respread, that is, bacterial cells were moved around allowing single cells in some colony to form their own new colonies. If colonies contained resistant bacterial cells before entering into contact with the phage virus, one would expect that some of these cells would form new resistant colonies on the respread dish and so to find a higher number of surviving bacteria there. When both plates were incubated for growth, there were actually as much as 50 times greater number of bacterial colonies on the respread dish. This showed that bacterial mutations to virus resistance had randomly occurred during the first incubation. Once again, the mutations occurred before selection was applied. More recently, the results of Luria and Delbrück were questioned by Cairns and others, who studied mutations in sugar metabolism as a form of environmental stress. Some scientists suggest that this result may have been caused by selection for gene amplification and/or a higher mutation rate in cells unable to divide. Others have defended the research and propose mechanisms which account for the observed phenomena consistent with adaptive mutagenesis. This distribution appears to have been first determined by Haldane. An unpublished manuscript was discovered in 1991 at University College London describing this distribution. The derivation is different but the results are difficult to compute without the use of a computer. Description of the test A small number of cells are used to inoculate parallel cultures in a non-selective medium. The cultures are grown to saturation to obtain equal cell densities. The cells are plated onto selective media to obtain the number of mutants (r). Dilutions are plated onto rich medium to calculate the total number of viable cells ( Nt ). The number of mutants that appear in the saturated culture is a measure of both the mutation rate and when the mutants arise during the growth of the culture: mutants appearing early in the growth of the culture will propagate many more mutants than those that arise later during growth. These factors cause the frequency ( r / Nt ) to vary greatly, even if the number of mutational events ( m ) is the same. Frequency is not a sufficiently accurate measure of mutation and the mutation rate (m / Nt) should always be calculated. The estimation of the mutation rate (μ) is complex. Luria and Delbruck estimated this parameter from the mean of the distribution but this