cv. Napier. [...] No segundo experimento, o tratamento das silagens de milho com L. buchneri (LB), exclusivamente, ou associado com L. plantarum (LB+LP) não afetou as variáveis relacionadas com o valor nutritivo das silagens. As concentrações de ácido acético nas silagens de milho foram superiores no tratamento controle. Nas silagens de cana-de-açúcar, o tratamento LB apresentou maiores teores de MS e DVIVMO. O tratamento controle apresentou maior população de bactérias ácido láticas, e menor valor de pH. A silagem de cana-de-açúcar sem aditivo atingiu o pico de temperatura após o tratamento LB, sendo esse pico superior na silagem aditivada. Foram observadas menores perdas de matéria seca e produção de gases nas silagens aditivadas, resultando assim em maior recuperação de MS no tratamento LB. Foi observado maior tempo para atingir a temperatura máxima e menor acúmulo de temperatura até o quinto dia de exposição aeróbia no tratamento controle. A temperatura máxima atingida durante a exposição aeróbia foi superior no tratamento LB. Nas silagens de capim, as silagens do tratamento LB+LP apresentaram maiores teores de MS, e menor teor de PB. Foi observada maior população de bactérias ácido láticas nas silagens do tratamento LB+LP. A inoculação com L. buchneri, sozinho, ou em associação com L. plantarum não foi eficiente em melhorar a qualidade e estabilidade aeróbia das forragens avaliadas
Two trials were conducted for the evaluation of microbial additives on tropical forages ensiling. At the first trial the effect of increasing L. buchneri doses, and different storage periods on the fermentation profile, aerobic stability and nutritive value of corn silage were evaluated. At the second trial the effect of L. buchneri with or without L. plantarum was evaluated on the fermentation profile, aerobic stability and nutritive value of corn, sugarcane and tropical grass silages. [...] In the second trial, inoculation of corn silages with L. buchneri (LB), alone or in association with L. plantarum (LB+LP) did not affect the variables related to the silage nutritive value. Acetic acid content of corn silages was higher for the control treatment. In sugarcane silages, LB treatment showed higher DM content and IVTOMD. Control treatment showed higher lactic acid bacteria counts, and lower pH. Sugarcane silage with no additive reached temperature peak after LB treatment, with this peak being higher at the additivated silage. There were observed lower dry matter losses and gas production in LB silages, resulting in higher dry matter recovery. It was observed higher time to reach maximum temperature and lower temperature accumulation until fifth day of aerobic exposure for the control treatment. Maximum temperature reached during aerobic exposure was higher for LB treatment. In tropical grass silages, LB+LP silages showed higher DM and lower CP contents. It was observed higher lactic acid bacteria in LB+LP treatment. Inoculation with L. buchneri, alone or in association with L. plantarum was not efficient in improving quality and aerobic stability of evaluated forages
Assuntos
Fermentação , Pennisetum/microbiologia , Saccharum/microbiologia , Zea mays/microbiologia , Armazenamento de Alimentos , Concentração de Íons de Hidrogênio , LactobacillusRESUMO
O trabalho analisa o efeito do inoculante bacteriano em silagens de capim Elefante (Pennisetum purpureum cv. Roxo) em concentrações crescentes de inclusão. O experimento realizado no Departamento de Zootecnia da Universidade Federal Rural da Amazônia (UFRA) consistiu em cinco tratamentos e quatro repetições (Controle, LP/PC 5x104 , LP/PC 1x105 , LP/PC 5x105 , LP/PC 1x106 ) que se encontravam em baldes plásticos com tampa e vedada com fita plastica, onde foram pesadas semanalmente ate o final do experimento. Foram realizadas analises químicas e bromatológicas da forragem e das silagensonde foi retirada amostras no momento da abertura (Dia 0) e no ultimo dia de aerobiose (Dia 10), alem da contagem de fungos e leveduras nas silagens para avaliar a qualidade do material ensilado. No período de aerobiose foi inseridos data loggers nos silos para a observar a temperatura na pós fermentação. A utilização do inoculante bacteriano Lactobacillus plantarum e Pediococcus acidilactici pode ser recomendável, pois melhorou a estabilidade aeróbia. Em termos de perfil fermentativo e composição bromatológica, seus efeitos foram mínimos