Lacticaseibacillus parahuelsenbergensis sp. nov., Lacticaseibacillus styriensis sp. nov. and Lacticaseibacillus zeae subsp. silagei subsp. nov., isolated from different grass and corn silage.

Four rod-shaped, non-motile, non-spore-forming, facultative anaerobic, Gram-stain-positive lactic acid bacteria, designated as EB0058T, SCR0080, LD0937T and SCR0063T, were isolated from different corn and grass silage samples. The isolated strains were characterized using a polyphasic approach and EB0058T and SCR0080 were identified as Lacticaseibacillus zeae by 16S rRNA gene sequence analysis. Based on whole-genome sequence-based characterization, EB0058T and SCR0080 were separated into a distinct clade from Lacticaseibacillus zeae DSM 20178T, together with CECT9104 and UD2202, whose genomic sequences are available from NCBI GenBank. The average nucleotide identity (ANI) values within the new subgroup are 99.9 % and the digital DNA-DNA hybridization (dDDH) values are 99.3-99.9 %, respectively. In contrast, comparison of the new subgroup with publicly available genomic sequences of L. zeae strains, including the type strain DSM 20178T, revealed dDDH values of 70.2-72.5 % and ANI values of 96.2-96.6 %. Based on their chemotaxonomic, phenotypic and phylogenetic characteristics, EB0058T and SCR0080 represent a new subspecies of L. zeae. The name Lacticaseibacillus zeae subsp. silagei subsp. nov. is proposed with the type strain EB0058T (=DSM 116376T=NCIMB 15474T). According to the results of 16S rRNA gene sequencing, LD0937T and SCR0063T are members of the Lacticaseibacillus group. The dDDH value between the isolates LD0937T and SCR0063T was 67.6 %, which is below the species threshold of 70 %, clearly showing that these two isolates belong to different species. For both strains, whole genome-sequencing revealed that the closest relatives within the Lacticaseibacillus group were Lacticaseibacillus huelsenbergensis DSM 115425 (dDDH 66.5 and 65.9 %) and Lacticaseibacillus casei DSM 20011T (dDDH 64.1 and 64.9 %). Based on the genomic, chemotaxonomic and morphological data obtained in this study, two novel species, Lacticaseibacillus parahuelsenbergensis sp. nov. and Lacticaseibacillus styriensis sp. nov. are proposed and the type strains are LD0937T (=DSM 116105T=NCIMB 15471T) and SCR0063T (=DSM 116297T=NCIMB 15473T), respectively.

Cleaner anaerobic fermentation and greenhouse gas reduction of crop straw.

Rice anaerobic fermentation is a significant source of greenhouse gas (GHG) emissions, and in order to efficiently utilize crop residue resources to reduce GHG emissions, rice straw anaerobic fermentation was regulated using lactic acid bacteria (LAB) inoculants (FG1 and TH14), grass medium (GM) to culture LAB, and Acremonim cellulolyticus (AC). Microbial community, GHG emission, dry matter (DM) loss, and anaerobic fermentation were analyzed using PacBio single-molecule real-time and anaerobic fermentation system. The epiphytic microbial diversity of fresh rice straw was extremely rich and contained certain nutrients and minerals. During ensiling, large amounts of GHG such as carbon dioxide are produced due to plant respiration, enzymatic hydrolysis reactions, and proliferation of aerobic bacteria, resulting in energy and DM loss. Addition of FG1, TH14, and AC alone improved anaerobic fermentation by decreasing pH and ammonia nitrogen content (P < 0.05) and increased lactic acid content (P < 0.05) when compared to the control, and GM showed the same additive effect as LAB inoculants. Microbial additives formed a co-occurrence microbial network system dominated by LAB, enhanced the biosynthesis of secondary metabolites, diversified the microbial metabolic environment and carbohydrate metabolic pathways, weakened the amino acid metabolic pathways, and made the anaerobic fermentation cleaner. This study is of great significance for the effective utilization of crop straw resources, the promotion of sustainable livestock production, and the reduction of GHG emissions.IMPORTANCETo effectively utilize crop by-product resources, we applied microbial additives to silage fermentation of fresh rice straw. Fresh rice straw is extremely rich in microbial diversity, which was significantly reduced after silage fermentation, and its nutrients were well preserved. Silage fermentation was improved by microbial additives, where the combination of cellulase and lactic acid bacteria acted as enzyme-bacteria synergists to promote lactic acid fermentation and inhibit the proliferation of harmful bacteria, such as protein degradation and gas production, thereby reducing GHG emissions and DM losses. The microbial additives accelerated the formation of a symbiotic microbial network system dominated by lactic acid bacteria, which regulated silage fermentation and improved microbial metabolic pathways for carbohydrates and amino acids, as well as biosynthesis of secondary metabolites.

Ensiling pretreatment fortified with laccase and microbial inoculants enhances biomass preservation and bioethanol production of alfalfa stems.

This study investigated the potential of ensiling pretreatment fortified with laccase and a lactic acid bacteria (LAB) inoculant on improving the utilization of alfalfa stems for bioethanol production. The alfalfa stems were ensiled with no additives (Con), 0.04 % laccase (LA), a LAB inoculant containing Pediococcus pentosaceus at 1 × 106 fresh weight (FW) and Pediococcus acidilactici at 3 × 105 cfu/g FW (PP), and a combination of LA and PP (LAP) for 120 days. By reshaping the bacterial community structure of alfalfa stem silages toward a higher abundance of Lactobacillus, the addition of laccase and LAB inoculant either alone or in combination facilitated lactic acid fermentation to reduce fermentation losses, as evidenced by low concentrations of ammonia nitrogen (53.7 to 68.9 g/kg total nitrogen) and ethanol (2.63 to 3.55 g/kg dry matter). All additive treatments increased lignocellulose degradation and soluble sugars concentrations of alfalfa stem silages. Due to delignification and polyphenol removal, glucan and xylan conversion (70.3 % vs. 35.7 % and 51.6 % vs. 27.9 %, respectively) and ethanol conversion efficiency (53.9 % vs. 26.4 %) of alfalfa stems were greatly increased by ensiling fortified with LA versus Con, and these variables (79.8 % for glucan, 58.7 % for xylan, and 60.1 % for ethanol conversion efficiency) were further enhanced with a synergistic effect of LA and PP fortification. The spearman correlation analysis revealed that bioethanol fermentation of silage biomass was closely related to ensiling parameters and total phenols. In conclusion, ensiling pretreatment with LA and PP combination offered a feasible way to efficient utilization of alfalfa stems for bioethanol production.

A novel species of lactic acid bacteria, Ligilactobacillus pabuli sp. nov., isolated from alfalfa silage.

In this study, we isolated a novel strain of lactic acid bacteria, AF129T, from alfalfa silage prepared locally in Morioka, Iwate, Japan. Polyphasic taxonomy was used to characterize the bacterial strain. The bacterium was rod-shaped, Gram-stain-positive, non-spore-forming and catalase-negative. The strain grew at various temperatures (15-40°C) and pH levels (4.0-8.0). The optimum growth conditions were a temperature of 30°C and a pH of 6.0. AF129T exhibited growth at salt (NaCl) concentrations of up to 6.5 % (w/v). The G+C content of the strain's genomic DNA was 41.5 %. The major fatty acids were C16 : 0, C18 : 1ω9c, C19 : 0cyclo ω8c and summed feature 8. 16S rRNA gene sequencing revealed that AF129T represents a member of the genus Ligilactobacillus and it has higher sequence similarities with Ligilactobacillus pobuzihii (98.4 %), Ligilactobacillus acidipiscis (97.5 %) and Ligilactobacillus salitolerans (97.4 %). The digital DNA-DNA hybridization values for AF129T and phylogenetically related species of the genus Ligilactobacillus ranged from 19.8% to 24.1%. The average nucleotide identity of the strain with its closely related taxa was lower than the threshold (95 %-96 %) used for species differentiation. In the light of the above-mentioned physiological, genotypic, chemotaxonomic and phylogenetic evidence, we confirm that AF129T represents a member of the genus Ligilactobacillus and constitutes a novel species; we propose the name Ligilactobacillus pabuli sp. nov. for this species. The type strain is AF129T =MAFF 518002T =JCM 34518T=BCRC 81335T.

Microbial enrichment evaluation during the fermentation of ensiling pruned branches from tea plants.

To investigate new disposal methods for agricultural waste from tea plantations, the dynamics of chemical composition and microbial populations of tea plant pruning waste during 60 days of ensiling were studied. The results showed that the pH value decreased markedly in the initial stage and then stabilized after Day 21 (approximately 3.9). At the end of ensiling, lactic and acetic acids were enriched in the silage, while most of the characteristic components of tea, including polyphenols, theanine, and caffeine, were well preserved. Microbiological analysis showed that the relative abundance of Lactobacillus increased rapidly from Day 3 (73.2%), reaching 98.6%. In addition, the dynamics of the microbial community during fermentation were related to its chemical composition and fermentation metabolites. In summary, silage can be an effective approach for treating and utilizing agricultural waste from tea plant pruning.

Ambient ultraviolet radiation: A new factor affecting anaerobic fermentation of oat and subsequent methane emissions.

To investigate the effect of ambient ultraviolet (UV) radiation on the Qinghai-Tibetan Plateau on anaerobic fermentation and subsequent methane emissions, oats grown under different ambient UV conditions (UV1, 100% ambient UV radiation; UV2, 50% ambient UV radiation) were fermented with molasses and Lactobacillus plantarum (LP) inoculant treatments. The UV2 treatment increased (P < 0.05) epiphytic bacterial counts and reduced (P < 0.05) the water-soluble carbohydrate contents of oat. Both the UV2 treatment and the LP inoculant substantially increased the abundance of Lactobacillus and inhibited detrimental microorganisms (enterobacteria and yeasts) during anaerobic fermentation, resulting in the absence of butyric acid. The UV2 treatment increased (P < 0.05) the acid detergent fiber contents after anaerobic fermentation and indirectly increased (P < 0.05) methane emissions by 8.52-14.69% during in vitro ruminal digestion. This study demonstrated that low ambient UV radiation during cultivation facilitated anaerobic fermentation and consequently enhanced subsequent methane emissions from the rumen.

Evaluation of biological and chemical additives on microbial community, fermentation characteristics, aerobic stability, and in vitro gas production of SuMu No. 2 elephant grass.

BACKGROUND: The study was conducted to evaluate the effects of biological and chemical additives on microbial community, fermentation characteristics, aerobic stability, and in vitro gas production of SuMu No. 2 elephant grass. RESULTS: Aerobic bacteria and yeast were not affected on days 5 and 7 but were significantly (P < 0.224) reduced on days 14, 30, and 60, whereas lactic acid and lactic acid bacteria were significantly (P > 0.001) higher in all ensiling days within all treatment groups. During the ensiling days, the pH, acetic acid, butyric acid, and yeast were decreased in all treatment groups, whereas the Lactobacillus plantarum group and L. plantarum + natamycin group were highly significantly (P > 0.001) decreased. During air exposure, the water-soluble carbohydrates, ammonia nitrogen, lactic acid, and acetic acid were not affected on days 1-4, whereas pH and aerobic bacteria (were significantly (P < 0.05) increased on days 2-4. The addition of Lactobacillus plantarum and natamycin increased the gas production, in vitro dry matter digestibility, and in vitro neutral detergent fiber of SuMu No. 2 elephant grass silages. CONCLUSIONS: The addition of biological and chemical additives, such as L. plantrum alone and the combination with natamycin, affected the undesirable microbial community, fermentation characteristics, aerobic stability, and in vitro gas of SuMu No. 2 elephant grass. © 2021 Society of Chemical Industry.

Silage Fermentation: A Potential Biological Approach for the Long-Term Preservation and Recycling of Polyphenols and Terpenes in Globe Artichoke (Cynara scolymus L.) By-Products.

An economic and effective method for storage is necessary to make full use of the nature of active components in artichoke by-products and ease environmental pressure. In this paper, the potential of silage fermentation for the preservation and recycling of polyphenols and terpenes in artichoke by-products is evaluated. The silage of artichoke by-products is characterized by lactic acid bacteria fermentation. Silage distinctly increases the abundance of lactic acid bacteria in artichoke by-products, such as Lactobacillus, Lactococcus, Serratia, and Weissella, and greatly increases the abundance of Firmicutes. The improvement of the microorgan structure and composition is of great significance for the quality of artichoke by-products. Polyphenols in the stems and leaves of artichokes are preserved well in silage. Among the 18 polyphenol compounds detected by high performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry (HPLC-QqQ-MS/MS), the contents of 11 phenolic acids and four flavonoids increased significantly. For terpenes detected by gas chromatography-mass spectrometry (GC-MS), the contents of four pentacyclic triterpenoids increased significantly, while two sterols were kept stable in the silage process. Silage is a potential biotechnology for the long-term preservation of bioactive components, such as polyphenols and terpenes in artichoke by-products, and the results provide a scientific basis for the efficient utilization of by-products.

Lactobacillus buchneri subsp. silagei subsp. nov., isolated from rice grain silage.

Two Gram-stain-positive, rod-shaped, non-motile, non-spore-forming, catalase-negative bacteria, designated strains SG162T and NK01, were isolated from Japanese rice grain silage and total mixed ration silage, respectively. They were initially identified as Lactobacillus buchneri based on the 16S rRNA gene sequence similarities. However, the two strains were separated into a distinct clade from L. buchneri DSM 20057T (=JCM 1115T) through whole-genome sequence-based characterization, forming an infraspecific subgroup together with strains CD034 and S42, whose genomic sequences were available in the public sequence database. Strains within the subgroup shared 99.4-99.7 % average nucleotide identity (ANI) and 97.5-99.0 % digital DNA-DNA hybridization (dDDH) with each other, albeit 96.9-97.0 % ANI and 76.0-76.6 % dDDH against DSM 20057T. Strains SG162T and NK01 could utilize more substrates as sole carbon sources than DSM 20057T, potentially owing to the abundance of genes involved in carbon metabolism, especially the Entner-Doudoroff pathway. The inability of γ-aminobutyric acid (GABA) production was evidenced by the lack of glutamate decarboxylase and glutamate/GABA antiporter genes in the new subgroup strains. Strain SG162T grew at 10-45 °C (optimum, 30 °C), pH 3.5-8.0, and 0-8 % (w/v) NaCl. Its genomic DNA G+C content was 44.1 mol%. The predominant fatty acids were C16 : 0, C19 : 0 cyclo ω8c, and summed feature 8. On the basis of the polyphasic characterization findings, strains SG162T and NK01 represent a novel subspecies of L. buchneri, for which the name Lactobacillus buchneri subsp. silagei subsp. nov. is proposed. The type strain is SG162T (=JCM 32599T=DSM 107969T), and strains CD034 and S42 are also transferred to L. buchneri subsp. silagei.

Encapsulación de Weissella paramesenteroides y su aplicación en residuos de pescado / Weissella paramesenteroides encapsulation and its application in the use of fish wastes

The goal of the present study was to evaluate the encapsulation of Weissella paramesenteroides, isolated from bee bread, as a technological tool for its use in biological fish silage. The pH decrease in fish silages using the bacteria encapsulated and in a non-encapsulated form was compared. W. paramesenteroides showed a good performance in the development of bioEnc ap su lati on; logical fish silage. The alginate encapsulation method showed an encapsulation efficacy of 85% and provides a reliable technological application.

El objetivo del presente estudio fue evaluar la encapsulación de Weissella paramesenteroides, aislada a partir del pan de polen, como herramienta tecnológica para su uso en la elaboración de ensilado biológico de pescado. Se comparó el descenso de pH para los ensilados utilizando la bacteria encapsulada y no encapsulada. W. paramesenteroides mostró un buen desempeño en el desarrollo de ensilado biológico de pescado. El método de encapsulación con alginato mostró una eficacia del 85% y puede ser utilizado para su aplicación tecnológica.

The occurrence of aflatoxigenic Aspergillus spp. in dairy cattle feed in Southern Brazil

ABSTRACT The presence of mycotoxins or related fungi in animal feed is a major problem for animal and human health. Silage and concentrated feed samples were collected from 21 dairy farms in the Western part of Paraná state in Southern Brazil. Water activity and pH of all samples were measured, and each sample was analyzed to check for the presence of aflatoxigenic Aspergillus. Water activity was observed to be lower in the concentrated feed samples. The pH was lower in the silage samples, indicating fermentation processes. Two silage samples and four concentrated feed samples were contaminated with Aspergillus spp. Seven isolates of Aspergillus spp. were obtained and their potential to produce aflatoxins was evaluated. Four of the isolates, two from the silage samples and two from the concentrated feed samples, produced the aflatoxins B1, B2, G1, and G2 in culture media. These isolates were identified as Aspergillus parasiticus and Aspergillus nomius. The presence of aflatoxigenic isolates of Aspergillus spp. in silage and concentrated feed samples is a matter of concern, because of the risk of aflatoxin production and contamination of the animal feed.

Microbiological and chemical profiles of elephant grass inoculated with and without Lactobacillus plantarum and Pediococcus acidilactici.

The study was conducted to evaluate the microbiological and chemical profiles of elephant grass inoculated with and without different wild strains of lactic acid bacteria. Silage was prepared of four treatments and one control with three replicates as control (EKC, adding 2 ml/kg sterilizing water), Lactobacillus plantarum (USA commercial bacteria) (EKP), Lactobacillus plantarum (EKA), Pediococcus acidilactici (EKB), and Pediococcus acidilactici (SKD) isolated from King grass. Silage were prepared using polyethylene terephthalate bottles, and incubated at room temperature for different ensiling days. The pH and acetic acid (AA) were significantly (P < 0.05) reduced and lactic acid (LA), butyric acid (BA), and ethanol were significantly increased (P < 0.05) at 3, 5, 7, and 14 days in treatment groups as compared to control. Water-soluble carbohydrate (WSC) and NH3-N concentration was not affected at days 3, 5, and 7, but significantly (P < 0.05) reduced at 14 days in treatment groups as compared to control. The LA, BA, and ethanol were significantly (P < 0.05) increased and AA, WSC NH3-N, and yeast were significantly (P < 0.05) decreased at 30 days of ensiling in treatment groups as compared to control. It is recommended that the inoculation of LAB could improve the fermentation quality of elephant grass silage and further effort is needed to evaluate these effects on silage produced on farm scale and on animal production performance.

Fermentation profile and microbial population in soybean silages with inoculant and powdered molasses / Perfil fermentativo e população microbiana em silagens de soja com inoculante e melaço em pó

Fermentation profile and microbial population were assessed in soybean silages without any additive (control), with inoculant (I), with I + powdered molasses (I+M), and with powdered molasses only (M). Soybean plants were harvested at the R6 stage and ensiled in 2kg-capacity laboratory silos. The additives were added to the natural matter base of silages. The assessed fermentation periods were 1, 3, 7, 14, 28, and 56 days. A 4 × 6 factorial arrangement (4 additives × 6 fermentation periods) in a completely randomized design with 3 replicates was used. Lactic, acetic, and butyric acids concentrations were influenced by additives and periods (P< 0.05). It was observed higher lactic acid values to control silages, on the 56th day. Lower average values of acetic and butyric acids were observed to I+M and M silages. It was observed quadratic effect to pH values with a reduction estimated of 0.5504, 0.5358, 0.6312 and 0.6680 units to pH values to control, I, I+M, and M silages in the first 10 days. A maximum lactic acid bacteria population was observed at the 28th day of fermentation in silages with inoculant. The inoculant and powdered molasses improve the fermentation profile of soybean silages.(AU)

Avaliou-se o perfil fermentativo e a composição microbiana de silagens de soja sem aditivos (controle), com inoculante (I), com I + melaço em pó (I+M) e com melaço em pó (M). As plantas de soja foram colhidas no estádio R6 e ensiladas em silos laboratoriais com capacidade de 2kg. Os aditivos foram adicionados às silagens na matéria natural. Os períodos de fermentação avaliados foram: 1, 3, 7, 14, 28 e 56 dias. Utilizou-se um esquema fatorial 4 × 6 (quatro aditivos × seis períodos), em um delineamento completamente ao acaso, com três repetições. Observou-se efeito de aditivos e de período (P<0,05) sobre os ácidos láctico, acético e butírico. Foi constatado maior teor de ácido láctico na silagem controle aos 56 dias. Para os teores dos ácidos acético e butírico, registraram-se menores valores médios nas silagens I+M e M. Observou-se efeito quadrático para o pH das silagens, com estimativa de redução de pH nos primeiros 10 dias igual a 0,5504; 0,5358; 0,6312 e 0,6680 nas silagens controle, I, I+M e M, respectivamente. A máxima população de bactéria ácido-láctica foi observada aos 28 dias nas silagens inoculadas. O inoculante e o melaço em pó melhoram o perfil fermentativo de silagens de soja.(AU)

Composição química da forragem e do ensilado de azevém anual em função de diferentes tempos de secagem e estádios fenológicos / Chemical composition of annual ryegrass forage and silage for different drying times and phenological phases

O objetivo do presente estudo foi determinar como o tempo de secagem e o estádio fenológico da planta influenciam na composição química e qualidade da silagem de azevém (Lolium multiflorum, Lam.). O delineamento experimental foi de blocos ao acaso, com seis tratamentos (vegetativo: cortar e ensilar; cortar + pré-secagem de 4 horas e ensilar; cortar + pré-secagem de 7 horas e ensilar; pré-florescimento: ensilagem, cortar e ensilar e pré-secagem de 4 horas; florescimento: cortar e ensilar) e quatro repetições, com quatro meses de conservação da silagem. Foram determinados: desaparecimento de massa após o corte, rendimento de massa seca, proteína bruta e frações fibrosas. Além disso, foram determinados na silagem: pH, lipídios totais, nitrogênio insolúvel em detergente neutro e ácido. O manejo empregado auxilia na desidratação da forragem, com perda na quantidade de nitrogênio no vegetativo. No pré-florescimento, essa perda não ocorre. O avanço do ciclo da forrageira ocasiona diminuição na quantidade de nutrientes na biomassa, entretanto a produção de forragem é aumentada. A silagem apresentou composição bromatológica semelhante à observada na massa verde de azevém, o que demonstra a eficiência do método de conservação. A ensilagem em estádios mais avançados é aconselhada quando se busca maior quantidade de biomassa ensilada, porém com qualidade inferior.(AU)

The aim was how to define the effect of pre-drying time and the plant phase's influence on chemical composition and quality of ryegrass silage. The experimental design was randomized blocks with six treatments (Vegetative: Cut and ensiled; cut + pre-drying 4 hours and ensiled and; cut + pre-drying of 7 hours and ensiled; Pre-flowering: Silage cutting and ensiled and pre-drying 4 hours Flowering:. cut and ensiled) and four repetitions. The silage was stowed for four months. It were determined the mass disappearance after cutting, dry matter yield, crude protein and fiber fractions. Moreover, it was determined on silage pH, total lipids and insoluble protein content. The management used aids in dehydration of fodder, resulting in losses on the amount of nitrogen on vegetative phase. For pre-flowering, this loss has not occur. The advance of fodder cycle causes a decrease in the amount of nutrients in biomass, but fodder yield increased. Ryegrass silage presented chemical composition similar to that observed on green mass of ryegrass, evinced the efficiency of fodder conservation method. The silage in more advanced phases is advised when seeking greater amount of ensiled biomass, but lower quality.(AU)

Isolation and molecular identification of lactic acid bacteria from King grass and their application to improve the fermentation quality of sweet Sorghum.

The aim of the present study was isolation and molecular identification of lactic acid bacteria from King grass and their application to improve the fermentation quality of sweet Sorghum. Seventy-six strains of LAB were isolated; five strains were selected for Physiological and morphological tests and 16S rRNA sequencing. All five strains grew at different pH 3.5-8.0, different temperature 35, 40, 45, 50 °C and different NaCl concentrations 3, 6.5, 9.5%. Strains HDASK were identified Lactobacillus plantarum and SK3907, SK2A32, SK3A42 and ASKDD Pediococcus acidilactici. Three isolated strains and one commercial strain were added to sweet sorghum. Silage was prepared of four treatments and one control with three replicates as control (SKC, adding 2 ml/kg sterilizing water), L. plantarum commercial bacteria (SKP), L. plantarum (HDASK) isolated from King grass (SKA), P. acidilactici (SK3907) isolated from King grass (SKB) and P. acidilactici (ASKDD) isolated from King grass (SKD). All silage were prepared using polyethylene terephthalate bottles, and incubated at room temperature for different ensiling days. The level of pH, acetic acid, NH3-N, water soluble carbohydrate and butyric acid was significantly (P < 0.05) decreased. Lactic acid, ethanol and propionic acid (PA) was significantly (P < 0.05) increased in treatments compared to control. The dry matter, propionic acid neutral detergent fiber, acid detergent fiber did not significantly (P < 0.05) differ among the treatments but the values were increased and decreased. The number of yeast, mold and LAB were significantly (P < 0.05). It is suggested that the supplementation of LAB could enhanced the fermentation quality of sweet Sorghum silage.

Lactobacillus silagincola sp. nov. and Lactobacillus pentosiphilus sp. nov., isolated from silage.

Three Gram-stain positive, non-motile, non-spore-forming, catalase-negative and rod-shaped bacterial strains (IWT5T, IWT25T and IWT140), isolated from silage, were investigated by using a polyphasic taxonomic approach. Strains IWT5T and IWT25T grew at 10-37 °C and 30-37 °C, and at pH 4.0-7.5 and 4.0-7.0, respectively. The G+C contents of genomic DNA of strains IWT5T and IWT25T were 43.2 and 44.4 mol%, respectively. Strains IWT5T and IWT25T contained C16 : 0, C18 : 1 ω9c and summed feature 7 (unknown 18.846/C19 : 1 ω6c/C19 : 0cyclo ω10c) as the major fatty acids. Strain IWT5T was most closely related to the type strains of Lactobacillus mixtipabuli (99.9 % 16S rRNA gene sequence similarity) and Lactobacillus silagei (99.5 %). For IWT25T, the 16S rRNA gene sequence similarities with the closely related neighbour type strains L. mixtipabuli and L. silagei were 99.5 and 99.5 %, respectively. The 16S rRNA gene sequence similarities among the three novel isolates were 99.5-99.9 %. The average nucleotide identities of strains IWT5T and IWT25T to other neighbours of the genus Lactobacillus were less than 82 % and the genomes of IWT25T and IWT140 shared 97.3 % average nucleotide identity, demonstrating that the three strains were allocated to two different novel species of the genus Lactobacillus. Together with multilocus sequence analysis, phenotypic and chemotaxonomic characteristics, strains IWT5T (=JCM 31144T=DSM 102973T) and IWT25T (=JCM 31145T=DSM 102974T) are proposed as the type strains of novel species of the genus Lactobacillus, with the names Lactobacillus silagincola sp. nov. and Lactobacillus pentosiphilus sp. nov., respectively.

Dose-response effects of dietary pequi oil on fermentation characteristics and microbial population using a rumen simulation technique (Rusitec).

The effect of increasing the concentration of commercial pequi (Caryocar brasiliense) oil on fermentation characteristics and abundance of methanogens and fibrolityc bacteria was evaluated using the rumen simulation technique (Rusitec). In vitro incubation was performed over 15 days using a basal diet consisting of ryegrass, maize silage and concentrate in equal proportions. Treatments consisted of control diet (no pequi oil inclusion, 0 g/kg DM), pequi dose 1 (45 g/kg DM), and pequi dose 2 (91 g/kg DM). After a 7 day adaptation period, samples for fermentation parameters (total gas, methane, and VFA production) were taken on a daily basis. Quantitative real time PCR (q-PCR) was used to evaluate the abundance of the main rumen cellulolytic bacteria, as well as abundance of methanogens. Supplementation with pequi oil did not reduce overall methane production (P = 0.97), however a tendency (P = 0.06) to decrease proportion of methane in overall microbial gas was observed. Increasing addition of pequi oil was associated with a linear decrease (P < 0.01) in dry matter disappearance of maize silage. The abundance of total methanogens was unchanged by the addition of pequi oil, but numbers of those belonging to Methanomassiliicoccaceae decreased in liquid-associated microbes (LAM) samples (P < 0.01) and solid-associated microbes (SAM) samples (P = 0.09) respectively, while Methanobrevibacter spp. increased (P < 0.01) only in SAM samples. Fibrobacter succinogenes decreased (P < 0.01) in both LAM and SAM samples when substrates were supplemented with pequi oil. In conclusion, pequi oil was ineffective in mitigating methane emissions and had some adverse effects on digestibility and selected fibrolytic bacteria.

Effect of grape pomace on fermentation quality and aerobic stability of sweet sorghum silage.

The objective of this study was to evaluate the effect of grape pomace (GP) with different adding levels (0%, 5%, 10% and 15%, fresh matter basis), alone (GP-LAB) or in combine with an inoculant LAB (GP+LAB), on the fermentation quality and aerobic stability of sweet sorghum silage. After 90 days of ensiling in vacuumized mini-silos, silages were subject to a 7-day aerobic stability test, in which chemical, microbial and polyphenol composition were measured. In the GP-LAB group, adding GP decreased (P < 0.05) concentrations of water-soluble carbohydrate (WSC) and butyric acid in silage. In the GP+LAB group, adding GP increased (P < 0.05) concentrations of lactic acid, WSC and crude protein, decreased (P < 0.05) final pH value, NH3 -N ratio and butyric acid concentration in silage. Polyphenol level was reduced (P < 0.05) after silage fermentation. During aerobic exposure, the fungi count, pH value and silage temperature increased (P < 0.05), the levels of lactic acid, acetic acid and polyphenols (quercetin 3-O-glucoside and quercetin 3-O-glucuronid) decreased (P < 0.05) in silage. GP+LAB treated silage had a lag phase for aerobic spoilage. When the fermentation products, microbial counts, chemical and polyphenol composition were considered, the use of 10% GP+LAB at ensiling could provide a valuable source for improved fermentation quality and aerobic stability of sweet sorghum silage.

In-vitro antibacterial, antifungal, antioxidant and functional properties of Bacillus amyloliquefaciens.

BACKGROUND: Food born pathogenic bacteria and filamentous fungi are able to grow on most foods, including natural foods, processed foods, and fermented foods and create considerable economic loss. The aim of this study was to determine the antibacterial, antifungal, antioxidant and functional properties of Bacillus amyloliquefaciens recovered from silage. METHODS: Minimum Inhibitory Concentration (MIC) of the compounds was assessed by using broth micro dilution method. The 1,1-diphenyl-2-picrylhydrazyl (DPPH)-radical scavenging and hydroxyl radical-scavenging abilities were measured to evaluate antioxidant activity of the strain. RESULTS: Primary antimicrobial compound production screening revealed that B. amyloliquefaciens exhibited significant activity against all the tested bacteria and fungi compared to other strains. The 16S rRNA and gyrase A gene sequence analysis determined using molecular biological tools confirmed that the strain was 99% similarity towards B. amyloliquefaciens. The Minimum Inhibitory Concentration (MIC) of ethyl acetate extract against Bacillus subtilis, Enterococcus cloacae and Staphylococcus aureus were 25.0 µg ml-1, and S, epidermidis were 12.5 µg ml-1, respectively. Filamentous fungi Aspergillus clavatus, A. fumigates, A. niger and Gibberella moniliformis showed 25 µg ml-1. VJ-1 was able to survive the gastrointestinal conditions simulating the stomach and duodenum passage with the highest percentage of hydrophobicity. In addition, its resistance to hydrogen peroxide and highest hydroxyl radical and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activities, with inhibition rates of 56.84% and 67.12% respectively, were its advantage. An antimicrobial susceptibility pattern was an intrinsic feature of this strain, and thus, consumption does not represent a health risk to humans. CONCLUSION: Bacillus amyloliquefaciens might be a promising candidate for new pharmaceutical agents and probiotics.

Lactobacillus mixtipabuli sp. nov. isolated from total mixed ration silage.

Using a polyphasic taxonomic approach, we investigated three bacterial strains - IWT30T, IWT8 and IWT75 - isolated from total mixed ration silage prepared in Hachimantai, Iwate, Japan. The isolates comprised Gram-stain positive, non-motile, non-spore-forming, catalase-negative, rod-shaped bacteria. Good growth occurred at 15-45 °C and at pH 4.0-7.5. Their major cellular fatty acids were C18:1ω9c and C19:1 cyclo 9,10.The G+C content of genomic DNA of strain IWT30T was 44.6 mol%. Comparative 16S rRNA gene sequence analysis showed that these novel strains belonged to the genus Lactobacillus. These strains shared 100 % 16S rRNA gene sequence similarity and were most closely related to the type strains of Lactobacillus silagei, Lactobacillus odoratitofui, Lactobacillus similis, Lactobacillus collinoides, Lactobacillus paracollinoides and Lactobacillus kimchicus, with sequence similarity values of 99.5, 98.8, 98.7, 97.8, 97.8 and 96.8 %, respectively. The level of DNA-DNA relatedness between these strains and their closest phylogenetic neighbours was less than 30 %. On the basis of additional phylogenetic analysis of pheS and rpoA gene sequences and phenotypic and chemotaxonomic characteristics, we conclude that these three strains represent a novel species of the genus Lactobacillus, for which we propose the name Lactobacillus mixtipabuli sp. nov. The type strain is IWT30T ( = JCM 19805T = DSM 28580T).