Michael D. Flythe

Effect of Dietary Starch Source and Concentration on Equine Fecal Microbiota

Starch from corn is less susceptible to equine small intestinal digestion than starch from oats, and starch that reaches the hindgut can be utilized by the microbiota. The objective of the current study was to examine the effects of starch source on equine fecal microbiota. Thirty horses were assigned to treatments: control (hay only), HC (high corn), HO (high oats), LC (low corn), LO (low oats), and LW (low pelleted wheat middlings). Horses received an all-forage diet (2 wk; d -14 to d -1) before the treatment diets (2 wk; d 1 to 14). Starch was introduced gradually so that horses received 50% of the assigned starch amount (high = 2 g starch/kg BW; low = 1 g starch/kg BW) by d 4 and 100% by d 11. Fecal samples were obtained at the end of the forage-only period (S0; d -2), and on d 6 (S1) and d 13 (S2) of the treatment period. Cellulolytics, lactobacilli, Group D Gram-positive cocci (GPC), lactate-utilizers and amylolytics were enumerated. Enumeration data were log transformed and analyzed by repeated measures ANOVA. There were sample day × treatment interactions (P < 0.0001) for all bacteria enumerated. Enumerations from control horses did not change during the sampling period (P > 0.05). All treatments except LO resulted in increased amylolytics and decreased cellulolytics, but the changes were larger in horses fed corn and wheat middlings (P < 0.05). Feeding oats resulted in increased lactobacilli and decreased GPC (P < 0.05), while corn had the opposite effects. LW had increased lactobacilli and GPC (P < 0.05). The predominant amylolytic isolates from HC, LC and LW on S2 were identified by 16S RNA gene sequencing as Enterococcus faecalis, but other species were found in oat fed horses. These results demonstrate that starch source can have a differential effect on the equine fecal microbiota.

Interaction of Isoflavones and Endophyte-Infected Tall Fescue Seed Extract on Vasoactivity of Bovine Mesenteric Vasculature

It was hypothesized that isoflavones may attenuate ergot alkaloid-induced vasoconstriction and possibly alleviate diminished contractility of vasculature after exposure to ergot alkaloids. The objective of this study was to determine if prior incubation of bovine mesenteric vasculature with the isoflavones formononetin (F), biochanin A (B), or an ergovaline-containing tall fescue seed extract (EXT) and their combinations affect ergotamine (ERT)-induced contractility. Multiple segments of mesenteric artery and vein supporting the ileal flange of the small intestine were collected from Angus heifers at slaughter (n = 5, bodyweight = 639 ± 39 kg). Duplicates of each vessel type were incubated in tissue culture flasks at 37°C with a 50-mL volume of Krebs–Henseleit buffer containing: only buffer (control); or 1 × 10−6 M EXT; F; or B; and combinations of 1 × 10−6 M EXT + F; 1 × 10−6 M EXT + B; 1 × 10−6 M F + B; or 1 × 10−6 M EXT + F + B. After incubation for 2 h, sections were mounted in a multimyograph chamber. The ERT dose responses were normalized to 0.12 M KCl. Pretreatment with F, B, and F + B without EXT resulted in similar contractile responses to ERT in mesenteric artery and all incubations containing EXT resulted in a complete loss of vasoactivity to ERT. In mesenteric artery pretreated with EXT, treatments that contained B had higher contractile responses (P < 0.05) at ERT concentrations of 1 × 10−7 and 5 × 10−7 M. Also, treatments containing B tended (P < 0.1) to have greater responses than treatments without B at ERT concentrations of 1 × 10−6, 5 × 10−6, and 5 × 10−5 M. In mesenteric vein pretreated with EXT, treatments containing F had greater contractile responses to ERT at 1 × 10−5, 5 × 10−5, and 1 × 10−4 M (P < 0.05). These data indicated that F and B at 1 × 10−6 M and their combination did not impact the overall contractile response to ERT in mesenteric vasculature. However, F and B may offset some of the vasoconstriction caused by prior exposure to ergot alkaloids.

Vasoconstrictive responses by the carotid and auricular arteries in goats to ergot alkaloid exposure

A fungal endophyte (Neotyphodium coenophialum) infects most plants of “Kentucky 31” tall fescue (Lolium arundinaceum) and produces ergot alkaloids that cause persistent constriction of the vascular system in grazing livestock. Consequently, animals undergoing this toxicosis cannot regulate core body temperature and are vulnerable to heat and cold stresses. An experiment was conducted to determine if the caudal and auricular arteries in goats (Capra aegagrus hircus) vasoconstrict in response to ergot alkaloids. Seven, rumen fistulated goats were fed ad libitum orchardgrass (Dactylis glomeratia) hay and ruminally infused with endophtye-free seed (E−) for a 7-day adjustment period. Two periods followed with E− and endophyte-infected (E+) seed being randomly assigned to the 2 goat groups in period 1 and then switching treatments between groups in period 2. Infused E+ and E− seed were in equal proportions to the hay such that concentrations of ergovaline and ergovalanine were 0.80 μg per g dry matter for the E+ treatment. Cross-sections of both arteries were imaged using Doppler ultrasonography on days 0, 2, 4, 6, 8, and 12 in period 1 and on days 0, 1, 2, 3, 6, 7, and 9 in period 2. Differences from average baseline areas were used to determine presence or absence of alkaloid-induced vasoconstriction. Carotid arteries initiated constriction on imaging day 2 in both periods, and auricular arteries initiated constriction on imaging day 2 in period 1 and on day 6 in period 2. Luminal areas of the carotid arteries in E+ goats were 46% less than baseline areas in both periods after vasoconstriction occurred, whereas auricular arteries in E+ goats were 52% less than baseline areas in period 1 and 38% in period 2. Both arteries in E+ goats in period 1 relaxed relative to baseline areas by imaging day 2 after they were switched to the E− treatment. Results indicated that goats can vasoconstrict when exposed to ergot alkaloids that could disrupt their thermoregulation.

Ruminal tryptophan-utilizing bacteria degrade ergovaline from tall fescue seed extract

The objectives of this study were to evaluate degradation of ergovaline in a tall fescue [ (Schreb.) Darbysh.] seed extract by rumen microbiota ex vivo and to identify specific bacteria capable of ergovaline degradation in vitro. Rumen cell suspensions were prepared by harvesting rumen fluid from fistulated wether goats ( = 3), straining, and differential centrifugation. Suspensions were dispensed into anaerobic tubes with added Trypticase with or without extract (∼10 μg kg ergovaline). Suspensions were incubated for 48 h at 39°C. Samples were collected at 0, 24, and 48 h for ergovaline analysis and enumeration of hyper-ammonia producing (HAB) and tryptophan-utilizing bacteria. Ergovaline values were analyzed by repeated measures using the mixed procedure of SAS. Enumeration data were log transformed for statistical analysis. When suspensions were incubated with extract, 11 to 15% of ergovaline disappearance was observed over 48 h ( = 0.02). After 24 h, suspensions with added extract had 10-fold less HAB than controls ( = 0.04), but treatments were similar by 48 h ( = 1.00). However, after 24 h and 48 h, suspensions with extract had 10-fold more tryptophan-utilizing bacteria ( < 0.01) that were later isolated and identified by their 16S RNA gene sequence as . The isolates and other known rumen pure cultures ( JB1, B159, HD4, B, F, MD1, SR) were evaluated for the ability to degrade ergovaline in vitro. Pure culture cell suspensions were incubated as described above and samples were taken at 0 and 48 h for ergovaline analysis. Data were analyzed using the ANOVA procedure of SAS. All HAB, including the isolates, tested degraded ergovaline (54 to 75%; < 0.05). B14 was also able to degrade ergovaline but to a lesser capacity (12%; < 0.05), but all other bacteria tested did not degrade ergovaline. The results of this study indicate which rumen bacteria may play an important role in ergovaline degradation and that microbiological strategies for controlling their activity could have ramifications for fescue toxicosis and other forms of ergotism in ruminants.

Effect of starch source in pelleted concentrates on fecal bacteria in pre- and postpartum mares

&NA; Dietary starch source has been shown to affect fecal bacterial communities of horses fed minimally processed cereal grains. However, processing may increase foregut starch digestibility, reducing effects of starch source on fecal bacterial communities. This study aimed to determine the effect of starch source in pelleted concentrates on fecal Lactobacillus spp., amylolytic bacteria, and cellulolytic bacteria in broodmares mares, during the prepartum and postpartum period. Thoroughbred mares (n = 18) were paired by last breeding date then randomly assigned to either an oat‐based or a corn and wheat middlings‐based pelleted concentrate fed with forage. Mares were fed their assigned concentrates beginning on 310 days of gestation, and fecal samples were collected at 324 days of gestation, before parturition, 1 day, 14 days, and 28 days postpartum. Fecal samples were enumerated by serial dilution and inoculation into selective, enriched media for Lactobacillus spp., amylolytic bacteria, and cellulolytic bacteria. Data were log transformed then analyzed using a mixed model ANOVA with repeated measures (SAS 9.3) to test the main effects of treatment, time of sample, and treatment by time interaction. Starch source did not affect enumerated bacterial communities (P > .05); thus, pelleting concentrates may alter some of the effects of starch sources on the hindgut microbiota. Sample date did not affect amylolytic bacteria (P > .05); however, lactobacilli and cellulolytic bacteria decreased 1 day postpartum (P < .05). Although we did not observe an effect of starch source on fecal bacteria in mares, parturition did appear to alter the hindgut microbiota.

Effects of different levels of lespedeza and supplementation with monensin, coconut oil, or soybean oil on ruminal methane emission by mature Boer goat wethers after different lengths of feeding

ABSTRACT Mature Boer goat wethers were supplemented with 0.5% BW rolled corn and consumed pelleted alfalfa (CON), pelleted Sericea lespedeza (HSL; 6.4% condensed tannins), a 1:1 mixture of alfalfa and lespedeza (MSL), or alfalfa with monensin (ION; 22 mg/kg), coconut oil (CCO; 4%), or soybean oil (SBO; 4%). Total DM intake in the 20-wk study (3.86%, 3.75%, 3.52%, 3.69%, and 3.64% BW) and total tract OM digestibility determined every 5 wk (72.8%, 69.5%, 70.3%, 72.0%, and 71.1%) were not affected by treatment, although there were differences in nitrogen digestion (77.5%, 70.7%, 67.0%, 77.0%, 75.7%, and 73.6% for CON, MSL, HSL, ION, CCO, and SBO, respectively; SEM = 1.76). Ruminal methane emission was not influenced by period and was lowest among treatments for CON expressed as percentages of gross (10.3%, 6.8%, 6.3%, 7.2%, 6.5%, and 6.5%; SEM = 0.35) and digestible energy (14.8%, 10.2%, 9.3%, 10.6%, 9.8%, and 10.1% for CON, MSL, HSL, ION, CCO, and SBO, respectively; SEM = 0.62). In conclusion, both levels of lespedeza elicited similar depressions in ruminal methane emission, with a magnitude of change similar to that of an ionophore and coconut and soybean oils, and effects did not vary with week of the study.