Residues contribute to water pollution and pose a challenge to microalgal populations because numerous contaminants are toxic to microalgae, even in the micromolar range. Adaptation of microalgae from herbicide sensitivity to herbicide resistance was analysed by an ecological genetic approach, using the unicellular alga Dictyosphaerium chlorelloides (Chlorophyceae) as an experimental model. A dose-effect study showed that the Malthusian parameter under conditions of r selection in an uncrowded environment and the carrying capacity under conditions of K selection in an crowded environment were both restricted even by low concentrations (< 1 muM) of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) herbicide. When a culture was treated with 50 muM DCMU, it cleared after a few days, as a result of destruction of sensitive cells by the herbicide. However, after further incubation for several days, the culture sometimes regained colour, owing to the growth of cell variants resistant to the herbicide. A fluctuation analysis was carried out to distinguish between (1) herbicide-resistant cells arising by direct and specific acquired adaptation in response to the herbicide and (2) herbicide-resistant cells arising by rare spontaneous mutations occurring randomly during replication of organisms prior to the incorporation of the herbicide. The fluctuation analysis unequivocally demonstrated that DCMU is not facilitating the development of DCMU-resistant cells; rather, we found that DCMU-resistant cells occur spontaneously by mutation in nonselective conditions prior to the incorporation of the herbicide (preselective mutations). The rate of spontaneous mutation from DCMU sensitivity to DCMU resistance was similar to 2.2 X 10(-6) mutants per cell division. Mutation was recurrent from a normal wild-type DCMU-sensitive allele to an DCMU-resistant allele, but such herbicide-resistant alleles were detrimental in terms of fitness in the absence of the herbicide. A competition experiment between wild-type DCMU-sensitive cells and DCMU-resistant mutants showed that, in small populations, the DCMU-resistant mutants are driven to extinction. The resistant variants are maintained in the absence of the herbicide as the result of a balance between new resistant cells arising by rare spontaneous mutation and resistant cells eliminated by natural selection. In our case, the average frequency of DCMU-resistant mutants in the absence of the DCMU is about five DCMU-resistant mutants per million cells. The results of our experimental model suggest that spontaneous mutation from herbicide sensitivity to herbicide resistance is sufficient in itself to assure the survival of microalgae populations in herbicide-contaminated environments when the population size is large.
