Evaluation of subsampling and fixation procedures used for counting rumen protozoa.

Total numbers of protozoa can be significantly lower in rumen fluid than in whole rumen contents, depending on the time of sampling and the procedure used to separate the fluid and solid fractions. Moreover, generic distribution in rumen fluid was significantly affected in all cases tested. The percentage of Entodinium spp. increased, whereas percentages of Diplodinium spp. and Ophryoscolex spp. decreased. Microscopic observation of fresh and fixed rumen contents did not indicate any marked attachment of protozoa to particulate matter. In addition, dilution of whole rumen contents with water, 5 mM sucrose, or 0.1% Tween 80 before fixation did not affect total numbers or generic composition of protozoa. It was thus concluded that attachment to feed particles is probably not a problem in counting procedures. Blending of whole rumen contents to facilitate subsampling caused a decrease in numbers of protozoa. The concentration of formaldehyde used for preservation of rumen contents, 4, 10, or 18.5%, did not affect the total count.