The Regulatory Mechanism Of AI-2 Quorum Sensing Of Avian Pathogenic Escherichia Coli Chlortetracycline Drug Resistance

In China and the world,Avian Pathogenic Escherichia coli is one of the most common pathogenic bacteria in livestock industry,especially in the chicken industry.Its occurrence and widespread distribution result in high morbidity and mortality which not only seriously restrict the development of the poultry industry but also give a high expense.Up to now,the traditional method to prevent and cure avian Escherichia coli disease still is antibiotics.However,with the long-term used and abusive of the antibiotics,the drug resistance is increasing and the multidrug resistant strains appears.Therefore,it is particularly significant to search for new drug targets.As we all known,quorum sensing system is a bacterial control system which widely exists in gram negative and positive bacteria.Moveover,Autoinducer-2(AI-2)is the main signal molecule that was used to exchange inner information within species of Escherichia coli.But in the multi-resistant strains,the topic whether the AI-2 has an association with pathogenicity is rarely researched in academia.This experiment revealed the effect of AI-2 on Avian pathogenic Escherichia coli chlortetracycline resistance.And then it explains the regulation mechanism of AI-2 on chlortetracycline resistant gene tetA.To test the drug resistance on avian pathogenic Escherichia coli isolates,the method of two times dilution was used to assess Pathogenic Escherichia coli strains of APEC 28,APEC 32 and APEC 35 on drug resistance of chlortetracycline.The result showed that the minimum inhibitory concentration of APEC 28,APEC 32 and APEC 35 on Chlortetracycline was 120 ?g/ mL,which proves all these three strains are chlortetracycline resistant bacteria.Refer to the tetracycline resistant genes tetA,tetB and tetC in NCBI,and the primer sequence was designed to amplify the resistance gene by PCR amplification.In the result,the resistance gene of these three strains of avian Escherichia coli was tetA.In order to study the effect of exogenous AI-2 on the three strains of Escherichia coli,using colony counting method(CFU)to studying,the results showed that exogenous AI-2,AEPC 28 and APEC 32 on the drug resistance of aureomycin increased,on the contrary APEC 35 resistance reduced.Using the method of transcriptional real-time fluorescence quantitative PCR(RT-qPCR)technology to detect chlortetracycline resistant gene transcription level of tetA,under the pressure of Chlortetracycline,exogenous AI-2 lead to the gene transcription levels ofAPEC 28,APEC 32,APEC35 lsrR were increased by 1.5 times and 1.4 times and 1.5times;in the meantime,it also made the transcription level of APEC 28 tetA gene increased2.3 times,the transcription level of APEC 32 of the tetA gene increased 1.8 times,and APEC 35 of the gene transcription level of APEC tetA decreased 1.5 times.For further study,we applied gel retardation test to detect the relationship between tetA gene and LsrR protein which shows the added exogenous AI-2 did not affect tetA gene through LsrR protein.In this study,it has been shown that the quorum sensing molecule AI-2 has a regulatory effect on the tetA resistant gene,but its regulation is not carried out by the receptor LsrR of AI-2 intracellular.The further research of this study may provide new drug targets and treatment for the treatment of pathogenic Escherichia coli in resistant avian diseases.