| Staphylococcus aureus(S.aureus),widely found on the human body surface and the upper respiratory tract,especially the nasopharynx causes infections which associated with pneumonia.Since the contribution of penicillin in the 1940 s,ongoing ability to fleetly acquire drug-fast to antimicrobials has been a common trait of S.aureus.However,the abuse of antibiotics have led to the emergence mec A gene of drug resistant pathogens such as Methicillin-Resistant Staphylococcus aureus(MRSA),and caused MRSA to successfully encode penicillin-binding proteins(PBPs),prominently penicillin-binding protein 2a(PBP2a),its overproduction makes most of S.aureus acquired resistance to β-lactam antibiotics including methicillin.Although PBP2 a confers broad-spectrum resistance to β-lactams,some newer cephalosporins(β-lactam antibiotics)are effective against MRSA.Ceftaroline fosamil(CPT),as a fifth-generation cephalosporin,has high-affinity with PBP2 a and potent antibacterial activity against MRSA,which has novel structure prevents it from being degraded by some β-lactamase and distinguished from other β-lactams.Nevertheless,according to the antimicrobial susceptibility breakpoint specified by the Clinical and Laboratory Standards Association(CLSI),the minimum bacteriostatic concentration(MIC)of some MRSA isolates to CPT in clinical > 4 μg/m L,which is CPT-resistant.Therefore,the action to slow MRSA’s resistance to CPT is imminent.Combined natural plants with known β-lactams,is one strategy well-used to conquer the overflow of β-lactams resistance strains,which can restore the anti-MRSA activity of antibiotics or even reverse the drug-resistance of some MRSA isolates.In this study,we screened a small molecular compound that can enhance the anti-MRSA activity of CPT from approximately 300 natural compounds and found Corilagin(COR),which can synergize with CPT in vitro on MRSA resistant strain USA300 with a FICI of 0.156.COR can also synergize with 12 MRSA clinical strains and showed that COR,along with other β-lactams and Fosfomycin,also had a synergistic effect against S.aureus.We also proved that the combination of I/8 MIC(COR + CPT)can significantly inhibit the growth of USA300 and clinical strain 1B3 G.Then,by establishing a mouse pneumonia model,we verified the antibacterial effect of COR + CPT in the treatment of S.aureus infection in vivo.As a combination treatment group,it can significantly improve the survival rate of mice to 53%,and the lung damage of mice caused by S.aureus infection.At the same time,the clearance rate of bacteria in the lung shows a significant difference towards monotherapy.Comprehending the process of resistance mechanisms,as is known to all,can help prolong the life of current antibiotics and discover new targets,this helps to solve speedy drug resistance in clinical strains.We induced the resistance of MRSA bacteria to COR + CPT to explore the cause of the resistance mechanism of bacteria.The results showed that the combination of 1/128 MIC COR+ 1/4 MIC CPT could effectively alleviate the resistance of induced strains to antibiotics.Finally,we conducted proteomics analysis on strains 10th-COR + CPT(32 + 0.5)that have successfully induced resistance to COR + CPT,and found that 556 proteins had significant changes in the expression level,including 324 protein up-regulation,232 protein down-regulation.Through PPI network analysis combined with q PCR verification,it was found that the resistance of MRSA bacteria to COR + CPT may be related to the Eno,Ldh1 and Ldh2 proteins in the tricarboxylic acid cycle(TCA cycle);Dan K and Gro L proteins in Stress;Gap A1,Sdh A and Acn A proteins in TCA cycle — Stress.TCA cycle is an important metabolic pathway in the organism.Under external pressure,such as antibiotic stimulation,changes in microorganisms in the body require TCA cycle regulation.In addition to antibiotics,the accumulation of lactic acid produced by TCA cycle metabolism can also become a stimulus to activate the stress system in the bacteria to cope with the uncomfortable environment.However,our experiments prove that multiple TCA cycle and Stress related genes are down-regulated,so we infer that the inhibition of TCA cycle and Stress pathway may be the main mechanism of USA300 resistance to COR + CPT resistance.This study provides effective candidate compounds for CPT’s synergistic treatment plan,and a reliable theoretical basis for future CPT treatment of MRSA bacterial infections and delays MRSA resistance,and supplys potential targets for inhibiting MRSA resistance to COR + CPT. |
