Effects of ecological factors on the survival and physiology of Ralstonia solanacearum bv. 2 in irrigation water

The fate of Ralstonia solanacearum bv. 2, the causative agent of brown rot in potato, in aquatic habitats of temperate climate regions is still poorly understood. In this study, the population dynamics and the physiological r esponse of R. solanacearum bv. 2 were tested in sterile pure water and in a gricultural drainage water obtained from waterways near potato cropping fie lds in the Netherlands. The behaviour of five different biovar 2 isolates i n drainage water at 20 degreesC was very similar among strains. One typical isolate with consistent virulence (strain 1609) was selected for further s tudies. The effects of temperature, light, canal sediment, seawater salts, and the presence of competing microorganisms on the survival of strain 1609 were assessed. Moreover, the impacts of the physiological state of the ino culum and the inoculum density were analyzed. The population dynamics of st rain 1609 in sterile pure water were also characterized. In sterile pure wa ter, the fate of R. solanacearum 1609 cells depended strongly on temperatur e, irrespective of inoculum density or physiological state. At 4 degreesC a nd 44 degreesC, strain 1609 CFU numbers showed declines, whereas the strain was able to undergo several cell divisions at 12 degreesC, 20 degreesC, an d 28 degreesC. At 20 degreesC and 28 degreesC, repeated growth took place w hen the organism was serially transferred, at low inoculum density, from gr own water cultures into fresh water devoid of nutrients. Both at low and hi gh cell densities and regardless of physiological state, R. solanacearum 16 09 cells persisted as culturable cells for limited periods of time in drain age water. A major effect of temperature was found, with survival being max imal at 12 degreesC, 20 degreesC, and 28 degreesC. Temperatures of 4 degree sC, 36 degreesC, or 44 degreesC induced accelerated declines of the cultura ble cell numbers. The drainage water biota had a strong effect on survival at 12 degreesC, 20 degreesC, and 28 degreesC, as the persistence of strain 1609 was significantly enhanced in sterile drainage water systems. Furtherm ore, there was a negative effect of incident light, in a light:dark regime, on the survival of R. solanacearum 1609 in natural drainage water. Also, l evels of seawater salts realistic for drainage water in coastal areas were detrimental to strain survival. Ralstonia solanacearum 1609 showed consider able persistence in canal sediment saturated with drainage water, but died out quickly when this sediment was subjected to drying. Evidence was obtain ed for the conversion of R. solanacearum 1609 cells to nonculturable cells in water microcosms kept at 4 degreesC, but not in those kept at 20 degrees C. A substantial fraction of the cells found to be nonculturable were still viable, as evidenced by the direct viable count and by staining with the r edox dye 5-cyano-2,3-ditolyl tetrazolium chloride. The potential occurrence of viable-but-nonculturable cells in natural waters poses a problem for th e detection of R. solanacearum by cultivation-based methods.