Use of Recombinant DNA Technology| Microbiology

Genetic modification of rumen bacteria by using rDNA technology may improve the rate or extent of lignocellulolysis in the animal. The task of developing effective rDNA technology in rumen bacteria, particularly for improving lignocellulose digestion is complex.

After the rate limiting steps have been defined, it is necessary to identify and clone suitable heterologous genes for insertion into a rumen bacterium. It is then necessary to develop effective vectors and transformation system to develop methods to monitor both the survival of the gene in the host cell and its survival in the rumen and finally to measure its effect on in vivo digestibility of lignocellulose.

Besides rDNA technology, genetic mutation is another technique, which produces greater quantities of rate-limiting enzymes. Mutagenesis in Ruminococcus albus cause regulation of cellulose enzymes.

Genomic DNA libraries have been constructed of fragments from the cellulolytic rumen bacteria Bacteriodes succinogens, Ruminococcus albus, Butyrivibrio fibrisolvens and others. Gene cloning in rumen bacteria may provide valuable insights into our understanding of the process operating in rumen.

Rumen Microflora:

It is very much similar to the chemostat or a fermenter in which cellulosic biomass in the form of fodder is digested. Actually, it varies in size from cow to sheep wherein it has a capacity of approximately 30 litres to 6 litres, respectively.

The temperature remains 30°C and pH 6.5. The anaerobic atmosphere persists in rumen. It runs continuously. Since rumen contains microorganisms (symbionts and parasites), the fodder is digested in this region.

The ruminants animals do not have cellulolytic enzymes to digest cellulose but mammals can metabolise them due to presence of cellulose decomposers namely, Fibrobacter succinogenes, Butyrivibrous fibrisolvens, Ruminobacter albus, Clostridium lochheadii; starch decomposers such as Bacteriodes ruminicola, Ruminobacter amylophilus, Selenomonas luminatium, Succinomonas amyolyticus.

Strep­tococcus, bovis; lactate decom­posers e.g. Selenomonas lactilytica and Megasphaera elsdenii, pectin decomposers (Lachnospora multiparus) and lastly methanogens, Methanobrevibacter ruminantium and Methanomicrobium mobile.

The rumen has about 10²/ml microbial population of anaer­obes. The concept of microbial consortium or biofilm is an ex­ample wherein combined activity of several bacterial species and protozoa occur. For instance, a fermentative bacterium produced hydrogen gas and another species of methanogens uses it for production of methane gas.

Rumen also consists of ciliate protozoal fauna of obligate anaerobes. They eat and ingest rumen bacteria, hence maintain the bacterial densities. Rumen also possesses some anaerobic fungi, which ferment cellulose and volatile fatty acids. They also play a role in degradation of other polysaccharides including a partial degradation of lignin, hemicellulose and pectin.

The microflora and their action depend upon the type of fodder (carbohydrate, lipids, proteins, etc.) available in rumen. For example, if an animal is fed legume hay rich in pectin, the bacterial genes (Lachnospira multiparus) will be more active.

Cer­tain rumen bacteria pro­duce alcohol as a fermen­tation production, yet ethanol rarely accumulated in the rumen due to the fact that it can be fermented to acetate and H₂. This hy­drogen never accumulates because it is quickly used to reduce CO₂ to CH₄ by methanogens.

Acetate is not converted to CH₄ in the rumen because the re­tention time is too short for development of acetolastic methanogens, which typically grow very slowly.

Sometimes a change in the microbial composition of the rumen causes illness or even death of the animal. If an animal is given grain diet, a high density and population of Streptococcus bovis occurs. S. bovis produces lactate from starchy grains, which acidifies the rumen called acidosis (Fig. 34.2.).

Certain fungi (anaerobic) are present in the symbiotic gut community. The uniflagellates spores of these fungi have been noted as motile spores move around the rumen until they come in contact with a piece of swallowed vegetational matter. They then grow long projections called rhizoids, into the food. The rhizoids produce enzymes that digest the cell wall of the food and disintegrate in to pulp.

Eventually, a sporangium develops and more flagel­late spores are released. These microorganisms also seem able to use N₂, which has been secreted or ingested into the rumen in the form of urea. They use this to synthesize proteins. Rumen fungi also degrade cellulose, hemicellulose, lignin and pectate substances of woody-herbaceous plants used as fodder for ani­mals.