CRISPR-Cas 9 to generate deletions in the organism Enterococcus faecium

Abstract

Enterococci became a major hospital-acquired infection. vanA and vanB genes make Enterococcus faecium resistant to many antibiotics. AvrA and other non-transposable antibiotic genes do not spread between bacteria. To determine the effect of the avrA gene on E. faecium's antibiotic resistance, bioinformatics tools like BLAST, Predict Protein, and ProteInfer were used, and the Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-Cas9 genome editing tool was used to generate targeted mutants. Minimum Inhibitory Concentrations (MIC) of vancomycin, ampicillin, and daptomycin were also checked for E.faecium susceptibility. BLAST, Predict Protein, and ProteInfer found that E. faecium has an avrA gene, Beta Pleated sheet, and a structural role in its cell wall and membrane. CRISPR-Cas9 failed to delete the avrA gene, although MIC analysis revealed E. faecium's resistance to ampicillin (MIC: >128μg/mL) compared to vancomycin (MIC: 1μg/mL) and daptomycin (MIC: 4μg/mL).