Effect of pig manure to grass silage ratio on methane production in batch anaerobic co-digestion of concentrated pig manure and grass silage

Anaerobic co-digestion of concentrated pig manure (PM) with grass silage (GS) at five different PM to GS volatile solid (VS) ratios of 1:0, 3:1, 1:1, 1:3 and 0:1 was evaluated by examining operation stability and methane (CH₄) production potentials. The highest specific CH₄ yields were 304.2 and 302.8 ml CH₄/g VS at PM to GS ratios of 3:1 and 1:1, respectively. The digestion systems failed at the ratio of 0:1. The lag phase lasted 29.5, 28.1, 24.6 and 21.3 days at the ratios of 1:0, 3:1, 1:1 and 1:3, respectively. The daily methane yield was linearly correlated with the acetic acid concentration, indicating methane production was probably associated with acetoclastic methanogenesis. The hydrolysis constant linearly decreased with increasing the fraction of GS in the feedstock. This study recommends applying the PM to GS ratio of 1:1 in practice due to a high specific methane yield and a short lag phase.     

Investigation of mircroorganisms colonising activated zeolites during anaerobic biogas production from grass silage

The colonisation of activated zeolites (i.e. clinoptilolites) as carriers for microorganisms involved in the biogas process was investigated. Zeolite particle sizes of 1.0-2.5 mm were introduced to anaerobic laboratory batch-cultures and to continuously operated bioreactors during biogas production from grass silage. Incubation over 5-84 days led to the colonisation of zeolite surfaces in small batch-cultures (500 ml) and even in larger scaled and flow-through disturbed bioreactors (28 l). Morphological insights were obtained by using scanning electron microscopy (SEM). Single strand conformation polymorphism (SSCP) analysis based on amplification of bacterial and archaeal 16S rRNA fragments demonstrated structurally distinct populations preferring zeolite as operational environment. via sequence analysis conspicuous bands from SSCP patterns were identified. Populations immobilised on zeolite (e.g. Ruminofilibacter xylanolyticum) showed pronounced hydrolytic enzyme activity (xylanase) shortly after re-incubation in sterilised sludge on model substrate. In addition, the presence of methanogenic archaea on zeolite particles was demonstrated.     

Effects of thermo-chemical pre-treatment of grass silage on methane production by anaerobic digestion

Dried grass silage (GS) was pre-treated at different NaOH loading rates (1%, 2.5%, 5% and 7.5% by volatile solids (VS) mass in grass silage) and temperatures (20 °C, 60 °C, 100 °C and 150 °C) to determine effects on its bio-degradability in terms of the hydrolysis yield and degradation of ligno-cellulosic materials for biogas production. At 100 °C and the four NaOH loadings, up to 45% of the total COD was solubilised and up to 65.6%, 36.1% and 21.2% of lignin, hemicellulose and cellulose were removed, respectively; biological methane production potentials obtained were 359.5, 401.8, 449.5 and 452.5 ml CH?/g VS added, respectively, being improved by 10-38.9% in comparison with untreated GS. VS removals following anaerobic digestion were 67.6%, 76.9%, 85.3%, 95.2% and 96.7% for untreated GS and GS treated at the four NaOH loadings, respectively. 100 °C and the NaOH loading rate of 5% is recommended as a proper GS pre-treatment condition.     

Variation of ADM1 by using temperature-phased anaerobic digestion (TPAD) operation

The objective of the study was to examine the application of the Anaerobic Digestion Model No. 1 (ADM1) developed by the IWA task group for mathematical modelling of anaerobic process. Lab-scale temperature-phased anaerobic digestion (TPAD) process were operated continuously, and were fed with co-substrate composed of dog food and flour. The model platform implemented in the simulation was a derivative of the ADM1. Sensitivity analysis showed that k_m.process (maximum specific uptake rate) and K_S.process (half saturation value) had high sensitivities to model components. Important parameters including maximum uptake rate for propionate utilisers (k_m.pro) and half saturation constant for acetate utilisers (K_S.ac) in the thermophilic digester and maximum uptake rate for acetate utilisers (k_m.ac) in the mesophilic digester were estimated using iterative methods, which optimized the parameters with experimental results. Simulation with estimated parameters showed good agreement with experimental results in the case of methane production, uptake of acetate, soluble chemical oxygen demand (SCOD) and total chemical oxygen demand (TCOD). Under these conditions, the model predicted reasonably well the dynamic behavior of the TPAD process for verifying the model.     

Analysis and application of ADM1 for anaerobic methane production

An anaerobic model for the serum bottle test was developed and analyzed with sensitivities of stoichiometric and kinetic parameters to the components in order to establish a basis for appropriate application of the model. Anaerobic glucose degradation in a serum bottle was selected as an example. The anaerobic model was developed based on the anaerobic digestion model no. 1 (ADM1), which had five processes with 17 kinetic and stoichiometric parameters. Sensitivity analysis showed that the yield of product on the substrate (f) has high sensitivities to model components, and that the methane concentration was the most sensitive component. Important parameters including yield of product on the substrate (f), yield of biomass on the substrate (Y), and half-saturation values (K) were estimated using genetic algorithms, which optimized the parameters with experimental results. The Monod maximum specific uptake rate (k) was, however, so strongly associated with the concentration of biomass, that values could not be estimated individually. Simulation with estimated parameters showed good agreement with experimental results in the case of methane production. However, there were some differences in acetate and propionate concentrations.     

Electricity generation in single-chamber microbial fuel cells using a carbon source sampled from anaerobic reactors utilizing grass silage

Production of electricity from samples obtained during anaerobic digestion of grass silage was examined using single-chamber air-cathode mediator-less microbial fuel cells (MFCs). The samples were obtained from anaerobic reactors at start-up conditions after 3 and 10 days of operation under psychrophilic (15 °C) and mesophilic (37 °C) temperatures. Electricity was directly produced from all samples at a concentration of 1500 mg COD L⁻¹. Power density obtained from the samples, as a sole carbon source, ranged from 56 ± 3 W m⁻³ to 31 ± 1 W m⁻³ for the mesophilic and psychrophilic samples, respectively. Coulombic efficiencies ranged from 18 ± 1% to 12 ± 1% for the same samples. The relationship between the maximum voltage output and initial COD concentration appeared to follow saturation kinetics at the external resistance of 217 Ω. Chemical oxygen demand (COD) removal was over 90% and total phenolics removal was in the range of 30-75% for all samples tested, with a standard amount of 60 mg L⁻¹ total phenolics removed for every sample. Our results indicate that generating electricity from solution samples of anaerobic reactors utilizing grass silage is possible, opening the possibility for combination of anaerobic digestion with MFC technology for energy generation.     

Intake, rumen fermentation and nutrient flow to the omasum in beef cattle fed grass silage fortified with sucrose and/or supplemented with concentrate

The objective was to determine the relative effects of a specific increase in grass silage sucrose concentration, or a specific supplement of a starch-based concentrate, on rumen fermentation and nutrient supply to the omasum in beef cattle. Four ruminally cannulated Holstein–Friesian steers were fed grass silage only (G), G plus 3 kg concentrates/day (GC), G plus 90 g sucrose/kg dry matter (DM) (GS) and G plus 90 g sucrose/kg DM plus 3 kg concentrates/day (GCS) in a 4 × 4 Latin Square design experiment. Omasal flow was estimated using Co-EDTA, Yb-acetate and indigestible neutral detergent fibre (INDF) as digesta flow markers and purine bases as microbial markers. Concentrate supplementation reduced (P < 0.01) silage and increased (P < 0.001) total DM intake whereas sucrose had no effect. There was a sucrose × concentrate interaction (P < 0.05) for rumen pH whereby addition of sucrose to grass silage alone decreased pH and to grass silage plus concentrate had no effect. Rumen ammonia N (P < 0.01), total volatile fatty acid (VFA) concentration (P < 0.05) and the molar proportions of valerate (P < 0.05) and butyrate (P < 0.001) increased with concentrate supplementation whereas, sucrose supplementation had no effect on rumen fermentation parameters. Organic matter (OM) intake, omasal OM flow, the quantities of OM apparently (OMAD) and truly digested (OMTD) in the rumen (P < 0.001) and total tract OM digestibility (P < 0.01) increased, and apparent and true ruminal OM digestibility decreased (P < 0.05) with concentrate supplementation. Supplementation with concentrate decreased (P < 0.05) ruminal neutral detergent fibre (aNDFom) digestibility and increased (P < 0.05) aNDFom omasal flow. There was a tendency for addition of sucrose to increase (P < 0.1) ruminal OMAD and OMTD, while there was no effect of sucrose addition on intake or digestion of aNDFom. Concentrate supplementation increased (P < 0.001) N intake, flows of N, non-ammonia N (NAN), microbial N (MN) (P < 0.05) and non-ammonia non-microbial N (NANMN) (P < 0.01) and apparent total tract digestibility of N (P < 0.01), whereas sucrose reduced (P < 0.05) N intake and apparent ruminal N digestibility. There was no effect of concentrate or sucrose on N use efficiency or efficiency of microbial protein synthesis. Concentrate supplementation increased (P < 0.001) plasma β-hydroxybutyrate levels. In comparison to supplementing unwilted, well preserved grass silage of moderate digestibility with 3 kg starch-based concentrate per day, the limited response to the rate of sucrose supplementation employed suggests that increasing the water-soluble carbohydrate (WSC) concentration of grass silage through agronomic and/or ensiling practices will have relatively little effect on intake, rumen digestion or efficiency of microbial N synthesis.     

Modeling anaerobic digestion of microalgae using ADM1

The coupling between a microalgal pond and an anaerobic digester is a promising alternative for sustainable energy production by transforming carbon dioxide into methane using solar energy. In this paper, we demonstrate the ability of the original ADM1 model and a modified version (based on Contois kinetics for the hydrolysis steps) to represent microalgae anaerobic digestion. Simulations were compared to experimental data of an anaerobic digester fed with Chlorella vulgaris. The modified ADM1 fits adequately the data for the considered 140 day experiment encompassing a variety of influent load and flow rates. It turns out to be a reliable predictive tool for optimising the coupling of microalgae with anaerobic digestion processes.     

Modeling the dynamic performance of full-scale anaerobic primary sludge digester using Anaerobic Digestion Model No. 1 (ADM1)

Bioprocess and Biosystems Engineering - Anaerobic digestion is one of the most commonly accepted processes applied for the stabilization and treatment of primary sludge generated in municipal...     

Effect of maize silage to grass silage ratio and feed particle size on ruminal fermentation in vitro

The effect of the forage source on ruminal fermentation in vitro was investigated for fine (F) and coarse (C) milled diets, using a modified Hohenheim gas production test and a semi-continuous rumen simulation technique (Rusitec). It was hypothesised that the replacement of maize silage by grass silage might lead to associative effects and that interactions related to particle size variation could occur. Five diets with a maize silage to grass silage ratio of 100 : 0, 79 : 21, 52 : 48, 24 : 76 and 0 : 100 differed in their content of CP and carbohydrate fractions, as well as digestible crude nutrients, derived from a digestibility trial with wether sheep. For in vitro investigations, the diets were ground to pass a sieve of either 1 mm (F) or 4 mm (C) perforation. Cumulative gas production was recorded during 93 h of incubation and its capacity decreased with increasing proportion of grass silage in the diet. Across all diets, gas production was delayed in C treatments compared with F treatments. Degradation of crude nutrients and detergent fibre fractions was determined in a Rusitec system. Daily amounts of NH3-N and short-chain fatty acids (SCFA) were measured in the effluent. Degradation of organic matter (OM) and fibre fractions, as well as amounts of NH3-N, increased with stepwise replacement of maize silage by grass silage. Degradability of CP was unaffected by diet composition, as well as total SCFA production. In contrast to the results of the gas production test, degradation of OM and CP was higher in C than in F treatments, accompanied by higher amounts of NH3-N and SCFA. Interactions of silage ratio and particle size were rare. It was concluded that the stepwise replacement of maize silage by grass silage might lead to a linear response of most fermentation characteristics in vitro. This linear effect was also supported by total tract digestibility data. However, further investigations with silages of variable quality seem to be necessary.     

Comparative evaluation of biogas production from dairy manure and co‐digestion with maize silage by CSTR and new anaerobic hybrid reactor

This study aimed to investigate potential methane production through anaerobic digestion of dairy manure and co‐digestion with maize silage. Two different anaerobic reactor configurations (single‐stage continuously stirred tank reactor [CSTR] and hybrid anaerobic digester) were used and biogas production performances for each reactor were compared. The HR was planned to enable phase separation in order to improve process stability and biogas production under higher total solids loadings (≥4%). The systems were tested under six different organic loading rates increased steadily from 1.1 to 5.4 g VS/L.d. The CSTR exhibited lower system stability and biomass conversion efficiency than the HR. The specific biogas production of the hybrid system was between 440 and 320 mL/gVS with 81–65% volatile solids (VS) destruction. The hybrid system provided 116% increase in specific biogas production and VS destruction improved by more than 14%. When MS was co‐digested together with dairy manure, specific biogas production rates increased about 1.2‐fold. Co‐digestion was more beneficial than mono‐material digestion. The hybrid system allowed for generating methane enriched biogas (>75% methane) by enabling phase separation in the reactor. It was observed that acidogenic conditions prevailed in the first two compartments and the following two segments as methanogenic conditions were observed. The pH of the acidogenic part ranged between 4.7 and 5.5 and the methanogenic part was between 6.8 and 7.2.     

Flow pattern visualization in a mimic anaerobic digester using CFD

Three-dimensional steady-state computational fluid dynamics (CFD) simulations were performed in mimic anaerobic digesters to visualize their flow pattern and obtain hydrodynamic parameters. The mixing in the digester was provided by sparging gas at three different flow rates. The gas phase was simulated with air and the liquid phase with water. The CFD results were first evaluated using experimental data obtained by computer automated radioactive particle tracking (CARPT). The simulation results in terms of overall flow pattern, location of circulation cells and stagnant regions, trends of liquid velocity profiles, and volume of dead zones agree reasonably well with the experimental data. CFD simulations were also performed on different digester configurations. The effects of changing draft tube size, clearance, and shape of the tank bottoms were calculated to evaluate the effect of digester design on its flow pattern. Changing the draft tube clearance and height had no influence on the flow pattern or dead regions volume. However, increasing the draft tube diameter or incorporating a conical bottom design helped in reducing the volume of the dead zones as compared to a flat-bottom digester. The simulations showed that the gas flow rate sparged by a single point (0.5 cm diameter) sparger does not have an appreciable effect on the flow pattern of the digesters at the range of gas flow rates used.     

Effect of volatile compounds in grass silage on voluntary intake by growing cattle

Twenty-four low dry matter (DM) silages differing in fermentation quality were harvested at the same time from a crop that consisted mainly of timothy (Phleum pratense), and meadow fescue (Festuca pratensis). The silage samples were analysed by gas chromatography (GC) - mass spectrometry and gas chromatography - flame ionisation detection in order to determine and quantify volatiles present in silage. The voluntary intake of the 24 silages had been measured in a previous feeding trial with growing steers of Norwegian Red. Thirteen esters, five aldehydes, three alcohols, and one sulphide were identified and quantified. A total of 51 variables describing the chemical composition of the silages were included in a partial least-squares regression, and the relationship of silage fermentation quality to voluntary intake was elucidated. The importance of variables describing silage fermentation quality in relation to intake was judged from a best combination procedure, jack-knifing, and empirical correlations of the variables to intake. The GC-analysed compounds were mainly present in poorly fermented silages. However, compared with other explanatory chemical variables none of these compounds was of importance for the voluntary intake as evaluated by partial least-squares regression. A validated variance of 71% in silage DM intake was explained with the selected variables: total acids (TA), total volatile fatty acids (TVFA), lactic acid/total acid ratio and propionic acid. In this study extent (by the variable TA) and type of silage fermentation (by TVFA) influenced intake. Further, it is suggested that by restricting the fermentation in low DM grass silages the potential intake of silage DM is maximised.     

Effects of silage from maize crops differing in maturity at harvest,grass silage feed value and concentrate feed level on performance of finishing lambs

The effects of (i) medium and high feed value (MFV and HFV) maize silages and (ii) MFV and HFV grass silages, each in combination with a range of concentrate feed levels, on the performance of finishing lambs were evaluated using 280 Suffolk-X lambs (initial live weight 36.1 kg). The MFV and HFV maize silages represented crops with dry matter (DM) concentrations of 185 and 250 g/kg, respectively, at harvest, and had starch and metabolisable energy (ME) concentrations of 33 and 277 g/kg DM and 9.6 and 11.0 MJ/kg, respectively. HFV and MFV grass silages had DM and ME concentrations of 216 and 294 g/kg and 11.0 and 11.5 MJ/kg DM, respectively. A total of 13 treatments were involved. The four silages were offered ad libitum with daily concentrate supplements of 0.2, 0.5 or 0.8 kg per lamb. A final treatment consisted of concentrate offered ad libitum with 0.5 kg of the HFV grass silage daily. Increasing the feed value of grass silage increased (P < 0.001) forage intake, daily carcass and live weight gains, final live weight and carcass weight. Increasing maize silage feed value tended to increase (P = 0.07) daily carcass gain. Increasing concentrate feed level increased total food and ME intakes, and live weight and carcass gains. There was a significant interaction between silage feed value and the response to concentrate feed level. Relative to the HFV grass silage, the positive linear response to increasing concentrate feed level was greater with lambs offered the MFV grass silage for daily live weight gain (P < 0.001), daily carcass gain (P < 0.01) and final carcass weight (P < 0.01). Relative to the HFV maize silage, there was a greater response to increasing concentrate feed level from lambs offered the MFV maize silage in terms of daily carcass gain (P < 0.05) and daily live weight gain (P = 0.06). Forage type had no significant effect on the response to increased concentrate feed level. Relative to the MFV grass silage supplemented with 0.2 kg concentrate, the potential concentrate-sparing effect of the HFV grass silage, and the MFV and HFV maize silages was 0.41, 0.09 and 0.25 kg daily per lamb, respectively. It is concluded that increasing forage feed value increased forage intake and animal performance, and maize silage can replace MFV grass silage in the diet of finishing lambs as performance was equal to or better (depending on maturity of maize at harvest) than that for MFV grass silage.     

Digestion, rumen fermentation and circulating concentrations of insulin, growth hormone and IGF-1 in steers fed diets based on different proportions of maize silage and grass silage

Replacing grass silage with maize silage results in a fundamental change in the ratio of structural to non-structural carbohydrates with commensurate changes in rumen fermentation patterns and nutrient utilisation. This study investigated the effects of feeding four forage mixtures, namely grass silage (G); 67 g/100 g grass silage + 33 g/100 g maize silage (GGM); 67 g/100 g maize silage + 33/100 g grass silage (MMG); maize silage (M) to four ruminally and duodenally canulated Holstein Friesian steers. All diets were formulated to be isonitrogenous (22.4 g N/kg DM) using a concentrate mixture. Dietary dry matter (DM) and organic matter (OM) digestibility increased with ascending maize silage inclusion (P < 0.1) whereas starch and neutral detergent fibre digestibility declined (P < 0.05). Ratio of non-glucogenic to glucogenic precursors in the rumen fluid increased with maize silage inclusion (P < 0.01) with a commensurate reduction in rumen pH (P < 0.05). Mean circulating concentrations of insulin were greatest and similar in diets MMG and GGM, lower in diet M and lowest in diet G (P < 0.01). There were no effects of diet on the mean circulating concentration of growth hormone (GH), or the frequency, amplitude and duration of GH pulses, or the mean circulating concentrations of IGF-1. Increasing levels of DM, OM and starch intakes with the substitution of grass silage with maize silage affected overall digestion, nutrient partitioning and subsequent circulating concentrations of insulin.     

Modelling of the mesophilic anaerobic co-digestion of olive mill wastewater with olive mill solid waste using anaerobic digestion model No. 1 (ADM1)

The anaerobic digestion model No. 1 (ADM1), conceived by the international water association (IWA) task group for mathematical modelling of anaerobic digestion processes is a structured generic model which includes multiples steps describing biochemical and physicochemical processes encountered in the anaerobic degradation of complex organic substrates and a common platform for further model enhancement and validation of dynamic simulations for a variety of anaerobic processes. In this study the ADM1 model was modified and applied to simulate the mesophilic anaerobic co-digestion of olive mill wastewater (OMW) with olive mill solid waste (OMSW). The ADM1 equations were coded and implemented using the simulation software package MATLAB/Simulink. The most sensitive parameters were calibrated and validated using updated experimental data of our previous work. The results indicated that the ADM1 model could simulate with good accuracy: gas flows, methane and carbon-dioxide contents, pH and total volatile fatty acids (TVFA) concentrations of effluents for various feed concentrations digested at different hydraulic retention times (HRTs) and especially at HRTs of 36 and 24 days. Furthermore, effluent alkalinity and ammonium nitrogen were successfully predicted by the model at HRTs of 12 and 24 days for some feed concentrations.     

Effect of maize silage to grass silage ratio and feed particle size on protein synthesis and amino acid profile in different microbial fractions in a semi-continuous rumen simulation

The objective was to investigate the effect of variation in forage source and feed particle size of a diet, including interactions, on the amount and the composition of microbial crude protein (CP) in a semi-continuous culture system (Rusitec). Different microbial CP fractions were compared. Five diets with mean forage proportion of 0.88 and different maize silage to grass silage ratios (100 : 0, 79 : 21, 52 : 48, 24 : 76 and 0 : 100) were used. Diets were ground through sieves with a pore size of either 1 or 4 mm, matching the particle size of fine (F) and coarse (C), respectively. Diets were characterised by increasing concentrations of CP and fibre fractions, and decreasing concentrations of starch with ascending inclusion rates of grass silage. Microbial mass was isolated from feed residues after incubation from the liquid phase of the fermenter and from the liquid effluent. The amount of synthesised microbial CP was determined on the basis of 15N balance. It increased quite linearly by the stepwise replacement of maize silage by grass silage, and was higher in C treatments compared to F treatments. Efficiency of microbial CP synthesis (EMPS) was improved from 29 to 43 mg microbial N/g degraded organic matter (OM) by increasing the proportion of grass silage in the diet, but was unaffected by particle size. The N content as well as the profiles of amino acids of the three microbial fractions was affected by diet composition and particle size. The ratio of solid- to liquid-associated microbes was affected by diet composition and feed particle size. The amount and EMPS seemed to be improved by degradation of OM from grass silage and an increasing availability of N. Moreover, the results of this study indicated a shift in the composition of the microbial community caused by variation in forage composition and feed particle size.     

Influence of the maize silage to grass silage ratio and feed particle size of rations for ruminants on the community structure of ruminal Firmicutes in vitro

Aims: To investigate the effect of the forage source and feed particle size (FPS) in ruminant rations on the composition of the ruminal Firmicutes community in vitro. Methods and Results: Three diets, varying in maize silage to grass silage ratio and FPS, were incubated in a rumen simulation system. Microbial samples were taken from the liquid fermenter effluents. Microbial community analysis was performed by 16S rRNA‐based techniques. Clostridia‐specific single‐strand conformation polymorphism profiles revealed changes of the community structure in dependence on both factors tested. The coarse grass silage–containing diets seemed to enhance the occurrence of different Roseburia species. As detected by real‐time quantitative PCR, Ruminococcus albus showed a higher abundance with decreasing FPS. A slightly lower proportion of Bacilli was found with increasing grass silage to maize silage ratio by fluorescence in situ hybridization (FISH). In contrast, a slightly higher proportion of bacterial species belonging to the Clostridium‐clusters XIV a and b was detected by FISH with increasing grass silage contents in the diet. Conclusions: The ruminal Firmicutes community is affected by the choice of the forage source and FPS. Significance and Impact of the Study: This study supplies fundamental knowledge about the response of ruminal microbial communities to changing diets. Moreover, the data suggest a standardization of grinding of feeds for in vitro studies to facilitate the comparison of results of different laboratories.     

Influence of nitrogen fertilization on tropical‐grass silage assessed by ensiling process monitoring using chemical and microbial community analyses

Aims: Utilization of silage in livestock farming is expected to increase in developing countries in the tropical and subtropical parts of the world. The aim of this study was to investigate the influence of nitrogen fertilization on the chemical composition of herbage, ensiling process and silage quality, and to contribute to the improvement of tropical‐grass silage preparation. Methods and Results: Guinea grass grown under two different nitrogen‐fertilizer application conditions [1·5 kg N a^?1 (high‐N) and 0·5 kg N a^?1 (low‐N)] was packed in plastic bags, and its ensiling process was investigated by chemical and microbial‐community analyses. Relatively well‐preserved silage was obtained from high‐N herbage, which accumulated a high nitrate concentration. Denaturing gradient gel electrophoresis analysis revealed that Lactobacillus plantarum dominated throughout the ensiling of high‐N herbage and in the early phase of that of low‐N herbage. In low‐N silages prepared from ammonium sulfate‐ and urea‐fertilized herbage, Lact. plantarum was replaced by clostridia after 40 and 15 days of ensiling, respectively. Conclusions: Nitrate content of herbage is an important factor that influences silage quality, and careful fertilization management can facilitate stable and successful fermentation of tropical‐grass silage without any pretreatment. Significance and Impact of the Study: The positive effect of nitrate on the ensiling process of tropical‐grass was proved by microbial‐community analysis.