Microniches in biofilm depth are hot-spots for antibiotic resistance acquisition in response to in situ stress

Abstract

Class 1 integrons play a major role in antibiotic resistance dissemination among Gram-negative bacteria. They are genetic platforms able to capture, exchange and express antibiotic resistance gene cassettes. The integron integrase, whose expression is regulated by the bacterial SOS response, is the key element of the integron catalyzing insertion/excision/shuffling of gene cassettes. We previously demonstrated that the basal level of integrase expression and in consequence, its activity, is increased via the starvation-induced stringent response in the biofilm population. However, biofilms are heterogeneous environments where bacteria are under various physiological states. Here we thus analyzed at the bacterial level, the SOS response and integrase expression within the biofilm, using confocal microscopy and flow cytometry. We showed that in the absence of exogenous stress, only a small number of bacteria (~ 1%) located in the depth of the biofilm induce the SOS-response leading to a high level of integrase expression, through both a stringent response-dependent and -independent manner. Our results thus indicate that few bacteria located in microniches of the biofilm depth undergo sufficient endogenous stress to promote the acquisition of antibiotic resistance, forming a reservoir of bacteria ready to rapidly resist antibiotic treatments.

Intermittent antibiotic treatment of bacterial biofilms favors the rapid evolution of resistance

“…The capacity of structured biofilm environments to maintain high cost mutations was observed in several biofilm evolution studies (Ahmed et al, 2020;Ahmed et al, 2018;France et al, 2019;Santos-Lopez et al, 2019), in which rare beneficial mutations could be protected from competitions in biofilm microniches, increasing their odds to be selected and propagated. In support of this hypothesis, it was recently shown that the SOS response induced in E. coli biofilm microniches leads to increased rate of mutagenesis (Tlili et al, 2021) and that beneficial resistance mutations had reduced probability to be lost by genetic drift in structured environments (Chakraborty et al, 2021b). Alternatively, due to diffusion limitation, the concentration of antibiotics could drop more slowly in biofilms than in planktonic conditions.…”