To investigate the in vivo and in vitro antibacterial activity of doxycycline combined with levofloxacin on carbapenem-resistant Klebsiella pneumoniae (CRKP).

Eight clinically isolated CRKP strains were collected in Yichun People's Hospital, and the minimum inhibitory concentration (MIC) of 8 CRKPs was determined using microbroth dilution method in vitro. The antibacterial effect of the two drugs was judged by checkerboard dilution method. The time-sterilization curve was further applied to evaluate the combined bactericidal effect of the two drugs. Crystal violet staining semi-quantitative biofilm method was used to determine the inhibitory effect and elimination effect of the combination of two drugs on biofilm formation. In vivo, a model of acute lung infection of mice was established by nasal instillation of CRKP bacteria, and the mental state of mice was recorded. The bacterial load of lung tissues, the levels of serum inflammatory factor C-reactive protein (CRP), IL-6 and pathological morphological changes were evaluated and analyzed.

The MIC of doxycycline to 8 CRKP strains was 4~256 ug·mL^-1, levofloxacin to 8 strains of CRKP MIC of 16~256 ug·mL^-1. The checkerboard method showed that doxycycline combined with levofloxacin had an 80% synergistic or additive effect on 8 CRKP strains. The time-sterilization curve showed that after the treatment of doxycycline combined with levofloxacin on CRKP2 and CRKP7 for 24 hours, the number of bacteria decreased by ≥2 lg CFU·mL^-1 compared with the initial bacterial number, showing bacteriostatic or bactericidal effects. The results of crystal violet staining showed that the combination of doxycycline and levofloxacin had inhibitory and destructive effects on CRKP biofilms. Animal experiments showed that compared with the model group, the combined group mice had a good mental state, and the loading levels of lung tissue, C-reactive protein (CRP) and IL-6 significantly reduced (P<0.001). The results of HE staining showed that the alveolar structure was clear, the inflammatory cells reduced, and the hyperemia area reduced in the drug administration group compared with the model group, and the differences were statistically significant (P<0.001).

In vitro experiments show that doxycycline combined with levofloxacin has a synergistic antibacterial or bactericidal effect on CRKP, and in vivo experiments show that doxycycline combined with levofloxacin can reduce inflammatory indexes, effectively treat bacterial infections in lung tissue, exerting good antibacterial effect in vivo.