Mechanisms for pseudoalteromonas piscicida-induced killing of vibrios and other bacterial pathogens

Authors: Richards, Gary; Watson, Michael; Needleman, David; Uknalis, Joseph; BOYD, E. FIDELMA - University Of Delaware; FAY, JOHNNA - University Of Delaware

Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/1/2017
Publication Date: 6/1/2017
Citation: Richards, G.P., Watson, M.A., Needleman, D.S., Uknalis, J., Boyd, E., Fay, J.P. 2017. Mechanisms for pseudoalteromonas piscicida-induced killing of vibrios and other bacterial pathogens. Applied and Environmental Microbiology. doi: 10.1128/AEM.00175-17.

Interpretive Summary: Pseudoalteromonas are marine bacteria that are known to secrete antimicrobial compounds which inhibit and kill competing bacteria in the marine environment. We isolated three strains of Pseudoalteromonas piscicida from the Delaware Bay and characterized their secreted enzymes. We also identified a second method by which some P. piscicida kill competing bacteria. It involves the direct transfer of digestive vesicles from the surface of the Pseudoalteromonas to the surface of competitors, digestion of holes in the competitor’s cell wall by compounds associated with the vesicles, and apparent feeding of the Pseudoalteromonas off the nutrients released by the digested bacterium in a predatory fashion. Pseudoalteromonas piscicida inhibited and killed human pathogens, suggesting that it may produce important antibacterial compounds and have practical probiotic applications in aquaculture and seafood safety, and in reducing biofilm formation.

Technical Abstract: Pseudoalteromonas piscicida is a Gram-negative gammaproteobacterium found in the marine environment. Three strains of pigmented P. piscicida were isolated from seawater and partially characterized by inhibition studies, electron microscopy, and analysis for proteolytic enzymes. Growth inhibition and death occurred around colonies of P. piscicida on lawns of the naturally occurring marine pathogens Vibrio vulnificus, Vibrio parahaemolyticus, Vibrio cholerae, Photobacterium damselae, and Shewanella algae. Inhibition also occurred on lawns of Staphylococcus aureus but not on Escherichia coli O157:H7 or Salmonella enterica serovar Typhimurium. Inhibition was not pH associated, but it may have been related to the secretion of a cysteine protease with strong activity, as detected with a synthetic fluorogenic substrate. This diffusible enzyme was secreted from all three P. piscicida strains. Direct overlay of the Pseudoalteromonas colonies with synthetic fluorogenic substrates demonstrated the activity of two aminopeptidase Bs, a trypsin-like serine protease, and enzymes reactive against substrates for cathepsin G-like and caspase 1-like proteases. In seawater cultures, scanning electron microscopy revealed numerous vesicles tethered to the outer surface of P. piscicida and a novel mechanism of direct transfer of these vesicles to V. parahaemolyticus. Vesicles digested holes in V. parahaemolyticus cells, while the P. piscicida congregated around the vibrios in a predatory fashion. This transfer of vesicles and vesicle-associated digestion of holes were not observed in other bacteria, suggesting that vesicle binding may be mediated by host-specific receptors. In conclusion, we show two mechanisms by which P. piscicida inhibits and/or kills competing bacteria, involving the secretion of antimicrobial substances and the direct transfer of digestive vesicles to competing bacteria.