LONG-TERM SURVIVAL OF MICROORGANISMS IN FLUID INCLUSIONS IN HALITE

Halophilic microbes are readily incorporated within brine inclusions in modern halite precipitated from shallow (<1 m) brines and in surface halite crusts from Death Valley and Saline Valley, California, collected between 1980 and 2005. Salt cores from Death Valley (drilled to 186 m, 200 ka) and Saline Valley (100 m, 150 ka) preserve a well-dated record of saline paleoenvironments. Systematic analysis of fluid inclusions and trapped microbes in halites from the salt cores (microscopic imaging, culturing of microorganisms in basin specific brine-broth chemistries, purification and polymerase chain reaction (PCR) amplification of trapped intracellular DNA with archaea- and bacteria-specific 16S rDNA primers) is testing the limits of microbial survivability and DNA preservation. Fluid inclusions in halite crystals from these cores contain numerous trapped microorganisms (up to 106-107 cells per ml), that are notably most abundant in halite formed in perennial saline lakes. Halophilic and halotolerant organisms have been cultured from fluid inclusions in 7 stratigraphic intervals, Death Valley core 8 to 85 m (10 ka to 100 ka), in addition to the haloarchaeal species from 86 m (100 ka) isolated by Mormile et al. (2003). PCR amplification of archaeal DNA from fluid inclusions in halite (13 m, 22 ka, Death Valley core) was achieved from the same samples in which halophilic archaea were cultured independently at two laboratories (BU and WCU). Phylogenetic analysis (16S rDNA) of cultured microorganisms from halite (13 m and 18 m; 22 ka and 31 ka), Death Valley core, shows that they are not closely related to existing halophilic archaea (94% or less similarity), which rules out laboratory contamination by modern microbes. These results collectively indicate that fluid inclusions in halite originally crystallized in Saline Valley and Death Valley paleolakes preserve viable halophilic archaea and ancient DNA on times scales of 10 ka to 100 ka.