L. Jin

Expression of a fungal ferulic acid esterase in alfalfa modifies cell wall digestibility.

BackgroundAlfalfa (Medicago sativa) is an important forage crop in North America owing to its high biomass production, perennial nature and ability to fix nitrogen. Feruloyl esterase (EC 3.1.1.73) hydrolyzes ester linkages in plant cell walls and has the potential to further improve alfalfa as biomass for biofuel production.ResultsIn this study, faeB [GenBank:AJ309807] was synthesized at GenScript and sub-cloned into a novel pEACH vector containing different signaling peptides to target type B ferulic acid esterase (FAEB) proteins to the apoplast, chloroplast, endoplasmic reticulum and vacuole. Four constructs harboring faeB were transiently expressed in Nicotiana leaves, with FAEB accumulating at high levels in all target sites, except chloroplast. Stable transformed lines of alfalfa were subsequently obtained using Agrobacterium tumefaciens (LBA4404). Out of 136 transgenic plants regenerated, 18 independent lines exhibited FAEB activity. Subsequent in vitro digestibility and Fourier transformed infrared spectroscopy (FTIR) analysis of FAEB-expressing lines showed that they possessed modified cell wall morphology and composition with a reduction in ester linkages and elevated lignin content. Consequently, they were more recalcitrant to digestion by mixed ruminal microorganisms. Interestingly, delignification by alkaline peroxide treatment followed by exposure to a commercial cellulase mixture resulted in higher glucose release from transgenic lines as compared to the control line.ConclusionModifying cell wall crosslinking has the potential to lower recalcitrance of holocellulose, but also exhibited unintended consequences on alfalfa cell wall digestibility due to elevated lignin content. The combination of efficient delignification treatment (alkaline peroxide) and transgenic esterase activity complement each other towards efficient and effective digestion of transgenic lines.

Anti-Escherichia coli O157:H7 properties of purple prairie clover and sainfoin condensed tannins.

Condensed tannins (CT) from purple prairie clover (PPC; Dalea purpurea Vent.) and sainfoin (SF; Onobrychis viciifolia) were assessed for anti-Escherichia coli activity by comparing their ability to react with proteins and liposome, cause cell aggregation, and alter outer membrane (OM) morphology and permeability. The PPC CT had greater (P < 0.01) protein-precipitating capacity than SF CT using either bovine serum albumin or ribulose 1,5-disphosphate carboxylase as model proteins. Minimum inhibitory concentration of PPC CT for two strains of E. coli and five strains of E. coli O157:H7 was four to six times lower than that of SF CT. E. coli exposed to 10 µg/mL of both CT had higher (P < 0.05) OM permeability than controls and was greater (P < 0.05) for PPC than for SF CT. Addition of both CT at 50 and 200 µg/mL caused cell aggregation which was more evident (P < 0.05) for PPC than for SF CT. Transmission electron microscopy showed electron dense material on the cell surface when cells were exposed to 50 µg/mL of PPC CT. The greater anti-E. coli activity of PPC than SF CT was due to its enhanced ability to precipitate protein that increased OM permeability and promoted cell aggregation.

Effect of grain type and processing index on growth performance, carcass quality, feeding behavior, and stress response of feedlot steers.

One hundred sixty crossbred steers (538 ± 36 kg BW) were used in an 84-d experiment with a randomized block design to study the effects of wheat or barley grain processed to 2 different indices on growth performance, feeding behavior, carcass characteristics, stress, and temperament of finishing beef cattle. Treatments were a wheat-based diet (88.4% of diet DM; WH) and a barley-based diet (89% of diet DM; BA), processed to an index of either 75% (HI) or 85% (LO) of their original volume weight. Cattle were allocated to 16 feedlot pens (10 animals per pen, 4 pens per treatment), 8 of which were equipped with the GrowSafe system for monitoring feeding behavior. Flight speed, hair, and saliva samples were collected on d 1, 28, 56, and 84 to determine temperament, acute, and chronic stress. All steers were slaughtered at the end of the experiment, and carcass quality was evaluated. Cattle fed WH had a lower (P < 0.05) meal length and frequency of visits per meal and tended (P = 0.10) to have a lower DMI, meal size, and feeding time than those fed BA. The LO processing index increased (P = 0.05) DMI and reduced (P < 0.05) the G:F and the percentage of saleable meat of the carcass compared to HI. There was a trend (P = 0.09) for a grain × processing index interaction, where cattle fed BA-LO had a lower incidence of severe liver abscesses compared with cattle fed other treatments. Cattle fed WH had greater hair cortisol concentrations (P = 0.01) and flight speed (P < 0.01) than those fed BA. There was a trend (P = 0.07) for a grain × processing index interaction, where heifers fed WH-LO had a lower salivary cortisol than those fed other treatments. Results suggest that a LO processing index had a negative effect on feed efficiency and carcass performance and that the WH diet caused a range of effects on feed intake and behavior indicative of steers with greater excitability and chronic stress.

Impact of adding Saccharomyces strains on fermentation, aerobic stability, nutritive value, and select lactobacilli populations in corn silage.

Bacterial inoculants can improve the conservation and nutritional quality of silages. Inclusion of the yeast Saccharomyces in the diet of dairy cattle has also been reported to be beneficial. The present study assessed the ability of silage to be used as a means of delivering Saccharomyces strains to ruminants. Two strains of Saccharomyces cerevisiae (strain 1 and 3)and 1 strain of Saccharomyces paradoxus (strain 2) were inoculated (10(3) cfu/g) individually onto corn forage that was ensiled in mini silos for 90 d. Fermentation characteristics, aerobic stability, and nutritive value of silages were determined and real-time quantitative PCR (RT-qPCR) was used to quantify S. cerevisiae, S.paradoxus, total Saccharomyces, fungal, and bacterial populations. Fermentation characteristics of silage inoculated with S1 were similar to control silage. Although strain 3 inoculation increased ash and decreased OM contents of silage (P = 0.017), no differences were observed in nutrient composition or fermentation profiles after 90 d of ensiling. Inoculation with Saccharomyces had no detrimental effect on the aerobic stability of silage. In vitro DM disappearance, gas production, and microbial protein synthesis were not affected by yeast inoculation.Saccharomyces strain 1 was quantified throughout ensiling, whereas strain 2 was detected only immediately after inoculation. Saccharomyces cerevisiae strain 3 was quantified until d 7 and detectable 90 d after ensiling. All inoculants were detected and quantified during aerobic exposure. Inoculation with Saccharomyces did not alter lactobacilli populations. Saccharomycetales were detected by RT-qPCR throughout ensiling in all silages. Both S. cerevisiae and S. paradoxus populations increased during aerobic exposure, demonstrating that the density of these yeast strains would increase between the time that silage was removed from storage and the time it was fed.

Effects of essential oils from medicinal plants acclimated to Benin on in vitro ruminal fermentation of Andropogon gayanus grass.

BACKGROUND Plants from West Africa commonly used in both human and veterinary medicine contain various secondary metabolites. However, their potential in mitigating ruminal methane production has not been explored. This study examined the effects of seven essential oils (EOs) from plants acclimated to Benin at four dosages (100, 200, 300 and 400 mg L(-1)), on in vitro rumen microbial fermentation and methane production using Andropogon gayanus grass as a substrate. RESULTS Compared to control, Laurus nobilis (300-400 mg L(-1) ), Citrus aurantifolia (300-400 mg L(-1)) and Ocimum gratissimum (200-400 mg L(-1)) decreased (P < 0.05) methane production (mL g(-1) DM) by 8.1-11.8%, 11.9-17.8% and 7.9-30.6%, respectively. Relative to the control, reductions in methane (mL g(-1) DM) of 11.4%, 13.5% and 14.2% were only observed at 400 mg L(-1) for Eucalyptus citriodora, Ocimum basilicum and Cymbopogon citratus, respectively. These EOs lowered methane without reducing concentrations of total volatile fatty acids or causing a shift from acetate to propionate production. All EOs (except M. piperita) reduced (P < 0.05) apparent dry matter (DM) disappearance of A. gayanus. CONCLUSIONS The current study demonstrated that EOs from plants grown in Benin inhibited in vitro methane production mainly through a reduction in apparent DM digestibility.

Condensed tannins affect bacterial and fungal microbiomes and mycotoxin production during ensiling and upon aerobic exposure

ABSTRACT Purple prairie clover (PPC; Dalea purpurea Vent.) containing 84.5 g/kg dry matter (DM) of condensed tannin (CT) was ensiled without (control) or with polyethylene glycol (PEG) for 76 days, followed by 14 days of aerobic exposure. Changes in fermentation characteristics were determined, and the composition of bacterial and fungal communities were assessed using metagenomic sequencing. The addition of polyethylene glycol (PEG) that deactivated CT at ensiling increased (P < 0.05 to ∼0.001) soluble N, nonprotein N, lactic acid, total volatile fatty acids, ammonia N, deoxynivalenol (DON), and ochratoxin A (OTA) but decreased (P < 0.001) pH and water-soluble carbohydrates. The concentrations of DON and OTA increased (P < 0.001) for both silages, with the extent of increase being greater for control than for PEG-treated silage during aerobic exposure. The PEG-treated silage exhibited higher (P < 0.01 to ∼0.001) copy numbers of total bacteria, Lactobacillus, yeasts, and fungi than the control. The addition of PEG decreased (P < 0.01) bacterial diversity during both ensiling and aerobic exposure, whereas it increased (P < 0.05) fungal diversity during aerobic exposure. The addition of PEG at ensiling increased (P < 0.05) the abundances of Lactobacillus and Pediococcus species but decreased (P < 0.01) the abundances of Lactococcus and Leuconostoc species. Filamentous fungi were found in the microbiome at ensiling and after aerobic exposure, whereas Bacillus spp. were the dominate bacteria after aerobic exposure. In conclusion, CT decreased protein degradation and improved the aerobic stability of silage. These desirable outcomes likely reflect the ability of PPC CT to inhibit those microorganisms involved in lowering silage quality and in the production of mycotoxins. IMPORTANCE The present study reports the effects of condensed tannins on the complex microbial communities involved in ensiling and aerobic exposure of purple prairie clover. This study documents the ability of condensed tannins to lower mycotoxin production and the associated microbiome. Taxonomic bacterial community profiles were dominated by Lactobacillales after fermentation, with a notable increase in Bacillus spp. as a result of aerobic exposure. It is interesting to observe that condensed tannins decreased bacterial diversity during both ensiling and aerobic exposure but increased fungal diversity during aerobic exposure only. The present study indicates that the effects of condensed tannins on microbial communities lead to reduced lactic acid and total volatile fatty acid production, proteolysis, and mycotoxin concentration in the terminal silage and improved aerobic stability. Condensed tannins could be used as an additive to control unfavorable microbial development and maybe enhanced feed safety.

Effects of essential oils from African basil on fermentation of Andropogon gayanus grass in the Artificial Rumen (RUSITEC)

Kouazounde, J. B., Gbenou, J. D., He, M., Jardim, T., Jin, L., Wang, Y., Beauchemin, K. A. and McAllister, T. A. 2015. Effects of essential oils from African basil on fermentation of Andropogon gayanus grass in the Artificial Rumen (RUSITEC). Can. J. Anim. Sci. 95: 425-431. Essential oils (EO) from African basil (Ocimum gratissimum) have shown the potential to modify rumen microbial fermentation and reduce ruminal methane production from grass forages in in vitro batch cultures. However, it is not known whether the effects of EO on rumen microbial fermentation attenuate over time. The objective of this study was to examine the effects of African basil EO at 0 (control), 100, 200 and 400 mg L-1 incubation medium on microbial fermentation and methane production in the Rumen Simulation Technique (RUSITEC) using Andropogon gayanus grass as a substrate. African basil EO quadratically affected (P<0.05) methane production gas production and the pH of fermenter liquid. Total volatile fatty acid (VFA) production was linearly decreased (P<0.05) by African basil EO along with a shift in VFA profile towards less propionate and more acetate and butyrate. African basil EO quadratically altered (P<0.05) apparent dry matter, neutral detergent fiber digestibility, 15N incorporation into total microbial protein and the total production of microbial protein. This study confirms that EO from African basil quadratically affected methane emissions arising from the ruminal fermentation of A. gayanus grass mainly by reducing overall digestibility of the forage.