Search

Abstract MicroRNAs (miRNAs), as important post-transcriptional regulators, are ubiquitous in various tissues. The aim of this exploratory study was to determine the presence of miRNAs in rumen fluid, and to investigate the possibility of miRNA-mediated cross-talk within the ruminal ecosystem. Rumen fluid samples from four cannulated Holstein cows were collected during two feeding regimes (forage and high-grain diet) and DNA and RNA were extracted for amplicon and small RNA sequencing. Epithelial biopsies were simultaneously collected to investigate the co-expression of miRNAs in papillae and rumen fluid. We identified 377 miRNAs in rumen fluid and 638 in rumen papillae, of which 373 were shared. Analysis of microbiota revealed 20 genera to be differentially abundant between the two feeding regimes, whereas no difference in miRNAs expression was detected. Correlations with at least one genus were found for 170 miRNAs, of which, 39 were highly significant (r > |0.7| and P

Agroecosystems are comprised of environmental compartments where associated microbial communities interact with one another. These microbial communities, called microbiomes, inhabit livestock, insects, and plants. Microbiomes are also present in the soil and watersheds. Clarifying the nature and extent of microbial interactions between compartments both at intra-farm and global scales can promote sustainable production systems, healthier animals, increased crop yields, and safer meat products. Early research on microbiomes was hindered by a lack of expertise and the high cost of molecular sequencing. However, these limitations have been largely resolved with advances in and reduced costs of sequencing technologies. In this paper, we summarize sequencing and bioinformatics approaches, and review the crucial roles of diverse microbiomes in livestock, plants and soil, as well as pollinators and pest insects. These crucial roles include nutrient cycling, nutrient acquisition, metabolism of toxins and enhanced host immune function. Additionally, we examine potentially undesirable effects of microbiomes associated with climate change and agri-food production such as their role in the release of greenhouse gases from cattle and their impact on meat safety and spoilage. By increasing the awareness of microbiomes and the growing ease with which they can be studied, we hope to foster a greater adoption of microbiome research. Further understanding of the diverse effects and interactions of microbiomes will advance our efforts to increase agricultural production while reducing its negative environmental footprint, thus making the agroecosystems more sustainable.

Switching diets from forage to a high-grain (HG) diet increases the risk of rumen fermentation disorders in cattle. However, the effects of the duration of the HG feeding, after the diet switch, on animal behavior and health have received considerably less attention. This experiment primarily aimed to assess the effects of the duration of an HG diet on the chewing, eating, and lying behavior and salivation dynamics in a control group (CON) and a group of cows receiving a phytogenic feed supplement (TRT) at 0.04% (DM basis), which included L-menthol, thymol, eugenol, mint oil, and cloves powder. The experiment was a crossover design with nine non-lactating cows, and two experimental periods with an intermediate washout of four weeks. In each period, the cows were first fed a forage diet for a week to collect baseline measurements representing week 0; then, the diet was switched over a week to HG (65% concentrate), which was fed for four continuous weeks (week 1, week 2, week 3, and week 4 on an HG diet, respectively). The cows were divided in two groups of four and five animals and were randomly allocated to CON or TRT. The data analysis revealed that at the start of the HG feeding, the dry matter intake and the cows’ number of lying bouts increased, but the eating time, rumination time, and meal frequency decreased, resulting in a greater eating rate. We also found that an advanced duration on an HG diet further decreased the rumination time, total chewing time, chewing index, and sorting in favor of short feed particles, with the lowest values in week 4. The feed bolus size increased but feed the ensalivation decreased in week 4 compared to week 0. The dietary switch increased salivary lysozyme activity, and the advanced duration on the HG diet increased salivary pH, but salivary phosphate decreased in weeks 1 and 2 on the HG diet. Supplementation with TRT increased sorting in favor of physically effective NDF (peNDF) in week 2 and increased salivary pH in week 4 on an HG diet. Overall, the negative effects of the HG diet in cattle are more pronounced during the initial stage of the HG feeding. However, several detrimental effects were exacerbated with the cows’ advanced duration on feed, with host adaptive changes still observed after 3 and 4 weeks following the diet switch. The TRT mitigated some of the negative effects through the temporal improvement of the salivary properties and the intake of peNDF, which are known to modulate rumen fermentation.

ABSTRACT Differences in chickens’ feed intake may be the underlying factor influencing feed-efficiency (FE)-associated variation in intestinal microbiota and physiology. In chickens eating the same amount of feed, quantitative feed restriction may create similar intestinal conditions and help clarify this cause-and-effect relationship. This study investigated the effect of ad libitum versus restrictive feeding (85% of ad libitum) on ileal and cecal microbiota, concentrations of short-chain fatty acids, visceral organ size, intestinal morphology, permeability, and expression of genes related to nutrient uptake, barrier function, and innate immune response in broiler chickens with divergent residual feed intake (RFI; metric for FE). On day 30 posthatch, 28 low-RFI (good FE) and 29 high-RFI (poor FE) chickens across both feeding-level groups (n = 112) were selected. Supervised multigroup data integration and relevance network analyses showed that especially Lactobacillus (negative) in ileal digesta, Turicibacter (positive) in cecal digesta, and Enterobacteriaceae (positive) in both intestinal segments depended on chicken’s feed intake, whereas the level of Anaerotruncus in cecal digesta was most discriminative for high RFI. Moreover, shallower crypts and fewer goblet cells in ceca indicated host-related energy-saving mechanisms with low RFI, whereas greater tissue resistance suggested a stronger jejunal barrier function in low-RFI chickens. Values corresponding to feed intake level × RFI interactions indicated larger pancreas and lower levels of ileal and cecal short-chain fatty acids in restrictively fed high-RFI chickens than in the other 3 groups, suggesting host physiological adaptations to support greater energy and nutrient needs of high-RFI chickens compensating for the restricted feeding. IMPORTANCE The impact of the FE-associated differences in feed intake on intestinal bacterial and host physiological parameters has so far not been clarified. Understanding the underlying principles is essential for the development of cost-effective strategies to improve FE in chicken production. Under conditions of quantitative feed restriction, low- and high-RFI chickens ate the same amount of feed. Therefore, this research helps in distinguishing intestinal bacterial taxa and functions that were highly reliant on feed intake from those that were associated with physiological adaptations to RFI-associated differences in host nutritional needs and intestinal nutrient availability. This work provides a background for further research to assess manipulation of the intestinal microbiota, host physiology, and FE in chickens by dietary intervention.

There is a great interest to understand the impact of the gut microbiota on host's nutrient use and FE in chicken production. Both chicken's feed intake and gut bacterial microbiota differ between high and low-feed efficient chickens. To evaluate the impact of the feed intake level on the feed efficiency (FE)-associated variation in the chicken intestinal microbiota, differently feed efficient chickens need to eat the same amount of feed, which can be achieved by feeding chickens restrictively. Therefore, we investigated the effect of restrictive vs. ad libitum feeding on the fecal microbiome at 16 and 29 days posthatch (dph), FE and nutrient retention in chickens of low and high residual feed intake (RFI; metric for FE). Restrictively fed chickens were provided the same amount of feed which corresponded to 85% of the ad libitum fed group from 9 dph. FE was determined for the period between 9 and 30 dph and feces for nutrient retention were collected on 31 to 32 dph. From the 112 chickens (n = 56 fed ad libitum, and n = 56 fed restrictively), 14 low RFI and 15 high RFI ad libitum fed chickens, and 14 low RFI (n = 7 per sex) and 14 high RFI restrictively fed chickens were selected as the extremes in RFI and were retrospectively chosen for data analysis. Bray-Curtis dissimilarity matrices showed significant separation between time points, and feeding level groups at 29 dph for the fecal bacterial communities. Relevance networking indicated positive associations between Acinetobacter and feed intake at 16 dph, whereas at 29 dph Escherichia/Shigella and Turicibacter positively and Lactobacillus negatively correlated to chicken's feed intake. Enterobacteriaceae was indicative for low RFI at 16 dph, whereas Acinetobacter was linked to high RFI across time points. However, restrictive feeding-associated changes in the fecal microbiota were not similar in low and high RFI chickens, which may have been related to the higher nutrient retention and thus lower fecal nutrient availability in restrictively fed high RFI chickens. This may also explain the decreased RFI value in restrictively fed high RFI chickens indicating improved FE, with a stronger effect in females.

Dietary inclusion of resistant starches can promote host health through modulation of the gastrointestinal microbiota, short-chain fatty acid (SCFA) profiles, and lipid metabolism. This study investigated the impact of a transglycosylated cornstarch (TGS) on gastric, ileal, cecal, proximal-colonic, and mid-colonic bacterial community profiles and fermentation metabolites using a growing pig model. It additionally evaluated the effect of TGS on the expression of host genes related to glucose and SCFA absorption, incretins, and satiety in the gut as well as host genes related to lipid metabolism in hepatic and adipose tissue. Sixteen growing pigs (4 months of age) were fed either a TGS or control (CON) diet for 11 days. Bacterial profiles were determined via Illumina MiSeq sequencing of the V3–5 region of the 16S rRNA gene, whereas SCFA and gene expression were measured using gas chromatography and reverse transcription-quantitative PCR. Megasphaera, which was increased at all gut sites, began to benefit from TGS feeding in gastric digesta, likely through cross-feeding with other microbes, such as Lactobacillus. Shifts in the bacterial profiles from dietary TGS consumption in the cecum, proximal colon, and mid colon were similar. Relative abundances of Ruminococcus and unclassified Ruminococcaceae genus were lower, whereas that of unclassified Veillonellaceae genus was higher in TGS- compared to CON-fed pigs (p < 0.05). TGS consumption also increased (p < 0.05) concentrations of SCFA, especially propionate, and lactate in the distal hindgut compared to the CON diet which might have up-regulated GLP1 expression in the cecum (p < 0.05) and mid colon compared to the control diet (p < 0.10). TGS-fed pigs showed increased hepatic and decreased adipocyte expression of genes for lipid synthesis (FASN, SREBP1, and ACACA) compared to CON-fed pigs, which may be related to postprandial portal nutrient flow and reduced systemic insulin signaling. Overall, our data show that TGS consumption may affect gastrointestinal bacterial signaling, caused by changes in gut bacterial profiles and the action of propionate, and host lipid metabolism.

Microbial composition and activity in the gastrointestinal tract (GIT) of cattle has important implications for animal health and welfare, driving the focus of research toward ways to modify their function and abundance. However, our understanding of microbial adaption to nutritional changes remains limited. The aim of this study was to examine the progressive mechanisms of adaptation in the rumen and hindgut of cattle receiving increasing amounts of starch with or without dietary supplementation of a blended phytogenic feed additive (PFA; containing menthol, thymol and eugenol). We used 16S rRNA gene amplicon sequencing to assess the microbial composition and predicted metabolic pathways in ruminal solid and liquid digesta, and feces. Furthermore, we employed targeted liquid chromatography-mass spectrometry methods to evaluate rumen fluid metabolites. Results indicated a rapid microbial adaptation to diet change, starting on the second day of starch feeding for the particle associated rumen liquid (PARL) microbes. Solid rumen digesta- and feces-associated microbes started changing from the following day. The PARL niche was the most responsive to dietary changes, with the highest number of taxa and predicted pathways affected by the increase in starch intake, as well as by the phytogenic supplementation. Despite the differences in the microbial composition and metabolic potential of the different GIT niches, all showed similar changes toward carbohydrate metabolism. Metabolite measurement confirmed the high prevalence of glucose and volatile fatty acids (VFAs) in the rumen due to the increased substrate availability and metabolic activity of the microbiota. Families Prevotellaceae, Ruminococcaceae and Lachnospiraceae were found to be positively correlated with carbohydrate metabolism, with the latter two showing wide-ranging predicted metabolic capabilities. Phytogenic supplementation affected low abundant taxa and demonstrated the potential to prevent unwanted implications of feeding high-concentrate diet, such as reduction of microbial diversity. The inclusion of 50% concentrate in the diet caused a major shift in microbial composition and activity in the GIT of cattle. This study demonstrated the ability of microorganisms in various GIT niches to adjust differentially, yet rapidly, to changing dietary conditions, and revealed the potential beneficial effects of supplementation with a PFA during dietary adaptation.

Saliva facilitates feed ingestion, nutrient circulation, and represents an important pH buffer for ruminants, especially for cattle fed high-concentrate diets that promote rumen acidification. This experiment evaluated the short-term effects of nine phytogenic compounds on salivation, saliva physico-chemical composition as well as ingested feed boli characteristics in cattle. A total of nine ruminally cannulated Holstein cows were used. Each compound was tested in four of these cows as part of a high-concentrate meal (2.5 kg of total mixed ration in dry matter basis for 4 h) in low or high dose, and was compared to a control meal without compound. Saliva was sampled orally (unstimulated saliva) for physico-chemical composition analysis. Composition of the ingested saliva (stimulated saliva), salivation and feed boli characteristics were assessed from ingesta collected at the cardia during the first 30 min of the meal. Analysis of unstimulated saliva showed that supplementation with capsaicin and thyme oil increased buffer capacity, while supplementation with thymol, L-menthol and gentian root decreased saliva pH. In addition, supplementing angelica root decreased saliva osmolality. Regression analysis on unstimulated saliva showed negative associations between mucins and bicarbonate as well as with phosphate when garlic oil, thyme oil or angelica root was supplemented. Analysis of stimulated saliva demonstrated that supplementation with garlic oil increased phosphate concentration, thyme oil tended to increase osmolality, capsaicin and thymol increased buffer capacity, and ginger increased phosphate content. Furthermore, salivation rate increased with ginger and thymol, and tended to increase with garlic oil, capsaicin, L-menthol and mint oil. Feed ensalivation increased with capsaicin. A positive association was found between feed bolus size and salivation rate when any of the phytogenic compounds was supplemented. Overall, our results demonstrate positive short-term effects of several phytogenic compounds on unstimulated and stimulated saliva physico-chemical properties, salivation or feed boli characteristics. Thus, the phytogenic compounds enhancing salivary physico-chemical composition have the potential to contribute to maintain or improve ruminal health in cattle fed concentrate-rich rations.

Feeding with high-concentrate diets increases the risk of subacute ruminal acidosis (SARA). This experiment was conducted to evaluate whether supplementing a phytogenic feed additive based on L-menthol, thymol, eugenol, mint oil (Mentha arvensis) and cloves powder (Syzygium aromaticum) (PHY) can amend the ruminal fermentation profile, modulate the risk of SARA and reduce inflammation in cattle. The experiment was designed as a crossover design with nine non-lactating Holstein cows, and was conducted in two experimental runs. In each run, cows were fed a 100% forage diet one week (wk 0), and were then transitioned stepwise over one week (0 to 65% concentrate, wk adapt.) to a high concentrate diet that was fed for 4 weeks. Animals were fed diets either with PHY or without (CON). The PHY group had an increased ruminal pH compared to CON, reduced time to pH < 5.8 in wk 3, which tended to decrease further in wk 4, reduced the ruminal concentration of D-lactate, and tended to decrease total lactate (wk 3). In wk 2, PHY increased acetate, butyrate, isobutyrate, isovalerate, and the acetate to propionate ratio compared to CON. Phytogenic supplementation reduced inflammation compared to CON in wk 3. Overall, PHY had beneficial effects on ruminal fermentation, reduced inflammation, and modulated the risk of SARA starting from wk 3 of supplementation.

Starch-rich diets can cause subacute ruminal acidosis (SARA) in dairy cows with potentially different susceptibility according to lactation number. We wanted to evaluate the bacterial community and the fermentation end products in feces to study susceptibility to hindgut acidosis and dysbiosis. Sixteen dairy cows received a medium-concentrate diet (MC, 40% concentrate, 18.8% starch) for one week and a high-concentrate diet (HC, 60% concentrate, 27.7% starch, DM) for four weeks. Milk yield, dry-matter intake, chewing activity, ruminal pH, milk constituents, and fecal samples for short-chain fatty acids (SCFA), pH, and 16S rRNA-gene sequencing were investigated. The HC feeding caused a reduction in fecal pH, bacterial diversity and richness, an increase in total SCFA, and a separate phylogenetic clustering of MC and HC samples. Ruminal and fecal pH had fair correlation (r = 0.5). Cows in the second lactation (2ndL) had lower dry matter intake (DMI) than cows of third or fourth or more lactations (3rdL; ≥4 L), whereas DMI/kg body weight was lower for ≥4 L than for 2ndL and 3rdL cows. The mean ruminal pH was highest in ≥4 L, whereas the time spent below the SARA threshold was highest for 3rdL cows. The latter also had higher total SCFA in the feces. Our results suggest that hindgut dysbiosis is caused by increased substrate flow to the hindgut, but further investigations are needed to define hindgut acidosis. The 3rdL cows were most susceptible to rumen acidosis and hindgut dysbiosis due to high DMI level, but missing counter regulations, as suggested happening in 2ndL and ≥4 L cows.

Extruded flaxseed (25%) and ground hay (75%) were each fed (DM basis) either together in a total mixed ration (TMR) or as flaxseed first followed by hay (non-TMR) to three pens of eight crossbred steers (n = 24 per diet) for 240 days. Compared to TMR, feeding non-TMR enriched subcutaneous fat with α-linolenic acid (ALA, 18:3n-3) and its biohydrogenation intermediates including vaccenic acid [trans(t)11-18:1], rumenic acid [cis(c)9,t11-conjugated linoleic acid] and conjugated linolenic acid (CLnA). Rumen microbial analysis using QIIME indicated that 14 genera differed (P ≤ 0.05) between TMR and the non-TMR. Azoarcus and Streptococcus were the only genera which increased in relative abundance in the TMR fed steers, whereas Methanimicrococcus, Moryella, Prevotella, Succiniclasticum, Succinivibrio, Suttenella, and TG5 decreased as compared to steers fed the non-TMR. Among these, Moryella, Succiniclasticum, and Succinivibrio, spp. were correlated with fatty acid profiles, specifically intermediates believed to be components of the major biohydrogenation pathway for ALA (i.e., t11, c15-18:2, c9, t11, c15-18:3, and total CLnA). In addition, negative correlations were found between the less abundant Ruminoccocus-like OTU60 and major ALA biohydrogenation intermediates, as well as positive correlations with several intermediates from alternative pathways that did not involve the formation of trans 11 double bonds. The present results suggest a number of pathways for ALA biohydrogenation are operating concurrently in the rumen, with their balance being influenced by diet and driven by less abundant species rather than members of the core bacterial population.