Seasonal changes in the cecal microflora of the high-arctic Svalbard reindeer (Rangifer tarandus platyrhynchus).

The dominant cecal bacteria in the high-arctic Svalbard reindeer were characterized, their population densities were estimated, and cecal pH was determined in summer, when food quality and availability is good, and in winter, when it is very poor. In summer the total culturable viable bacterial population was (8.9 +/- 5.3) X 10(8) cells ml-1, whereas in winter it was (1.5 +/- 0.7) X 10(8) cells ml-1, representing a decrease to 17% of the summer population density. Of the dominant species of cultured bacteria, Butyrivibrio fibrisolvens represented 23% in summer and 18% in winter. Streptococcus bovis represented 17% in summer and 5% in winter. Bacteroides ruminicola represented 10% in summer and 26% in winter. In summer and winter, respectively, the proportion of the viable population showing the following activities was as follows: fiber digestion, 36 and 48%; cellulolysis, 10 and 6%; xylanolysis, 33 and 48%; and starch utilization, 77 and 71%. The most abundant cellulolytic species in summer was Butyrivibrio fibrisolvens, representing 62% of the total cellulolytic population, and in winter it was Ruminococcus albus, representing 80% of the total cellulolytic population. The most abundant xylanolytic species in summer was Butyrivibrio fibrisolvens, and in winter it was Bacteroides ruminicola, representing 59 and 54% of the xylanolytic isolates in summer and winter, respectively. The cecal bacterial of the Svalbard reindeer have the ability to digest starch and the major structural carbohydrates of the diet that are not digested in the rumen. The cecum in these animals has the potential to contribute very substantially to the digestion of the available plant material in both summer and winter.     

Seasonal changes in the ruminal microflora of the high-arctic Svalbard reindeer (Rangifer tarandus platyrhynchus).

The dominant rumen bacteria in high-arctic Svalbard reindeer were characterized, their population densities were estimated, and ruminal pH was determined in summer, when food quality and availability are good, and in winter, when they are poor. In summer the total cultured viable population density was (2.09 +/- 1.26) X 10(10) cells ml-1, whereas in winter it was (0.36 +/- 0.29) X 10(10) cells ml-1, representing a decrease to 17% of the summer population density. On culture, Butyrivibrio fibrisolvens represented 22% of the bacterial population in summer and 30% in winter. Streptococcus bovis represented 17% of the bacterial population in summer but only 4% in winter. Methanogenic bacteria were present at 10(4) cells ml-1 in summer and 10(7) cells ml-1 in winter. In summer and winter, respectively, the proportions of the viable population showing the following activities were as follows: starch utilization, 68 and 63%; fiber digestion, 31 and 74%; cellulolysis, 15 and 35%; xylanolysis, 30 and 58%; proteolysis, 51 and 28%; ureolysis, 40 and 54%; and lactate utilization, 13 and 4%. The principal cellulolytic bacterium was B. fibrisolvens, which represented 66 and 52% of the cellulolytic population in summer and winter, respectively. The results indicate that the microflora of the rumen of Svalbard reindeer is highly effective in fiber digestion and nitrogen metabolism, allowing the animals to survive under the austere nutritional conditions typical of their high-arctic habitat.     

Seasonal changes in the ruminal microflora of the high-arctic Svalbard reindeer (Rangifer tarandus platyrhynchus)

The dominant rumen bacteria in high-arctic Svalbard reindeer were characterized, their population densities were estimated, and ruminal pH was determined in summer, when food quality and availability are good, and in winter, when they are poor. In summer the total cultured viable population density was (2.09 +/- 1.26) X 10(10) cells ml-1, whereas in winter it was (0.36 +/- 0.29) X 10(10) cells ml-1, representing a decrease to 17% of the summer population density. On culture, Butyrivibrio fibrisolvens represented 22% of the bacterial population in summer and 30% in winter. Streptococcus bovis represented 17% of the bacterial population in summer but only 4% in winter. Methanogenic bacteria were present at 10(4) cells ml-1 in summer and 10(7) cells ml-1 in winter. In summer and winter, respectively, the proportions of the viable population showing the following activities were as follows: starch utilization, 68 and 63%; fiber digestion, 31 and 74%; cellulolysis, 15 and 35%; xylanolysis, 30 and 58%; proteolysis, 51 and 28%; ureolysis, 40 and 54%; and lactate utilization, 13 and 4%. The principal cellulolytic bacterium was B. fibrisolvens, which represented 66 and 52% of the cellulolytic population in summer and winter, respectively. The results indicate that the microflora of the rumen of Svalbard reindeer is highly effective in fiber digestion and nitrogen metabolism, allowing the animals to survive under the austere nutritional conditions typical of their high-arctic habitat.     

Characterization of the cecal bacteria of normal pigs.

One hundred ninety-two isolates from cecal contents of three normal weaned pigs were obtained by means of anaerobic roll tube methods and were characterized. Seventy-eight percent of the isolates were gram-negative. The most numerous species isolated from each of the pigs was Bacteroides ruminicola. This species accounted for 35% of the isolates that were characterized, and Selenomonas ruminantium accounted for 21% of the isolates. Other gram-negative bacteria isolated from all three pigs were Butyrivibrio fibrisolvens (6.0%) and Bacteroides uniformis (3.0%); predominant gram-positive isolates were Lactobacillus acidophilus (7.6%), Peptostreptococcus productus (3.0%), and Eubacterium aerofaciens (2.5%). The other 42 isolates were placed in 14 other species, and 5 additional isolates that did not fit well into existing species were not placed taxonomically. Fifteen of the isolates (representing nine species) produced urease.     

Interaction of ruminal bacteria in the production and utilization of maltooligosaccharides from starch.

The degradation and utilization of starch by three amylolytic and one nonamylolytic species of ruminal bacteria were studied. Pure cultures of Streptococcus bovis JB1, Butyrivibrio fibrisolvens 49, and Bacteroides ruminicola D31d rapidly hydrolyzed starch and maltooligosaccharides accumulated. The major starch hydrolytic products detected in S. bovis cultures were glucose, maltose, maltotriose, and maltotetraose. In addition to these oligosaccharides, B. fibrisolvens cultures produced maltopentaose. The products of starch hydrolysis by B. ruminicola were even more complex, yielding glucose through maltotetraose, maltohexaose, and maltoheptaose but little maltopentaose. Selenomonas ruminantium HD4 grew poorly on starch, digested only a small portion of the available substrate, and generated no detectable oligosaccharides as a result of cultivation in starch containing medium. S. ruminantium was able to grow on a mixture of maltooligosaccharides and utilize those of lower degree (less than 10) of polymerization. A coculture system containing S. ruminantium as a dextrin-utilizing species and each of the three amylolytic bacteria was developed to test whether the products of starch hydrolysis were available for crossfeeding to another ruminal bacterium. Cocultures of S. ruminantium and S. bovis contained large numbers of S. bovis but relatively few S. ruminantium and exhibited little change in the pattern of maltooligosaccharides observed for pure cultures of S. bovis. In contrast, S. ruminantium was able to compete with B. fibrisolvens and B. ruminicola for these growth substrates. When grown with B. fibrisolvens, S. ruminantium grew to high numbers and maltooligosaccharides accumulated to a much lesser degree than in cultures of B. fibrisolvens alone. S. ruminantium-B. ruminicola cultures contained large numbers of both species, and maltooligosaccharides never accumulated in these cocultures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enumeration and presumptive identification of some functional groups of bacteria in the rumen of dairy cows fed grass silage-based diets

Abstract Samples of rumen ingesta from two rumen-fistulated dairy cows fed grass silage-based diets were examined for numbers and types of bacteria that developed colonies on rumen fluid-agar media designated to support the growth of (a) a wide range of species, (b) cellulolytic bacteria, (c) lactate-fermenting bacteria, (d) non-fermentative bacteria. The most numerous species was Bacteroides ruminicola followed by Butyrivibrio fibrisolvens . The most abundant cellulolytic species were Eubacterium cellulosolvens and Ruminococcus flavefaciens. Megasphaera elsdenii and Selenomonas ruminantium were important lactate fermenters but an unidentified bacterium that grew poorly on maintenance medium was by far the most numerous among bacteria isolated from lactate-containing medium. One strain remained sufficiently viable to show that it fermented lactate to propionate and acetate.     

Interaction of ruminal bacteria in the production and utilization of maltooligosaccharides from starch.

The degradation and utilization of starch by three amylolytic and one nonamylolytic species of ruminal bacteria were studied. Pure cultures of Streptococcus bovis JB1, Butyrivibrio fibrisolvens 49, and Bacteroides ruminicola D31d rapidly hydrolyzed starch and maltooligosaccharides accumulated. The major starch hydrolytic products detected in S. bovis cultures were glucose, maltose, maltotriose, and maltotetraose. In addition to these oligosaccharides, B. fibrisolvens cultures produced maltopentaose. The products of starch hydrolysis by B. ruminicola were even more complex, yielding glucose through maltotetraose, maltohexaose, and maltoheptaose but little maltopentaose. Selenomonas ruminantium HD4 grew poorly on starch, digested only a small portion of the available substrate, and generated no detectable oligosaccharides as a result of cultivation in starch containing medium. S. ruminantium was able to grow on a mixture of maltooligosaccharides and utilize those of lower degree (less than 10) of polymerization. A coculture system containing S. ruminantium as a dextrin-utilizing species and each of the three amylolytic bacteria was developed to test whether the products of starch hydrolysis were available for crossfeeding to another ruminal bacterium. Cocultures of S. ruminantium and S. bovis contained large numbers of S. bovis but relatively few S. ruminantium and exhibited little change in the pattern of maltooligosaccharides observed for pure cultures of S. bovis. In contrast, S. ruminantium was able to compete with B. fibrisolvens and B. ruminicola for these growth substrates. When grown with B. fibrisolvens, S. ruminantium grew to high numbers and maltooligosaccharides accumulated to a much lesser degree than in cultures of B. fibrisolvens alone. S. ruminantium-B. ruminicola cultures contained large numbers of both species, and maltooligosaccharides never accumulated in these cocultures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Some rumen bacteria degrading fructan

Degradation of fructan obtained from timothy ( Phleum pratense L.) by the following six species of bacteria isolated from sheep rumen was studied: Streptococcus bovis, Bacteroides ruminicola, Selenomonas ruminantium, Butyrivibrio fibrisolvens, Treponema bryantii and Treponema saccharophilum. The enzymatic activity of the bacteria was analysed by TLC. The highest activity was found in whole cells and in the strains B. fibrisolvens No. 3 and T. saccharophilum S.     

Interactions between rumen amylolytic and lactate-utilizing bacteria in growth on starch

The growth and metabolism of the rumen amylolytic bacteria Streptococcus bovis, Butyrivibrio fibrisolvens and Bacteroides ruminicola, growing in pure cultures and co-cultures with the rumen lactilytic bacteria Megasphaera elsdenii and Veillonella alcalescens were followed. The interaction of amylolytic bacteria with V. alcalescens represents a simple food chain. The interaction with M. elsdenii is more complex, since there is a simultaneous competition for products of the starch degradation.     

Interactions between rumen amylolytic and lactate-utilizing bacteria in growth on starch

The growth and metabolism of the rumen amylolytic bacteria Streptococcus bovis, Butyrivibrio fibrisolvens and Bacteroides ruminicola , growing in pure cultures and co-cultures with the rumen lactilytic bacteria Megasphera elsdenii and Veillonella alcalescens were followed. The interaction of amylolytic bacteria with V. alcalescens represents a simple food chain. The interaction with M. elsdenii is more complex, since there is a simultaneous competition for products of the starch degradation.     

Amylolytic activity of selected species of ruminal bacteria.

A variety of species of ruminal bacteria were screened for the ability to grow in starch-containing medium and produce amylase. Of those tested, the highest levels of amylase were produced by Streptococcus bovis JB1 and Ruminobacter amylophilus H18. Other strains that grew well on starch and produced amylase included Butyrivibrio fibrisolvens A38 and 49 and Bacteroides ruminicola 23 and B14. Varying the carbohydrate source provided for growth resulted in changes in the growth rate and level of amylase produced by these strains. All strains grew rapidly in starch-containing medium, and the rates of growth were generally more rapid than those observed for maltose-grown cultures. For S. bovis JB1, B. ruminicola 23 and B14, and B. fibrisolvens 49 and A38, amylase was produced when growth was on maltose or starch, but this activity was greatly reduced in glucose-grown cultures. The distribution of amylolytic activity between cellular and extracellular fractions was sometimes affected by the carbohydrate provided for growth. If S. bovis JB1 and B. fibrisolvens 49 were grown on starch, amylase was largely associated with cell pellets; however, if grown on maltose these strains produced activities that were almost entirely present in the extracellular fluid fractions. Although not as dramatic, a similar shift in the location of amylase activities was noted for the two B. ruminicola strains when grown on the same substrates. Growth on maltose or starch had little influence on either the predominantly cell-associated activity of B. fibrisolvens A38 or the activity of R. amylophilus H18, which was equally divided between cell pellet and extracellular fluid fractions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Amylolytic activity of selected species of ruminal bacteria

A variety of species of ruminal bacteria were screened for the ability to grow in starch-containing medium and produce amylase. Of those tested, the highest levels of amylase were produced by Streptococcus bovis JB1 and Ruminobacter amylophilus H18. Other strains that grew well on starch and produced amylase included Butyrivibrio fibrisolvens A38 and 49 and Bacteroides ruminicola 23 and B14. Varying the carbohydrate source provided for growth resulted in changes in the growth rate and level of amylase produced by these strains. All strains grew rapidly in starch-containing medium, and the rates of growth were generally more rapid than those observed for maltose-grown cultures. For S. bovis JB1, B. ruminicola 23 and B14, and B. fibrisolvens 49 and A38, amylase was produced when growth was on maltose or starch, but this activity was greatly reduced in glucose-grown cultures. The distribution of amylolytic activity between cellular and extracellular fractions was sometimes affected by the carbohydrate provided for growth. If S. bovis JB1 and B. fibrisolvens 49 were grown on starch, amylase was largely associated with cell pellets; however, if grown on maltose these strains produced activities that were almost entirely present in the extracellular fluid fractions. Although not as dramatic, a similar shift in the location of amylase activities was noted for the two B. ruminicola strains when grown on the same substrates. Growth on maltose or starch had little influence on either the predominantly cell-associated activity of B. fibrisolvens A38 or the activity of R. amylophilus H18, which was equally divided between cell pellet and extracellular fluid fractions.(ABSTRACT TRUNCATED AT 250 WORDS)     

A survey of peptidase activity in rumen bacteria.

Twenty-nine strains of 14 species of rumen bacteria were screened for their ability to hydrolyse Ala2, Ala5, GlyArg-4-methoxy-2-naphthylamide (GlyArg-MNA) and Leu-MNA. Several species, notably Megasphaera elsdenii, were active against Ala2, and a smaller number, including Bacteroides ruminicola, Butyrivibrio fibrisolvens, Ruminococcus flavefaciens, Lachnospira multipara and Ruminobacter amylophilus, broke down Ala5. Streptococcus bovis had an exceptionally high leucine arylamidase activity. However, only Ba. ruminicola hydrolysed GlyArg-MNA. Further investigation revealed that only Ba. ruminicola and Bu. fibrisolvens hydrolysed Ala5 to Ala3 and Ala2, with little ALa4 being produced, in a manner similar to rumen fluid. The activity of Ba. ruminicola against synthetic peptidase substrates, including GlyArg-MNA, LysAla-MNA, ArgArg-MNA, GlyPro-MNA, LeuVal-MNA, and Ala3-p-nitroanilide, was similar to that of rumen fluid, whereas the activity of Bu. fibrisolvens was quite different. Since the main mechanism by which peptides are broken down in the rumen is similar to dipeptidyl aminopeptidase type I, for which GlyArg-MNA is a diagnostic substrate, it was concluded that Ba. ruminicola was the most important single species in peptide breakdown in the rumen.     

Carbon Dioxide Requirement of Various Species of Rumen Bacteria

The carbon dioxide requirement of 32 strains of rumen bacteria, representing 11 different species, was studied in detail. Increasing concentrations of CO(2) were added as NaHCO(3) to a specially prepared CO(2)-free medium which was tubed and inoculated under nitrogen. Prior depletion of CO(2) in the inoculum was found to affect the level of requirement; however, the complexity and buffering capacity of the medium did not appear to be involved. An absolute requirement for CO(2) was observed for eight strains of Bacteroides ruminicola, three strains of Bacteroides succinogenes, four strains of Ruminococcus flavefaciens, two strains of Lachnospira multiparus, one strain of Succinimonas amylolytica, and two strains of Butyrivibrio fibrisolvens. Inconsistent growth responses were obtained in CO(2)-free media with one strain each of B. fibrisolvens, Ruminococcus albus, and Selenomonas ruminantium. Growth of six additional strains of B. fibrisolvens, and single strains of Eubacterium ruminantium and Succinivibrio dextrinosolvens was markedly increased or stimulated by increasing concentrations of CO(2). Peptostreptococcus elsdenii B159 was the only organism tested which appeared to have no requirement, either absolute or partial, for CO(2). Higher concentrations of CO(2) were required for the initiation of growth, as well as for optimal growth, by those species which produce succinic acid as one of their primary end products.     

Utilization of xylooligosaccharides by selected ruminal bacteria.

The ability of ruminal bacteria to utilize xylooligosaccharides was examined. Xylooligosaccharides were prepared by partially hydrolyzing oat spelt xylan in phosphoric acid. This substrate solution was added (0.2%, wt/vol) to a complex medium containing yeast extract and Trypticase that was inoculated with individual species of ruminal bacteria, and growth and utilization were monitored over time. All of the xylanolytic bacteria examined were able to utilize this oligosaccharide mixture as a growth substrate. Butyrivibrio fibrisolvens, Eubacterium ruminantium, and Ruminococcus albus used xylooligosaccharides and whole, unhydrolyzed xylan to similar extents, while Prevotella ruminicola used twice as much xylooligosaccharides as xylan (76 versus 34%). Strains of Selenomonas ruminantium were the only nonxylanolytic species that were able to grow on xylooligosaccharides. The ability of individual S. ruminantium strains to utilize xylooligosaccharides was correlated with the presence of xylosidase and arabinosidases activities.     

Effect of pH on the efficiency of growth by pure cultures of rumen bacteria in continuous culture.

A total of 10 strains of rumen bacteria, Selenomonas ruminantium HD4, Megasphaera elsdenii B159, Butyrivibrio fibrisolvens A38, Streptococcus bovis JB1, Lactobacillus vitulinus GA1, Bacteroides ruminicola B14, B. ruminicola GA33, Ruminococcus albus 7, Ruminococcus flavefaciens C94, and Bacteroides succinogenes S85, were grown in energy-limiteH of the medium reservoir was lowered approximately 0.3 pH units, and the energy source concentration remaining in the culture vessel, optical density, cell mass, and pH were determined. A low pH appeared to have a detrimental effect on cell yields. Large variations were seen among strains in both the magnitude of yield depressions at lower pH values and in the pH at which the culture washed out. Lactate analysis indicated ta are discussed in relation to the effect of pH on the efficiency of protein synthesis in the rumen and rumen microbial ecology.     

Microbiology and ration digestibility in the hindgut of the ovine

Contents of the terminal ilea, ceca-proximal colons, and terminal recta were obtained from nine sheep, three of which were fed 100% orchardgrass hay, three of which were fed 60% cracked corn-40% orchardgrass hay, and three of which were fed 80% cracked corn-20% orchardgrass hay. The digestibility of dry matter in the ceca was greatest when the all-hay diet was fed, whereas the percentage of cellulose digestion in the ceca increased with increasing levels of concentrate. For all diets, the total volatile fatty acid concentrations were higher in the ceca than in the other two sites. The cecal pH levels decreased with increased corn levels in the diet. The total microbiol numbers per gram of ileal and cecal contents increased in response to feeding of concentrate; however, across all diets, the ileal counts were 8% or less of the cecal counts. In contrast, the cellulolytic microbial numbers in the ilea were 50% or more of those in the ceca and were highest with the all-hay diet. Both bacterial and end product concentrations in the ceca were equivalent to those occurring in rumina. A total of 16 cellulolytic cultures were isolated and characterized from ileal and cecal contents of animals fed all three diets. Seven gram-negative-rod-shaped organisms were identified as Butyrivibrio fibrisolvens, whose capacity to digest cellulose exceeded that of several rumen strains. The guanine-plus-cytosine content of the DNA of one strain of B. fibrisolvens was 38.8 mol%, compared with the only previously reported value for this species of 41.2 mol%.(ABSTRACT TRUNCATED AT 250 WORDS)     

Expression of a modified Neocallimastix patriciarum xylanase in Butyrivibrio fibrisolvens digests more fibre but cannot effectively compete with highly fibrolytic bacteria in the rumen

AIMS: This study investigated the competitive abilities of two Neocallimastix patriciarum-derived xylanases constructs in Butyrivibrio fibrisolvens H17c (xynA and pUMSX) and their ability to compete in vivo. METHODS AND RESULTS: The digestibility of neutral detergent fibre (NDF) increased during co-culture of xynA or pUMSX and weakly cellulolytic, but not with highly cellulolytic, ruminococci. Competition studies among xynA, pUMSX and cellulolytic consortia demonstrated that xynA was the fittest. XynA did not persist at high levels in the rumen and was undetectable after 22 days. CONCLUSION: The construction of recombinant xylanolytic B. fibrisolvens does improve the digestibility of fibre above that of the native, but digestibility is still less than that of the most potent fibre digesters such as ruminococci. SIGNIFICANCE AND IMPACT OF THE STUDY: Fibre digestion may be improved by genetic manipulation of ruminal bacteria but ecological parameters, such as persistence in vivo and the niche of the organism, must be taken into account.     

Microbiology and ration digestibility in the hindgut of the ovine.

Contents of the terminal ilea, ceca-proximal colons, and terminal recta were obtained from nine sheep, three of which were fed 100% orchardgrass hay, three of which were fed 60% cracked corn-40% orchardgrass hay, and three of which were fed 80% cracked corn-20% orchardgrass hay. The digestibility of dry matter in the ceca was greatest when the all-hay diet was fed, whereas the percentage of cellulose digestion in the ceca increased with increasing levels of concentrate. For all diets, the total volatile fatty acid concentrations were higher in the ceca than in the other two sites. The cecal pH levels decreased with increased corn levels in the diet. The total microbiol numbers per gram of ileal and cecal contents increased in response to feeding of concentrate; however, across all diets, the ileal counts were 8% or less of the cecal counts. In contrast, the cellulolytic microbial numbers in the ilea were 50% or more of those in the ceca and were highest with the all-hay diet. Both bacterial and end product concentrations in the ceca were equivalent to those occurring in rumina. A total of 16 cellulolytic cultures were isolated and characterized from ileal and cecal contents of animals fed all three diets. Seven gram-negative-rod-shaped organisms were identified as Butyrivibrio fibrisolvens, whose capacity to digest cellulose exceeded that of several rumen strains. The guanine-plus-cytosine content of the DNA of one strain of B. fibrisolvens was 38.8 mol%, compared with the only previously reported value for this species of 41.2 mol%.(ABSTRACT TRUNCATED AT 250 WORDS)     

The Effect of Monensin on Pure and Mixed Cultures of Rumen Bacteria

The antibiotic monensin was added to pure cultures of Bacteroides ruminicola, Selenomonas ruminantium, Anaerovibrio lipolytica and Megasphaera elsdenii. These organisms, representing succinate- and propionate-producing rumen bacteria, were not affected by monensin up to 10 μg/ml. Methanobacterium ruminantium was slightly inhibited by monensin, Butyrivibrio fibrisolvens, Ruminococcus albus and Streptococcus bovis were inhibited to differing extents by monensin at concentrations between 0.1 and 10 μg/ml. Bacteroides succinogenes was inhibited at first by monensin at >0.5 μg/ml but after a prolonged lag phase adapted to grow in the presence of monensin at concentrations below 5 μg/ml.
Monensin (1 μg/ml) almost completely stopped the digestion of chopped straw and dewaxed cotton fibres by rumen contents incubated in vitro. The digestion of grass and powdered filter paper was not significantly reduced under these conditions, but when the concentration of monensin was increased to between 3 and 5 μg/ml, the digestion of these substrates was reduced.