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Appl Environ Microbiol. 1971 July; 22(1): 120-130
Copyright © 1971 American Society for Microbiology. All Rights Reserved.

Improved Microscopy of Mycoplasma In Vitro ¹

Departments of Pathology and Microbiology, State University of New York at Buffalo, Buffalo, New York 14214

ABSTRACT

Techniques were developed for continuous microscopic observation of mycoplasmata growing in vitro in Rose chambers by using an inverted phase microscope. The methods permitted direct microscopic observation of undisturbed growth of mycoplasmata in liquid medium. Inocula of mycoplasmata were passed through 0.22-µm filters before culture to provide a suspension of discrete particles. The sequential growth of Mycoplasma pneumoniae was followed from points or single straight lines, with development of branching, a net-like confluence of filaments, large bodies occurring in the center of developing colonies, and finally coccoid forms. Other species of Mycoplasma which did not attach as readily to glass could be observed also by inverted phase microscopy. Umbonation of colonies (a "friedegg" appearance) occurred in liquid medium, indicating that this appearance was not due simply to interaction with the agar medium, but may reflect a qualitative difference in growth patterns between center and periphery. For growth on solid medium, direct observation of colonies in uncovered plates of agar medium was made by using inverted phase microscopy. This was found helpful in detecting small colonies and in observing relationships between colonies.

¹ Presented in part at the Annual Meeting of the Tissue Culture Association in San Juan, Puerto Rico, 10-13 June 1968.