Protozoa involved in butyric rather than lactic fermentative pattern during latent acidosis in sheep

We used six ruminally cannulated Texel wethers to study the relative role of protozoa and lactate-metabolizing bacteria in ruminal fermentative patterns during an induced latent acidosis. The sheep were fed an alfalfa hay diet (H) and latent acidosis was induced, following a short transition period of one week, with a grain-rich acidotic diet (W, 60% wheat + 40% alfalfa hay). Ruminal pH, ruminal volatile fatty acids (VFA), lactate and NH3 concentrations, protozoa and lactate-utilizing bacterial counts, the relative proportions of three main bacteria implicated in lactate metabolism (a lactate-producing species, Streptococcus bovis, and two lactate-utilizing species, Selenomonas ruminantium, and Megasphaera elsdenii) using specific 16S-rRNA-targeting oligonucleotide probes, and lactate dehydrogenase (LDH) activity were determined for both diets. The pH parameters (mean, minimum, maximum, time and area under pH 6.0 and 5.5) measured with the W diet were indicative of a latent (i.e., subacute and maintained) acidosis. However, a butyric rather than lactic latent acidosis was observed in this study. Total ruminal lactate concentration remained at low levels with the acidotic diet (< 4 mmol x L(-1)), but changes were observed in VFA composition, which was oriented towards butyrate at the expense of acetate (P < 0.05), while propionate remained constant. In agreement with the low ruminal lactate concentration, no changes in the proportion of S. bovis 16S-rRNA were observed. The lactate-metabolizing bacterial population also remained fairly constant in number, proportion and activity. The increase in butyrate concentration was accompanied by a proliferation of entodiniomorphs (P < 0.01). These results suggest that the protozoa limited lactate accumulation and possibly also the decrease in pH during latent acidosis. Experiments with defaunated and faunated sheep could provide further evidence of the role of protozoa in the development of rumen latent acidosis.     

Effects of varying proportions of concentrates on ruminal-reducing power and bacterial community structure in dry dairy cows fed hay-based diets

The objectives of this study were to evaluate the influence of diet composition on ruminal parameters, more particularly redox potential (Eh). Four Holstein dry dairy cows, fitted with ruminal cannulas, were allocated in a 4 × 4 Latin square design. They were given four experimental hay-based diets D0, D25, D42 and D56 consisting of 0%, 25%, 42% and 56% of ground wheat and barley concentrate mixture, respectively. They were fed at a daily feeding rate of 8.0 kg DM per cow during a 24-day experimental period (a 21-day diet adaptation, three consecutive days of measurement and sampling). The physicochemical parameters, such as pH and Eh, were measured and Clark's exponent (rH) was calculated from 1 h before feeding to 8 h after feeding at 1-h interval. Samples of ruminal fluid were taken at 0, 1, 2, 4, 6 and 8 h after feeding for the determination of volatile fatty acid (VFA) and ammonia N (NH3-N) concentrations. Ruminal bacterial populations were also studied by means of capillary electrophoresis single-strand conformation polymorphism (CE-SSCP) technique to focus on the structure of the ruminal microbiota and the diversity index was calculated. Mean ruminal Eh and rH were not modified by the concentrate-to-forage ratio and averaged - 210 mV and 6.30, respectively, across diets. The pH decreased slightly by 0.10 pH unit between treatments D0 and D56 with an average of 6.58. Nevertheless, the time during which physicochemical parameters remained at nadir value after feeding varied with diets: 2 and 7 h for D0 and 6 and 5 h for D56, respectively for pH and Eh. Moreover, fermentative parameters were altered by treatments: total VFA and NH3-N were greater in D56 (72.2 mM and 17.5 mg/100 ml) compared with D0 (65.2 mM and 14.2 mg/100 ml). However, neither the structure of bacterial populations of the rumen nor the diversity index (Shannon) was altered by treatments.     

Effects of the acid–base treatment of corn on rumen fermentation and microbiota,inflammatory response and growth performance in beef cattle fed high-concentrate diet

Beef cattle are often fed high-concentrate diet (HCD) to achieve high growth rate. However, HCD feeding is strongly associated with metabolic disorders. Mild acid treatment of grains in HCD with 1% hydrochloric acid (HA) followed by neutralization with sodium bicarbonate (SB) might modify rumen fermentation patterns and microbiota, thereby decreasing the negative effects of HCD. This study was thus aimed to investigate the effects of treatment of corn with 1% HA and subsequent neutralization with SB on rumen fermentation and microbiota, inflammatory response and growth performance in beef cattle fed HCD. Eighteen beef cattle were randomly allocated to three groups and each group was fed different diets: low-concentrate diet (LCD) (concentrate : forage = 40 : 60), HCD (concentrate : forage = 60 : 40) or HCD based on treated corn (HCDT) with the same concentrate to forage ratio as the HCD. The corn in the HCDT was steeped in 1% HA (wt/wt) for 48 h and neutralized with SB after HA treatment. The animal trial lasted for 42 days with an adaptation period of 7 days. At the end of the trial, rumen fluid samples were collected for measuring ruminal pH values, short-chain fatty acids, endotoxin (or lipopolysaccharide, LPS) and bacterial microbiota. Plasma samples were collected at the end of the trial to determine the concentrations of plasma LPS, proinflammatory cytokines and acute phase proteins (APPs). The results showed that compared with the LCD, feeding the HCD had better growth performance due to a shift in the ruminal fermentation pattern from acetate towards propionate, butyrate and valerate. However, the HCD decreased ruminal pH and increased ruminal LPS release and the concentrations of plasma proinflammatory cytokines and APPs. Furthermore, feeding the HCD reduced bacterial richness and diversity in the rumen. Treatment of corn increased resistant starch (RS) content. Compared with the HCD, feeding the HCDT reduced ruminal LPS and improved ruminal bacterial microbiota, resulting in decreased inflammation and improved growth performance. In conclusion, although the HCD had better growth performance than the LCD, feeding the HCD promoted the pH reduction and the LPS release in the rumen, disturbed the ruminal bacterial stability and increased inflammatory response. Treatment of corn with HA in combination with subsequent SB neutralization increased the RS content and helped counter the negative effects of feeding HCD to beef steers.     

Rumen microbial changes in cattle fed diets with or without salinomycin.

Four rumen-fistulated steers, randomly assigned to two groups (control and salinomycin fed) were used to monitor the changes in rumen microbial populations and volatile fatty acids (VFA) concentrations associated with feeding salinomycin (0.22 mg X kg-1 X day-1). Steers were adapted to an alfalfa hay and grain (80:20) diet before supplementing the diet with salinomycin, and then the diet was changed to 50:50 and 20:80 ratios of alfalfa hay to grain at 2-week intervals. Rumen samples for total and selective enumeration of anaerobic bacteria. VFA analysis, and enumeration of protozoa were collected during the 80:20 alfalfa hay-to-grain diet before salinomycin feeding, and during the 80:20, 50:50, and 20:80 hay-to-grain diets with salinomycin. At each sampling period, rumen samples were collected at 3 h after feeding on three consecutive days. Salinomycin feeding had no effect on rumen pH and total VFA concentration. The acetate-to-propionate ratio was significantly lower in salinomycin-fed steers than in the control. The molar proportion of butyrate increased in both control and salinomycin-fed steers. Total anaerobic bacterial counts were lower in salinomycin-fed steers than in the control steers after 8 weeks of salinomycin feeding. Salinomycin-resistant bacteria increased from 7.6 to 15.6% in salinomycin-fed steers but remained unchanged in control steers. Salinomycin had no effect on cellulolytic and lactate-utilizing bacteria, but the proportion of amylolytic bacteria was higher in salinomycin-fed steers than in control steers. The total number of protozoa decreased initially in salinomycin-fed steers. The initial reduction was due to reduced numbers of Entodinium species. Holotrichs were unaffected by salinomycin feeding.     

Rumen microbial changes in cattle fed diets with or without salinomycin

Four rumen-fistulated steers, randomly assigned to two groups (control and salinomycin fed) were used to monitor the changes in rumen microbial populations and volatile fatty acids (VFA) concentrations associated with feeding salinomycin (0.22 mg X kg-1 X day-1). Steers were adapted to an alfalfa hay and grain (80:20) diet before supplementing the diet with salinomycin, and then the diet was changed to 50:50 and 20:80 ratios of alfalfa hay to grain at 2-week intervals. Rumen samples for total and selective enumeration of anaerobic bacteria. VFA analysis, and enumeration of protozoa were collected during the 80:20 alfalfa hay-to-grain diet before salinomycin feeding, and during the 80:20, 50:50, and 20:80 hay-to-grain diets with salinomycin. At each sampling period, rumen samples were collected at 3 h after feeding on three consecutive days. Salinomycin feeding had no effect on rumen pH and total VFA concentration. The acetate-to-propionate ratio was significantly lower in salinomycin-fed steers than in the control. The molar proportion of butyrate increased in both control and salinomycin-fed steers. Total anaerobic bacterial counts were lower in salinomycin-fed steers than in the control steers after 8 weeks of salinomycin feeding. Salinomycin-resistant bacteria increased from 7.6 to 15.6% in salinomycin-fed steers but remained unchanged in control steers. Salinomycin had no effect on cellulolytic and lactate-utilizing bacteria, but the proportion of amylolytic bacteria was higher in salinomycin-fed steers than in control steers. The total number of protozoa decreased initially in salinomycin-fed steers. The initial reduction was due to reduced numbers of Entodinium species. Holotrichs were unaffected by salinomycin feeding.     

Impact of subacute ruminal acidosis on the diversity of liquid and solid-associated bacteria in the rumen of goats

This study was aimed to investigate the impact of subacute ruminal acidosis (SARA) on the diversity of liquid (LAB) and solid-associated bacteria (SAB) following high-grain feeding. Six ruminally cannulated goats were divided into two groups: one group was fed a hay diet (COD), and the other group was fed a high grain diet (SAID). Rumen liquids and rumen solids were sampled after 2 weeks adaption. SARA was diagnosed with a pH below 5.8 for 8 h. SAID decreased ruminal pH (P < 0.001) and increased the acetate (P = 0.017), propionate (P = 0.001), butyrate (P < 0.001) and total volatile fatty acid (P < 0.001) concentration in rumen compared with the COD. Denaturing gradient gel electrophoresis fingerprints analysis revealed a clear separation between both the diet and the fraction of rumen digesta in bacterial communities. Pyrosequencing analysis showed that the proportion of phylum Bacteroidetes in the SAID-LAB and SAID-SAB communities was less than in the COD group, whereas the SAID group had a greater percentage of Firmicutes in both the LAB and SAB libraries. UniFrac analyses and a Venn diagram revealed a large difference between the two diets in the diversity of rumen bacterial communities. Overall, our findings revealed that SARA feeding did alter the community structure of rumen liquids and rumen solids. Thus, manipulation of dietary factors, such as ratio of forage to concentrate may have the potential to alter the microbial composition of rumen liquid and rumen solid.     

Diurnal variations in bacterial numbers and fluid parameters in ruminal contents of animals fed low- or high-forage diets.

Differential carbohydrate media and anaerobic replica plating techniques were used to assess the degrees of diurnal variations in the direct and viable cell counts as well as the carbohydrate-specific subgroups within the mixed rumen bacterial populations in cattle fed maintenance (metabolizable energy) levels of either a high-forage or a high-concentrate diet once daily. The rumen was sampled at 1 h before feeding and 2, 4, 8, 12, and 16 h after feeding, and selected microbiological parameters of the isolated bacterial populations were assessed. Corresponding samples of ruminal fluid were assayed for fermentation acids, carbohydrate, ammonia, and pH changes. The data showed that regardless of diet, total bacterial numbers remained fairly constant throughout the day. The number of viable bacteria declined 40 to 60% after feeding and then increased to a maximum at 16 h postfeeding. Changes occurred in the carbohydrate-specific subgroups within the bacterial populations, and some of the changes were consistent with a predicted scheme of ruminal feedstuff carbohydrate fermentation. Regardless of diet, however, soluble-carbohydrate-utilizing bacteria predominated at all times. Xylan-xylose and pectin subgroups respectively comprised about one-half and one-third of the population when the high-forage diet was given. These subgroups, along with the cellulolytics, constituted lesser proportions of the population when the high-concentrate diet was given. The cellulolytic subgroup was the least numerous of all subgroups regardless of diet but followed a diurnal pattern similar to that predicted for cellulose fermentation. There were few diurnal variations or differences in bacterial cell compositions and ruminal fluid parameters between diets. The observed similarities and dissimilarities of the rumen bacterial populations obtained when the two diets were given are discussed. The data are consistent with the versatility and constancy of the rumen as a stable, mature microbial system under the specific low-level feeding regimens used.     

Effect of disodium fumarate on ruminal metabolism and rumen bacterial communities as revealed by denaturing gradient gel electrophoresis analysis of 16S ribosomal DNA

The aim of the present study was to examine the effects of feeding diets with addition of disodium fumarate (DF) to goats on ruminal metabolism and changes of rumen bacterial communities. Four cannulated goats were used in a 4 × 4 Latin square design. The results showed that ruminal pH increased linearly (P<0.01)as the amount of DF added increased, while lactate production decreased linearly (P<0.01). DF addition did not affect the production of acetate, propionate, butyrate, TVFA and NH₃-N. The effect of DF on the changes in rumen bacterial-community structure of goats was analyzed using 16S rDNA-based approaches. Amplicons of the V6-V8 variable regions of bacterial 16S rDNA were analyzed by denaturing gradient gel electrophoresis (DGGE), cloning and sequencing. Differences in rumen bacterial community structure were determined based on the Shannon index of diversity for pairwise comparison of the DGGE fingerprints and revealed significant changes in rumen microbiota after DF addition. As compared with those fed with the control diet, goats fed on the diets with DF addition showed a higher bacterial diversity. The sequences of seven amplicons in total 11 clones showed less than 97% similarity with those of previously identified or unidentified bacteria, suggesting that most bacteria in the gastrointestinal tract have not been cultured or identified. Amplicons related to Succinivibrio dextrinosolvens species were found in most DGGE fingerprints derived from goats on the diet containing DF, but not in goats on the control diet. These results demonstrated the ability of DF to improve the metabolism of rumen lactate fermentation and to influence the bacterial composition of the rumen in goats.     

Effects of expander-treating a barley-based concentrate on ruminal fermentation, bacterial N synthesis, escape of dietary N, and performance of dairy cows

Three primiparous dairy cows in early lactation with cannulas in rumen, duodenum and ileum were used in a 3×3 Latin square design to study effects of expander treatment of a barley-based concentrate. The concentrate was either pelleted at 75–80°C or expander treated at 125–130°C prior to pelleting. The diets consisted of 6.7 kg DM of grass silage and 10 kg DM of (1) 100% pelleted, (2) 50% pelleted and 50% expanded or (3) 100% expanded concentrate. The diets were offered as a mixed ration in four equal meals daily. Ruminal fermentation, bacterial N synthesis, duodenal, ileal and faecal flow of nutrients, and animal performance were monitored. Expander treatment numerically increased ruminal digestion of starch, which explained the observed increase in ruminal VFA concentration and the lowered ruminal pH (P<0.05). The proportion of butyrate in rumen liquid increased, whereas the proportion of propionate decreased in the expanded compared to the pelleted treatment (P<0.05). Expander treatment tended to increase rumen volume and rumen NDF pool size. Ruminal digestion of NDF was numerically lower in the expanded than in the pelleted treatment. No differences in bacterial N synthesis or efficiency of synthesis were observed among treatments. Expander treatment numerically increased the duodenal flow of non-ammonia N (NAN) and amino acid N (AAN), and seemed to increase the flow of non-ammonia non-bacterial N (NANBN) to the duodenum to a similar extent as was indicated by nylon bag studies. Milk production and milk fat and protein content were increased by the expander treatment (P<0.05), indicating that expander treatment increased the supply of nutrients for milk production.     

Comparison of microbial fermentation of high- and low-forage diets in Rusitec, single-flow continuous-culture fermenters and sheep rumen

Eight Rusitec and eight single-flow continuous-culture fermenters (SFCCF) were used to compare the ruminal fermentation of two diets composed of alfalfa hay and concentrate in proportions of 80 : 20 (F80) and 20 : 80 (F20). Results were validated with those obtained previously in sheep fed the same diets. Rusitec fermenters were fed once daily and SFCCF twice, but liquid dilution rates were similar in both types of fermenters. Mean values of pH over the 12 h postfeeding were higher (P < 0.001) in Rusitec than in SFCCF, with diet F80 showing higher values (P < 0.001) in both types of fermenters. Concentrations of total volatile fatty acids (VFA) were higher (P < 0.001) in SFCCF than in Rusitec, and in both systems were higher (P = 0.002) for diet F20 than for diet F80. There were significant differences between systems in the proportions of the main VFA, and a fermentation system × diet interaction (P < 0.001) was detected for all VFA with the exception of valerate. No differences (P = 0.145) between the two types of fermenters were detected in dry matter (DM) digestibility, but NDF, microbial N flow and its efficiency were higher (P = 0.001) in SFCCF compared to Rusitec. Whereas pH values and VFA concentrations remained fairly stable through the day in both in vitro systems, pH dropped and VFA increased shortly after feeding in sheep rumen reaching the minimum and maximal values, respectively, about 4 h after feeding. Both in vitro systems detected differences between diets similar to those found in sheep for liquid dilution rate, pH values, DM digestibility, microbial N flow and growth efficiency. In contrast, acetate/propionate ratios were lower for diet F20 than for F80 in sheep rumen (2.73 and 3.97) and SFCCF (3.07 and 4.80), but were higher for diet F20 compared to F80 (4.29 and 3.40) in Rusitec, with values considered to be unphysiological for high-concentrate diets. In vivo NDF digestibility was affected (P = 0.017) by diet, but no differences between diets (P > 0.05) were found in any in vitro system. A more precise control of pH in both types of fermenters and a reduction of concentrate retention time in Rusitec could probably improve the simulation of in vivo fermentation.     

Effects of dietary forage particle size and concentrate level on fermentation profile, in vitro degradation characteristics and concentration of liquid- or solid-associated bacterial mass in the rumen of dairy cows

This study investigated effects of dietary forage particle size (PS) and concentrate level (CL) on fermentation profiles of particle-associated rumen liquid (PARL) and free rumen liquid (FRL), in vitro degradation characteristics and concentration of bacterial mass attached to the solid or fluid rumen digesta phase in dairy cows. The experiment was a 4 × 4 Latin square design with four late-lactation dairy cows in four 23 day periods. Cows were restrictively fed (17 kg dry matter (DM)/d) one of four diets varying in the theoretical PS (6 and 30 mm) of grass hay and in the levels (approximately 200 and 550 g/kg, DM basis) of a cereal-based concentrate. Proportion of large particles (>6 mm) and the content of structural fibre in the diet increased by reducing dietary CL and, particularly, by increasing hay PS. This effect was not reflected by changes in mean total volatile fatty acid concentration or pH in the rumen. However, cows fed high concentrate diets had pH of 5.28 and 5.37 in PARL at 3 h after the last meal, when fine or long chopped hay was offered. The low pH may indicate a depression of the capacity of PARL to degrade fibre in vitro. Gas production in vitro of concentrate increased with the high concentrate diet at 12 h, suggesting that amylolytic capacity was affected only in early phases of fermentation. In addition, elevating dietary CL appeared to shift ruminal fermentation outputs from propionate to butyrate and valerate. Inclusion of coarsely chopped hay to a high concentrate diet does not appear to bring advantages due to increased structure in restrictively fed dairy cows. In addition, results suggest that the response of pH in PARL is more sensitive to dietary changes (i.e., forage PS and CL) than the response in FRL, and so PARL might be better to evaluate the risk of ruminal disfunction in dairy cows.     

Soybean oil and linseed oil supplementation affect profiles of ruminal microorganisms in dairy cows

The objective of this study was to evaluate changes in ruminal microorganisms and fermentation parameters due to dietary supplementation of soybean and linseed oil alone or in combination. Four dietary treatments were tested in a Latin square designed experiment using four primiparous rumen-cannulated dairy cows. Treatments were control (C, 60 : 40 forage to concentrate) or C with 4% soybean oil (S), 4% linseed oil (L) or 2% soybean oil plus 2% linseed oil (SL) in a 4 × 4 Latin square with four periods of 21 days. Forage and concentrate mixtures were fed at 0800 and 2000 h daily. Ruminal fluid was collected every 2 h over a 12-h period on day 19 of each experimental period and pH was measured immediately. Samples were prepared for analyses of concentrations of volatile fatty acids (VFA) by GLC and ammonia. Counts of total and individual bacterial groups (cellulolytic, proteolytic, amylolytic bacteria and total viable bacteria) were performed using the roll-tube technique, and protozoa counts were measured via microscopy in ruminal fluid collected at 0, 4 and 8 h after the morning feeding. Content of ruminal digesta was obtained via the rumen cannula before the morning feeding and used immediately for DNA extraction and quantity of specific bacterial species was obtained using real- time PCR. Ruminal pH did not differ but total VFA (110 v. 105 mmol/l) were lower (P < 0.05) with oil supplementation compared with C. Concentration of ruminal NH3-N (4.4 v. 5.6 mmol/l) was greater (P < 0.05) due to oil compared with C. Compared with C, oil supplementation resulted in lower (P < 0.05) cellulolytic bacteria (3.25 × 108 v. 4.66 × 108 colony-forming units (CFU)/ml) and protozoa (9.04 × 104 v. 12.92 × 104 cell/ml) colony counts. Proteolytic bacteria (7.01 × 108 v. 6.08 × 108 CFU/ml) counts, however, were greater in response to oil compared with C (P < 0.05). Among oil treatments, the amount of Butyrivibrio fibrisolvens, Fibrobacter succinogenes and Ruminococcus flavefaciens in ruminal fluid was substantially lower (P < 0.05) when L was included. Compared to C, the amount of Ruminococcus albus decreased by an average of 40% regardless of oil level or type. Overall, the results indicate that some ruminal microorganisms, except proteolytic bacteria, are highly susceptible to dietary unsaturated fatty acids supplementation, particularly when linolenic acid rich oils were fed. Dietary oil effects on ruminal fermentation parameters seemed associated with the profile of ruminal microorganisms.     

Effect of rumen ciliates on the digestion of different carbohydrates in sheep. I.--Utilization of cell wall carbohydrates (cellulose and hemicellulose) and of starch

Two diets rich in cell-wall carbohydrates or starch were given to 10 rumen-fistulated sheep; two sheep were defaunated and the others were inoculated with Polyplastron multivesiculatum (P) or Entodinium sp. (E), or both (P + E), or with conventional fauna. Ciliate biomass was greater when the animals were fed a high starch diet than when the diet was rich in cell-wall carbohydrates (table 2). With both diets, the Entodinium genus in the mixed fauna sampling predominated. We showed that Polyplastron was directly involved in cell-wall carbohydrate breakdown, while Entodinium capacity to digest cellulose remained low. We noted that with a diet rich in cellulose and hemicellulose, bacterial cellulolytic activity was improved by the presence of ciliates in the rumen but was decreased with the "starch" diet (table 3). The greater VFA concentration observed in the faunated animals expressed ciliate effect on the fermentations as well as activation of bacterial metabolism. With a high starch diet, the Entodinium sp. ciliates may have a buffering effect on the pH values in the rumen by limiting bacterial fermentation after food intake and by prolonging starch digestion during the day (table 4). The composition of the VFA mixture was modified by ciliate inoculation. The molar proportion of butyric acid always increased, while that of acetic and propionic acids evolved differently according to the diets and the ciliates (table 4). The higher ammonia concentration in the rumen liquor observed in faunated animals (table 4) could be explained either by the breakdown of both feed and bacterial proteins ingested by ciliates or by a lower ammonia nitrogen incorporation by fewer bacteria. Statistical analyses were used to explain the specific effect of P and E and also the interactions between them and between each of them and the diets.     

Effects of Dietary Linseed Oil and Propionate Precursors on Ruminal Microbial Community,Composition, and Diversity in Yanbian Yellow Cattle

The rumen microbial ecosystem is a complex system where rumen fermentation processes involve interactions among microorganisms. There are important relationships between diet and the ruminal bacterial composition. Thus, we investigated the ruminal fermentation characteristics and compared ruminal bacterial communities using tag amplicon pyrosequencing analysis in Yanbian yellow steers, which were fed linseed oil (LO) and propionate precursors. We used eight ruminally cannulated Yanbian yellow steers (510 ± 5.8 kg) in a replicated 4 × 4 Latin square design with four dietary treatments. Steers were fed a basal diet that comprised 80% concentrate and 20% rice straw (DM basis, CON). The CON diet was supplemented with LO at 4%. The LO diet was also supplemented with 2% dl-malate or 2% fumarate as ruminal precursors of propionate. Dietary supplementation with LO and propionate precursors increased ruminal pH, total volatile fatty acid concentrations, and the molar proportion of propionate. The most abundant bacterial operational taxonomic units in the rumen were related to dietary treatments. Bacteroidetes dominated the ruminal bacterial community and the genus Prevotella was highly represented when steers were fed LO plus propionate precursors. However, with the CON and LO diet plus malate or fumarate, Firmicutes was the most abundant phylum and the genus Ruminococcus was predominant. In summary, supplementing the diets of ruminants with a moderate level of LO plus propionate precursors modified the ruminal fermentation pattern. The most positive responses to LO and propionate precursors supplementation were in the phyla Bacteriodetes and Firmicutes, and in the genus Ruminococcus and Prevotella. Thus, diets containing LO plus malate or fumarate have significant effects on the composition of the rumen microbial community.     

Changes in bacterial diversity associated with epithelial tissue in the beef cow rumen during the transition to a high-grain diet

Our understanding of the ruminal epithelial tissue-associated bacterial (defined as epimural bacteria in this study) community is limited. In this study, we aimed to determine whether diet influences the diversity of the epimural bacterial community in the bovine rumen. Twenty-four beef heifers were randomly assigned to either a rapid grain adaptation (RGA) treatment (n = 18) in which the heifers were allowed to adapt from a diet containing 97% hay to a diet containing 8% hay over 29 days or to the control group (n = 6), which was fed 97% hay. Rumen papillae were collected when the heifers were fed 97%, 25%, and 8% hay diets. PCR-denaturing gradient gel electrophoresis (DGGE) and quantitative real-time PCR analysis were used to characterize rumen epimural bacterial diversity and to estimate the total epimural bacterial population (copy numbers of the 16S rRNA gene). The epimural bacterial diversity from RGA heifers changed (P = 0.01) in response to the rapid dietary transition, whereas it was not affected in control heifers. A total of 88 PCR-DGGE bands were detected, and 44 were identified from phyla including Firmicutes, Bacteroidetes, and Proteobacteria. The bacteria Treponema sp., Ruminobacter sp., and Lachnospiraceae sp. were detected only when heifers were fed 25% and 8% hay diets, suggesting the presence of these bacteria is the result of adaptation to the high-grain diets. In addition, the total estimated population of rumen epimural bacteria was positively correlated with molar proportions of acetate, isobutyrate, and isovalerate, suggesting that they may play a role in volatile fatty acid metabolism in the rumen.     

Effects of sampling location and time,and host animal on assessment of bacterial diversity and fermentation parameters in the bovine rumen

Aims: To investigate, using culture‐independent methods, whether the ruminal bacterial structure, population and fermentation parameters differed between sampling locations and time. Methods and Results: The detectable bacteria and fermentation parameters in the digesta from five locations in the rumen of three cows at three time points were analysed. The PCR‐denaturing gradient gel electrophoresis (PCR‐DGGE) profiles were similar among digesta samples from five locations (95·4%) and three time points (93·4%) within cows; however, a lower similarity was observed for samples collected from different host animals (85·5%). Rumen pH and concentration of volatile fatty acids (VFA) were affected by time points of sampling relative to feeding. Conclusions: The detectable bacterial structure in the rumen is highly conserved among different locations and over time, while the quantity of individual bacterial species may change diurnally in response to the feeding. Significance and impact of the study: This study supplies the fundamental understanding of the microbial ecology in the rumen, which is essential for manipulation of ruminal microflora and subsequent improvement in animal production.     

Feeding behavior, rumination, reticulo-ruminal fermentation and plasma volatile fatty acids, compared in goats and sheep; influence of the diet

Two-year old Alpine she-goats (n = 3) and Texel ewes (n = 3) were compared as to eating behaviour, rumination and volatile fatty acids (VFA) in rumen and blood. The animals were fed once daily with two different proportions (20 and 80%) of barley and hay. Dry matter intake was fixed at 48 g D.M/kg P0.75 per day. In similar feeding and environmental conditions, eating behaviour and rumination of goats and sheep did not differ much: the goats tended to eat faster and there were more rumination periods in the sheep. Latency time, mean duration of rumination periods, daily rumination time and circadian pattern of rumination did not differ significantly between the two species (fig. 1). With both diets we observed a higher VFA concentration and a lower acetate/propionate ratio in the rumen of the ewes; however, rumen pH was lower only in those eating the 80% barley ration (fig. 2). Blood VFA in the jugular vein did not differ between sheep and goats (fig. 3). The proportion of cereals and hay in the diets affected rumen fermentation and rumination pattern in both species. With a higher concentrate/roughage ratio, rumen and plasma VFA increased, while the pH and acetate/propionate ratio in the lumen juice, the number of rumination periods and daily rumination time decreased. When the animals were fed the 80% barley ration, there was practically no rumination in the first 9 h after the single meal. During this time, rumen pH was minimal and VFA levels in the rumen and blood were maximal.     

Effect of rumen ciliates on the digestive utilization of various carbohydrate-rich diets and on the end-products formed in the rumen. II. Utilization of inulin, saccharose and lactose

Three diets rich in inulin, saccharose and lactose, respectively, were given to 10 rumen-fistulated sheep. Two animals were defaunated, two were inoculated with either Polyplastron multivesibulatum or Entodinium sp., and two others were inoculated with both. The latter two were bred in conventional conditions. All animals ingested the same amounts of carbohydrates in the three diets (21-22 g/kg P0.75/day). Dietary nitrogen content was similar (table 1). The ciliate population was improved with the inulin diet (fig. 1; table 2). With a mixed population, the Entodinium improved with the inulin diet (fig 1; table 2). With a mixed population, the Entodinium sp. genus was always predominant. Holotrich protozoa (mainly Isotricha) in the rumen of the conventional sheep represented 15 to 30% of the total ciliate biomass, indicating that they were able to metabolize these soluble sugars. We also observed that P. multivesiculatum can ferment cellulose and all the soluble carbohydrates proposed in these diets. However, Entodinium sp. development occurred mainly in the presence of the sugard produced during carbohydrate hydrolysis by other ciliates or bacteria. The highest organic matter digestibility, noted in faunated animals (table 3) was confirmed by the VFA concentration in the rumen (table 4). This could be explained either by an activation of bacterial metabolism due to predation or by the direct effect of ciliates on fermentations, or both. Modifications in the VFA composition varied with ciliate inoculation, showing that ciliate metabolism may vary with the nature of the energy in the diet or that the observed results depended on various opposite effects in which the intensity of each component was influenced by the diet. In general, the acetic acid molar proportion increased and propionic acid decreased when there was a considerable Entodinium sp. population. The effect on butyric acid was low with these diets. Higher ammonia and lactic acid concentrations were observed in the rumen of faunated than defaunated sheep, irrespective of the ciliate inoculum.     

Moderation of ruminal fermentation by ciliated protozoa in cattle fed a high-grain diet.

The objective of this study was to assess the influence of ciliated protozoa on ruminal fermentation in cattle fed high-grain diets. Six ruminally cannulated steers fed a corn-based grain diet (85% concentrate plus 15% alfalfa hay) at 12-h intervals were assigned randomly to two groups, ciliate free and faunated, in a crossover design. Defaunation was by ruminal emptying, omasal flushing, and treatment with sodium sulfosuccinate. Two to 3 weeks after defaunation, the ruminal contents of all steers were sampled before the morning feeding (0 h) and at 1, 2, 4, 6, 8, and 12 h after feeding to measure pH, analyze fermentation products, and monitor counts of ciliated protozoa and lactic acid-producing and -fermenting bacterial groups. Total numbers of ciliated protozoa in the faunated steers averaged 4.3 x 10(5)/g, and the protozoa consisted of nine genera. Ciliate-free steers had lower (P less than 0.01) ruminal pHs (pH 5.97) than faunated cattle (pH 6.45); however, the treatment-time interaction was not significant. Ruminal lactate and ammonia concentrations were similar in both groups. The total volatile fatty acid concentration was higher (P less than 0.05) in the ciliate-free steers than in the faunated steers and exhibited a treatment-time interaction (P less than 0.05). The acetate-to-propionate ratio was higher (P less than 0.05) in the faunated group than in the ciliate-free group and showed a treatment-time interaction (P less than 0.05). Total anaerobic bacterial counts were about fourfold higher in the ciliate-free group than in the faunated group.(ABSTRACT TRUNCATED AT 250 WORDS)