Biology subjects

Nong, L.

Publications and source records attributed to Nong, L..

3 recordsLinked to original sources

bioRxiv2025

Driving factors for beta-lactam resistance gene amplification during de novo resistance evolution in E. coli

Long-term exposure of E. coli to non-lethal step-wise increasing concentrations of beta-lactam antibiotics induces high levels of resistance that can be accompanied by amplification of a chromosomal fragment around the ampC gene. We compared the amplification of the ampC fragment in the wildtype, an ampC knockout mutant, a mutant in which the ampC gene was replaced by a tetracycline resistance gene tet(B)and a strain in which the ampC has been translocated. When ampC was removed, no amplification occurred at the original ampC location, but DNA fragments were amplified around the genes coding for efflux pump AcrAB and the multiple antibiotic resistance operon MarRAB. When tet(B) replaced ampC, exposure to tetracycline induced amplification of comparable fragments, while exposure to amoxicillin induced duplication of a larger fragment elsewhere. When ampC was translocated, a fragment around it at the new location was amplified. The importance of the presence but not of the location within the chromosome of the resistance genes for the amplification process indicates that the mechanisms are neither gene nor location specific. Without the relatively efficient resistance gene ampC, duplication and amplification occur around acrAB and marRAB that code for amoxicillin and tetracycline resistance factors. These duplications and amplifications are prevented by ampC amplification.

microbiology↗

bioRxiv2024

Collateral sensitivity and cross resistance in six species of bacteria exposed to six classes of antibiotics

De novo resistance can be developed in bacteria because of exposure to sublethal concentrations of antibiotics. Once the strain has become resistant to an initial antibiotic, this can cause cross-resistance or collateral sensitivity to a second antimicrobial. Specific collateral sensitivity is rarely conserved across species because the mechanisms triggered in different microorganisms to resist antibiotics are often different. In this study, we explored which collateral sensitivity or cross resistance networks are present in six species of bacteria with induced de novo-resistance. These six species were induced to become resistant to amoxicillin/cefepime, enrofloxacin, kanamycin, tetracycline, erythromycin and chloramphenicol(1). After that, the collateral sensitivity and the cross-resistance networks were evaluated by measuring increase or decrease of MIC of thirteen antibiotics that are often used in the clinic. Collateral sensitivity for kanamycin occurred in five species of strains resistant to chloramphenicol and tetracycline and for {beta}-lactam in strains of five species resistant to kanamycin. Further genetic analysis clarified that fusA consistently mutated in five species of bacteria made de novo resistant against kanamycin, suggesting that fusA operates in parallel with the other mechanisms related to antimicrobial resistance Based on considerations of resistance, a treatment protocol starting with chloramphenicol/tetracycline, followed by kanamycin and ending with amoxicillin may eliminate bacteria that have developed resistance against the initial treatment.

microbiology↗

bioRxiv2024

Progression of ampC amplification during de novo amoxicillin resistance development in E. coli

Beta-lactam antibiotics are the most applied antimicrobials in human and veterinarian health care. Hence, beta-lactam resistance is a major health problem. Gene amplification of AmpC beta-lactamase is a main contributor to de novo {beta}-lactam resistance in E. coli. However, the time course of amplification and the accompanying DNA mutations are unclear. Here, we study the progression of ampC amplification and ampC promoter mutations in the evolution of resistance by stepwise increasing amoxicillin concentration. AmpC promoter mutations occur by day two, while the amplification by a factor of approximately eight occurs after more than six days of amoxicillin exposure. The combination of amplification and promoter mutations increase ampC mRNA level by an average factor of 200 after 22 days. An IS1 insertion was identified in the amplification junction, suggesting the amplification is facilitated by mobile genetic elements transposition. In order to identify the essential genes for ampC amplification, the chromosomal mutations of strains with induced amoxicillin resistance were compared a similarly evolved resistant {Delta}ampC knockout. The evolved {Delta}ampC contained several resistance mutations that were absent in the WT, which accumulated more mutations in stress response genes. The amoxicillin evolved {Delta}ampC does not show amplification of the fragment around the original ampC position but exhibits a large duplication or triplication at another position, suggesting selection of genes to amplify is essential for resistance adaption. IMPORTANCEAmoxicillin is one of the most used antimicrobial against bacterial infections. DNA fragments containing ampC are amplified upon prolonged and stepwise increasing exposure to amoxicillin. These ampC amplification fragments have been identified in extended-spectrum beta-lactamases (ESBLs) plasmids, which are considered the main cause of beta- lactam resistance. Understanding the progression of ampC amplification enables amoxicillin resistance prevention. In this study, we show the time course of two important factors for ampC transcription enhancement, ampC amplification and ampC promoter mutations, during de novo amoxicillin resistance evolution. We propose that the transposon IS1 contributes to the amplification and that the sigma factor 70 regulates ampC overexpression.

microbiology↗