Listeria monocytogenes relAandhptMutants Are Impaired in Surface-Attached Growth and Virulence

Abstract

We describe here the identification and characterization of two Listeria monocytogenes (Tn917-LTV3) relA and hpt transposon insertion mutants that were impaired in growth after attachment to a model surface. Both mutants were unable to accumulate (p)ppGpp in response to amino acid starvation, whereas the wild-type strain accumulated (p)ppGpp within 30 min of stress induction. The induction of transcription of the relA gene after adhesion was demonstrated, suggesting that the ability to mount a stringent response and undergo physiological adaptation to nutrient deprivation is essential for the subsequent growth of the adhered bacteria. The absence of (p)ppGpp in the hpt mutant, which is blocked in the purine salvage pathway, is curious and suggests that a functional purine salvage pathway is required for the biosynthesis of (p)ppGpp. Both mutants were avirulent in a murine model of listeriosis, indicating an essential role for the stringent response in the survival and growth of L. monocytogenes in the host. Taken as a whole, this study provides new information on the role of the stringent response and the physiological adaptation of L. monocytogenes for biofilm growth and pathogenesis.The ubiquitous gram-positive pathogen L. monocytogenes is responsible for the clinical syndromes of listeriosis in humans and animals (27, 28). The principle mode of transmission of the organism to humans is believed to be the consumption of contaminated food (44). Normally, the consumption of foodborne L. monocytogenes does not result in overt gastrointestinal disease. Rather, in serious cases, listeriosis presents as bacteremia, meningitis, or miscarriage (30).The ability of L. monocytogenes to adhere to and colonize surfaces during food preparation and storage is important in the contamination of food products prior to consumption. While much is known about the virulence of L. monocytogenes (reviewed in references 7 and 24), detailed knowledge about the mechanisms of attachment and proliferation of L. monocytogenes on surfaces is scant. Previously, the adhesion of L. monocytogenes to plant surfaces has been shown to be a consequence of hydrophobic bonds between the plant surface and outer surface components of the bacterium rather than a specific ligand-receptor binding process (3).It is known that biofilm formation is a complex process, and a number of cell surface structures have been implicated in the initial stages of biofilm formation. In Pseudomonas aeruginosa these include flagella (motility) (35) and the expression of type IV pili for microcolony formation (36). In streptococci a number of cell surface adhesins, together with autolysins, have been identified as being important in biofilm development (12,17,19,21,26). The subsequent development and differentiation of an ordered three-dimensional biofilm requires the expression of extracellular polysaccharides (9) and may involve cell-to-cell signaling molecules (11,26). During the initial establishment of a sessile community, it is likely that bacteria have to adapt their...

The analysis of the intramacrophagic virulome ofBrucella suisdeciphers the environment encountered by the pathogen inside the macrophage host cell

“…In E. coli, the two proteins RelA and SpoT regulate the concentration of (p)ppGpp, a global transcriptional activator of numerous metabolic pathways under starvation conditions (19). RelA is important for the virulence of Listeria monocytogenes (20), for the activation of quorum sensing in Pseudomonas aeruginosa (21), and for long-term survival of Mycobacterium tuberculosis (22). In Brucella spp.…”

Comparative proteogenomic analysis of the Leptospira interrogans virulence-attenuated strain IPAV against the pathogenic strain 56601

“…Previous studies demonstrated that the ability of bacteria to synthesize (p)ppGpp and mount a stringent response is an essential physiological adaptation required for stages of pathogenesis, such as initial growth of adherent bacteria [38], toxin production [39], motility [40] and expression of other virulence regulators [41]. Comparative analysis revealed that the IPAV spoT gene (LIF_A2491) has two missense mutations at the amino-acid residues, Val293 and Lys407, resulting in replacement by Ala and Glu, respectively.…”

The arsenal of virulence factors deployed byListeria monocytogenesto promote its cell infection cycle

“…150,151 A L. monocytogenes mutant for relA, a gene encoding a (p)ppGpp synthetase, is unable to accumulate (p)ppGpp in response to amino acid starvation. 152 In addition to be required for full virulence in mice, RelA was shown to play a critical role for bacterial intracellular growth in non-phagocytic cells and macrophages. These results suggest that, after phagosomal escape, the capacity to mount a stringent response is pivotal for maximal Listeria intracellular replication.…”