| The discovery and widespread use of antibiotics has not only saved countless lives,but has also taken a leading role in the prevention and control of bacterial diseases in livestock and poultry farming,However,the unregulated use of antibiotics has led to increasing antibiotic resistance(AMR).Polymyxins are considered to be the antibiotics of last resort against most gram-negative infections.However,since 2015,our scientists have reported for the first time a polymyxin resistance gene mcr-1located on a plasmid,which makes clinical treatment more difficult.Vancomycin has been considered a last resort for the treatment of severe MRSA and other drug-resistant gram-positive bacteria,However,vancomycin-resistant Staphylococcus aureus has also been clinically isolated,so new strategies are urgently needed to address these problems.It is worth noting that due to the advantages of short development time and high safety of "old drugs for new use" drugs,they are now widely used.Fenticonazole nitrate is clinically used to fight fungal infection by using inhibiting the synthesis of ergosterol in fungal cell membranes.However,the potential of fenticonazole nitrate against bacteria remains unclear.Based on this,this study mainly carried out the following work:1.Combination screening based on whole bacteria and study of antibacterial activity and mechanism of action of fenticonazole nitrate against Escherichia coliUsing multi-drug resistant E.coli 42 containing the mcr-1 gene,a preliminary screening of small molecule compounds from the FDA drug library,represented by fenticonazole nitrate,which can reduce the use of polymyxin E by 8-32 fold,was performed.The results of growth curve and time sterilization curve proved that the combination of fenticonazole nitrate and polymyxin E had a significant synergistic antibacterial effect,and the combination of the two could interfere with the biofilm.Mechanistic investigation showed that fenticonazole nitrate binds directly to MCR-1 protein,thereby inhibiting MCR-1 activity;Fenticonazole nitrate significantly enhanced the ability of polymyxin E to disrupt the bacterial outer membrane of Escherichia coli,but had no effect on the bacterial inner membrane;Further investigation revealed that fenticonazole nitrate could disrupt membrane potential,and the combination with polymyxin E could inhibit ATP production and thus exert antibacterial activity.In addition,the outer membrane component LPS and the efflux pump help protect the bacteria from damage caused by fenticonazole nitrate.A mouse model of E.coli 42 intraperitoneal infection showed that the combination of fenticonazole nitrate and polymyxin E increased the survival rate of mice by 70%.In addition,the combination therapy also reduced the levels of IL-1β,TNF-α,ALT,AST and CK.2.Study on the antibacterial activity and mechanism of Fenticonazole nitrate against Staphylococcus aureusGrowth curve and bactericidal curve test proved that fenticonazole nitrate had good antibacterial activity against Staphylococcus aureus.Crystal violet staining and CLSM method showed that fenticonazole nitrate could significantly inhibit the formation of biofilm of MRSA.Mechanism research confirmed that Fenticonazole nitrate induced membrane damage by damaging membrane permeability,membrane potential and increasing membrane fluidity of Staphylococcus aureus.The model of mellonella mellonalis infection showed that fenticonazole nitrate could play a significant antibacterial role in vivo.In conclusion,Fenticonazole nitrate can enhance the antibacterial activity of Colistin in vitro and in vivo,and it additionally has a desirable antibacterial effect on Staphylococcus aureus.These results provide preliminary research basis and lead compounds for the clinical development of new drug strategies against drug-resistant bacterial infections,and provide new solutions to alleviate the increasingly serious bacterial resistance crisis. |
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