A method for discriminating between live and dead microbes in a sample, by discriminating between transcriptionally-active and inert microbial nucleic acid sequences in the sample. In particular, the method is based on the comparison of levels of nucleotide substitution in a sample cultured in presence of an RNA-labelling agent. Also, a diagnosis method of microbial infections in a subject; and methods of assessing the risk of contamination of a sample, implementing the method for discriminating between live and dead microbes in a sample.