Evaluation of a new method for the preparation of homogeneous mycelial suspensions.

Guidry, D. J. (Louisiana State University, New Orleans) and G. H. Trelles. Evaluation of a new method for the preparation of homogeneous mycelial suspensions. J. Bacteriol. 83:53-60. 1962.-An all-glass tissue homogenizer (providing both conical and cylindrical grinding surfaces) was employed in the preparation of homogeneous mycelial suspensions. Pellets of mycelium from constant shake liquid culture were submitted to grinding for various periods of time. The effect of grinding time on optical density of the homogenate, hyphal fragment size, and viability of the fungus was determined. A grinding time of 4 min was found to yield suspensions of mycelium in which 89% of the hyphal fragments contained four or less than four cells. Optical density was used as the basis for both preparation of standard samples of inoculum and measurement of growth. Preparation of a light transmittance curve facilitates the preparation of suspensions of any desired concentration. Although plate colony counts on samples from the same flask showed satisfactory agreement, there was appreciable variation in counts on samples from different flasks. The variation in growth from flasks receiving equal samples of inoculum was also determined. Results indicate that, at concentrations of mycelium equal to an optical density of 0.301, a difference of 15% or more in the optical density of two samples (10 flasks per sample) can be accepted as significant. Some of the factors influencing variation in viability of inoculum as well as variation in flask growth have been determined. Ways in which experimental errors can be kept to a minimum are discussed.