Bacteriological Examination of Oral Material by a Differential Counting Technique

¹ Unilever Research Laboratory, Colworth House, Sharnbrook, Bedford, England

A technique has been developed to enable a differential bacterial count to be performed on small samples under conditions where an absolute count is not practicable. A concentrated inoculum was spread on the surface of a non-selective medium and incubated for a short time so that microcolonies developed. Impressions of these microcolonies were transferred to the surface of a glass microscope slide by chemical fixation, were stained, and the different types were counted under the microscope. Impressions of individual microcolonies were identified by their cellular and colonial appearance, as compared with similar preparations of known species.

The technique had advantages of ease and speed over conventional methods when more than one type of organism was to be counted, and it was not necessary to standardize the size of inoculum. The count was proportional, but could be made absolute if a separate count was performed on one of the major groups present. It was not satisfactory for mixtures of strict and facultative anaerobes or for organisms such as the Gram-negative rods whose cellular and colonial appearances were insufficiently characteristic.

Results of pilot experiments performed on oral material indicated wide proportional variations between different sites in the same mouth and at the same sites at different times. Examination of excavated carious material suggested a significant proportional increase in lactic acid bacteria toward the advancing edge of the lesion.