Background: Fluoroquinolone antibiotics are important bactericidal drugs that combat the bacterial infections globally. The emergence of bacterial resistance to this antibiotic class is an overwhelming serious problem in Egypt. Aim of the study: To determine the bacterial genes responsible for the the occurrence resistance to fluoroquinolone antibiotics in different hospitals in Egypt. Methodology: Samples were withdrawn one hundred urinary tract infection (UTI) patients as well as on hundred lower respiratory infection (LRTI) patients in different hospitals in Egypt who were previously administrated different dosage regimens of fluoroquinolone antibiotics. Using antibiotic susceptibility tests along with the bacterial growth spectrophotometric techniques at 600 nm wavelength, the leading bacterial resistant isolates were identified. The major genes responsible for this pattern of resistance were determined exploiting the third generation high throughput sequencing technique using PacBio RS II sequencer with catalog number PacBio RS II manufactured in Pacific Bio-sciences company, USA. Results: The major bacterial isolates along with their fluoroquinolone resistant genes were detected to be Pseudomonas aeruginosa strain GPS containing GPS resistant gene encoding DNA gyrase subunit B, Serratia Marcescens strain GSM containing GSM resistant gene encoding DNA gyrase subunit B and Streptococcus pneumonae strain TAP comprising TAP resistant gene encoding DNA gyrase subunit A. Conclusion: The current study resulted in the discovery of novel fluoroquinolone antibiotic resistant genes in different hospital in Egypt.