Release of Genetic Transforming Agent From Pneumococcal Cultures During Growth and Disintegration

Abstract

Genetic transformations of pneumococcus mediated by streptomycin-induced lysates were studied to gain some insight into the nature of freshly released transforming principle, and the influence of the physiologic state of the donor population on the transformation process. It was found that streptomycin could make the DNA of sensitive cells available for transformation of other cells. Living cultures of pneumococcus growing exponentially in ordinary media were also found to discharge significant quantities of genetically active DNA. Such cultures, not treated with any drug, showed no evidence of concomitant cell disintegration or death. Both single markers and small linkage groups could be transferred in transformations mediated by drug-induced lysates and by filtrates of living cultures. The quantity of DNA liberated is small (less than 0.1 µg per ml), but these transformations are at least as efficient as transformations mediated by purified DNA, when compared on the basis of total DNA available. Up to 1 per cent of the cells in an average recipient culture can be transformed by a small quantity of culture fluid. Both in drug-treated and in untreated cultures the amount of transforming activity increased and then decreased during growth of the culture. Although the source of transforming DNA in growing cultures could not be established, the decline in the transforming activity of aging drug-treated or untreated cultures was attributed to the presence of deoxyribonuclease. The release of this nuclease by pneumococcal cultures midway in exponential growth is sufficient to result in a mild degradation of the low concentration of freshly released transforming agent present. Maximal release of active transforming agent by a living culture coincided in time with the development of maximal receptivity to exogenous DNA by that culture. As a result, recombinants could be recovered from appropriately genetically marked strains growing in each other's presence. In view of these results, it seems possible that DNA-mediated transformations might provide, or might have provided, a mechanism of genetic recombination in nature for some bacterial species in which sexual mechanisms may not be available.

Gene Expression Analysis of the Streptococcus pneumoniae Competence Regulons by Use of DNA Microarrays

“…lytA, the major autolysin gene, has long been known to be dispensable for transformation (22), casting doubt on any important role in competence or in cell wall remodeling for DNA uptake. However, it has also long been known that extracellular DNA is found in competent cultures (18), and the regulation of lytA does suggest a possible role of this gene in its release. Two other genes in the recA operon, cinA and dinF, are induced at competence but are dispensable for transformation (17).…”

Characterization of the Streptococcus pneumoniae NADH oxidase that is required for infection

“…S. pneumoniae human clinical isolate 0100993 (serotype 3) was used as the parent strain for the construction of a nox allelic-replacement mutant. S. pneumoniae strain R6 is a single-colony isolate of strain R36A obtained from Avery's strain D39 (Avery et al, 1944 ;Ottolenghi & Hotchkiss, 1962). S. pneumoniae strains were grown at 37 mC with 5 % CO # in Todd-Hewitt broth with 0n5 % yeast extract or on tryptic soy agar plates containing 5 % sheep blood, supplemented with 1 µg erythromycin ml − " when appropriate.…”

Modulation of Competence for Genetic Transformation in Streptococcus pneumoniae

“…Various laboratories have reported critical cell densities varying between 2 x lo6 ml-l and 2 x lo8 ml-l (Ottolenghi & Hotchkiss, 1962;Tomasz & Hotchkiss, 1964;Tomasz, 1966;Gurney & Fox, 1968;Porter & Guild, 1969;Morrison, 1977). Competence appears in cultures of pneumococci growing in appropriate media at a specific cell density.…”