This chapter will deal mainly with the characteristics of bacteria from the rumen that have been successfully cultivated in the laboratory. For some ecosystems, particularly those dominated by slow-growing or specialized microorganisms, it has become clear that only a very small fraction (often <1%) of the total microbial diversity has been recovered by cultural methods (Amann et al., 1995) and that descriptions of the ecosystem based on the available isolated strains can be highly misleading. These discrepancies are apparent both from comparison of direct microscopic and culturable counts, and from direct analyses of ribosomal RNA sequence diversity. In the rumen, organisms surviving in significant numbers must have growth rates sufficient to counteract the constant dilution due to turnover of rumen contents, and there are indications that the discrepancies may be less extreme. Leedle et al (1982) found that the culturable count fluctuated with time after feeding between 14% and 74% of the direct microscopic count in the rumens of animals fed on two different diets. Since the viability of several rumen species is known to change upon starvation, the lower figure could partly reflect changes in the viability of known, culturable species. Thus it is to be hoped that the major obstacles to cultivation of the most numerous rumen bacteria have been overcome by the development of sufficiently rigorous anaerobic methods and of suitable isolation media. It remains likely, however, that some functionally important groups (e.g. obligate syntrophs) may not have been recovered; Mclnerney et al (1981) used co-culture with Desulfovibrio in the presence of sulphate to isolate a fatty acid-oxidizing bacterium similar to Syntrophomonas wolfei from bovine rumen contents.

The rumen bacteria

Regulation of feed intake rumen microbiology and fermentation nutrient absorption and splanchnic metabolism endogenous regulatory and pregnancy lactation tissue growth host resistance against parasites and pathogens.

Ruminant Physiology: Digestion, Metabolism, Growth and Reproduction

The use of chemically defined media is gaining popularity in some commercial fermentations, particularly for the preparation of biological products. Although these media are still not frequently developed for industrial processes, they do exhibit favorable characteristics at large scale that are not observed with traditional complex media. This review focuses on the application, development, and practical considerations, especially process economics, of fermentations in chemically defined media in an industrial environment.

Chemically defined media for commercial fermentations

https://people.uleth.ca/~selibl/Brentspapers/AnaerobeReview.pdf

The rumen: a unique source of enzymes for enhancing livestock production.

Polyphenolics and tannins effect on in vitro digestibility of selected Acacia species leaves

A completely defined growth medium has been developed to determine the nitrogen requirements for several species of ruminal bacteria, and has revealed two strains which are impaired in de novo biosynthesis of certain amino acids. Using NH4Cl as a sole nitrogen source, the medium supported growth of Butyrivibrio, Selenomonas, Prevotella and Streptococcus species. One strain of B. fibrisolvens (E14) and one strain of P. ruminicola (GA33) did not grow in the presence of NH4Cl until the medium was supplemented with amino acids or peptides. For B. fibrisolvens strain E14, methionine was identified as the specific growth-limiting amino acid although methionine alone did not support growth in the absence of NH4Cl. For P. ruminicola strain GA33, any individual amino acid other than methionine or cysteine could supplement the medium and support growth. Enzyme assays confirmed a lack of NADH and NADPH-dependent glutamate dehydrogenase (GDH) activities in this strain.

A defined medium for rumen bacteria and identification of strains impaired in de novo biosynthesis of certain amino acids

J.D. BROOKER, D.K. LUM, A.M. THOMSON AND H.M. WARD. 1995. A gene-targeting suicide vector for Streptococcus bovis has been constructed using the Escherichia coli/Streptococcus shuttle plasmid, pMU1328, and a region derived from the broad host-range, Gram-positive transposon, Tn916. This suicide plasmid replicates autonomously in E. coli, but not in Strep, bovis or Strep, bovis Tn916. Under positive selection, the plasmid was shown to integrate into Strep, bovis Tn916 chromosomal DNA at a frequency of 3 × 10-8cell-1and was stably maintained for at least 100 generations in the absence of selection. This is the first report of a recombination system in ruminal bacteria. The ability to target genes, knock out specific functions or introduce novel genes into these micro-organisms will allow ruminal species to be manipulated and may eventually lead to improved animal production.

A gene-targeting suicide vector for Streptococcus bovis

Streptococcus bovis strain SB3 was genetically marked by conjugal transfer of the tetracycline-resistant transposon, Tn916, from Enterococcus faecalis to Strep. bovis. The transposon was stable in the Strep. bovis chromosome in the presence or absence of tetracycline. Streptococcus bovis: Tn916 was introduced into the rumen of experimental sheep and was maintained for at least 76 d. The population was stable in the presence of a grain-based ration but rapidly declined when sheep were transferred to pasture. On return to the grain-based diet, the Strep. bovis: Tn916 population reappeared. These data demonstrate the potential of this technique in studies of microbial interactions in the rumen.

Use of transposon Tn916 as a genetic marker in the rumen

Ruminant production is a major focus of Australian agriculture. The ability of ruminant animals such as sheep and cattle to make productive use of low quality plant materials depends on the activity and efficiency of the anaerobic microbial population that resides in the rumen. Factors that affect ruminant production include the ability of cellulolytic microorganisms to digest plant structural polysaccharides (primarily cellulose and hemicellulose), the capacity of microorganisms to metabolise and detoxify otherwise inhibitory plant products and the efficiency of nitrogen utilisation by ruminal organisms. This review will consider some current Australian research programs aimed at improving ruminant production efficiency by genetic engineering of ruminal bacteria.

Improved animal production by genetic engineering of ruminal bacteria.

Butyrovibrio fibrisolvens strain E14 has an absolute requirement for methionine. Metabolism of L-[β- 14 C]-serine to methionine occurred in the methionine-independent B, fibrisolvens strain H17c but not in strain E14. The absolute requirement for methionine in strain E14 could be met by addition of S-adenosylmethionine to the medium, but incorporation was not due to the presence of free methionine in the S-adenosylmethionine preparation. The results show that B. fibrisolvens strain E14 is unable to synthesize methionine de novo, probably due to a lack of methionine synthase. Butyrivibrio fibrisolvens may also possess an alternative pathway of methionine biosynthesis from S-adenosylmethionine.

J.D. Brooker

Papers

A defined medium for rumen bacteria and identification of strains impaired in de novo biosynthesis of certain amino acids

A gene-targeting suicide vector for Streptococcus bovis

Use of transposon Tn916 as a genetic marker in the rumen

Improved animal production by genetic engineering of ruminal bacteria.

Lack of methionine biosynthesis de novo in Butyrivibrio fibrisolvens strain E14