The expression of aminoglycoside resistance genes in integron cassettes is not controlled by riboswitches

Abstract

Regulation of gene expression is a key factor influencing the success of antimicrobial resistance determinants. A variety of determinants conferring resistance against aminoglycosides (Ag) are commonly found in clinically relevant bacteria, but whether their expression is regulated or not is controversial. The expression of several Ag resistance genes has been reported to be controlled by a riboswitch mechanism encoded in a conserved sequence. Yet this sequence corresponds to the integration site of an integron, a genetic platform that recruits genes of different functions, making the presence of such a riboswitch counterintuitive. We provide, for the first time, experimental evidence against the existence of such Ag-sensing riboswitch. We first tried to reproduce the induction of the well characterized aacA5 gene using its native genetic environment, but were unsuccessful. We then broadened our approach and analyzed the inducibility of all AgR genes encoded in integrons against a variety of antibiotics. We could not observe biologically relevant induction rates for any gene in the presence of several aminoglycosides. Instead, unrelated antibiotics produced mild but consistently higher increases in expression, that were the result of pleiotropic effects. Our findings rule out the riboswitch control of aminoglycoside resistance genes in integrons.

The expression of integron arrays is shaped by the translation rate of cassettes

“…To assess if cassettes can modulate the expression of downstream genes in the array (i.e. : if they have polar effects on the array) we took advantage of the recently generated pMBA collection 20,21 . pMBA is a p15a replicon designed to provide cassettes with an appropriate genetic context: it contains a class 1 integron platform encoding the Pc, the Pint, and the attI site where cassettes are cloned in first position mimicking an integrase mediated attI x attC reaction.…”

“…An expression gradient is then generated: the first cassette displays the highest expression while expression gradually decreases for cassettes further away in the array. (B) 136 different antibiotic resistance cassettes were individually cloned in first position in pMBA vector 20,21 , allowing to study how each ARC affects the expression gradient.…”

Anaerobiosis modulates the performance of antimicrobial resistance genes inEnterobacterales

“…Plasmid-borne Mobile Integrons (MIs) carry ca. 200 antimicrobial resistance cassettes (ARCs) against various antibiotic families 5 , and are extremely prevalent among clinical isolates of Escherichia coli and Klebsiella pneumoniae 6,7 , pathogens that cause more than 1 million deaths every year 8 . In this work we show that AMR genes may confer different resistance levels in anaerobic conditions, which has potential clinical and eco-epidemiological consequences.…”

Identification of Promoter Activity in Gene-Less Cassettes fromVibrionaceaeSuperintegrons

“…Integrons are known for their key role in the rise and spread of antibiotic resistance (AR) genes. Carried on conjugative plasmids, m obile integrons (MIs) circulate among the most important Gram-negative pathogens, disseminating more than 170 resistance determinants against most antibiotics 4 . Yet, integrons are naturally found in almost 20% of bacterial genomes available in the databases.…”