Further Studies on Polysaccharide Production by Bovine Rumen Bacteria

In recent years the iodophilic microorganisms of the rumen have attracted the interest of some ruminologists.Oxford (12) and Masson and Oxford (11) found that holotrich ciliates from the rumen of sheep were active in depositing intracellular polysaccharide granules.These workers have extracted the granules from the cells and found them to be similar to an amylopectin, the main component of which was believed to be glucose.Heald (9) has shown that the carbohydrate content of tureen microorganisms increases sharply after feeding, followed by a rapid decline.Elsden (6) has demonstrated the same phenomenon after feeding glucose to sheep.Doetsch et al. (4) reported that an iodine staining substance, designated ISS, was produced by mixed suspensions of rumen bacteria in vitro from a variety of carbohydrates, including maltose, cellobiose, glucose, and xylose.Finally, Robinson et al. (17) have further determined some of the conditions which govern the formation and utilization of ISS using an empirical method of estimation.This group has proposed that ISS formation represents one "of several normal processes characteristic to a majority of authentic rumen microorganisms.It was therefore suggested that ISS formation may be used as a criterion for estimating the "normalcy" of the physiological state of the bacterial flora of ruminants.The purpose of the work reported here is to characterize ISS chemically and to note further the effect of some conditions which might occur in the rumen upon the in vitro production of ISS with a more sensitive method of estimation. EXPERIMENTAL PROCEDURERumen liquor samples were obtained from a 7-year-old Jersey cow fitted with a permanent rumen fistula.The diet consisted of alfalfa hay and a 16% protein grain mixture.Cell suspensions were prepared by filtering rumen liquor through four layers of cheesecloth, the filtrate being centrifuged for 10 minutes at 771 × G.The resulting supernatant liquor was centrifuged at 12,500 × G for 10 minutes, the clear supernatant liquid decanted off, and the cells suspended in M/15 phosphate (pH 6.9) buffer as previously described (4).This buffered suspension was then centrifuged in 15-ml.conical centrifuge tubes for 2 minutes at 1,200 × G.This last centrifuging removed almost all of the residual plant fragments and protozoa, the bacteria remaining in suspension.This suspension was standardized (4), flushed with nitrogen, and incubated at 39 ° C. for 11/2 hours to obtain utilization of stored ISS which might be present in the cells.