Kan, J., B. Flood, J.P. McCrow, J.S. Kim, L. Tan, and K.H. Nealson. 2011. Journal of Microbiological Methods 86:62–68.
One of the big operational problems facing laboratories today is the ability to rapidly distinguish between strains of bacteria that, while physiologically distinct, are nearly impossible to separate based on 16S rRNA gene sequence differences. Here we demonstrate that ITS–DGGE provides a convenient approach to distinguishing between closely related strains of Shewanella, some of which were impossible to separate and identify by 16 rRNA gene sequence alone. Examined Shewanella genomes contain 8–11 copies of rrn (ribosomal RNA gene) operons, and variable size and sequence of 16S–23S ITS (intergenic transcribed spacer) regions which result in distinct ITS–DGGE profiles. Phylogenetic constructions based on ITS are congruent with the genomic trees generated from concatenated core genes as well as with those based on conserved indels, suggesting that ITS patterns appear to be linked with evolutionary lineages and physiology. In addition, three new Shewanella strains (MFC 2, MFC 6, and MFC 14) were isolated from microbial fuel cells enriched from wastewater sludge and identified by ITS–DGGE. Subsequent physiological and electrochemical studies of the three isolates confirmed that each strain is phenotypically/genotypically distinct. Thus, this study validates ITS–DGGE as a quick fingerprint approach to identifying and distinguishing between closely related but novel Shewanellaecotypes.