Lactic Acid Bacteria Inoculants Research Articles

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.

Improvements in fermentation and nutritive quality of elephant grass [Cenchrus purpureus (Schumach.) Morrone] silages: a review.

Elephant grass [Pennisetum purpureum Schumach. syn. Cenchrus purpureus (Schumach.) Morrone], also known as Napier grass and King grass, includes varieties Taiwán, Gigante, Merkerón, Maralfalfa, and others. The grass achieves high biomass production in tropical-subtropical, temperate, and arid areas. The high-water concentration of elephant grass suggests that ensiling could offer an alternative way to preserve the nutritional quality of the grass during storage, however, some considerations should be addressed because of the particularities of the grass. Ensiling elephant grass may produce adequate fermentation but could suffer effluent losses and subsequent losses of nutrients due to leaching. To improve fermentation and nutrient characteristics of elephant grass silages, several studies were conducted with the inclusion of additives. Lactic acid bacteria inocula have reduced pH and increased crude protein content of elephant grass silage, but aerobic stability of silages could be affected by the bacterial inoculation. There is limited information, however, on the potential of different silage inoculants to reduce growth of spoilage microorganisms during the aerobic phase of silage prepared with elephant grass. Exogenous fibrolytic enzymes also may improve elephant grass silage quality by enhancing microbial fiber-degradation with subsequent increase in lactic acid and its associated pH reduction. Another study approach to improve fermentation and nutritional quality of elephant grass silages involved the addition of different feeds at ensiling, including conventional feeds such corn, wheat, rice bran, and molasses or alternative feeds such as different dehydrated by-products obtained from the food industries of juice and jelly. In the manuscript, the presented scientific information shows the great potential of the different manipulations to improve the quality of elephant grass silages and with possible enhance of the economic profit and sustainability of livestock farming in the tropical areas.

Insights into fermentation with lactic acid bacteria on the flavonoids biotransformation of alfalfa silage

BackgroundOxidative stress is currently one of the main threats to animal health, and flavonoids in forage have good antioxidant activity. However, the impact of fermentation on flavonoids and their antioxidant activity in forage is still unclear. This study aims to investigate the effect of lactic acid bacteria inoculation on the biological transformation of flavonoids in alfalfa silage fermentation and its relationship with antioxidant activity.ResultsCompared with the raw materials, silage fermentation can increase the total flavonoid content of alfalfa. The addition of Pediococcus pentosaceus (CP115739.1) and Lactiplantibacillus plantarum (CP115741.1) can significantly increase the total flavonoid content in alfalfa silage (P < 0.05). The addition of lactic acid bacteria significantly improved the antioxidant capacity of alfalfa silage (P < 0.05). Pearson correlation analysis showed a significant correlation between total flavonoids and DPPH (R = 0.62, P < 0.05), and a highly significant correlation between total flavonoids and FRAP (R = 0.70, P < 0.01). Compared with natural silage fermentation, the addition of lactic acid bacteria leads to changes in the biological transformation process of flavonoids in alfalfa. Its unique products, 3,7,4′-trioxyflavonoids, as well as acacetin and taxifolin 7-O-rhamnoside, are significantly positively correlated with antioxidant activity.ConclusionsSilage fermentation contributes to the transformation of flavonoids, and inoculation with certain lactic acid bacteria can increase the content of flavonoids (including apigenin, luteolin, and other free flavonoids). It is worth noting that after fermentation, the antioxidant capacity of alfalfa is significantly improved, which may be attributed to the biotransformation of flavonoids related to acacetin, 3,7,4′-trihydroxyflavonoids, and taxifolin 7-O-rhamnoside. This study provides a potential pathway for obtaining value-added silage fermentation products by selecting specific lactic acid bacteria inoculants.Graphical

Effects of harvest date and additives on maize silage quality under boreal conditions

There is increasing interest in Finland to cultivate maize for silage, although the climatic conditions are in the borderline for maize due to the short and cool growing season. This may result in an immature crop that differs from typical maize for ensiling by having low dry matter (DM) and starch concentrations. We evaluated the preservation characteristics of forage maize during 2019 and 2020 harvested in Helsinki, Finland, at two stages of maturity. The DM concentration of the crops ranged from 230 to 360 g kg-1, and starch concentration from 179 to 283 g kg-1 DM. The crops were ensiled in laboratory scale using four different chemical organic acid based additives or a heterofermentative lactic acid bacteria inoculant. A control silage without additive treatment was also included. All silages were well fermented with low pH (average 3.75) and proportion of ammonia N in total N (average 47 g kg-1). Formic acid based additives restricted silage fermentation and most chemical additives improved the aerobic stability of maize silages compared to the control and inoculant treated silages that, under the conditions of the current study, did not differ from each other.

Fermentation Quality and In Vitro Digestibility of Sweet Corn Processing Byproducts Silage Mixed with Millet Hull or Wheat Bran and Inoculated with a Lactic Acid Bacteria

The aim of the experiment was to investigate the effect of different ratios of excipient (millet hull or wheat bran) and LAB inoculation on the fermentation quality and in vitro digestibility of a mixed silage of SCPBs. The preliminary experimental results showed that inoculating with lactic acid bacteria (LAB) directly in the fresh sweet corn processing byproduct (SCPBs) silage had a higher ammonia nitrogen/total nitrogen (AN/TN) ratio and lower silage fermentation quality due to high moisture content. Subsequently, millet hull or wheat bran were mixed with SCPBs in a 7:3 (T1), 8:2 (T2), and 9:1 (T3) ratio and ensiled with LAB. Under the condition of each mixing ratio, the silage treatments were categorized into groups without any additives (control) and with LAB. Fermentation quality, in vitro digestibility, chemical composition, and energy values were determined after 45 days of silage. The pH, AN/TN, neutral detergent fiber, acid detergent fiber, and acid detergent lignin were lowest in the SCPBs and millet hull mixed silage (SMH) group under the T3 treatment, whereas they were lowest in the SCPBs and wheat bran mixed silage (SWB) group under the T2 treatment. The mean lactic acid and acetic acid values were higher in the SWB group than in the SMH group (6.92, 6.81 vs. 4.00, 4.52). Under the T3 treatment in the SMH group, AN/TN was significantly reduced with the addition of LAB (4.52 vs. 4.37, p &lt; 0.05). The SMH group had the highest crude protein (CP) under the T3 treatment, whereas the SWB group had the highest CP under the T2 treatment. The mean CP in the SWB group was higher than that of the SMH group (18.17, 19.44 vs. 10.55, 10.55). Under the T1 treatment, in the SWB group, the addition of LAB resulted in a significant increase in in vitro crude protein digestibility (p &lt; 0.05). The results showed that silage fermentation quality and in vitro digestibilitv55y improved with the addition of LAB. The optimum mixing ratio for the SWB group was 9:1 and 8:2 for the SMH group.

Characterization of differences in physicochemical properties, volatile organic compounds and non-volatile metabolites of prune wine by inoculation of different lactic acid bacteria during malolactic fermentation

To investigate the effects of inoculating with three strains of lactic acid bacteria on prune wine quality during malolactic fermentation, this study determined its antioxidant activity, phenolic compounds, organic acids, and volatile/non-volatile metabolites. The results showed that inoculation with Lactobacillus paracasei SMN-LBK improved the antioxidant activity and phenolic compounds of prune wine. 73 VOCs were detected in prune wine by HS-SPME-GC–MS, and VOC content increased by 4.3% and 9.1% in MLFS and MLFB, respectively. Lactobacillus delbrueckii subsp. Bulgaricus showed better potential for winemaking, and citral and 5-nonanol, were detected in the MLF samples. 39 shared differential metabolites were screened and their metabolic pathways were investigated based on nontargeted metabolomics. Differences in amino acid and flavonoid content between strains reflected their specificity in flavonoid biosynthesis and amino acid biosynthesis. These findings will provide useful information for the biochemical study and processing of prune wine.

Effect isolated lactic acid bacteria inoculation on the quality, bacterial composition and metabolic characterization of Caragana korshinskii silage

Feed shortage has hindered the development of the world’s livestock industry, and the rational use of non-conventional forages can help resolve this issue. To improve the fermentation quality of Caragana korshinskii silage, this study isolated lactic acid bacteria (LAB) from fresh Caragana korshinskii and silage samples, as alternatives to traditional commercial LAB inoculants. Utilizing Single Molecule, Real-Time (SMRT) sequencing technology and metabolomics analysis, the microbial community structure and metabolome were examined after 60 days of fermentation to elucidate the mechanism by which isolated LAB affect the fermentation quality of Caragana korshinskii silage. After 60 days of ensiling, the dominant microbial population in the silage shifted from harmful bacteria such as Erwinia, Pantoea, and Enterobacter to LAB. Compared to commercial LAB, the isolated Lactiplantibacillus plantarum significantly reduced the microbial diversity of Caragana korshinskii silage. The species composition of LAB became simpler with Lentilactobacillus parakefiri dominating and achieving a higher relative abundance. By down-regulating Butanoate metabolism and the Pentose phosphate pathway and up-regulating the Ascorbate and aldarate metabolism pathway, the isolated Lactiplantibacillus plantarum significantly lowered the pH of the silage, increased the content of water-soluble carbohydrates (WSC) and lactic acid (LA), reduced the neutral detergent fiber content (NDF), and improved in vitro neutral detergent fiber digestibility (IVNDFD), thereby enhancing the fermentation quality of Caragana korshinskii silage. On the other hand, the isolated Limosilactobacillus fermentum had the opposite effect, with its ensiling outcome being less effective than that of commercial LAB. The results of the study indicate that the addition of the isolated Lactiplantibacillus plantarum improved the quality of Caragana korshinskii silage by regulating metabolic pathways and the composition of microbes within the ensiled feed.Graphical

Effects of lactic acid bacteria inoculants on the nutrient composition, fermentation quality, and microbial diversity of whole-plant soybean-corn mixed silage.

The mixture of whole-plant soybean and whole-plant corn silage (WPSCS) is nutrient balanced and is also a promising roughage for ruminants. However, few studies have investigated the changes in bacterial community succession in WPSCS inoculated with homofermentative and heterofermentative lactic acid bacteria (LAB) and whether WPSCS inoculated with LAB can improve fermentation quality by reducing nutrient losses. This study investigated the effect of Lactobacillus plantarum (L. plantarum) or Lactobacillus buchneri (L. buchneri) on the fermentation quality, aerobic stability, and bacterial community of WPSCS. A 40:60 ratio of whole-plant soybean corn was inoculated without (CK) or with L. plantarum (LP), L. buchneri (LB), and a mixture of LP and LB (LPB), and fermented for 14, 28, and 56 days, followed by 7 days of aerobic exposure. The 56-day silage results indicated that the dry matter content of the LP and LB groups reached 37.36 and 36.67%, respectively, which was much greater than that of the CK group (36.05%). The pH values of the LP, LB, and LPB groups were significantly lower than those of the CK group (p < 0.05). The ammoniacal nitrogen content of LB was significantly lower than that of the other three groups (p < 0.05), and the ammoniacal nitrogen content of LP and LPB was significantly lower than that of CK (p < 0.05). The acetic acid content and aerobic stability of the LB group were significantly greater than those of the CK, LP, and LPB groups (p < 0.05). High-throughput sequencing revealed a dominant bacteria shift from Proteobacteria in fresh forage to Firmicutes in silage at the phylum level. Lactobacillus remained the dominant genus in all silage. Linear discriminant analysis effect size (LEFSe) analysis identified Lactobacillus as relatively abundant in LP-treated silage and Weissella in LB-treated groups. The results of KEGG pathway analysis of the 16S rRNA gene of the silage microbial flora showed that the abundance of genes related to amino acid metabolism in the LP, LB, and LPB groups was lower than that in the CK group (p < 0.05). In conclusion, LAB application can improve the fermentation quality and nutritional value of WPSCS by regulating the succession of microbial communities and metabolic pathways during ensiling. Concurrently, the LB inoculant showed the potential to improve the aerobic stability of WPSCS.

Impact of lactic acid bacteria inoculation on fungal diversity during Spanish-style green table olive fermentations

This study delved into the evolution of fungal population during the fermentation of Spanish-style green table olives (Manzanilla cultivar), determining the influence of different factors such as fermentation matrix (brine or fruit) or the use of a lactic acid bacteria inoculum, on its distribution. The samples (n = 24) were directly obtained from industrial fermentation vessels with approximately 10.000 kg of fruits and 6.000 L of brines. Our findings showcased a synchronized uptick in lactic acid bacteria counts alongside fungi proliferation. Metataxonomic analysis of the Internal Transcribed Spacer (ITS) region unearthed noteworthy disparities across different fermentation time points (0, 24, and 83 days). Statistical analysis pinpointed two Amplicon Sequence Variants (ASV), Candida and Aureobasidium, as accountable for the observed variances among the different fermentation time samples. Notably, Candida exhibited a marked increase during 83 days of fermentation, opposite to Aureobasidium, which demonstrated a decline. Fungal biodiversity was slightly higher in brines than in fruits, whilst no effect of inoculation was noticed. At the onset of fermentation, prominently detected genera were also Mycosphaerella (19.82 %) and Apohysomyces (16.31 %), hitherto unreported in the context of table olive processing. However, their prevalence dwindled to nearly negligible levels from 24th day fermentation onwards (<2 %). On the contrary, they were replaced by the fermentative yeasts Saccharomyces and Isstachenkia. Results obtained in this work will be useful for designing new strategies for better control of table olive fermentations.

Optimization of the fermentation process and antioxidant activity of mixed lactic acid bacteria for honeysuckle beverage.

The aim of the research was to obtain a high healthcare honeysuckle beverage with strong antioxidant activity. Honeysuckle (Lonicera japonica Thunb) was used as the raw material in this experiment. The effects of fermentation temperature, fermentation time, lactic acid bacteria inoculation amount, and sugar addition amount on the sensory quality of honeysuckle beverage were investigated by single factor test and orthogonal test, and the best process was obtained. The physicochemical indexes and antioxidant activity of honeysuckle beverages fermented with lactic acid bacteria were studied. The results showed that the fermentation temperature of the beverage was 37 °C, the fermentation time was 24 h, the inoculation amount of Lactiplantibacillus plantarum and Lactobacillus acidophilus mixed starter (1:1) was 3%, and 8% white granulated sugar was added. The highest sensory score was 87.30 ± 0.17, which was the optimal process. The honeysuckle liquid mixed inoculation with Lactiplantibacillus plantarum and Lactobacillus acidophilus was fermented for 24 h. The number of viable bacteria reached 9.84 ± 0.02 lg cfu/mL, the pH value was 3.10 ± 0.01, and the total polyphenol content was 7.53 ± 0.03 mg GAE/g. The number of lactic acid bacteria, pH, total polyphenol content, and free radical scavenging rate were significantly increased (p < 0.05) compared with the non-inoculated and single-inoculated lactic acid bacteria. To sum up, it was concluded that a better quality beverage could be obtained by fermenting a solution of honeysuckle with Lactiplantibacillus plantarum and Lactobacillus acidophilus mixed fermentation agent, providing a new approach and new ideas for the development of deep processing and fermented beverages using honeysuckle.

Потенціал росту стерляді прісноводної (Acipenser ruthenus Linnaeus, 1758) в ювенальний період за використання пробіотичного препарату «Емпробіо»

Purpose. To study the effect of the complex probiotic “Emprobio” on the survival and growth rates of larvae and fry of the sterlet (Acipenser ruthenus) as well as to assess its effect on the morphological structure of the intestine and liver. Methodology. The study of the effect of the probiotic was carried out on larvae and fry of sterlet. 6-hour enrichment of live feed organisms was carried out in 2 variants. In the variant 1, the concentration of the probiotic preparation was 0.4 cm3/dm3 of the incubation mixture. In the variant 2, similarly, the preparation concentration was 4.0 cm3/dm3. Larvae and fry were fed with Artemia nauplii, which were the transfer of the probiotic. Every 7 days, the average fish weight, the growth and survival rates were analyzed. Determination of the number of colonies of lactic acid bacteria in the intestines of fish was carried out by the method of deep seeding in Petri dishes. Determination of unicellular fungi was carried out by the method of serial dilutions of intestinal contents in a peptone-buffer solution with subsequent inoculation on a sterile wort-agar medium. Light-optical examination of cells and tissues was carried out in penetrating light using the equipment “E. Leitz “diaplan” Wetzlan” (Germany), halogen illuminator “Linvatec-2” (USA). General morphometric studies of tissue structures were performed with the help of a built-in eyepiece-micrometer as well as with the use of an eyepiece-micrometer overlay grid (eyepiece 7x (Huygens). In-depth studies of individual tissue areas were carried out using a 100x objective, “I-Plan” with a built-in iris diaphragm of 5.5 units of a compensation 20x eyepiece “K. Zeiss”. The data were statistically processed in MS Excel. Findings. During the 1st 7 days of the experiment, there were no significant differences in the growth rate of larvae between the groups.In the 2nd week of the experiment, an increase in the average daily weight gain of fish was observed in the experimental groups, which exceeded the control by 47.0% and 24.7%, respectively.At the end of the experiment, the average weight of fry of the 1st group was higher by 26.9%, and of the 2nd — by 27.9%, compared to the control group. As a result of the use of the probiotic preparation for 28 days, the survival rate of sterlet juveniles in the 1st experimental group was 75.0%. That is, it was 47.1% higher than the results of the control group. In the 2nd experimental group, the survival rate of sterlet juveniles was 80.1%, which was 6.8% higher than in the 1st group, and 57.1% higher than in the control group. The use of “Emprobio” in the feed at a concentration of 4.0 cm3/dm3 contributed to a faster process of microflora redistribution in the intestine. Therefore, already after the 1st 5 days of exogenous feeding, the average daily growth rates of the juveniles in this experimental group had higher values (by 8.8%) compared to the control group. The results of inoculation the intestine contents in the experimental groups showed that the use of the probiotic preparation in the incubation mixture in the amount of 0.4 cm3/dm3 ensured the inoculation of probiotic lactic acid bacteria in the intestines of sterlet juveniles. Their functioning was reflected in fish productive parameters, starting from the 14th day of the experiment. However, the lack of intestinal inoculation by one of the components of the preparation, namely saccharomycetes, indicated the insufficiency of such a concentration to show the full potential of the probiotic. In the 2nd experimental group, positive growth dynamics of the entire microbiome can be seen. Histological studies showed a positive effect of the probiotic preparation at a concentration of 4.0 cm3/dm3 on the morphological structure of the liver. In particular, the area of hepatocytes, which increased by 42.9% after 7 days of the experiment and by 63.0% after 21 days of the experiment. The sizes of the nuclei and the ratio of the diameter of the nucleolus/nucleus of hepatocytes also increased. At the same time, the probiotic did not cause probable changes in the morphological structure of the intestinal wall. Originality. For the first time, the possibility of using the complex probiotic “Emprobio”, introduced by enrichment in live feed for growing sterlet in the juvenile period was analyzed.A positive effect of probiotics at a concentration of 4.0 cm3/dm3 on growth processes and survival of starlet juveniles was shown. The effect of microorganisms on the histostructure of the intestine and liver of the sterlet was clarified. Practical value. The obtained results will be useful for improving the technology of growing sturgeon larvae and fry. Keywords: sterlet, growth rate, survival, enrichment, probiotic, Emprobio, intestine, liver.

Forage type and additive effects on fermentation quality and biorefinery performance of silages

AbstractThe concept of a green biorefinery has recently gained interest and can be defined as a process where green biomass is processed to a variety of products. Most green biorefineries rely on fresh green biomass as the feedstock, but using a stable ensiled biomass could provide benefits. We evaluated the effects of silage additive treatments on silage fermentation quality and performance in a laboratory scale liquid–solid separation to simulate the first step of a green biorefinery. In Experiment 1, red clover and fresh and wilted grass were ensiled without additive or treated with a lactic acid bacteria inoculant or a formic acid based additive. In Experiment 2, grass or red clover were treated with a fibrolytic enzyme, formic acid or a combination of them, and a control without additive was also included. Silage fermentation quality was improved by additive use. Biomass dry matter concentration was negatively related to liquid yield, but effects of additive treatments on the biorefinery performance were minor and inconsistent between different forages. Optimizing agronomic and feedstock conservation management plays an important role for the success and sustainability of the biorefinery process. Good silage management practices with minimal losses during storage should be targeted, but no clear patterns in biorefinery outputs were observed in the current study when different types of additives were used in grass and clover silage production.

Heat-resistant lactic acid bacteria inoculants modulated the bacterial microbiota and fermentation quality of whole plant maize silage after long-term storage in the subtropical area

In tropical or subtropical areas, high temperatures and prolonged storage time usually result in poor fermentation quality, poor aerobic stability and dry matter loss of silages. This study explored the effect of heat-resistant Lactobacillus buchneri TSy1–3 (LB), Lactobacillus rhamnosus BDy3–10 (LR), and their combination (M) on the nutritional characteristics, fermentation parameters and microbiota of whole plant maize silage during long-duration ensiling in the subtropical area. The results showed that long-term ensiling naturally led to a decrease in water-soluble carbohydrate, neutral detergent fiber and acid detergent fiber concentrations. In contrast to the control treatment, LB, LR and M reduced the coliform bacteria population and the nutrient loss, while increased lactic acid bacteria population and acetic acid and propionic acid concentrations after 180 d of fermentation. Moreover, they helped to increase Lactobacillus abundance and made it still dominate the bacterial community during long-term storage. The negative/positive ratio was higher in the LR and LB treatments when compared with the control treatment, suggesting that LR and LB increased the stability of the bacterial community networks. The heat-resistant inoculants rapidly produced lactic acid to modulate the bacterial community composition during 60 d of ensiling, while generated more acetic acid and propionic acid to alter the microflora during 180 d of ensiling. This may be caused by predicted functions indicating that metabolism pathways were modulated by different inoculations and storage times. Overall, heat-resistant lactic acid bacteria improved the nutritional and fermentation quality of ensiled forage and regulated the bacterial community during long-term storage in the subtropical region.

MICROBIOLOGICAL PROCESSES IN THE LUCERNE HAYLAGE UNDER INOCULATION WITH LACTIC ACID BACTERIA

Objective. To characterize the effect of lactic acid bacteria (LAB) on the quantitative and qualitative composition of the groups of microorganisms in the lucerne haylage after their introduction. Methods. Microbiological (cultivation of microorganisms on various media, determination of their viability, measurement of osmotolerance, determination of the amount of lactic acid during cultivation on skimmed sterile milk), microscopic, laboratory experiments on the lucerne haying with a dry matter (DM) content of 46 % and 37 % with and without inoculation with LAB strains, statistical. Results. Lucerne was inoculated with monocultures of Lactobacillus sp. LH42, Lactobacillus sp. LH43, Lactobacillus sp. LH47, L. plantarum КТ-L18/1 osmotolerant strains, which showed tolerance to 2 %, 4 %, 6.5 % sodium chloride in 100 % of cases. The best indicators in terms of active and titrated acidity were found in cultures of Lactobacillus sp. LН47, L. plantarum КТ-L18/1. The inoculation of LAB into the haylage ecosystem at Day 15 of fermentation resulted in an increase in the number of LAB compared to the untreated control. At Day 45 of lucerne haying, the number of LAB decreased and was registered at the level from 1 to 100 billion CFU/g at a 46 % DM content and at the level from 0.01 to 10 billion CFU/g at a 37 % DM content. The greatest development of clostridia in the lucerne haylage in experiments was reported at Day 3 with a sharp decrease in their number at Day 45. The influence of Lactobacillus sp. LH 43, Lactobacillus sp. LH 47, L. plantarum KT-L18/1 on the reduction of the growth of the number of clostridia both at a 46 % and 37 % DM content at the end of the fermentation period was shown. A higher number of yeasts was established at a lower (37 %) content of DM in the haylage. A fungistatic effect was established in variants with LAB inoculation versus the control variant. The relationship between the dynamics of decreasing active acidity (pH) and the dynamics of microbiota development has been shown. Conclusion. Inoculation of the studied strains of lactic acid bacteria into high-protein lucerne haylage mass contributed to an order of magnitude increase in the number of LAB versus the control variant, suppression of the development of clostridia, yeasts and a rapid decrease in active acidity, which confirms data about the effect of LAB-based inoculants on the fermentation of plant raw materials by adjusting the composition of groups of microorganisms

The Effects of Grass Silage Additive Type and Barley Grain Preservation Method on Rumen Function, Microbial Ecology, and Energy Metabolism of Dairy Cows

The effects of grass silage and barley grain preservation methods on dairy cows were evaluated using four Nordic Red dairy cows placed in respiration chambers in a 4 × 4 Latin square. Silage was conserved using a formic acid-based product (AS) or a homofermentative lactic acid bacteria inoculant (IS), while grains were dried (DB) or crimped and ensiled (EB). Fermentation profile of silages and the chemical composition of the mixed diets were very similar. The dietary treatments did not affect feed intake, milk production, and rumen fermentation except molar proportion of butyrate, and energy metabolism. Digestibility of dry matter and organic matter were higher (p &lt; 0.05) and that of crude protein was lower (p &lt; 0.05) for AS than IS. Feeding EB compared to DB decreased (p &lt; 0.05) diet organic matter and starch digestibility. The cows receiving AS tended (p = 0.06) to emit more methane per day than those receiving IS, but methane yield and intensity were not different between dietary treatments. Bacteria alpha diversity was higher (p &lt; 0.01) in barley samples than grass silages and was not affected by the diet in rumen samples. All freshly prepared rations were dominated by Lactobacillaceae, Erwiniaceae, and Pseudomonadaceae but rations based on AS than IS remained more stable over 2 days. In conclusion, grass silage and barley grain preservation methods did not affect the measured parameters in dairy cows and the preservation method can be selected based on practical on-farm factors.

Microbial count, chemical composition and fatty acid profile of biological silage obtained from pacu and spotted sorubim fish waste using lactic acid bacteria fermentation

With the increasing production and processing of fish, managing the generated waste is a growing concern. Fish processing by-products, like viscera, are rich in energy and nutrients and can be transformed into valuable raw materials, including fish silage, which represents an alternative to fishmeal. This study evaluated the quality of biological silage obtained from the fermentation of the waste of two fish species, pacu and spotted sorubim, by seven lactic acid bacteria inoculants. Cultivations were conducted using either 100% fish viscera or mixture of 90% fish viscera and 10% sugarcane molasses as the substrate at 30 or 35 °C, for 120 h, with an inoculum concentration of 6% (v/w) in the bioreactors. Results showed variations (P < 0.05) in microbiological count, chemical composition, and fatty acid profile in the silages after 5 days of fermentation. Spotted sorubim-based media supported better lactic acid bacteria growth, and the silages exhibited varying fatty acid profiles, depending on the formulation. Media inoculated with Lactobacillus brevis NRRL-B 4527 containing 90% spotted sorubim viscera and 10% sugarcane molasses at 30 °C had the highest protein content (44.6 ± 0.7% dry basys). Oleic (C18:1n-9), palmitic (C16:0), and stearic (C18:0) acids were the most abundant fatty acids. The LAB/MES ratio of 0.96 and pH final of 4.21 underline the microbial quality of this silage.

Effects of exogenous lactic acid bacteria and maize meal on fermentation quality and microbial community of Orychophragmus violaceus silage.

Orychophragmus violaceus is a local Brassicaceae in China, while most of it is directly mowed and discarded after the ornamental period. In order to develop forage resources, this study firstly evaluated the potential preservation of O. violaceus silage. O. violaceus was harvested at full-bloom stage, and ensiled without (CK) or with maize meal (Y5), lactic acid bacteria inoculant (Z) and compound additive (Y5Z) for 60 d. Results of chemical and microbiological analysis showed that a large amount of lactic acid was produced and the final pH value was below 4.1 in silages regardless of additive application. CK silage was well preserved as indicated by the low levels of dry matter loss and butyric acid content, and the predominant genus were identified as Enterococcus and Pediococcus. Y5 silage had potential health risks for humans and animals as seen by frequent occurrence of pathogenic bacteria Clostridium and Achromobacter. Z and Y5Z silages were poorly preserved, resulting in great dry matter loss and butyric acid content. Considering the abundant acetic acid production, the dominant Lactobacillus might possess a heterofermentative pathway in Z and Y5Z silages. In conclusion, O. violaceus has the potential to be long stored as silage because of its sufficient water-soluble carbohydrates, while exogenous lactic acid bacteria and maize meal generally provided little positive effect. In future research, efficient homofermentative Lactobacillus strains were suggested to be screened to further enhance the ensiling process of O. violaceus silage.

Effect of cutting height, a bacterial inoculant and a fibrolytic enzyme on corn (Zea mays L.) silage quality

This study aimed to evaluate cutting height (CH) effects on ensiled corn without additives (C), with a lactic acid bacteria inoculant (L), a fibrolytic enzyme (F), or a mixture of both (FL), considering chemical composition and both in vitro digestibility dry matter (IVDMD) and in vitro neutral detergent fiber (IVNDFD). Corn was harvested at three different cutting heights (12, 25 or 42 cm above the soil) and ensiled with or without additives (AD). Data was analyzed according to a factorial design, with a 3 x 4 arrangement of treatments and three repeats. Dry matter content was highest in C12 and lowest in F12 (P&lt;0.05) silages. As cutting height was higher, cell wall content was lower (P&lt;0.05). Even considering it increased after the use of additives (P&lt;0.05), the highest values occurred with FL silages. Crude protein was equal (P˃0.05) between CH and increased (P&lt;0.05) with AD. The highest IVDMD was observed for 42 cm CH, while IVDMD and IVNDFD were higher in C and F, but lower with FL. None of the inoculation treatments, alone or combined improved corn silage quality. In fact, FL combination decreased such quality. Highlights: The efficacy of lactic acid bacteria (LAB) and exogenous fibrolytic enzyme (EFE) is a funtion of the amount of cell walls and sugars present in the maize plant at harvest. Adding LAB and EFE to maize forage harvested at different cutting heights did not improve silage quality, in fact, the combination of both additives decreased silage quality. Further studies are needed to prove the efficacy of EFE and its combination with LAB on maize silage quality.

Comprehensive profiling of the metabolome in corn silage inoculated with or without Lactiplantibacillus plantarum using different untargeted metabolomics analyses

ABSTRACTSilage fermentation is a complicated biochemical process involving interactions between microbes and metabolites. However, the overall metabolome feature of ensiled forage and its response to lactic acid bacteria inoculation is poorly understood. Hence, in this study metabolome profiles of whole-plant corn silage inoculated with or without Lactiplantibacillus plantarum were characterised via solid-phase microextraction/gas chromatography/mass spectrometry (SPME-GC-MS), gas chromatography/time-of-flight mass spectrometry (GC-TOF-MS), and Liquid chromatography/Q Exactive HFX mass spectrometry (LC-QE-MS/MS) analysis. There were 2087 identified metabolites including 1143 reliably identified metabolites in fresh and ensiled whole-plant corn. After ensiling, the increased metabolites in whole-plant corn were mainly composed of organic acids, volatile organic compounds (VOC), benzene and substituted derivatives, carboxylic acids and derivatives, fatty acyls, flavonoids, indoles and derivatives, organooxygen compounds (including amines and amides), phenols, pyridines and derivatives, and steroids and steroid derivatives, which includes neurotransmitters and metabolites with aromatic, antioxidant, anti-inflammatory, and antimicrobial activities. Phenylacetaldehyde was the most abundant aromatic metabolite after ensiling. L-isoleucine and oxoproline were the major free amino acids in silage. Ensiling markedly increased the relative abundances of 3-phenyllactic acid, chrysoeriol, 6-O-acetylaustroinulin, acetylcholine, γ-aminobutyric acid, pyridoxine, and alpha-linoleic acid. Inoculation with L. plantarum remarkably changed silage VOC composition, and essential amino acids, 3-phenyllactic acid, and cinnamaldehyde compared with untreated silage. The present study does not only provide a deeper insight into metabolites of the ensiled whole-plant corn but also reveals metabolites with specific biological functions that could be much helpful in screening novel lactic acid bacteria to well ensile forages. Inoculation with L. plantarum significantly affects the metabolome in ensiled whole-plant corn.

Autumn grass treated with a hydrolysable tannin extract versus lactic acid bacteria inoculant: Effects on silage fermentation characteristics and nutritional value and on performance of lactating dairy cows.

Hydrolysable tannins (HT) show potential as silage additive for autumn herbage silages, high in (rumen degradable) protein, as they may reduce proteolysis.Additionally, they have abilities to form pH-reversible tannin-protein complexes, non-degradable in the rumen but degradable in theabomasum and intestines of ruminants. Therefore they can improve milk N efficiency and shift N excretions from urine to faeces, possibly mitigating the environmental impact of ruminants. In this study, two small bunker silos were filled with autumn grass. One was treated with 20 g/kg DM HT extract (TAN) (TannoSan-L), the other with 8 mg/kg DM inoculant containing lactic acid bacteria (INO) (Bonsilage Fit G). Secondly, micro-silos (2.75 L) were filled with four treatments; (1) grass without additive (CON) (n = 5); (2) TAN (n = 5); (3) INO (n = 5); and (4) TAN + INO (n = 5). The bunker silos were used in a cross-over feeding experiment with periods of 4 weeks involving 22 lactating Holstein cows (average ± SD: 183 ± 36.3 days in milk, 665 ± 71.0 kg body weight, and 33.8 ± 3.91 kg/day milk yield). The HT dose was insufficient to reduce proteolysis or alter chemical composition and nutritional value in the micro- and bunker silages. Including grass silage added with TAN (3.2 g HT/kg DM) in the diet, did not affect feed intake nor fat and protein corrected milk yield in comparison to feeding the grass silage added with INO in a similar diet. The TAN-fed cows had an increased faecal N excretion and decreased apparent total-tract N and organic matter digestibility, but no improvement in the cows' N utilization could be confirmed in milk and blood urea levels. Overall, feeding an autumn grass silage treated with 20 g/kg chestnut HT extract did not affect the performance of dairy cows in comparison to feeding an autumn grass silage treated with a lactic acid bacteria inoculant.