Klebsiella pneumoniaetype VI secretion system-mediated microbial competition is PhoPQ controlled and reactive oxygen species dependent

Abstract

ABSTRACTKlebsiella pneumoniaeis recognized as an urgent threat to human health due to the increasing isolation of multidrug resistant strains. Hypervirulent strains are a major concern due to their ability to cause life-threating infections in healthy hosts. The type VI secretion system (T6SS) is widely implicated in microbial antagonism, and it mediates interactions with host eukaryotic cells in some cases. In silico search for genes orthologous to T6SS component genes and T6SS effector genes across 700K. pneumoniaegenomes shows extensive diversity in T6SS genes across theK. pneumoniaespecies. Temperature, oxygen tension, pH, osmolarity, iron levels, and NaCl regulate the expression of the T6SS encoded by a hypervirulentK. pneumoniaestrain. Polymyxins and human defensin 3 also increase the activity of the T6SS. A screen for regulators governing T6SS uncover the correlation between the transcription of the T6SS and the ability to killE. coliprey. Whereas H-NS represses the T6SS, PhoPQ, PmrAB, Hfq, Fur, RpoS and RpoN positively regulate the T6SS.K. pneumoniaeT6SS mediates intra and inter species bacterial competition. This antagonism is only evident when the prey possess an active T6SS. The PhoPQ two component system governs the activation ofK. pneumoniaeT6SS in bacterial competitions. Mechanistically, PhoQ periplasmic domain, and the acid patch within, is essential to activateK. pneumoniaeT6SS.KlebsiellaT6SS also mediates anti-fungal competition. We have delineated the contribution of each of the individual VgrGs in microbial competition, and identified VgrG4 as a T6SS effector. Structurally, domain DUF2345 of VgrG4 is sufficient to intoxicate bacteria and yeast. ROS generation mediates the antibacterial effects of VgrG4, and the antitoxin Sel1E protects against the toxic activity of VgrG4. Our findings provide a better understanding of the regulation of the T6SS in bacterial competitions, and place ROS as an early event in microbial competition.AUTHOR SUMMARYKlebsiella pneumoniaehas been singled out as an “urgent threat to human health” due to extremely drug resistant strains. Numerous studies investigate the molecular mechanisms underlying antibiotic resistance inK. pneumoniae, while others dissect the virulence strategies of this pathogen. However, there is still limited knowledge on the fitness ofKlebsiellain the environment, and particularly the competition ofKlebsiellawith other species. Here, we demonstrated thatKlebsiellaexploits the type VI secretion system (T6SS) nanoweapon to kill bacterial competitors and fungi.K. pneumoniaeperceives T6SS attacks from bacterial competitors, resulting in retaliation against the aggressive cell. The perception of the attack involved the sensor PhoPQ and led to the up-regulation of the T6SS. We identified one of the toxins deployed by the T6SS to antagonize other microbes, and revealed howKlebsiellaprotects itself from this toxin. Our findings provide a better understanding of the T6SS role in microbial competition and uncover new aspects on how bacteria regulate T6SS-mediated microbial antagonism.