Resuscitation of Tenacibaculum sp., the causative bacterium of spotting disease of sea urchin Strongylocentroutus intermedius, from the viable but non-culturable state

Abstract

Tenacibaculum sp., the causative bacterium of spotting disease of sea urchin Strongylocentroutus intermedius, has been known to enter into the viable but non-culturable (VBNC) state in 75% artificial seawater (ASW) at 5°C. From this fact it is suspected that this bacterium probably resuscitates from the VBNC state in summer. To examine whether non-culturable cells of Tenacibaculum sp. are able to regain the culturable state, VBNC cells in 75% ASW and 75% ASW containing homogenated sea urchin (HSU) were treated with temperature upshift (5°C to 25°C) and addition of iron chloride. Resuscitation from VBNC cells with temperature upshift was observed only in 75% ASW containing HSU for 3 days incubation after entry into the VBNC state. Resuscitation of VBNC cells treated with iron chloride was observed in each microcosm. Resuscitation from the VBNC state was confirmed by dilution experiments and measurements of generation time. Resuscitated cells regained original characteristics, agglutination against anti-Tenacibaculum sp. strain F-2, pathogenicity to sea urchin and morphology.