Bacterial Leaf Spot Susceptibility Screening of Chili Pepper Cultivars Using qPCR Determination of Xanthomonas euvesicatoria pv. euvesicatoria Titers.

Bacterial leaf spot is a serious disease of chili pepper (Capsicum spp.) caused by Xanthomonas euvesicatoria pv. euvesicatoria. Conventional resistance screening is time and resource intensive. It was considered that a quick and simple determination of cultivar susceptibility could be achieved through estimating bacterial titers of inoculated plants. A SYBR quantitative polymerase chain reaction (qPCR)-based assay was compared with conventional PCR, then used to detect and enumerate pathogen titers in serial dilutions and DNA extracted from infected plant leaves. The qPCR detection limit was approximately 1 CFU µl-1, 10 times more sensitive than conventional PCR. A linear correlation (R2 = 0.994) was obtained from the standard curve comparing plate-truthed serial dilutions of the pathogen with the qPCR cycle threshold. Six strains were used to inoculate cultivars Hugo and Warlock. One strain, X. euvesicatoria pv. euvesicatoria BRIP62403, was consistently the most virulent based on visual symptoms and pathogen titers in planta inferred by qPCR performed on DNA extracted from infected leaves 2 and 6 weeks postinoculation. Visual observations 6 weeks after inoculation were highly correlated (R2 = 0.8254) to pathogen titers. The qPCR method was used to categorize 20 chili pepper cultivars 2 weeks after inoculation. A high positive correlation (R2 = 0.6826) was observed between visual scoring and pathogen titers from 20 chili pepper cultivars, facilitating categorization of susceptible, intermediate, and resistant cultivars. The qPCR approach developed here facilitates susceptibility screening of chili pepper cultivars at an early stage of selection and could be readily adapted to a range of other pathosystems.

Recent Advances in Enteric Methane Mitigation and the Long Road to Sustainable Ruminant Production.

Mitigation of methane emission, a potent greenhouse gas, is a worldwide priority to limit global warming. A substantial part of anthropogenic methane is emitted by the livestock sector, as methane is a normal product of ruminant digestion. We present the latest developments and challenges ahead of the main efficient mitigation strategies of enteric methane production in ruminants. Numerous mitigation strategies have been developed in the last decades, from dietary manipulation and breeding to targeting of methanogens, the microbes that produce methane. The most recent advances focus on specific inhibition of key enzymes involved in methanogenesis. But these inhibitors, although efficient, are not affordable and not adapted to the extensive farming systems prevalent in low- and middle-income countries. Effective global mitigation of methane emissions from livestock should be based not only on scientific progress but also on the feasibility and accessibility of mitigation strategies.

Feeding an unsalable carrot total-mixed ration altered bacterial amino acid degradation in the rumen of lambs.

The objective of this study was to determine the influence of a total-mixed ration including unsalable carrots at 45% DM on the rumen microbiome; and the plasma, rumen and liver metabolomes. Carrots discarded at processing were investigated as an energy-dense substitute for barley grain in a conventional feedlot diet, and improved feed conversion efficiency by 25%. Here, rumen fluid was collected from 34 Merino lambs at slaughter (n = 16 control; n = 18 carrot) after a feeding period of 11-weeks. The V4 region of the 16S rRNA gene was sequenced to profile archaeal and bacterial microbe communities. Further, a comprehensive, targeted profile of known metabolites was constructed for blood plasma, rumen fluid and biopsied liver metabolites using a gas chromatography mass spectrometry (GC-MS) metabolomics approach. An in vitro batch culture was used to characterise ruminal fermentation including gas and methane (CH4) production. In vivo rumen microbial community structure of carrot fed lambs was dissimilar (P < 0.01; PERMANOVA), and all measures of alpha diversity were greater (P < 0.01), compared to those fed the control diet. Unclassified genera in Bacteroidales (15.9 ± 6.74% relative abundance; RA) were more abundant (P < 0.01) in the rumen fluid of carrot-fed lambs, while unclassified taxa in the Succinivibrionaceae family (11.1 ± 3.85% RA) were greater (P < 0.01) in the control. The carrot diet improved in vitro ruminal fermentation evidenced as an 8% increase (P < 0.01) in DM digestibility and a 13.8% reduction (P = 0.01) in CH4 on a mg/ g DM basis, while the control diet increased (P = 0.04) percentage of propionate within total VFA by 20%. Fourteen rumen fluid metabolites and 27 liver metabolites were influenced (P ≤ 0.05) by diet, while no effect (P ≥ 0.05) was observed in plasma metabolites. The carrot diet enriched (impact value = 0.13; P = 0.01) the tyrosine metabolism pathway (acetoacetic acid, dopamine and pyruvate), while the control diet enriched (impact value = 0.42; P ≤ 0.02) starch and sucrose metabolism (trehalose and glucose) in rumen fluid. This study demonstrated that feeding 45% DM unsalable carrots diversified bacterial communities in the rumen. These dietary changes influenced pathways of tyrosine degradation, such that previous improvements in feed conversion efficiency in lambs could be explained.

Characterisation of the Faecal Microbiome of Foals from 0-5 Months of Age and Their Respective Mares across Five Geographic Locations.

BACKGROUND: A foal undergoes considerable growth and development from birth to weaning, progressing from a milk-based diet to complete herbivory. The symbiotic relationships between bacteria, archaea and fungi substantiate this energy demand by colonising the hindgut and remaining flexible throughout the diet transitions. METHODS: A total of 70 faecal samples were collected from 14 mares and their foals across five studs in NSW as they aged from 0 to 5 months old. DNA was extracted from faecal samples and underwent amplification and sequencing of the 16S rRNA gene V4 hypervariable region of archaea and bacteria, and the fungal internal transcribed spacer-1 (ITS1) region. The fungal and bacterial community structure was assessed using Bray-Curtis dissimilarities, and the effect of age at sampling and location was determined using PERMANOVA. RESULTS: Age at sampling had a substantial effect on the foal's archaeal and bacterial faecal microbiota (PERMANOVA: R2 = 0.16; p < 0.01), while the effect of geographical location was smaller but still significant (PERMANOVA: R2 = 0.07; p < 0.01). The overall abundance, diversity and richness of bacterial and archaeal populations increased (p < 0.01) as foals aged, most noticeably rising between foals 1 to 2 and 2 to 3 months of age. The 15 most relatively abundant fungal species were all environmental saprophytes, most strongly affected by geographical location (p < 0.01) rather than age at sampling. There was an effect of location on Preussia Africana (p = 0.02) and a location × age interaction for fungal species Preussia persica (p < 0.01), Acremonium furcatum (p = 0.04), and Podospora pseudocomata (p = 0.01). There was no effect of age, location, or location × age interaction on the relative abundance of the remaining fungal species. CONCLUSIONS: The faecal microbiome appeared to stabilise for most bacterial and archaeal genera by 2 to 3 months of age, resembling an adult mare. Bacterial genera isolated from faecal samples belonged mainly to the Firmicutes phylum. Age at sampling more strongly affected the archaeal and bacterial faecal microbiota than the effect of the geographical location where the horse was sampled. The lack of effect of location on microbe populations suggests that although environmental factors may influence population structure, there are distinct differences at each stage of foal maturation.

Effect of ergot alkaloids and a mycotoxin deactivating product on in vitro ruminal fermentation using the Rumen simulation technique (RUSITEC).

The rumen simulation technique (RUSITEC) was used to investigate the effect of ergot alkaloids (EA) and a mycotoxin deactivating product (Biomin AA; MDP) on nutrient digestion, ruminal fermentation parameters, total gas, methane, and microbial nitrogen production. Ruminal fermentation vessels received a feedlot finishing diet of 90:10 concentrate:barley silage (DM basis). Using a randomized complete block design, treatments were assigned (n = 4 vessels/treatment) within two RUSITEC apparatuses in a 2 × 2 factorial arrangement. Treatments included: (1) control (CON) diet (no EA and no MDP); (2) CON diet + 1 g/d MDP; (3) CON diet + 20 mg/kg EA; and (4) CON diet + 20 mg/kg EA + 1 g/d MDP. The study was conducted over 14 d with 7 d of adaptation and 7 d of sample collection. Data were analyzed in SAS using PROC MIXED including fixed effects of EA, MDP, and the EA×MDP interaction. Random effects included RUSITEC apparatus and cow rumen inoculum (n = 4). Ergot alkaloids decreased dry matter (DMD) (P = 0.01; 87.9 vs. 87.2%) and organic matter disappearance (OMD) (P = 0.02; 88.8 vs. 88.4%). Inclusion of MDP increased OMD (P = 0.01; 88.3 vs. 88.9%). Neutral detergent fiber disappearance (NDFD) was improved with MDP; however, an EA×MDP interaction was observed with MDP increasing (P < 0.001) NDFD more with EA diet compared to CON. Acetate proportion decreased (P = 0.01) and isovalerate increased (P = 0.03) with EA. Consequently, acetate:propionate was reduced (P = 0.03) with EA. Inclusion of MDP increased total volatile fatty acid (VFA) production (P < 0.001), and proportions of acetate (P = 0.03) and propionate (P = 0.03), and decreased valerate (P < 0.001), isovalerate (P = 0.04), and caproate (P = 0.002). Treatments did not affect (P ≥ 0.17) ammonia, total gas, or methane production (mg/d or mg/g of organic matter fermented). The inclusion of MDP reduced (P < 0.001) microbial nitrogen (MN) production in the effluent and increased (P = 0.01) feed particle-bound MN. Consequently, total MN decreased (P = 0.001) with MDP. In all treatments, the dominant microbial phyla were Firmicutes, Bacteroidota, and Proteobacteria, and the major microbial genus was Prevotella. Inclusion of MDP further increased the abundance of Bacteroidota (P = 0.04) as it increased both Prevotella (P = 0.04) and Prevotellaceae_UCG-003 (P = 0.001). In conclusion, EA reduced OMD and acetate production due to impaired rumen function, these responses were successfully reversed by the addition of MDP.

Ergot formed from a parasitic fungus (Claviceps purpurea) affects various types of grains (rye, wheat, or oats) and may contain several toxic ergot alkaloids (EA). Individual EA may impact the rumen microorganisms, and cattle feed intake, digestibility, health, and overall performance. A common method to alleviate toxicity in mycotoxin-contaminated feed is through the addition of mycotoxin binders (MDP); however, their efficacy against EA is unknown. To better understand the effect of EA in cattle, we performed an in vitro experiment to examine the impact of EA on the ruminal microbial populations and fermentation of a finishing feedlot diet using an artificial rumen (RUSITEC). Additionally, an MDP was added to test if it could reduce the detrimental effects of EA on rumen fermentation. MDP increased total volatile fatty acids (VFA) and reduced total microbial protein synthesis. Furthermore, EA reduced microbial diversity and the acetate:propionate ratio. Although EA reduced organic matter digestibility and acetate production, these negative effects were reversed by the addition of the MDP.

Unsalable Vegetables Ensiled With Sorghum Promote Heterofermentative Lactic Acid Bacteria and Improve in vitro Rumen Fermentation.

This study characterized the nutritive and microbial profiles and the fermentation characteristics of silage with the following compositions on a dry matter (DM) basis: (1) 100% sorghum, (2) 70% sorghum + 30% carrot or pumpkin, and (3) 40% sorghum + 60% carrot or pumpkin. The treatments were further divided based on the addition or no addition of a probiotic inoculant. After 70 days of ensiling, the silage was incubated for 48 h using the in vitro batch culture technique. Crude protein and non-fiber carbohydrates in the silage increased (P ≤ 0.01) by 5.7 percent point (pp) and 9.6 pp, respectively, with pumpkin at 60% DM. The V4 region of the 16S rRNA gene was sequenced to profile pre-ensiled and ensiled archeal and bacterial communities. Silages containing carrot or pumpkin strongly influenced the microbial structure (PERMANOVA: R 2 = 0.75; P < 0.001), despite the ensiled treatments being dominated by Lactobacillus spp., except for the control, which was dominated by Weissella and Pediococcus spp. (P < 0.01). Linear discriminant analysis indicated that carrot and pumpkin silages were responsible for the increased relative abundance of Lactobacillus and Acinetobacter spp. (log LDA score ≥ 2), respectively. After 48 h of incubation, carrot and pumpkin inclusion increased (P < 0.01) the in vitro DM digestibility by 22.5 and 31.3%, increased the total volatile fatty acids (VFAs) by 16 and 20.6% (P < 0.01), respectively, and showed a tendency (P = 0.07) to increase the gas production. Therefore, this study supports the use of carrot or pumpkin in sorghum silages to maximize feed digestibility and total VFA concentrations.

Ensiling sorghum with unsalable pumpkin improves feed digestibility with minimal influence on the rumen microbial population using the rumen simulation technique.

This study aimed to determine the influence of sorghum ensiled with unsalable pumpkin at 20 or 40% dry matter (DM) basis on rumen fermentation characteristics and rumen microbial communities using the rumen simulation technique (RUSITEC). The experiment used a completely randomised design including silages comprising (1) 100% sorghum; (2) 80% sorghum + 20% DM pumpkin; or (3) 60% sorghum + 40% DM pumpkin. Each RUSITEC run (n = 2) was 15 d long, including 6 d of adaptation and 9 d of sampling. Dry matter digestibility (DMD) was measured on d 8 and 10-13. Gas production was measured daily, whereas methane and volatile fatty acids (VFA) production were measured from d 7-15. Solid-associated microbes (SAM) were collected on d 5, 10 and 15, whereas liquid-associated microbes (LAM) were collected after 15-d incubation. The V4 region of the 16S rRNA gene and the ITS1 region were sequenced to identify archaeal, bacterial and fungal communities. Ensiling 40% DM pumpkin with sorghum increased DMD and decreased the ratio of acetate to propionate (P ≤ 0.01). Both bacterial SAM and LAM communities were dominated by Megasphaera, and had the highest relative abundance (P = 0.03) with 40% DM pumpkin after 5 d incubation in the SAM community, while species of the Aspergillus genus dominated fungal SAM and LAM communities with 20 or 40% DM unsalable pumpkin. Therefore, ensiling up to 40% DM unsalable pumpkin with sorghum produces a high-quality ruminant feed with minimal influence on the rumen microbial population. KEY POINTS: • Including 40% DM unsalable pumpkin decreased acetate:propionate • Ensiling unsalable pumpkin with sorghum increases digestibility in a RUSITEC • Rumen microbial communities were slightly influenced by unsalable pumpkin inclusion.

The use of live yeast to increase intake and performance of cattle receiving low-quality tropical forages.

The objective was to evaluate the effects of a specific strain of live yeast (LY) on growth performance, fermentation parameters, feed efficiency, and bacterial communities in the rumen of growing cattle fed low-quality hay. In experiment (exp.) 1, 12 Droughtmaster bull calves (270 ± 7.6 kg initial body weight [BW]) were blocked by BW into two groups, allocated individually in pens, and fed ad libitum Rhodes grass hay (8.4% of crude protein [CP]) and 300 g/bull of supplement (52% CP) without (Control) or with LY (8 × 109 colony-forming unit [CFU]/d Saccharomyces cerevisiae CNCM I-1077; Lallemand Inc., Montreal, Canada) for 28 d, followed by 7 d in metabolism crates. Blood and rumen fluid were collected before feeding and 4 h after feeding. In exp. 2, for assessment of growth performance, 48 Charbray steers (329 ± 20.2 kg initial BW) were separated into two blocks by initial BW and randomly allocated into 12 pens. The steers were fed Rhodes grass hay (7.3% CP) and 220 g/steer of supplement (60% CP) without or with LY (8 × 109 CFU/d) for 42 d, after a 2-wk adaptation period. In exp. 1, fiber digestibility was calculated from total fecal collection, and, in exp 2, indigestible neutral detergent fiber (NDF) was used as a marker. Inclusion of LY increased (P = 0.03) NDF intake by 8.3% in exp. 1, without affecting total tract digestibility. No changes were observed in microbial yield or in the efficiency of microbial production. There was a Treatment × Time interaction (P < 0.01) for the molar proportion of short-chain fatty acids, with LY increasing propionate before feeding. Inclusion of LY decreased rumen ammonia 4 h after feeding (P = 0.03). The addition of LY reduced rumen bacterial diversity and the intraday variation in bacterial populations. Relative populations of Firmicutes and Verrucomicrobia varied over time (P < 0.05) only within the Control group. At the genus level, the relative abundance of an unclassified bacterial genus within the order Clostridiales, a group of cellulolytic bacteria, was reduced from 0 to 4 h after feeding in the Control group (P = 0.02) but not in the LY group (P = 1.00). During exp. 2, LY tended to increase average daily gain (ADG) (P = 0.08) and feed efficiency (P = 0.10), with no effect on NDF intake or digestibility. In conclusion, S. cerevisiae CNCM I-1077 reduced the intraday variation of rumen bacteria and increased the amount of NDF digested per day. These observations could be associated with the tendency of increased ADG and feed efficiency in growing cattle fed a low-quality forage.

Meat quality characteristics of lot-fed Australian Rangeland goats are unaffected by live weight at slaughter.

The effects of live weight on carcass characteristics and meat quality of Australian Rangeland goats were determined. Fifty-two intact-male kid goats were fed Mitchell grass hay and finisher pellets ad libitum for 42 days. Prior to slaughter, kids were categorised into live weight groups: 'Heavy' (≈33.1 kg) or 'Light' ≈ 24.3 kg). Fifteen kids per group were randomly selected, slaughtered and carcass characteristics measured. The longissimus lumborum (LL) and the biceps femoris (BF) muscles were removed for quality measurements. The Heavy group had higher dressing, hindquarters, non-carcass component and offal percentages (P < 0.05). Live weight had no effect on proportion of carcass cuts, muscle meat colour, cooking loss, Warner-Bratzler shear force, total fat or ash (P > 0.05), but influenced the moisture and protein content in the LL and the moisture content in the BF (P < 0.05). The findings suggest that live weight of entire male Rangeland goats had no effect on meat quality characteristics.

Feeding unsaleable carrots to lambs increased performance and carcass characteristics while maintaining meat quality.

This study investigated the effect of feeding unsaleable carrots to lambs within a total-mixed ration (TMR) on performance, carcass characteristics, meat quality and sensory parameters. Thirty-six Australian Merino wether lambs were fed a control (barley-based) or carrot-based TMR for 11-weeks. Carrot-fed lambs had 2.7% higher cold dressing percentage (P = 0.03) while consuming less than control lambs. Subcutaneous fat of carrot-fed lambs contained less branch-chained, and more cis- and trans-monounsaturated fatty acids (FA; P ≤ 0.01) compared to control-fed lambs, which tended (P = 0.08) to have higher concentrations of polyunsaturated FA, despite the Longissimus lumborum (LL) muscle being unchanged by diet. Under retail display conditions, L* and hue values were lower (P ≤ 0.04) for 5 d aged LL samples from carrot-fed lambs. No differences were observed in other meat quality and sensory parameters between diets. Therefore, feeding unsaleable carrots at 45% DM in a TMR can improve lamb performance and carcass characteristics, while maintaining meat quality and FA composition.

Influence of probiotics on biofilm formation and diversity of bacteria colonising crop sorghum ensiled with unsalable vegetables.

The objective of this study was to characterise in situ digestion kinetics and bacterial colonisation of crop sorghum ensiled with unsalable carrot or pumpkin at 0, 20 or 40% dry matter (DM). Silages with or without the application of a commercial probiotic were incubated in situ for 0, 3, 6, 9, 24 and 48 h. Calculation of in situ digestion kinetics was conducted for DM, organic matter and neutral detergent fibre (aNDF). The V4 region of the 16S rRNA gene was sequenced to determine the composition and diversity of bacteria colonising the silage. Organic matter and DM digestion kinetics indicated that greater vegetable inclusion increased (P < 0.05) the soluble fraction and effective degradability. Bacterial richness at 48 h incubation was greater (P = 0.02) in 20% carrot and 40% pumpkin treatments, compared with the control. An effect of level × probiotic was observed with increased Shannon diversity (P = 0.01) for 40% carrot and 20% pumpkin probiotic treatments, respectively. Primary colonising bacteria were members of the Prevotella genus, dominating after 3 and 6 h of incubation. The abundance of Prevotella increased by 4.1% at 3 h (P < 0.01) and by 4.7% at 9 h incubation with probiotics, compared with the control. Secondary biofilm colonisers included members of Treponema, Saccharofermentans, Fibrobacter, Ruminobacter and Anaerosporobacter genera, dominant from 9 h incubation onward. This study demonstrated that including unsalable vegetables at 20 or 40% DM increases the soluble fraction and effective degradability of sorghum silage during in situ digestion and increases diversity of bacteria colonising ensiled vegetables within the rumen. KEY POINTS: • Ensiling unsalable vegetables is a viable strategy to reduce food waste. • Ensiled vegetables increased in situ soluble fraction and effective degradability. • Bacterial richness at 48 h incubation improved with 20% carrot or 40% pumpkin. • Diversity of colonising rumen bacteria increased with carrot or pumpkin inclusion.

Saccharomyces cerevisiae fermentation products (SCFP) stabilize the ruminal microbiota of lactating dairy cows during periods of a depressed rumen pH.

BACKGROUND: Effects of Saccharomyces cerevisiae fermentation products (SCFP) on rumen microbiota were determined in vitro and in vivo under a high and a depressed pH. The in vitro trial determined the effects of Original XPC and NutriTek (Diamond V, Cedar Rapids, IA) at doses of 1.67 and 2.33 g/L, respectively, on the abundances of rumen bacteria under a high pH (> 6.3) and a depressed pH (5.8-6.0) using quantitative PCR (qPCR). In the in vivo trial eight rumen-cannulated lactating dairy cows were used in a cross-over design. Cows were randomly assigned to SCFP treatments (Original XPC, Diamond V, Cedar Rapids, IA) or control (No SCFP) before two 5-week experimental periods. During the second period, SCFP treatments were reversed. Cows on the SCFP treatment were supplemented with 14 g/d of SCFP and 126 g/d of ground corn. Other cows received 140 g/d ground corn. During the first 4 wk. of each period, cows received a basal diet containing 153 g/kg of starch. During week 5 of both periods, the rumen pH was depressed by a SARA challenge. This included replacing 208 g/kg of the basal diet with pellets of ground wheat and barley, resulting in a diet that contained 222 g/kg DM of starch. Microbial communities in rumen liquid digesta were examined by pyrosequencing, qPCR, and shotgun metagenomics. RESULTS: During the in vitro experiment, XPC and NutriTek increased the relative abundances of Ruminococcus flavefaciens, and Fibrobacter succinogenes determined at both the high and the depressed pH, with NutriTek having the largest effect. The relative abundances of Prevotella brevis, R. flavefaciens, ciliate protozoa, and Bifidobacterium spp. were increased by XPC in vivo. Adverse impacts of the in vivo SARA challenge included reductions of the richness and diversity of the rumen microbial community, the abundances of Bacteroidetes and ciliate protozoa in the rumen as determined by pyrosequencing, and the predicted functionality of rumen microbiota as determined by shotgun metagenomics. These reductions were attenuated by XPC supplementation. CONCLUSIONS: The negative effects of grain-based SARA challenges on the composition and predicted functionality of rumen microbiota are attenuated by supplementation with SCFP.

Crop Sorghum Ensiled With Unsalable Vegetables Increases Silage Microbial Diversity.

Ensiling vegetables with forage crops is a suggested method of waste diversion and can be directly utilized as a livestock feed. Carrot or pumpkin, ensiled at 0, 20, or 40% dry matter (DM) with crop sorghum, and with or without a second-generation silage inoculant were assessed for nutritive composition, organic acid profiles, aerobic stability and in vitro rumen fermentation characteristics. The study was a completely randomized design, with the fixed effects consisting of vegetable type (carrot vs. pumpkin), level (i.e., the level of vegetables), inoculant (inoculant or non-inoculant) and the interactions, and mini-silos within treatment as the random effect. The experimental unit for sorghum treatments represented by each mini-silo (5 kg capacity). Silage was sampled after 70-days ensiling for nutrient composition, 14-day aerobic stability, organic acid profiles and microbial diversity. After 24 h in vitro incubation, rumen fermentation parameters were assessed, measuring gas and methane (CH4) production, in vitro digestibility and volatile fatty acid concentrations. Sorghum ensiled with carrot or pumpkin at 20% or 40% DM increased crude fat (P ≤ 0.01) and decreased (P ≤ 0.01) silage surface temperature upon aerobic exposure compared to the control. Bacterial communities analyzed through 16S rRNA gene sequencing linearly increased (P ≤ 0.01) in diversity across both vegetables when the vegetable proportion was increased in the silage; dominated by Lactobacillus species. ITS analysis of the fungal microbiota upon silage opening and after 14 days (aerobic stability) identified increased (P ≤ 0.03) fungal diversity with increasing vegetable proportions, predominantly populated by Fusarium denticulatum, Issatchenkia orientalis, Kazachstania humilis, and Monascus purpureus. Upon assessment in vitro, there was an increase (P ≤ 0.04) in in vitro digestibility and some CH4 parameters (% CH4, and mg CH4/g DM), with no effect (P ≥ 0.17) on remaining CH4 parameters (mL CH4/g DM, mg CH4/g digested DM), gas production or pH. However, increasing vegetable amount decreased percentage of acetic acid and increased percentage of propionic acid of the total VFA, decreasing A:P ratio and total VFA concentration as a result (P ≤ 0.01). The results from this study indicate including carrot or pumpkin at 20 or 40% DM in a sorghum silage can produce a highly digestible, microbially diverse and energy-rich livestock feed.

Microbial characterization and fermentative characteristics of crop maize ensiled with unsalable vegetables.

Incorporation of carrot or pumpkin at 0, 20 or 40% dry matter (DM-basis) with crop maize, with or without a silage inoculant was evaluated after 70 days ensiling for microbial community diversity, nutrient composition, and aerobic stability. Inclusion of carrots or pumpkin had a strong effect on the silage bacterial community structure but not the fungal community. Bacterial microbial richness was also reduced (P = 0.01) by increasing vegetable proportion. Inverse Simpson's diversity increased (P = 0.04) by 18.3% with carrot maize silage as opposed to pumpkin maize silage at 20 or 40% DM. After 70 d ensiling, silage bacterial microbiota was dominated by Lactobacillus spp. and the fungal microbiota by Candida tropicalis, Kazachstania humilis and Fusarium denticulatum. After 14 d aerobic exposure, fungal diversity was not influenced (P ≥ 0.13) by vegetable type or proportion of inclusion in the silage. Inoculation of vegetable silage lowered silage surface temperatures on day-7 (P = 0.03) and day-14 (P ≤ 0.01) of aerobic stability analysis. Our findings suggest that ensiling unsalable vegetables with crop maize can successfully replace forage at 20 or 40% DM to produce a high-quality livestock feed.

Effects of Hardwood Biochar on Methane Production, Fermentation Characteristics, and the Rumen Microbiota Using Rumen Simulation.

Biochar is a novel carbonized feed additive sourced from pyrolyzed biomass. This compound is known to adsorb gasses and carbon, participate in biological redox reactions and provide habitat biofilms for desirable microbiota proliferation. Therefore, biochar holds potential to modify rumen fermentation characteristics and reduce enteric CH4 emissions. The objective of this study was to investigate the effect of hardwood biochar supplementation on fermentation parameters, methane (CH4) production and the ruminal archaeal, bacterial, and fungal microbiota using the in vitro RUSITEC (rumen simulation technique) system. Treatments consisted of a control diet (oaten pasture: maize silage: concentrate, 35:35:30 w/w) and hardwood biochar included at 400 or 800 mg per day (3.6 and 7.2% of substrate DM, respectively), over a 15-day period. Biochar supplementation had no effect (P ≥ 0.37) on pH, effluent (mL/d), total gas (mL/d), dry matter (DM) digestibility or CH4 production (mg/d). The addition of 800 mg biochar per day had the tendency (P = 0.10) to lower the % of CH4 released in fermentation compared to 400 mg/d biochar treatment. However, no effect (P ≥ 0.44) was seen on total VFA, acetate, propionate, butyric, branched-chain VFA, valerate and caproate production and the ratio of acetate to propionate. No effect (P > 0.05) was observed on bacterial, archaeal or fungal community structure. However, biochar supplementation at 800 mg/d decreased the abundance of one Methanomethylophilaceae OTU (19.8-fold, P = 0.046) and one Lactobacillus spp. OTU (31.7-fold, P < 0.01), in comparison to control treatments. Two fungal OTUs classified as Vishniacozyma victoriae (5.4 × 107 increase) and Sporobolomyces ruberrimus (5.4 × 107-fold increase) were more abundant in the 800 mg/d biochar samples. In conclusion, hardwood biochar had no effects on ruminal fermentation characteristics and may potentially lower the concentration of enteric CH4 when included at higher dosages by manipulating ruminal microbiota abundances.

Fat accretion measurements strengthen the relationship between feed conversion efficiency and Nitrogen isotopic discrimination while rumen microbial genes contribute little.

The use of biomarkers for feed conversion efficiency (FCE), such as Nitrogen isotopic discrimination (Δ15N), facilitates easier measurement and may be useful in breeding strategies. However, we need to better understand the relationship between FCE and Δ15N, particularly the effects of differences in the composition of liveweight gain and rumen N metabolism. Alongside measurements of FCE and Δ15N, we estimated changes in body composition and used dietary treatments with and without nitrates, and rumen metagenomics to explore these effects. Nitrate fed steers had reduced FCE and higher Δ15N in plasma compared to steers offered non-nitrate containing diets. The negative relationship between FCE and Δ15N was strengthened with the inclusion of fat depth change at the 3rd lumbar vertebrae, but not with average daily gain. We identified 1,700 microbial genes with a relative abundance >0.01% of which, 26 were associated with Δ15N. These genes explained 69% of variation in Δ15N and showed clustering in two distinct functional networks. However, there was no clear relationship between their relative abundances and Δ15N, suggesting that rumen microbial genes contribute little to Δ15N. Conversely, we show that changes in the composition of gain (fat accretion) provide additional strength to the relationship between FCE and Δ15N.

Exploration of Biological Markers of Feed Efficiency in Young Bulls.

The efficiency with which ruminants convert feed to desirable products is difficult to measure under normal commercial settings. We explored the use of potential biological markers from easily obtainable samples, that is, blood, hair, and feces, to characterize potential causes of divergent efficiency when considered as residual feed intake (RFI) or feed conversion efficiency (FCE). A total of 54 Charolais bulls, 20 in period 1 and 34 in period 2, were examined for individual dry matter intake (DMI) and growth. Bulls were offered a diet of 70:30 wrapped grass silage to concentrate for 99 d. At the conclusion of the test period, blood samples were collected for the determination of vitamins B2 and B6, and plasma used for the determination of metabolites, natural isotopic 15N abundance (15N NIA, expressed as δ15N ‰) and fractionation (Δ15Nplasma proteins-diet and Δ13Cplasma proteins-diet) and near-infrared spectroscopy (NIRS). Feces were analyzed by NIRS. Bulls were slaughtered at 15-17 months of age and carcass characteristics determined. Bulls were ranked according to RFI with extremes (SD ± 0.5; n = 31) classified as either efficient (Neg-RFI) or inefficient (Pos-RFI). Extreme bulls were then classified for FCE (high vs low FCE), changing the groups. Pos-RFI bulls consumed 14% more feed than Neg-RFI bulls for the same level of weight gain. Low FCE bulls tended to eat more, but had lower weight gains than high FCE bulls. No differences were detected in carcass conformation, fat scores, hot carcass weight, or dressing percentage. Yet, heart and bladder weights were heavier in Pos-RFI, and rumen weight tended to be heavier in Pos-RFI bulls. RFI did not affect bulk 15N or 13C fractionation. A negative correlation was observed between FCE and Δ15Nplasma proteins-diet. Inefficient bulls (Pos-RFI) had higher δ15N in glycine compared to Neg-RFI bulls. Similarly, metabolomic analysis showed a tendency for concentrations of glycine and sarcosine to be elevated in Pos-RFI bulls, whereas aspartic acid and carnosine tended to be elevated, and serine tended to be lower in High FCE. Among vitamins, only flavin adenine dinucleotide concentration was higher in the blood of bulls with High FCE. These results suggest that the two feed efficiency metrics differ in the underlying mechanisms of metabolism, where RFI is driven by differences in the energetic requirements of visceral organs and the extent of AA catabolism.

The Structural and Functional Capacity of Ruminal and Cecal Microbiota in Growing Cattle Was Unaffected by Dietary Supplementation of Linseed Oil and Nitrate.

Microorganisms in the digestive tract of ruminants differ in their functionality and ability to use feed constituents. While cecal microbiota play an important role in post-rumen fermentation of residual substrates undigested in the rumen, limited knowledge exists regarding its structure and function. In this trial we investigated the effect of dietary supplementation with linseed oil and nitrate on methane emissions and on the structure of ruminal and cecal microbiota of growing bulls. Animals were allocated to either a CTL (control) or LINNIT (CTL supplemented with 1.9% linseed and 1.0% nitrates) diet. Methane emissions were measured using the GreenFeed system. Microbial diversity was assessed using amplicon sequencing of microbial genomic DNA. Additionally, total RNA was extracted from ruminal contents and functional mcrA and mtt genes were targeted in amplicon sequencing approach to explore the diversity of functional gene expression in methanogens. LINNIT had no effect on methane yield (g/kg DMI) even though it decreased methane production by 9% (g/day; P < 0.05). Methanobrevibacter- and Methanomassiliicoccaceae-related OTUs were more abundant in cecum (72 and 24%) compared to rumen (60 and 11%) irrespective of the diet (P < 0.05). Feeding LINNIT reduced the relative abundance of Methanomassiliicoccaceae mcrA cDNA reads in the rumen. Principal component analysis revealed significant differences in taxonomic composition and abundance of bacterial communities between rumen and cecum. Treatment decreased the relative abundance of a few Ruminococcaceae genera, without affecting global bacterial community structure. Our research confirms a high level of heterogeneity in species composition of microbial consortia in the main gastrointestinal compartments where feed is fermented in ruminants. There was a parallel between the lack of effect of LINNIT on ruminal and cecal microbial community structure and functions on one side and methane emission changes on the other. These results suggest that the sequencing strategy used here to study microbial diversity and function accurately reflected the absence of effect on methane phenotypes in bulls treated with linseed plus nitrate.

From pre- to postweaning: Transformation of the young calf's gastrointestinal tract.

The ruminant gastrointestinal tract (GIT) faces the challenge of protecting the host from luminal contents and pathogens, while supporting the absorption and metabolism of nutrients for growth and maintenance. The GIT of the calf in early life undergoes some of the most rapid microbial and structural changes documented in nature, and these adaptations in GIT function make the young calf susceptible to GIT diseases and disorders. Despite these challenges, the calf's GIT has a certain degree of plasticity and can sense nutrient supply and respond to bioactive ingredients. Calf GIT research has historically focused on the transition during weaning and characterizing ruminal papillae development using microscopy and digesta metabolite responses. Through the use of new molecular-based approaches, we have recently shown that delaying the age of weaning and providing a step-down weaning protocol is associated with a more gradual shift in ruminal microbiota to a postweaned state. In addition to ruminal adaptations during weaning, nutrient flow to the lower gut changes dramatically during weaning, coinciding with a wide array of structural and microbiological changes. Structural and gene expression changes suggest that the lower gut of the dairy calf undergoes alterations that may reduce barrier function when solid feeds are consumed. More recently, in vivo data revealed that the weaning transition increases total gut permeability of the calf. Interestingly, the lower gut may be able to communicate with the forestomach, meaning that a nutrient can be sensed in the lower gut and cause subsequent adaptations in the forestomach. An improved understanding of how diet, microbiota, and functional ingredients interact to affect growth and barrier function of the intestinal tract would greatly benefit the dairy calf industry. A mechanistic understanding of such adaptations would also aid in the formulation of specific management regimens and provision of functional ingredients required to characterize and enhance gut function in young calves.