Exploring Essential Genes in Staphylococcus aureus JE2-MRSA by: Transposition-Directed Insertion Site Sequencing (TraDIS)

Methicillin resistant Staphylococcus aureus (MRSA) is a major public health concerns worldwide causing a variety of clinical diseases including sepsis and abscess/skin infections. Transposon Directed Insertion-site Sequencing (TraDIS) is a genome-wide mutagenesis method, which allows simultaneous assaying of large transposon mutant libraries for exploring gene significance, functionality, and inter-genetic relationships in bacteria. In the current study, our goals were i) to construct a high-density Tn-mutants library in the JE2-MRSA strain of this species, and ii) and to identify the essential genes for in growth in vitro and for in vivo infection of skin and organs in a mouse model. We transducted the Tn-5 transposon/transposase complex into the strain by phage ø11 using an improved transduction protocol to create a high-density transposon library of 10^9 mutants. After the analysis by Bio-Tradis analysis pipeline, we found that the library containing 315,400 unique insertion sites (i.e. one insertion every 9 base pairs). We identified 213 essential genes with zero transposon insertion, suggesting that these genes were essential for growth on BHI agar medium, representing 8% of the total JE2 MRSA genome. Studies to identify essential genes for in vivo infection is ongoing and will be reported.