Low Methane Production Research Articles

Graphite-enhanced methanogenesis in coal measure shale anaerobic digestion: Implications for increasing gas yield and CO2 utilization

Anaerobic digestion (AD) of coal measure shale (CMS) for methane production is a promising technology for increasing yield of coal measure gas (CMG) wells. However, improving methane production efficiency of CMS remains a challenge. Conductive material has been shown to promote biogas production in different AD systems, but its effectiveness in CMS requires investigation. In this study, graphite was selected as conductive material due to its low cost and suitability for field applications, and its effects on methane production was investigated. The results indicate that as graphite concentration increases from 0 to 1.6 g/L, the methane production of CMS increases. However, when the graphite concentration reaches 2.0 g/L, the methanogenesis is inhibited, resulting in lower methane production compared to the control group. In the group with a graphite concentration of 1.6 g/L, the abundances of Romboutsia and Pseudomonas increase, which improves the compatibility of the microbial community with the CMS AD system and causes the amount of the substrates for methanogenesis to increase. Also, the direct interspecies electron transfer between Methanobacterium, Methanosaeta and Pseudomonas is strengthened, which enhances the methane production. However, excessive addition of graphite (2.0 g/L) intensifies the competitions from dissimilatory iron reduction and sulfate reduction reactions for substrates and electrons with the methanogenesis, causing the methane production to decrease. This study confirms the effectiveness of graphite in enhancing methane production of CMS, and the promoted hydrogenotrophic methanogenesis enhances the CO2 methanation, which offers an efficient pathway for increasing CMG well yield and enabling CO2 underground utilization.

NUTRIENT CONTENT, IN VITRO GAS PRODUCTION AND POST INCUBATION PARAMETERS OF INDIGENOUS BROWSE FODDERS IN DIFFERENT AGRO-ECOLOGICAL ZONES OF NORTHWEST ETHIOPIA

<p><strong>Background: </strong>In Ethiopia indigenous browse species have the potential to serve as a sustainable supplement for poor quality feeds and enhance ruminant livestock production. <strong>Objective:</strong> To analyze the nutrient content, in vitro gas production, and post incubation parameters of indigenous browse species in northwestern Ethiopia. <strong>Methodology: </strong>Eight indigenous browse fodder trees were collected from each agroecological zone for this experiment. Leaf and pod samples of the fodder trees were collected, dried and ground for laboratory analysis. Analysis of variance was carried out for nutrient content, in vitro gas, and methane production of the samples within agroecological zones using standard analytical procedures. The statistical design used to analyze the data was Completely Randomized Design (CRD). <strong> Results:</strong> The highest CP content was recorded from <em>Vernonia</em><em> amygdalina </em>(223.4 g/kg DM)<em>,</em> <em>Dodonaea viscosa </em>(207.7 g/kg DM), and <em>Acacia abyssinica </em>(216.0 g/kg DM) in the highland, midland and lowland agro-ecology respectively. In lowland area, NDF, ADF and ADL contents were highest for <em>Albezia amara</em> leaves and lowest for <em>Acacia brevispica</em> respectively. <em>V. amygdalina</em>, <em>Stereospermum kunthianum</em> and <em>Ficus vasta</em> had the highest gas volume at 24-hour incubation time in high, mid and lowland, respectively. The estimated organic matter digestibility, metabolizable energy and short chain fatty acid at 24 hour of incubation was the highest for <em>V. amygdalina</em>, <em>S kunthianum</em> and <em>F. vasta</em> in high, mid and lowland, respectively. <em>Ficus sycomorus</em> and <em>Myrica salicifolia</em> had the lowest methane production among the species in the low, mid and highland study areas, respectively. <strong>Implications: </strong>The present study provides a valuable resource for selecting suitable feed options for livestock in different agroecological zones. High CP and digestible browse species like <em>V. amygdalina,</em> <em>S. kunthianum</em>, and <em>F. vasta</em> could be prioritized for livestock feed. Low methane-producing species like <em>F. sycomorus</em> and <em>M. salicifolia</em> could be incorporated into diets to reduce enteric methane emissions. <strong>Conclusion</strong>: Based on their nutrient composition and in vitro gas production potential, these feed resources are of better quality to supplement grazing livestock during the seasons of critical feed shortage as the main feed resources are either limited in availability or lower in nutrient composition and digestibility in the study areas.</p>

The potential role of iron-carbon micro-electrolysis materials in curtailing lag-phase stimulates kitchen waste anaerobic digestion at different solid contents: Performance, synergistic effect and microbial response

High-solid anaerobic digestion (HSAD) of kitchen waste was generally faced to the common problems such as systemic acidification, prolonged lag-phase time and low methane production. Iron-carbon micro-electrolysis (ICME) materials exhibited advantages that porous structure, large specific surface area and excellent conductivity. It was beneficial for organic compounds to hydrolysis. Moreover, ICME materials could establish direct interspecies electron transfer (DIET) pathway between bacteria and methanogens. ICME materials were commonly used to enhance the AD of wastewater, but they were rarely applied to HSAD of kitchen waste. In this study, ICME materials were utilized to enhance HSAD of kitchen waste at different solid content conditions. The results showed that the highest cumulative biogas yield (705.23 mL/g VS) was obtained in the experimental group (TS = 10%), which was 94.15% higher than that of the control group. At the same time, the addiction of ICME could shorten lag-phase time. Electrochemical characteristics and XPS analysis showed that ICME materials promoted the release of Fe2+ in the AD system and acceleration of direct interspecies electron transfer between microorganisms. Microbial community analysis showed that ICME materials enriched electroactive bacteria (Proteiniphilum), Methanosarcina, Methanobrevibacter and Methanofollis. Functional gene prediction revealed that ICME materials increased the relative abundance of carbohydrate transport and metabolism and coenzyme transport and metabolism. It provided a potential measure to treat kitchen waste.

Exploring macroalgae biorefinery: Extraction of bioactive compounds and production of volatile fatty acids

Macroalgae have gained significant attention in recent research owing to their potential as novel food source and their noteworthy nutritional properties. However, a substantial amount of these macroalgae accumulates along the coast without being utilized, highlighting the need for proper treatment and disposal methods to mitigate secondary pollution effects. Previous studies on macroalgae have primarily focused on extracting bioactive compounds or anaerobic digestion processes to produce methane or volatile fatty acids (VFA), with observed improvements following different pre-treatments. In this study, three biorefinery options for macroalgae have been compared. Additionally, the extraction of bioactive compounds followed by VFA production is proposed as a promising new valorization strategy. Milled macroalgae exhibited a low methane production yield (138 ± 17 NmL CH4·g volatile solid−1), corresponding to 31 ± 4 % biodegradability, while the acidification percentage was higher (45 ± 1%). Among the three solvents applied (water, ethanol and acetone), ethanol (80%) at 25 °C was the most effective in recovering bioactive compounds, such as chlorophylls, sugars, and phenolic compounds with antioxidant activity. The extraction of chlorophylls and phenolic compounds was not influenced by particle size reduction. However, a more efficient extraction of sugars was observed with lower particle size. Moreover, ethanol treatment demonstrated the good efficiency in VFA production, reaching up to 3.6 ± 0.2 g VFA-(chemical oxygen demand, COD)·L−1, with a VFA spectrum (in COD basis) consisting of 51% acetic acid, 29% propionic acid, 5% i-butyric acid, 7% butyric acid, and 7% i-valeric acid. These findings highlight the potential of ethanol for efficient compound recovery and VFA production from macroalgae.

Effect of hydrochar from biogas slurry co-hydrothermal carbonization with biomass on anaerobic digestion performance of food waste

Anaerobic digestion is a promising method for converting food waste into renewable methane-rich biogas. However, challenges, such as significant biogas slurry generation, poor stability, and low methane production, limit its widespread adoption. This study produced 15 types of hydrochars by co-hydrothermal carbonization of biogas slurry with sugarcane leaves, cellulosic ethanol residue, or food waste digestate at 180–260 °C to enhance anaerobic digestion performance and methane yield. The methane yield was influenced by both the raw materials and hydrothermal carbonization temperature. The hydrochar derived from sugarcane leaves at 180 °C exhibited the highest methane yield of 461.59 mL/g VS, which is a 13.69 % increase over the food waste group. Additionally, different doses of SL-180 were tested for their effects on the anaerobic digestion performance in food waste. The cumulative methane yield increased from 379.68 mL/g VS to 543.26 mL/g VS with SL-180 dosages ranging from 5 g/L to 40 g/L, enhancing cumulative methane yield by 3.90–48.66 % compared to the food waste group. The primary factor influencing methane yield was hydrochar degradation, whereas secondary factors included maintaining system stability and enriching methane-producing microorganisms, particularly Methanosaeta and Methanomassiliicoccus. Energy assessment and economic analysis indicate that combining hydrothermal carbonization with anaerobic digestion can achieve high energy output and economic benefits. This study offers insights and recommendations for managing biogas slurry, resource recovery, and improving the stability and methane yield in the anaerobic digestion of food waste.

Evaluation of different operational conditions in a microbial electrolysis cell inoculated with a pure culture of Shewanella oneidensis for hydrogen production.

Hydrogen is a promising alternative to meet the world's energy demand in the future because of its energetic characteristics. Microbial electrolysis cell (MEC) produces hydrogen from organic matter using exoelectrogenic bacteria. Shewanella oneidensis stands out for having the capacity to produce hydrogen using different electron transfer mechanisms. The present research aims to evaluate the hydrogen production efficiency in a MEC inoculated with a pure culture of S. oneidensis in different operational conditions. Since the use of a catalyst accounts for most of the MEC cost, no catalyst was used for anode or cathode. Experiments were performed in semi-continuous and batch mode using different electrodes, voltages applied, and medium in aerobic and anaerobic conditions. The highest hydrogen production rate (HPR) was 0.107 m3 of H2/m3day obtained in a semi-continuous experiment using graphite plates and stainless steel electrodes. In batch experiments, a HPR occurred at 0.7V, with a value of 0.048 m3 of H2/m3day versus 0.037 m3 of H2/m3day with 0.9V. HPR was higher with carbon felt electrode (0.056 m3 of H2/m3day). However, current density dropped after 38h, with carbon felt electrodes, and did not recover. Results of the present research showed that the MEC using a pure culture of S. oneidensis can be considered an alternative for hydrogen production without using a catalyst. Also, S. oneidensis produced hydrogen in both anaerobic and aerobic conditions with low methane production. Optimization can be proposed to improve hydrogen production based on the operational conditions tested in these experiments.

Development of oriented multi-enzyme strengthens waste activated sludge disintegration and anaerobic digestion: Performance, components transformation and microbial communities

Low methane production and long retention time are the main dilemmas in current anaerobic digestion (AD) of waste activated sludge (WAS). This work used WAS as only substrate to prepare oriented multi-enzyme (ME) that directly used for WAS pretreatment. Under the optimal parameters, the highest activities of protease and amylase in ME could respectively reach 16.5 U/g and 580 U/g, and the corresponding methane production attained 197 mLCH4/g VS, which was increased by 70.4% compared to blank group. It was found that ME pretreatment could strengthen WAS disintegration and organic matters dissolution, lead to the soluble chemical oxygen demand (SCOD) was increased from the initial 486 mg/L to 2583 mg/L, and the corresponding volatile suspended solid (VSS) and extracellular polymeric substances (EPS) were reduced by 27% and 73.8%, respectively. The results of three-dimensional excitation-emission matrix (3D-EEM) and Fourier transform infrared spectroscopy (FTIR) indicated that protein disintegration may be the critical step during the process of WAS hydrolysis with ME, of which the release of tyrosine-like proteins achieved the better biodegradability of WAS, while the results of X-ray photoelectron spectroscopy (XPS) showed that the formation of protein derivatives was the main harmful factor that could extend the lag phase of AD process. Microbial communities analysis further suggested that ME pretreatment facilitated the enrichment of acetogenic bacteria and acetotrophic methanogens, which caused the transition of the methanogenesis pathway from hydrogenotrophic to acetotrophic. This study is expected to furnish valuable insight for ME pretreatment on enhancing WAS disintegration and methane production.

Characteristics of rumen microbiota and Prevotella isolates found in high propionate and low methane-producing dairy cows.

Ruminal methane production is the main sink for metabolic hydrogen generated during rumen fermentation, and is a major contributor to greenhouse gas (GHG) emission. Individual ruminants exhibit varying methane production efficiency; therefore, understanding the microbial characteristics of low-methane-emitting animals could offer opportunities for mitigating enteric methane. Here, we investigated the association between rumen fermentation and rumen microbiota, focusing on methane production, and elucidated the physiological characteristics of bacteria found in low methane-producing cows. Thirteen Holstein cows in the late lactation stage were fed a corn silage-based total mixed ration (TMR), and feed digestion, milk production, rumen fermentation products, methane production, and rumen microbial composition were examined. Cows were classified into two ruminal fermentation groups using Principal component analysis: low and high methane-producing cows (36.9 vs. 43.2 L/DMI digested) with different ruminal short chain fatty acid ratio [(C2+C4)/C3] (3.54 vs. 5.03) and dry matter (DM) digestibility (67.7% vs. 65.3%). However, there were no significant differences in dry matter intake (DMI) and milk production between both groups. Additionally, there were differences in the abundance of OTUs assigned to uncultured Prevotella sp., Succinivibrio, and other 12 bacterial phylotypes between both groups. Specifically, a previously uncultured novel Prevotella sp. with lactate-producing phenotype was detected, with higher abundance in low methane-producing cows. These findings provide evidence that Prevotella may be associated with low methane and high propionate production. However, further research is required to improve the understanding of microbial relationships and metabolic processes involved in the mitigation of enteric methane.

Interspecies electron transfer regulation in a stage-separated anaerobic digestion system: The role of inoculum strategies, activated carbon integration, and organic loading rate adjustment

Understanding and regulation of interspecies electron transfer, including hydrogen-mediated interspecies electron transfer (MIET) and direct interspecies electron transfer (DIET), is crucial for enhancing anaerobic digestion (AD). In this study, MIET and DIET were regulated in a multistage anaerobic hythane reactor (MAHR) via inoculum management, activated carbon (AC) addition, and organic loading rate (OLR) adjustment. The operational stability, organic removal, and methane production can be enhanced, with AC addition in the methane production zone (Mm) (RpCMm and RnpCMm). The reactors of RpCMm and RnpCMm achieved start-up within 20 days and showed a 20 % and 104 % increase in maximum methane production rates, respectively. Microbial community characteristics confirmed that AC in Mm could facilitate DIET establishment, but its addition in Mh could not. This assertion was supported by the observed tighter and more positive correlation between biofilm conductivity and DIET-related microbes in RpCMm and RnpCMm, as opposed to RpCMh and RnpCMh. Further, the dual drive of DIET and MIET was revealed through the comprehensive analysis of operational performances and microbial community with the structure equation model. The electron fluxes analysis highlighted a dynamic shift of DIET and MIET in the methanogenic stage with AC addition and OLR variation. The DIET was proved more efficient than MIET in Mm with AC addition and OLR below 40 g COD/L/d, while high OLR (80 g COD/L/d) favored MIET with lower methane production efficiency. The results of this study shed light on the nuanced characteristics of interspecies electron transfer, providing valuable insights for the further design and operation of stage-separated AD.

Metagenomics reveals the variations in functional metabolism associated with greenhouse gas emissions during legume-vegetable rotation process

Legume-based rotation is commonly recognized for its mitigation efficiency of greenhouse gas (GHG) emissions. However, variations in GHG emission-associated metabolic functions during the legume-vegetable rotation process remain largely uncharacterized. Accordingly, a soybean-radish rotation field experiment was designed to clarify the responses of microbial communities and their GHG emission-associated functional metabolism through metagenomics. The results showed that the contents of soil organic carbon and total phosphorus significantly decreased during the soybean-radish process (P < 0.05), while soil total potassium content and bacterial richness and diversity significantly increased (P < 0.05). Moreover, the predominant bacterial phyla varied, with a decrease in the relative abundance of Proteobacteria and an increase in the relative abundance of Acidobacteria, Gemmatimonadetes, and Chloroflexi. Metagenomics clarified that bacterial carbohydrate metabolism substantially increased during the rotation process, whereas formaldehyde assimilation, methanogenesis, nitrification, and dissimilatory nitrate reduction decreased (P < 0.05). Specifically, the expression of phosphate acetyltransferase (functional methanogenesis gene, pta) and nitrate reductase gamma subunit (functional dissimilatory nitrate reduction gene, narI) was inhibited, indicating of low methane production and nitrogen metabolism. Additionally, the partial least squares path model revealed that the Shannon diversity index was negatively correlated with methane and nitrogen metabolism (P < 0.01), further demonstrating that the response of the soil bacterial microbiome responses are closely linked with GHG-associated metabolism during the soybean-radish rotation process. Collectively, our findings shed light on the responses of soil microbial communities to functional metabolism associated with GHG emissions and provide important insights to mitigate GHG emissions during the rotational cropping of legumes and vegetables.

Metagenome reveals the possible mechanism that microbial strains promote methanogenesis during anaerobic digestion of food waste

For better understanding the mechanism of microbial strains promoting methane production, four strains Hungatella xylanolytica A5, Bacillus licheniformis B1, Paraclostridium benzoelyticum C2 and Advenella faeciporci E1 were inoculated into anaerobic digestion systems. After bioaugmentation, the cumulative methane production of A5, B1, C2 and E1 groups elevated by 11.68%, 8.20%, 18.21% and 15.67% compared to CK group, respectively. The metagenomic analysis revealed that the species diversity and uniformity of the experimental groups was improved, and hydrolytic acidifying bacteria, represented by Clostridiaceae, Anaerolineaceae and Oscillospiraceae, and methanogens, such as Methanotrichaceae and Methanobacteriaceae, were enriched. Meanwhile, the abundance of key genes in carbohydrate, pyruvate and methane metabolism was increased in the inoculated groups, providing reasonable reasons for more methane production. The strengthening mechanism of microbial strains in this study offered a theoretical foundation for selecting a suitable bioaugmentation strategy to solve the problems of slow start-up and low methane production in anaerobic digestion.

Toilet effluent separation and brown water treatment: Survey and initial feasibility testing in Mexico

Separation of domestic effluents at the source and the utilization of low-flush toilets offer alternative approaches for developing efficient wastewater treatment systems while promoting energy generation through anaerobic digestion. This study focused on assessing toilet usage in Mexico and exploring the potential of anaerobic co-digestion of brown water (feces) and toilet paper as influential factors in wastewater treatment systems. A survey was conducted on a representative sample of Mexicans to gather information on toilet usage frequency, toilet paper use and disposal practices, as well as the type and quantity of commercial disinfectants and pharmaceutical compounds they use or consume. The survey revealed that per capita toilet paper consumption is 2.9 kg annually, that 58 % of respondents do not dispose used paper in the toilet, and that about 47 % use two to three cleaning and disinfection products. Notably, 97 % of the sampled Mexican population expressed a willingness to transition to more eco-friendly toilet options. Subsequently, in a second step, the anaerobic co-digestion of brown water with toilet paper was evaluated, demonstrating a relatively high production of volatile fatty acids but low methane production. This suggests an efficient hydrolysis/acidogenesis process coupled with restrained methanogenesis, probably due to pH decrease caused by acidogenesis. This study underscores that toilet paper and brown water are potential suitable substrates for anaerobic co-digestion. Furthermore, it sheds light on the behaviors of Mexican society regarding bathroom use and cleaning, contributing to the establishment of foundations for wastewater treatment systems with effluent separation at the source.

Methane production from the biodegradation of lignite with different sizes by mixed fungi-methanogen microflora.

Biogenic coalbed methane (CBM) is a developing clean energy source. However, it is unclear how the mechanisms of bio-methane production with different sizes of coal. In this work, pulverized coal (PC) and lump coal (LC) were used for methane production by mixed fungi-methanogen microflora. The lower methane production from LC was observed. The aromatic carbon of coal was degraded slightly by 2.17% in LC, while 11.28% in PC. It is attributed to the proportion of lignin-degrading fungi, especially Penicillium, which was reached 67.57% in PC on the 7th day, higher than that of 11.38% in LC. The results suggested that the limited interaction area in LC led to microorganisms hardly utilize aromatics. It also led the accumulation of aromatic organics in the fermentation broth in PC. Increasing the reaction area of coal and facilitating the conversion of aromatic carbon are suggested means to increase methane production in situ.

Mixed ensiling plus nitrate destroy fiber structure of rape straw, increase degradation, and reduce methanogenesis through in vitro ruminal fermentation.

Better utilization of rape straw can provide alternative strategies for sustainable ruminant and food production. The research reported here investigated changes in the carbohydrate composition of rape straw as a result of mixed ensiling with whole-crop corn or inoculated with nitrate, and the consequent effects on ruminal fermentation through in vitro batch culture. The three treatments included: rape straw and corn silage (RSTC), and ensiling treatment of rape straw with whole-crop corn (RSIC) or with calcium nitrate inoculation (RSICN). Ensiling treatment of rape straw and whole-crop corn or plus nitrate enriched lactic acid bacteria and lactate. The treatments broke the fiber surface connections of rape straw, leading to higher neutral detergent soluble (NDS) content and lower fiber content. Ensiling treatments led to greater (P < 0.05) dry matter degradation (DMD), molar proportions of propionate and butyrate, relative abundance of the phylum Bacteroidetes and genus Prevotella, and lower (P < 0.05) methane production in terms of g kg-1 DMD, molar proportions of acetate, and lower acetate to propionate ratio than the RSTC treatment. The RSICN treatment led to the lowest (P < 0.05) hydrogen concentration and methane production among the three treatments. Ensiling treatments of rape straw and whole-crop corn destroy the micro-structure of rape straw, promote substrate degradation by enriching the phylum Bacteroidetes and the genus Prevotella, and decrease methane production by favoring propionate and butyrate production. Nitrate inoculation in the ensiling treatment of rape straw and whole-crop corn further decreases methane production without influencing substrate degradation by providing an additional hydrogen sink. © 2023 Society of Chemical Industry.

Reduction of soil methane emissions from croplands with 20–40 years of cultivation mediated by methane-metabolizing microorganisms

Reducing carbon emissions from cropland ecosystems is an important measure to achieve early carbon neutrality. However, how methane-metabolizing microorganisms affect methane emissions from cropland soils of different planting years is not known. In this study, we determined methane emissions from cropland soils of different tobacco-rice rotation years after rice harvesting. By mean of functional gene sequencing, we analyzed the relationship between methane-metabolizing microorganisms and methane emissions, and revealed the microbial mechanism of action for methane emissions from cropland soils. The results showed that the soil organic carbon content was higher (P < 0.05) and methane emissions were lower (P < 0.05) in cropland cultivated for 20–40 years (PY20). Soil methane emissions from PY20 cropland were reduced by 30.11% and 14.58% (P < 0.05) compared to cropland planted for 10–20 years and more than 40 years. The alpha diversity of methanogens communities was significantly lower in PY20 cropland soils, whereas the methanotrophs communities alpha diversity did not differ significantly between planting years. The methanogenic genus Methanocorpusculum was significantly and positively correlated with methane emissions. The relative abundance of Methanocorpusculum in PY20 cropland soil was lower (P < 0.05), and the relative abundance of Methylocystis, a type II methanotrophs with greater oxidizing capacity for methane, was higher (P < 0.05). Reducing PY20 cropland soil methane emissions by reducing methane production and enhancing methane oxidation. At the same time, more methanotrophs in PY20 cropland soil formed a molecular interactions network with methanogens. Planting years altered the diversity and composition of methane-metabolizing microbial communities and the interactions of different taxa, which in turn affected methane emissions. Cropland soils that have been cultivated for 20–40 years have lower methane production capacity and higher methane oxidation capacity, resulting in lower methane emissions. This study provides a scientific reference for the microbial action mechanism of soil methane emission from cropland under different planting years, with a view to contributing to carbon neutralization in cropland ecosystems.

Evaluation of several macroalgae species on methane emission and antioxidant activity based on in vitro rumen fermentation characteristics

Marine macroalgae are the prospective and promising Indonesian natural resources containing bioactive compounds. This study aimed at evaluating various seaweed species with optimal biological activity to mitigate in vitro rumen methane emission and to confirm their antioxidant properties. In addition, relationships among several phenolic fractions and methane (CH4) production of seaweeds were elucidated by Pearson Correlation to screen seaweeds for their capacity to reduce CH4 formation. For in vitro batch cultures experiment, the diets were 500 mg of a substrate of seaweed. They were incubated with 50 ml of buffered rumen fluid (1:2 ratio of rumen fluid to buffer media) in a 100 ml serum vial at 39°C for 72 hours incubation time. A randomized block design was carried out with 14 dried seaweed treatments and 5 different cattle rumen fluid groups as a block. For total phenol, flavonoid, and antioxidant activity, a completely randomized design with 14 treatments and 3 replications was carried out. Data were analyzed by Analysis of Variance (ANOVA) and Duncan analysis. The results showed that the lowest methane production (ml/g DM incubated) significantly (P<0.05) was reduced by Halymenia sp. (0.24) and Caulerpa racemosa (0.41). Caulerpa racemosa showed high flavonoid content (1.5 mg Quercetin/g) and Ulva lactuca had the highest phenol content (0.55 mg Gallic Acid/g). Gracilaria coronopifolia had strong antioxidant activity DPPH radical scavenging activity (77.5% inhibition). It was concluded that Halymenia sp., Caulerpa racemosa and Gracilaria coronopifolia could be used as methane inhibitors and antioxidants for ruminants.

211 Evaluation of Beef Heifer Variability in the Ability to Eat and Digest a High Forage Diet

Abstract This study evaluated beef heifers selected for high or low digestible fiber intake (DFI) and investigated its relationship with methane production, residual feed intake (RFI), and total tract digestibility. Sixteen recently weaned black Angus beef heifers (n = 8/treatment) were selected from a group 64 heifers (224 ± 17.2 kg) fed a high forage-based diet [70% barley silage:30% pelleted concentrate (DM basis)]. The 64 heifers were fed in 6 outdoor pens equipped with GrowSafe bunks to monitor feed intake for 60 d (14 d adaptation + 46 d for data collection). Individual fecal samples and BW, and feed samples were taken once weekly. Fecal samples were pooled by animal. Feed and fecal samples were analyzed for dry matter (DM), neutral detergent fibre (NDF), and undigested neutral detergent fiber (uNDF) content. The internal marker uNDF was used to estimate total tract diet DM and NDF digestibility. The 8 heifers with the greatest and the 8 with the least digestible NDF intake (g/kg BW0.75) were selected and used for methane measurements using the GreenFeed system (42 d) and in a total-tract digestibility trial with total fecal and urine collection using the same high forage based diet. Results from the GrowSafe selection trial indicated no differences between the average BW of low and high DFI heifers (264 vs. 274 kg, P = 0.20). Heifers selected for high DFI had greater (P &amp;lt; 0.01) DM and NDF intake (kg/d or % of BW), and digestibility (49.6 vs. 42.1% NDF digestibility). The heifer groups differed in RFI (P &amp;lt; 0.01), with high DFI categorized as inefficient (+0.84 RFI) and low DFI as efficient (-0.34 RFI). No differences in ADG were observed between low and high DFI heifers during this short 46 d study period (0.614 vs 0.773, P = 0.16). High DFI heifers had lower methane production than low DFI heifers (15.9 vs. 19.0 g/kg of DMI, P = 0.02). The results of the total-tract digestibility trial showed that the high DFI heifers had a greater DMI compared with the low DFI heifers (10.9 vs. 10.2 kg/d; P = 0.04). The DMI intake was not different between groups when expressed as a % of BW (2.06 vs 2.04, P = 0.65). There was also no difference observed for DM digestibility (73.0 vs 73.1%) between the two groups. It is important to note that in the digestibility study, the DMI was less than in the GrowSafe selection trial (2.5 vs 3.2% BW) and this may have influenced results. Heifers with high DFI were heavier than low DFI heifers (533 vs. 505, P = 0.02) during the digestibility trial. Results suggest that selecting heifers for high DFI may increase growth rate and reduce methane production.

The Dose-Dependent Role of Sage, Clove, and Pine Essential Oils in Modulating Ruminal Fermentation and Biohydrogenation of Polyunsaturated Fatty Acids: A Promising Strategy to Reduce Methane Emissions and Enhance the Nutritional Profile of Ruminant Products

The livestock industry significantly contributes to greenhouse gas emissions, with ruminant animals, including cows, sheep, and goats, being responsible for a substantial share of these emissions due to methane production. Reducing methane emissions from ruminants is crucial for mitigating the environmental impact of livestock production. Additionally, there has been a growing interest in improving the nutritional quality of ruminant products through modifying their profile of fatty acids. The current study aimed to investigate the potential of sage (SAG), pine (PIN), and clove (CLO) essential oils as natural additives for modulating in vitro ruminal fermentation characteristics and biohydrogenation of polyunsaturated fatty acids (PUFA). Within the current experiment, three dose levels (300, 600, and 900 mg/L) of essential oils were evaluated using rumen inoculum from three mature Dalagh ewes (58 ± 2.84 kg body weight). The results revealed that the essential oils had a significant impact on gas production, methane and carbon dioxide production, ruminal fermentation parameters, and ruminal biohydrogenation of dietary PUFAs. The essential oil treatments resulted in reduced gas production compared with the control group. Methane production was significantly reduced by all doses of the essential oils, with the highest dose of CLO resulting in the lowest methane production. In addition, the essential oils affected ruminal fermentation parameters, including pH, ammonia concentration, and production of total volatile fatty acids. Promising modifications in ruminal biohydrogenation of PUFAs and the profile of fatty acids were also observed in the current study. These findings suggest that SAG, Pin, and CLO hold promise in mitigating methane emissions and improve the nutritional value of ruminant products. Further investigation is required to evaluate their effectiveness in practical feeding strategies for livestock.

Impact of adding tannins or medium-chain fatty acids in a dairy cow diet on variables of in vitro fermentation using a rumen simulation technique (RUSITEC) system

This work aimed to evaluate the effect of including 2 functional feed ingredients in a diet with a high proportion of pasture silage on in vitro ruminal fermentation, nutrient disappearances, and ruminal microbiomes. In a rumen simulation technique (RUSITEC) system (6 fermentation units), 4 treatments were evaluated: 1) CONTROL: a diet based on corn and grass silage, regular soybean meal (SBM), and corn; 178 g/kg of crude protein (CP) (used as negative control), 2) TSBM: Replacement of SBM by Mervobest® (used as positive control), 3) PHY (phytogenic mixture (tannins from quebracho and chestnut) at 10 g/kg dry matter (DM) replacing SBM) and 4) MCFA (mixture of medium-chain fatty acids, Aromabiotic Cattle® at 10 g/kg DM replacing SBM). Four runs were performed with 3 diets incubated simultaneously in duplicate using a balanced incomplete block design, lasting 17 days each. The inoculum was obtained from 3 rumen fistulated cows, and artificial saliva was infused at 650 mL/day. Samples were taken from each fermentation unit to determine the gas production, pH, NH3-N, and volatile fatty acids concentrations. The apparent digestibility of diet compounds was determined. Microbial protein synthesis and microbial community diversity were estimated using 15N as an external isotopic marker and by sequencing part of the 16 S rDNA gene, respectively. The TSBM and PHY treatments decreased NH3-N concentrations (P < 0.001) and the DM disappearance (P = 0.004). The crude protein disappearance was the lowest for TSBM followed by PHY (P < 0.001). The true protein disappearance was lower for TSBM and PHY than for CONTROL and MCFA (P < 0.001). The isovaleric acid proportion was lower for TSBM, MCFA, and PHY than for CONTROL. Treatments TSBM and MCFA registered lower neutral detergent fibre disappearance (P < 0.001) than the other diets but without changes in the quantified fibrolytic genera in the ruminal microbiota. The PHY treatment increased the relative abundance of Succinivibrio spp. and reduced that of Methanobrevibacter relative to that of CONTROL and TSBM, which could be linked to lower methanogenesis. The MCFA treatment increased Succinivibrio compared with TSBM and Succiniclasticum spp. compared with CONTROL. These results indicate that PHY inclusion at 10 g/kg DM decreased rumen protein degradation, similar to TSBM; while MCFA decreased rumen protein degradation to a lesser extent. Furthermore, both additives generated changes in the ruminal microbiota that would be associated with lower methane production.

Fermentative hydrogen production enhancement by microbial community selection and enrichment through biostimulation

This paper presents a novel technique to increase hydrogen production from dark fermentation of cheese whey, conducted using open mixed cultures. Hydrogen (H2)-producing microbial pools were selected and enriched using an innovative biostimulation technique, performed by sequential re-inocula of the interstitial microbial community using porous supports in sequential batch reactors. The supports offered a spatial refuge to the microbial community, facilitated re-inoculum procedures and reduce operational disturbance. Two different re-inoculum times (i.e. 24 h and 96 h) were tested to verify the influence of timing on the selection/enrichment. Results showed that the H2 production rate was up to 80 times higher using a 24 h re-inoculum time than using a 96 h re-inoculum time. Under a 24 h re-inoculum time, a stable H2 production rate was reached after two sequential re-inocula, with values ranging between 4 and 6.1 mLH2 h−1. The cumulative production, detected after the last re-inoculum (8 days), was 2.2 LH2 L-1reactor, corresponding to 42.5 mL gCOD-1. A very low methane (CH4) production was observed and only in the first two re-inocula, as short re-inoculum intervals favoured H2-producing bacteria. Microbiological analyses confirmed that subsequent re-inocula contributed to an increase in total microbial abundance. The proposed approach provides a concentrated inoculum in liquid medium (which is easy to use) exploiting an enrichment technique that is conceptually new and more economical/environmentally friendly than conventional thermal, acid or alkaline enrichment techniques.