Role of LsrR in the regulation of antibiotic sensitivity in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is a specific group of extraintestinal pathogenic E. coli that causes a variety of extraintestinal diseases in chickens, ducks, pigeons, turkeys, and other avian species. These diseases lead to significant economic losses in the poultry industry worldwide. However, owing to excessive use of antibiotics in the treatment of infectious diseases, bacteria have developed antibiotic resistance. The development of multidrug efflux pumps is one important bacterial antibiotic resistance mechanism. A multidrug efflux pump, MdtH, which belongs to the major facilitator superfamily of transporters, confers resistance to quinolone antibiotics such as norfloxacin and enoxacin. LsrR regulates hundreds of genes that participate in myriad biological processes, including mobility, biofilm formation, and antibiotic susceptibility. However, whether LsrR regulates mdtH transcription and then affects bacterial resistance to various antibiotics in APEC has not been reported. In the present study, the lsrR mutant was constructed from its parent strain APECX40 (WT), and high-throughput sequencing was performed to analyze the transcriptional profile of the WT and mutant XY10 strains. The results showed that lsrR gene deletion upregulated the mdtH transcript level. Furthermore, we also constructed the lsrR- and mdtH-overexpressing strains and performed antimicrobial susceptibility testing, antibacterial activity assays, real-time reverse transcription PCR, and electrophoretic mobility shift assays to investigate the molecular regulatory mechanism of LsrR on the MdtH multidrug efflux pump. The lsrR mutation and the mdtH-overexpressing strain decreased cell susceptibility to norfloxacin, ofloxacin, ciprofloxacin, and tetracycline by upregulating mdtH transcript levels. In addition, the lsrR-overexpressing strain increased cell susceptibility to norfloxacin, ofloxacin, ciprofloxacin, and tetracycline by downregulating mdtH transcript levels. Electrophoretic mobility shift assays indicated that LsrR directly binds to the mdtH promoter. Therefore, this study is the first to demonstrate that LsrR inhibits mdtH transcription by directly binding to its promoter region. This action subsequently increases susceptibility to the aforementioned four antibiotics in APECX40.

Yu L ，Li W ，Li Q ，Chen X ，Ni J ，Shang F ，Xue T ... -  《POULTRY SCIENCE》

Role of McbR in the regulation of antibiotic susceptibility in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) causes a variety of bacterial infectious diseases known as avian colibacillosis leading to significant economic losses in the poultry industry worldwide and restricting the development of the poultry industry. The development of efflux pumps is one important bacterial antibiotic resistance mechanism. Efflux pumps are capable of extruding a wide range of antibiotics out of the cytoplasm of some bacterial species, including β-lactams, polymyxins, tetracyclines, fluoroquinolones, aminoglycosides, novobiocin, nalidixic acid, and fosfomycin. In the present study, we constructed the mcbR mutant and the mcbR-overexpressing strain of E. coli strain APECX40 and performed antimicrobial susceptibility testing, antibacterial activity assays, real-time reverse transcription PCR, and electrophoretic mobility shift assays (EMSA) to investigate the molecular regulatory mechanism of McbR on the genes encoding efflux pumps. Our results showed that McbR positively regulates cell susceptibility to 12 antibiotics, including clindamycin, lincomycin, cefotaxime, cefalexin, doxycycline, tetracycline, gentamicin, kanamycin, norfloxacin, ofloxacin, erythromycin, and rifampicin by activating the transcription of acrAB, acrD, emrD, and mdtD (P < 0.01). Additionally, EMSA indicated that McbR specifically binds to the promoter regions of acrAB, acrD, acrR, emrD, and mdtD. This study suggests that, in APECX40, McbR plays an important role in the regulation of bacterial susceptibility by directly activating the transcription of efflux pumps genes.

Yu L ，Li W ，Liu Z ，Yu J ，Wang W ，Shang F ，Xue T ... -  《-》

Regulatory Role of the Two-Component System BasSR in the Expression of the EmrD Multidrug Efflux in Escherichia coli.

Yu L ，Li W ，Xue M ，Li J ，Chen X ，Ni J ，Shang F ，Xue T ... -  《-》

Role of ArcA in the regulation of antibiotic sensitivity in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is one of the common extraintestinal infectious disease pathogens in chickens, geese, and other birds, inducing serious impediments to the development of the poultry industry. Hence, investigating how bacteria regulate themselves amidst different challenging conditions is immense essential in prevention and treatment for bacterial pathogen infections. The ArcA regulatory factor has been reported to regulate oxygen availability in strains, but its role in regulation of antibiotics resistance in APEC is unclear. This study delved into understanding how ArcA regulates antibiotic resistance in APEC. An E. coli APEC40 arcA knockout strain was constructed, and the regulatory mechanism of arcA on APEC antibiotic susceptibility was identified by drug sensitivity test, colony counting assay, real-time quantitative PCR, β-galactosidase assays and electrophoretic mobility shift assay (EMSA). The results showed that ArcA directly binds to the promoter region of the outer membrane protein OmpC/OmpW and regulates bacterial susceptibility to kanamycin and penicillin G. At the same time, the double knockout of ompW and ompW/arcA resulted in an increase in resistance to kanamycin compared to the deletion of the arcA gene. This outcome provided experimental proof suggesting that the outer membrane protein OmpW could serve as a crucial pathway for the ingress of kanamycin into cells. These results confirmed the important regulatory role of ArcA transcription factors under APEC antibiotic stress.

Ma K ，Chinelo OR ，Gu M ，Kong F ，Jiang Y ，Wang H ，Xue T ... -  《POULTRY SCIENCE》

Autoinducer2 affects trimethoprim-sulfamethoxazole susceptibility in avian pathogenic Escherichia coli dependent on the folate synthesis-associate pathway.

Avian pathogenic Escherichia coli (APEC) causes airsacculitis, polyserositis, septicemia, and other mainly extraintestinal diseases in chickens, ducks, geese, pigeons, and other avian species, and is responsible for great economic losses in the avian industry. The autoinducer 2 (AI-2) quorum sensing system is widely present in many species of gram-negative and gram-positive bacteria and has been proposed to be involved in interspecies communication. In clinical APEC strains, whether or not AI-2 affects the expression of antibiotic-related genes has not been reported. In this study, we have reported that exogenous AI-2 increase the susceptibility of APEC strains to trimethoprim-sulfamethoxazole (SXT) in a folate synthesis-dependent pathway but not in the LsrR-dependent manner. Our results further explained that exogenous AI-2 can down regulate the transcription of the folate synthetase encoding genes folA and folC, and the folate synthesis-related genes luxS, metE, and metH. Gel shift assays confirmed that LsrR, the AI-2 receptor, did not bind to the promoters of folA and folC, suggesting that exogenous AI-2 might influence folate metabolism by a feedback inhibition effect but not in the LsrR-dependent pathway. This study might provide further information in the search for potential drug targets for prophylaxis of avian colibacillosis and for auxiliary antibiotics in the treatment of avian colibacillosis.

Yu L ，Li W ，Zhang M ，Cui Y ，Chen X ，Ni J ，Yu L ，Shang F ，Xue T ... -  《MicrobiologyOpen》