RelQ-mediated alarmone signaling regulates growth, sporulation, and stress-induced biofilm formation inClostridioides difficile

Abstract

ABSTRACTThe bacterial stringent response (SR) is a conserved transcriptional reprogramming pathway mediated by the nucleotide signaling alarmones, (pp)pGpp. The SR has been implicated in antibiotic survival inClostridioides difficile, a biofilm- and spore-forming pathogen that causes resilient, highly recurrentC. difficileinfections. The role of the SR in other processes and the effectors by which it regulatesC. difficilephysiology are unknown.C. difficileRelQ is a clostridial alarmone synthetase. Deletion ofrelQdysregulatesC. difficilegrowth in unstressed conditions, affects susceptibility to antibiotic and oxidative stressors, and drastically reduces biofilm formation. While wild-typeC. difficiledisplays increased biofilm formation in the presence of sub-lethal stress, the ΔrelQstrain cannot upregulate biofilm production in response to stress. Deletion ofrelQslows spore accumulation in planktonic cultures but accelerates it in biofilms. This work establishes biofilm formation and sporulation as alarmone-mediated processes inC. difficileand reveals the importance of RelQ in stress-induced biofilm regulation.