Overview report on the application of double digest selective label (DDSL) bacteria genotyping technique for identification of strains and certification of commercially used bacteria.

The objective of this report is to demonstrate the potential of the proposed simple typing technique, double digest selective label (DDSL), which was initially developed to identify clinical isolates of Pseudomonas aeruginosa, for other bacterial species including Salmonella enterica, Clostridium difficile, Staphylococcus aureus, and Bacillus subtilis. The technique is based on digestion of bacterial genomic DNA with two restriction enzymes and simultaneous labeling fragments with biotinylated deoxycytidine triphosphate in fill-in reaction by Taq polymerase. The number and distribution of generated DNA fragments can be optimized by selecting restriction enzymes. DDSL is fast, reproducible, cost effective and sufficiently discriminatory typing method applicable for identification of bacterial strains at laboratories having no access to expensive sequencing equipment and with limited funding and lack of skilled personnel. Data concerning the potential of the technique for short-term epidemiological surveillance and bacterial strain certification are presented and discussed. Multiple locus variable number tandem repeat analysis performed on our set of Clostridium difficile isolates did not demonstrate sufficient discriminatory power both with TR6 and TR10 loci on a set of 24 isolates. In contrast, the DDSL analysis resolved all isolates into individual strains. © King Abdulaziz City for Science and Technology 2020.