Potential of Mulberry Leaf Biomass and Its Flavonoids to Improve Production and Health in Ruminants: Mechanistic Insights and Prospects

Leaf biomass from the mulberry plant (genus Morus and family Moraceae) is considered a potential resource for livestock feeding. Mulberry leaves (MLs) contain high protein (14.0–34.2%) and metabolizable energy (1130–2240 kcal/kg) with high dry matter (DM) digestibility (75–85%) and palatability. Flavonoid contents of MLs confer unique antioxidant properties and can potentially help alleviate oxidative stress in animals during stressful periods, such as neonatal, weaning, and periparturient periods. In addition, mulberry leaf flavonoids (MLFs) possess antimicrobial properties and can effectively decrease the population of ruminal methanogens and protozoa to reduce enteric methane (CH4) production. Owing to its rich flavonoid content, feeding MLs increases fiber digestion and utilization leading to enhanced milk production in ruminants. Dietary supplementation with MLFs alters ruminal fermentation kinetics by increasing total volatile fatty acids, propionate, and ammonia concentrations. Furthermore, they can substantially increase the population of specific cellulolytic bacteria in the rumen. Owing to their structural homology with steroid hormones, the MLFs can potentially modulate different metabolic pathways particularly those linked with energy homeostasis. This review aims to highlight the potential of ML and its flavonoids to modulate the ruminal microbiome, fermentation, and metabolic status to enhance productive performance and health in ruminants while reducing CH4 emission.

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Pomegranate seed oil rich in conjugated linolenic acids reduces in vitro methane production

South African Journal of Animal Science ◽

10.4314/sajas.v46i3.13 ◽

1970 ◽

Vol 46 (3) ◽

pp. 325-335

Author(s):

E. Maleki ◽

G.Y. Meng ◽

M. Faseleh Jahromi ◽

R. Jorfi ◽

A. Khoddami ◽

...

Keyword(s):

Methane Production ◽

Volatile Fatty Acids ◽

Seed Oil ◽

Control Diet ◽

Nutrient Utilization ◽

Gas Production ◽

Metabolizable Energy ◽

Ruminal Fermentation ◽

Ch4 Production

The objective of this study was to determine the effect of pomegranate (Punica granatum L.) seed oil (PSO) on gas and methane (CH4) production, ruminal fermentation and microbial populations under in vitro conditions. Three treatments consisting of a control diet containing 10 mg tallow (CON); the control diet with 5 mg PSO + 5 mg tallow (MPSO) and the control diet containing 10 mg PSO (HPSO) were compared. Ten mg of the experimental fat/oil samples were inserted into a gas-tight 100 mL plastic syringe containing 30 mL of an incubation inoculum and 250 mg of a basic substrate of a hay/concentrate (1/1, w/w) mixture. In vitro gas production was recorded over 0, 2, 4, 6, 8, 10, 12 and 24 h of incubation. After 24 hours, incubation was stopped, and methane production, pH, volatile fatty acids (VFAs) and microbial counts were measured in the inoculant. Gas production at 4, 6, 8, 10, 12 and 24 h incubation, metabolizable energy and in vitro organic matter disappearance increased linearly and quadratically as level of PSO increased. Furthermore, the 10 mg PSO (HPSO) decreased CH4 production by 21.0% compared with the control (CON) group. There were no significant differences in total and individual VFA concentrations between different levels of PSO, except for butyric acid. After 24 h of incubation, methanogenesis decreased in the HPSO compared with the MPSO and CON treatments. In addition, total bacteria and protozoa counts increased with rising PSO levels, while population methanogenesis declined significantly. These results suggested that PSO could reduce methane emissions, which might be beneficial to nutrient utilization and growth in ruminants.

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Metatranscriptomic analyses reveal ruminal pH regulates fiber degradation and fermentation by shifting the microbial community and gene expression of carbohydrate-active enzymes

10.21203/rs.3.rs-32565/v1 ◽

2020 ◽

Author(s):

MengMeng Li ◽

Robin R. White ◽

Le Luo Guan ◽

Laura Harthan ◽

Mark D. Hanigan

Keyword(s):

Microbial Community ◽

Volatile Fatty Acids ◽

Liquid Fraction ◽

Transcriptome Profiling ◽

Metabolizable Energy ◽

Ruminal Fermentation ◽

Bacterial Phyla ◽

Ruminal Ph ◽

Fiber Degradation ◽

Genes Encoding

Abstract Background Volatile fatty acids (VFA) generated from ruminal fermentation by microorganisms provide up to 75% of total metabolizable energy in ruminants. Ruminal pH is an important factor affecting the profile and production of VFA by shifting the microbial community. However, how ruminal pH affects the microbial community and its relationship with expression of genes encoding carbohydrate-active enzyme (CAZyme) for fiber degradation and fermentation are not well investigated. To fill in this knowledge gap, six cannulated Holstein heifers were subjected to a continuous 10-day intraruminal infusion of distilled water or a dilute blend of hydrochloric and phosphoric acids to achieve a pH reduction of 0.5 units in a cross-over design. RNA-seq based transcriptome profiling was performed using total RNA extracted from ruminal liquid and solid fractions collected on day 9 of each period, respectively. Results Metatranscriptomic analyses identified 19 bacterial phyla with 121 genera, 3 archaeal genera, 11 protozoal genera, and 97 CAZyme transcripts in sampled ruminal contents. Within these, 4 bacteria phyla (Proteobacteria, Firmicutes, Bacteroidetes, and Spirochaetes), 2 archaeal genera (Candidatus Methanomethylophilus and Methanobrevibacter), and 5 protozoal genera (Entodinium, Polyplastron, Isotricha, Eudiplodinium, and Eremoplastron) were considered as the core active microbes, and genes encoding for cellulase, endo-1,4-beta- xylanase, amylase, and alpha-N-arabinofuranosidase were the most abundant CAZyme transcripts distributed in the rumen. Rumen microbiota is not equally distributed throughout the liquid and solid phases of rumen contents, and ruminal pH significantly affect microbial ecosystem, especially for the liquid fraction. In general, 76 bacterial genera, 4 protozoal genera, and 48 genes encoding CAZyme were significantly correlated with metabolic pathways for fiber degradation and VFA production. Within them, 29 bacterial genera, 4 protozoal genera, and 6 genes encoding CAZyme could be regulated by ruminal pH. Conclusions The ruminal microbiome changed the expression of transcripts for biochemical pathways of fiber degradation and VFA production in response to reduced pH, and at least a portion of the shifts in enzyme transcripts was associated with altered microbial community structure.

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Metatranscriptomic analyses reveal ruminal pH regulates fiber degradation and fermentation by shifting the microbial community and gene expression of carbohydrate-active enzymes

10.21203/rs.3.rs-32565/v2 ◽

2020 ◽

Author(s):

MengMeng Li ◽

Robin R. White ◽

Le Luo Guan ◽

Laura Harthan ◽

Mark D. Hanigan

Keyword(s):

Microbial Community ◽

Volatile Fatty Acids ◽

Liquid Fraction ◽

Transcriptome Profiling ◽

Metabolizable Energy ◽

Ruminal Fermentation ◽

Bacterial Phyla ◽

Ruminal Ph ◽

Fiber Degradation ◽

Genes Encoding

Abstract Background: Volatile fatty acids (VFA) generated from ruminal fermentation by microorganisms provide up to 75% of total metabolizable energy in ruminants. Ruminal pH is an important factor affecting the profile and production of VFA by shifting the microbial community. However, how ruminal pH affects the microbial community and its relationship with expression of genes encoding carbohydrate-active enzyme (CAZyme) for fiber degradation and fermentation are not well investigated. To fill in this knowledge gap, six cannulated Holstein heifers were subjected to a continuous 10-day intraruminal infusion of distilled water or a dilute blend of hydrochloric and phosphoric acids to achieve a pH reduction of 0.5 units in a cross-over design. RNA-seq based transcriptome profiling was performed using total RNA extracted from ruminal liquid and solid fractions collected on day 9 of each period, respectively.Results: Metatranscriptomic analyses identified 19 bacterial phyla with 156 genera, 3 archaeal genera, 11 protozoal genera, and 97 CAZyme transcripts in sampled ruminal contents. Within these, 4 bacteria phyla (Proteobacteria, Firmicutes, Bacteroidetes, and Spirochaetes), 2 archaeal genera (Candidatus Methanomethylophilus and Methanobrevibacter), and 5 protozoal genera (Entodinium, Polyplastron, Isotricha, Eudiplodinium, and Eremoplastron) were considered as the core active microbes, and genes encoding for cellulase, endo-1,4-beta- xylanase, amylase, and alpha-N-arabinofuranosidase were the most abundant CAZyme transcripts distributed in the rumen. Rumen microbiota is not equally distributed throughout the liquid and solid phases of rumen contents, and ruminal pH significantly affect microbial ecosystem, especially for the liquid fraction. In total, 21 bacterial genera, 4 protozoal genera, and 6 genes encoding CAZyme were regulated by ruminal pH. Metabolic pathways participated in glycolysis, pyruvate fermentation to acetate, lactate, and propanoate were downregulated by low pH in the liquid fraction.Conclusions: The ruminal microbiome changed the expression of transcripts for biochemical pathways of fiber degradation and VFA production in response to reduced pH, and at least a portion of the shifts in transcripts was associated with altered microbial community structure.

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Effects of fibrolytic enzymes and isobutyrate on ruminal fermentation, microbial enzyme activity and cellulolytic bacteria in pre- and post-weaning dairy calves

Animal Production Science ◽

10.1071/an17270 ◽

2019 ◽

Vol 59 (3) ◽

pp. 471 ◽

Cited By ~ 3

Author(s):

C. Wang ◽

Q. Liu ◽

G. Guo ◽

W. J. Huo ◽

Y. X. Wang ◽

...

Keyword(s):

Enzyme Activity ◽

Volatile Fatty Acids ◽

Dairy Calves ◽

Ruminal Fermentation ◽

Cellulolytic Bacteria ◽

Fibrolytic Enzymes ◽

Ruminal Ph ◽

Bull Calves ◽

Microbial Enzyme ◽

Microbial Enzyme Activity

The objective of the present study was to evaluate the effects of fibrolytic enzymes (FE, containing 160 units of cellulase and 4000 units of xylanase) or isobutyrate (IB) supplementation on ruminal fermentation, microbial enzyme activity and cellulolytic bacteria in dairy calves. Forty-eight Holstein bull calves of 15 days of age and of 44.9 ± 0.28 kg of BW were randomly assigned to four groups in a 2 × 2 factorial arrangement. Two levels of FE (0 g (FE–) or 1.83 g per calf per day (FE+)) and IB (0 g (IB–) or 6 g per calf per day (IB+)) were added. Calves were weaned at 60-day-old and four calves were selected from each treatment at random and slaughtered at 45 and 90 days of age. There was no IB × FE interaction effect. Ruminal pH decreased with IB or FE supplementation for post-weaned calves, whereas concentrations of total volatile fatty acids and acetate increased with IB or FE supplementation for pre- and post-weaned calves. Acetate to propionate ratio increased with IB supplementation, but was unaffected by FE supplementation. Ammonia-N concentration decreased with IB or FE supplementation for pre- and post-weaned calves. For post-weaned calves, activities of CMCase increased with IB or FE supplementation, and activities of cellobiase, xylanase, pectinase, β-amylase and protease increased with IB supplementation. Populations of B. fibrisolvens and F. succinogenes for pre- and post-weaned calves and R. flavefaciens for post-weaned calves increased with IB or FE supplementation. It is suggested that ruminal fermentation and growth performance of calves was improved with IB and FE supplementation, and the combination of IB and FE has the potential to stimulate the growth of pre- and post-weaned dairy calves.

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Metatranscriptomic analyses reveal ruminal pH regulates fiber degradation and fermentation by shifting the microbial community and gene expression of carbohydrate-active enzymes

Animal Microbiome ◽

10.1186/s42523-021-00092-6 ◽

2021 ◽

Vol 3 (1) ◽

Author(s):

Meng M. Li ◽

Robin R. White ◽

Le Luo Guan ◽

Laura Harthan ◽

Mark D. Hanigan

Keyword(s):

Microbial Community ◽

Volatile Fatty Acids ◽

Liquid Fraction ◽

Transcriptome Profiling ◽

Metabolizable Energy ◽

Ruminal Fermentation ◽

Bacterial Phyla ◽

Ruminal Ph ◽

Fiber Degradation ◽

Genes Encoding

Abstract Background Volatile fatty acids (VFA) generated from ruminal fermentation by microorganisms provide up to 75% of total metabolizable energy in ruminants. Ruminal pH is an important factor affecting the profile and production of VFA by shifting the microbial community. However, how ruminal pH affects the microbial community and its relationship with expression of genes encoding carbohydrate-active enzyme (CAZyme) for fiber degradation and fermentation are not well investigated. To fill in this knowledge gap, six cannulated Holstein heifers were subjected to a continuous 10-day intraruminal infusion of distilled water or a dilute blend of hydrochloric and phosphoric acids to achieve a pH reduction of 0.5 units in a cross-over design. RNA-seq based transcriptome profiling was performed using total RNA extracted from ruminal liquid and solid fractions collected on day 9 of each period, respectively. Results Metatranscriptomic analyses identified 19 bacterial phyla with 156 genera, 3 archaeal genera, 11 protozoal genera, and 97 CAZyme transcripts in sampled ruminal contents. Within these, 4 bacteria phyla (Proteobacteria, Firmicutes, Bacteroidetes, and Spirochaetes), 2 archaeal genera (Candidatus methanomethylophilus and Methanobrevibacter), and 5 protozoal genera (Entodinium, Polyplastron, Isotricha, Eudiplodinium, and Eremoplastron) were considered as the core active microbes, and genes encoding for cellulase, endo-1,4-beta- xylanase, amylase, and alpha-N-arabinofuranosidase were the most abundant CAZyme transcripts distributed in the rumen. Rumen microbiota is not equally distributed throughout the liquid and solid phases of rumen contents, and ruminal pH significantly affect microbial ecosystem, especially for the liquid fraction. In total, 21 bacterial genera, 4 protozoal genera, and 6 genes encoding CAZyme were regulated by ruminal pH. Metabolic pathways participated in glycolysis, pyruvate fermentation to acetate, lactate, and propanoate were downregulated by low pH in the liquid fraction. Conclusions The ruminal microbiome changed the expression of transcripts for biochemical pathways of fiber degradation and VFA production in response to reduced pH, and at least a portion of the shifts in transcripts was associated with altered microbial community structure.

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Yield, chemical composition, fermentation characteristics, in vitro ruminal variables, and degradability of ensiled amaranth (Amaranthus hypochondriacus) cultivars compared with corn (Zea mays) silage

Translational Animal Science ◽

10.1093/tas/txaa180 ◽

2020 ◽

Vol 4 (4) ◽

Author(s):

Hossein Shadi ◽

Yousef Rouzbehan ◽

Javad Rezaei ◽

Hassan Fazaeli

Keyword(s):

Chemical Composition ◽

Crude Protein ◽

Nutritional Value ◽

Volatile Fatty Acids ◽

Block Design ◽

Metabolizable Energy ◽

Neutral Detergent Fiber ◽

Cellulolytic Bacteria

Abstract Silages from four amaranth varieties (A5, A12, A14, and A28) were compared with corn silage (CS) in terms of their yield, chemical composition, phenolic compounds, oxalic acid and nitrate levels, silage fermentation characteristics, in vitro methane production, organic matter disappearance (OMD), microbial crude protein (MCP), ruminal ammonia (NH3-N), pH, volatile fatty acids, cellulolytic bacteria numbers, protozoa counts, and in situ dry matter (DM) and crude protein (CP) degradability were determined. Forages were harvested 93 d after planting, chopped, and ensiled in plastic buckets for 60 d. The study was based on a randomized complete block design, and data were analyzed using SAS, general linear model (GLM) procedure for normal distribution. Compared with CS, amaranth silages (AMS) had lower ash-free neutral detergent fiber nitrate, OMD (P < 0.001), phosphorus (P = 0.003), and metabolizable energy (ME) (P = 0.043) but higher (P < 0.001) CP, calcium, non-fiber carbohydrates (NFC), acid detergent lignin, ether extract, ash, total phenolics, pH, NH3-N concentration, MCP, digestible undegradable protein (DUP), and metabolizable protein (MP). Fresh, OM, OMD, ME (P < 0.001), and DM (P = 0.032) yields of AMS from different varieties were higher than CS, with the exception of A5. Overall, amaranth made good quality silage, with some variation, and A28 had the highest yield and nutritional value (CP, NFC, MCP, DUP, and MP). The yield, CP concentration, and nutritional value of A28 silage were higher than CS. Although these in vitro results are promising, they also need to be validated with future in vivo research.

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Effects of brewers’ spent grain protein hydrolysates on gas production, ruminal fermentation characteristics, microbial protein synthesis and microbial community in an artificial rumen fed a high grain diet

Journal of Animal Science and Biotechnology ◽

10.1186/s40104-020-00531-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Tao Ran ◽

Long Jin ◽

Ranithri Abeynayake ◽

Atef Mohamed Saleem ◽

Xiumin Zhang ◽

...

Keyword(s):

Protein Synthesis ◽

Microbial Community ◽

Antioxidant Properties ◽

Value Added ◽

Gas Production ◽

Protein Hydrolysates ◽

Ruminal Fermentation ◽

Microbial Protein ◽

Microbial Protein Synthesis ◽

Spent Grain

Abstract Background Brewers’ spent grain (BSG) typically contains 20% – 29% crude protein (CP) with high concentrations of glutamine, proline and hydrophobic and non-polar amino acid, making it an ideal material for producing value-added products like bioactive peptides which have antioxidant properties. For this study, protein was extracted from BSG, hydrolyzed with 1% alcalase and flavourzyme, with the generated protein hydrolysates (AlcH and FlaH) showing antioxidant activities. This study evaluated the effects of AlcH and FlaH on gas production, ruminal fermentation characteristics, nutrient disappearance, microbial protein synthesis and microbial community using an artificial rumen system (RUSITEC) fed a high-grain diet. Results As compared to the control of grain only, supplementation of FlaH decreased (P < 0.01) disappearances of dry matter (DM), organic matter (OM), CP and starch, without affecting fibre disappearances; while AlcH had no effect on nutrient disappearance. Neither AlcH nor FlaH affected gas production or VFA profiles, however they increased (P < 0.01) NH3-N and decreased (P < 0.01) H2 production. Supplementation of FlaH decreased (P < 0.01) the percentage of CH4 in total gas and dissolved-CH4 (dCH4) in dissolved gas. Addition of monensin reduced (P < 0.01) disappearance of nutrients, improved fermentation efficiency and reduced CH4 and H2 emissions. Total microbial nitrogen production was decreased (P < 0.05) but the proportion of feed particle associated (FPA) bacteria was increased with FlaH and monensin supplementation. Numbers of OTUs and Shannon diversity indices of FPA microbial community were unaffected by AlcH and FlaH; whereas both indices were reduced (P < 0.05) by monensin. Taxonomic analysis revealed no effect of AlcH and FlaH on the relative abundance (RA) of bacteria at phylum level, whereas monensin reduced (P < 0.05) the RA of Firmicutes and Bacteroidetes and enhanced Proteobacteria. Supplementation of FlaH enhanced (P < 0.05) the RA of genus Prevotella, reduced Selenomonas, Shuttleworthia, Bifidobacterium and Dialister as compared to control; monensin reduced (P < 0.05) RA of genus Prevotella but enhaced Succinivibrio. Conclusions The supplementation of FlaH in high-grain diets may potentially protect CP and starch from ruminal degradation, without adversely affecting fibre degradation and VFA profiles. It also showed promising effects on reducing CH4 production by suppressing H2 production. Protein enzymatic hydrolysates from BSG using flavourzyme showed potential application to high value-added bio-products.

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Effects of Increasing Doses of Condensed Tannins Extract from Cistus ladanifer L. on In Vitro Ruminal Fermentation and Biohydrogenation

Animals ◽

10.3390/ani11030761 ◽

2021 ◽

Vol 11 (3) ◽

pp. 761

Author(s):

Olinda Guerreiro ◽

Susana P. Alves ◽

Mónica Costa ◽

Maria F. Duarte ◽

Eliana Jerónimo ◽

...

Keyword(s):

Fatty Acids ◽

Condensed Tannins ◽

Volatile Fatty Acids ◽

Secondary Compounds ◽

Cistus Ladanifer ◽

Ruminal Fermentation ◽

Experimental Conditions ◽

Ct Dose ◽

Linear Decrease

Cistus ladanifer (rockrose) is a perennial shrub quite abundant in the Mediterranean region, and it is a rich source in secondary compounds such as condensed tannins (CTs). Condensed tannins from C. ladanifer were able to change the ruminal biohydrogenation (BH), increasing the t11–18:1 and c9,t11–18:2 production. However, the adequate conditions of the C. ladanifer CTs used to optimize the production of t11–18:1 and c9,t11–18:2 is not yet known. Thus, we tested the effect of increasing the doses of C. ladanifer CT extract (0, 25, 50, 75 and 100 g/kg dry matter (DM)) on in vitro rumen BH. Five in vitro batch incubations replicates were conducted using an oil supplemented high-concentrate substrate, incubated for 24 h with 6 mL of buffered ruminal fluid. Volatile fatty acids (VFAs) and long chain fatty acids (FA) were analyzed at 0 h and 24 h, and BH of c9–18:1, c9, c12–18:2 and c9, c12, c15–18:3, and BH products yield were computed. Increasing doses of C. ladanifer CTs led to a moderate linear decrease (p < 0.001) of the VFA production (a reduction of 27% with the highest dose compared to control). The disappearance of c9–18:1 and c9,c12–18:2 as well as the production of t11–18:1 and c9, t11:18:2 was not affected by increasing doses of C. ladanifer CTs, and only the disappearance of c9, c12, c15–18:3 suffered a mild linear decrease (a reduction of 24% with the highest dose compared to control). Nevertheless, increasing the C. ladanifer CT dose led to a strong depression of microbial odd and branched fatty acids and of dimethyl acetals production (less than 65% with the highest dose compared to control), which indicates that microbial growth was more inhibited than fermentative and biohydrogenation activities, in a possible adaptative response of microbial population to stress induced to CTs and polyunsaturated fatty acids. The ability of C. ladanifer to modulate the ruminal BH was not verified in the current in vitro experimental conditions, emphasizing the inconsistent BH response to CTs and highlighting the need to continue seeking the optimal conditions for using CTs to improve the fatty acid profile of ruminant fat.

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Effects of Methanogenic Inhibitors on Methane Production and Abundances of Methanogens and Cellulolytic Bacteria inIn VitroRuminal Cultures

Applied and Environmental Microbiology ◽

10.1128/aem.02779-10 ◽

2011 ◽

Vol 77 (8) ◽

pp. 2634-2639 ◽

Cited By ~ 82

Author(s):

Zhenming Zhou ◽

Qingxiang Meng ◽

Zhongtang Yu

Keyword(s):

Methane Production ◽

Volatile Fatty Acids ◽

Denaturing Gradient Gel Electrophoresis ◽

Cellulolytic Bacteria ◽

Bacterial Populations ◽

Content Type ◽

Specific Pcr ◽

Methanogen Population ◽

Propynoic Acid

ABSTRACTThe objective of this study was to systematically evaluate and compare the effects of select antimethanogen compounds on methane production, feed digestion and fermentation, and populations of ruminal bacteria and methanogens usingin vitrocultures. Seven compounds, including 2-bromoethanesulphonate (BES), propynoic acid (PA), nitroethane (NE), ethyltrans-2-butenoate (ETB), 2-nitroethanol (2NEOH), sodium nitrate (SN), and ethyl-2-butynote (EB), were tested at a final concentration of 12 mM. Ground alfalfa hay was included as the only substrate to simulate daily forage intake. Compared to no-inhibitor controls, PA, 2NEOH, and SN greatly reduced the production of methane (70 to 99%), volatile fatty acids (VFAs; 46 to 66%), acetate (30 to 60%), and propionate (79 to 82%), with 2NEOH reducing the most. EB reduced methane production by 23% without a significant effect on total VFAs, acetate, or propionate. BES significantly reduced the propionate concentration but not the production of methane, total VFAs, or acetate. ETB or NE had no significant effect on any of the above-mentioned measurements. Specific quantitative-PCR (qPCR) assays showed that none of the inhibitors significantly affected total bacterial populations but that they did reduce theFibrobacter succinogenespopulation. SN reduced theRuminococcus albuspopulation, while PA and 2NEOH increased the populations of bothR. albusandRuminococcus flavefaciens. Archaeon-specific PCR-denaturing gradient gel electrophoresis (DGGE) showed that all the inhibitors affected the methanogen population structure, while archaeon-specific qPCR revealed a significant decrease in methanogen population in all treatments. These results showed that EB, ETB, NE, and BES can effectively reduce the total population of methanogens but that they reduce methane production to a lesser extent. The results may guide futureinvivostudies to develop effective mitigation of methane emission from ruminants.

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Comparison of Two Sampling Techniques for Evaluating Ruminal Fermentation and Microbiota in the Planktonic Phase of Rumen Digesta in Dairy Cows

Frontiers in Microbiology ◽

10.3389/fmicb.2020.618032 ◽

2020 ◽

Vol 11 ◽

Author(s):

Camila Flavia de Assis Lage ◽

Susanna Elizabeth Räisänen ◽

Audino Melgar ◽

Krum Nedelkov ◽

Xianjiang Chen ◽

...

Keyword(s):

Volatile Fatty Acids ◽

Sampling Method ◽

Bacterial Community Composition ◽

Sampling Technique ◽

Standard Diet ◽

Ruminal Fermentation ◽

Ph Change ◽

Ruminal Fluid ◽

Rumen Ph ◽

Dissolved Hydrogen

The objective of this experiment was to compare ruminal fluid samples collected through rumen cannula (RC) or using an oral stomach tube (ST) for measurement of ruminal fermentation and microbiota variables. Six ruminally cannulated lactating Holstein cows fed a standard diet were used in the study. Rumen samples were collected at 0, 2, 4, 6, 8, and 12 h after the morning feeding on two consecutive days using both RC and ST techniques. Samples were filtered through two layers of cheesecloth and the filtered ruminal fluid was used for further analysis. Compared with RC, ST samples had 7% greater pH; however, the pattern in pH change after feeding was similar between sampling methods. Total volatile fatty acids (VFA), acetate and propionate concentrations in ruminal fluid were on average 23% lower for ST compared with RC. There were no differences between RC and ST in VFA molar proportions (except for isobutyrate), ammonia and dissolved hydrogen (dH2) concentrations, or total protozoa counts, and there were no interactions between sampling technique and time of sampling. Bacterial ASV richness was higher in ST compared with RC samples; however, no differences were observed for Shannon diversity. Based on Permanova analysis, bacterial community composition was influenced by sampling method and there was an interaction between sampling method and time of sampling. A core microbiota comprised of Prevotella, S24-7, unclassified Bacteroidales and unclassified Clostridiales, Butyrivibrio, unclassified Lachnospiraceae, unclassified Ruminococcaceae, Ruminococcus, and Sharpea was present in both ST and RC samples, although their relative abundance varied and was influenced by an interaction between sampling time and sampling method. Overall, our results suggest that ruminal fluid samples collected using ST (at 180 to 200 cm depth) are not representative of rumen pH, absolute values of VFA concentrations, or bacterial communities >2 h post-feeding when compared to samples of ruminal fluid collected using RC. However, ST can be a feasible sampling technique if the purpose is to study molar proportions of VFA, protozoa counts, dH2, and ammonia concentrations.

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