This study investigated the effect of diet and host on the rumen bacterial microbiome and the impact of an acidotic challenge on its composition. Using parallel pyrosequencing of the V3 hypervariable region of 16S rRNA gene, solid and liquid associated bacterial communities of 8 heifers were profiled. Heifers were exclusively fed forage, before being transitioned to a concentrate diet, subjected to an acidotic challenge and allowed to recover. Samples of rumen digesta were collected when heifers were fed forage, mixed forage, high grain, during challenge (4 h and 12 h) and recovery. A total of 560,994 high-quality bacterial sequences were obtained from the solid and liquid digesta. Using cluster analysis, prominent bacterial populations differed (P≤0.10) in solid and liquid fractions between forage and grain diets. Differences among hosts and diets were not revealed by DGGE, but real time qPCR showed that several bacteria taxon were impacted by changes in diet, with the exception of Streptococcus bovis. Analysis of the core rumen microbiome identified 32 OTU's representing 10 distinct bacterial taxa including Bacteroidetes (32.8%), Firmicutes (43.2%) and Proteobacteria (14.3%). Diversity of OTUs was highest with forage with 38 unique OTUs identified as compared to only 11 with the high grain diet. Comparison of the microbial profiles of clincial vs. subclinical acidotic heifers found a increases in the relative abundances of Acetitomaculum, Lactobacillus, Prevotella, and Streptococcus. Increases in Streptococcus and Lactobacillus likely reflect the tolerance of these species to low pH and their ability to proliferate on surplus fermentable carbohydrate. The acetogen, Acetitomaculum may thereforeplay a role in the conversion of lactate to acetate in acidotic animals. Further profiling of the bacterial populations associated with subclinical and clinical acidosis could establish a microbial fingerprint for these disorders and provide insight into whether there are causative microbial populations that could potentially be therapeutically manipulated.

Little is known about the nature of the rumen epithelial adherent (epimural) microbiome in cattle fed different diets. Using denaturing gradient gel electrophoresis (DGGE), quantitative real-time PCR (qPCR), and pyrosequencing of the V3 hypervariable coding region of 16S rRNA, epimural bacterial communities of 8 cattle were profiled during the transition from a forage to a high-concentrate diet, during acidosis, and after recovery. A total of 153,621 high-quality gene sequences were obtained, with populations exhibiting less taxonomic variability among individuals than across diets. The bacterial community composition exhibited clustering (P < 0.03) by diet, with only 14 genera, representing >1% of the rumen epimural population, differing (P < 0.05) among diets. During acidosis, levels of Atopobium, Desulfocurvus, Fervidicola, Lactobacillus, and Olsenella increased, while during the recovery, Desulfocurvus, Lactobacillus, and Olsenella reverted to levels similar to those with the high-grain diet and Sharpea and Succinivibrio reverted to levels similar to those with the forage diet. The relative abundances of bacterial populations changed during diet transition for all qPCR targets except Streptococcus spp. Less than 5% of total operational taxonomic units (OTUs) identified exhibited significant variability across diets. Based on DGGE, the community structures of epithelial populations differed (P < 0.10); segregation was most prominent for the mixed forage diet versus the grain, acidotic challenge, and recovery diets. Atopobium, cc142, Lactobacillus, Olsenella, RC39, Sharpea, Solobacterium, Succiniclasticum, and Syntrophococcus were particularly prevalent during acidosis. Determining the metabolic roles of these key genera in the rumens of cattle fed high-grain diets could define a clinical microbial profile associated with ruminal acidosis.

This study was conducted to determine if the duration of time that beef cattle are fed a high-grain diet affects short-chain fatty acid (SCFA) absorption, saliva production, and blood metabolites before, during, and following an induced bout of ruminal acidosis. Sixteen Angus heifers were assigned to 1 of 4 blocks and within block to 1 of 2 treatments designated as long adapted (LA) or short adapted (SA). Long adapted and SA heifers were fed a backgrounding diet [forage:concentrate (F:C) = 60:40] for 33 and 7 d, respectively, and then transitioned over 20 d to a high-grain diet (F:C = 9:91) with the timing of dietary transition staggered such that the LA and SA heifers were fed the high-grain diet for 34 and 8 d, respectively, before inducing ruminal acidosis. Ruminal acidosis was induced by restricting feed to 50% of DMI:BW for 24 h followed by an intraruminal infusion of ground barley at 10% DMI:BW. Heifers were then given their regular diet allocation 1 h after the intraruminal infusion. Data were collected during an 8 d baseline period (BASE), on the day of the acidosis challenge (CHAL), and during 2 consecutive 8 d recovery periods (REC1 and REC2). When pooled across periods, the fractional rates of propionate (42 vs. 34%/h; P = 0.045) and butyrate (45 vs. 36%/h; P = 0.019) absorption, measured using the isolated and washed reticulorumen technique, were greater for LA than SA heifers. Moreover, overall, LA heifers tended to have greater absolute rates of butyrate absorption (94 vs. 79 mmol/h; P = 0.087) and fractional rates of total SCFA absorption (37 vs. 32%/h; P = 0.100). Treatment × period interactions for lactate absorption (P = 0.024) and serum D-lactate concentration (P = 0.003) were detected with LA heifers having greater D-lactate concentrations during CHAL and greater fractional rates of lactate absorption during REC1 than SA. The absolute and fractional absorption of acetate, propionate, and butyrate increased between REC1 and REC2, with intermediate values for BASE (P ≤ 0.05). Although fractional rates of SCFA absorption were low during REC1, saliva production (P = 0.018) increased between BASE and REC1, with intermediate values for REC2. These results suggest that the duration of time that animals are fed a high-grain diet may increase propionate, butyrate, and lactate absorption, and that cattle may decrease SCFA absorption and increase saliva production shortly after an acute bout of ruminal acidosis.

This study was conducted to determine if the duration of time cattle are fed a high-grain diet affects their susceptibility to and recovery from ruminal acidosis. Sixteen Angus heifers (BW ± SEM, 261 ± 6.1 kg) were assigned to 1 of 4 blocks and fed a backgrounding diet consisting of 60% barley silage, 30% barley grain, and 10% supplement (DM basis). Within block, cattle were randomly assigned to 1 of 2 treatments differing in the number of days they were fed the high-grain diet before an acidosis challenge: 34 d for long adapted (LA) and 8 d for short adapted (SA). All heifers were exposed to the same 20 d dietary transition to a high-grain diet containing 9% barley silage, 81% barley grain, and 10% supplement (DM basis). Ruminal acidosis was induced by restricting feed to 50% of DMI:BW for 24 h followed by an intraruminal infusion of ground barley at 10% DMI:BW. Heifers were then given their regular diet allocation 1 h after the intraruminal infusion. Data were collected during an 8-d baseline period (BASE), on the day of the acidosis challenge (CHAL), and during 2 consecutive 8-d recovery periods (REC1 and REC2). Acidosis induction increased daily duration (531 to 1,020 min/d; P < 0.001) and area (176 to 595 (min × pH)/d; P < 0.001) that ruminal pH was <5.5 relative to BASE. Relative to BASE, inducing acidosis also increased the daily mean (0.3 to 11.4 mM; P = 0.013) and maximum (1.3 to 29.3 mM; P = 0.008) ruminal fluid lactate concentrations. There was no effect of dietary treatment on ruminal pH, lactate, or short-chain fatty acid (SCFA) concentrations (P > 0.050). However, during BASE and CHAL, SA heifers experienced greater linear (P = 0.031), quadratic (P = 0.016), and cubic (P = 0.008) coefficients for the duration of time that pH was <5.5. In addition, a treatment × day interaction for the duration that pH was <5.5 during REC1 suggested that LA cattle tended to recover from the challenge more rapidly than SA cattle (P = 0.085). Regression analysis confirmed that the LA heifers experienced a quicker linear (P = 0.019) recovery from induced acidosis over time. These results indicate adaptation of the ruminal epithelium continues with advancing time as evidenced by more stable ruminal pH both before and after an induced bout of acute ruminal acidosis but does not affect susceptibility of cattle to ruminal acidosis.

The objective of this study was to determine how duration of time that cattle are fed a high-grain diet affects feed sorting, both before and after an episode of acute ruminal acidosis. Sixteen Angus heifers (261 ± 6.1 kg; BW ± SEM) were assigned to 1 of 4 blocks and fed a backgrounding (BG) diet (60% forage, DM basis). Within block, heifers were randomly assigned to 1 of 2 treatments differing in days fed a high-grain (HG; 9% forage, DM basis, fed ad libitum) diet before a ruminal acidosis challenge: 34 d for long adapted (LA) and 8 d for short adapted (SA). Ruminal acidosis was induced by restricting feed to 50% of DMI as a proportion of BW (determined individually for each heifer) for 24 h followed by an intraruminal infusion of ground barley at 10% of DMI as a proportion of BW measured before feed restriction. Feed and orts were sampled during the BG period, the first 26 d on the HG diet (only for LA cattle), the 8-d baseline (BASE) period, on the day of the ruminal acidosis challenge (CH), and during 2 consecutive 8-d recovery periods (REC1 and REC2) for each heifer and subjected to particle size analysis: 19-mm (long), 8-mm (medium), and 1.18-mm (short) screens and a pan (fine). On the BG diet, sorting for medium particles tended to be greater (104.2 vs. 102.1%; P = 0.07) for LA heifers than SA heifers, while sorting against short particles was greater (98.2 vs. 100.0%; P = 0.05) for LA heifers. During the first 26 d on the HG diet, LA cattle sorted for (P < 0.001) long (118.8%), medium (117.8%), and short (104.1%) particles and sorted against (P < 0.001) fine particles (45.3%). This sorting pattern was consistent for LA heifers during BASE period, CH day, and recovery periods, across which SA heifers exhibited less sorting (P ≤ 0.1). Greater duration of pH < 5.5 during the BASE period was associated with greater sorting for long particles (R(2) = 0.75, P = 0.01) in LA heifers and for long (R(2) = 0.49, P = 0.05) and medium (R(2) = 0.88, P < 0.001) particles in SA heifers. Long-adapted heifers linearly increased the extent of sorting for long (P = 0.007) and medium (P < 0.001) particles and against fine particles (P = 0.05) during the days following the challenge to a greater extent than SA heifers. Overall, the results demonstrate that longer-term exposure of beef heifers to a HG diet, which caused persistent low rumen pH, influenced feed sorting of heifers, both before and after an induced bout of acute ruminal acidosis, in a manner that would help attenuate the effects of acidosis.

The objective of this study was to determine how feed sorting changes in response to the severity of an induced episode of acute ruminal acidosis. Sixteen Angus heifers (261 ± 6.1 kg; bodyweight (BW) ± s.e.m.) were adapted to a high-grain diet (9% forage) before a ruminal acidosis challenge. Ruminal acidosis was induced by restricting feed to 50% of dry matter intake (DMI) as a proportion of BW for 24 h, followed by an intra-ruminal infusion of ground barley at 10% of DMI as a proportion of BW. Ruminal pH and feed sorting were monitored for 8 days (Base) before the challenge and on the challenge (CH) day. Ruminal pH data (duration (min) and area above the curve (min × pH) of pH <5.5) were recorded using an indwelling measurement system. To assess feed sorting, feed and orts were sampled daily and subjected to particle-size analysis. The particle separator had three screens (19, 8, 1.18 mm) and a bottom pan, resulting in four fractions (long, medium, short, fine). Sorting (%) was calculated as the actual intake/predicted intake of each particle fraction. During the Base period and the CH day, heifers sorted for long, medium and short particles, and sorted against fine particles. During the Base period, heifers experienced ruminal pH <5.5 for 489 ± 73 min/day with an area of 154 ± 29 (pH × min)/day; on the CH day, the duration increased to 1020 ± 75 min/day with an area of 631 ± 102 (pH × min)/day. A greater change in duration of pH <5.5 was associated with a greater increase in sorting for long and medium particles. A greater change in the area of pH <5.5 was associated with a greater increase in sorting for long particles and against fine particles. Overall, results show that cattle that experienced a greater degree of acidosis partially coped by sorting their ration more to consume a greater proportion of long, fibrous particles.

SUMMARYGoldfish acclimated to cold water (e.g. 7°C) experience a marked reduction in functional lamellar surface area owing to the proliferation of an interlamellar cell mass (ILCM), a phenomenon termed gill remodelling. The goal of the present study was to assess the consequences of the reduced functional surface area on the capacity of goldfish to excrete ammonia. Despite the expected impact of ambient temperature on functional surface area, fish acclimated to 7°C and 25°C exhibited similar rates of ammonia excretion (J net,amm ); the Q 10 values for fed and starved fish were 1.07 and 1.20, respectively. To control for possible temperature-related differences in rates of endogenous ammonia production, J net,amm was determined at the two acclimation temperatures after loading fish with 1.12molg -1 of NH 4 Cl. In the 3h post-injection period, J net,amm was elevated to a greater extent in the 25°C fish. To estimate the potential contribution of increased ventilation and cardiac output to ammonia clearance in the warmer fish, the ammonia loading experiment was repeated on the 7°C fish immediately after they were exercised to exhaustion. The rate of excretion of ammonia was significantly increased in the exercised 7°C fish (presumably experiencing increased ventilation and cardiac output for at least some of the measurement period) suggesting that differences in external and internal convection may at least partially explain the enhanced capacity of the 25°C fish to clear the ammonia load. To more specifically assess the contribution of the different functional surface areas on the differing rates of ammonia clearance at the two acclimation temperatures, the 7°C fish were exposed for 7days to hypoxia (P O2 10 mmHg1.33kPa), a treatment known to cause the disappearance of the ILCM. The results demonstrated that the hypoxia-associated loss of the ILCM was accompanied by a significant increase in the rate of ammonia clearance in the 7°C fish when returned to normoxic conditions. To determine whether compensatory changes in the ammonia transporting proteins might be contributing to sustaining J net,amm under conditions of reduced functional lamellar surface area, the relative expression and branchial distribution of four Rh proteins were assessed by western blotting and immunocytochemistry. Although the relative expression of the Rh proteins was unaffected by acclimation temperature, there did appear to be a change in the spatial distribution of Rhag, Rhbg and Rhcg1. Specifically, these three Rh proteins (and to a lesser extent Rhcg2) appeared to localize in cells on the outer edge of the ILCM that were enriched with Na + /K + -ATPase. Thus, we suggest that despite the impediment to ammonia excretion imposed by the ILCM, goldfish acclimated to 7°C are able to sustain normal rates of excretion owing to the redistribution of ammonia transporting cells.