ENTRAPMENT OF BACTERIA IN FLUID INCLUSIONS IN LABORATORY-GROWN HALITE

Cells of the bacterium, Pseudomonasa aeruginosa modified with green fluorescent protein, were entrapped in fluid inclusions in laboratory-grown halite. Sterile, saline (NaCl-saturated) solution was placed in 6 10-mL beakers and allowed to evaporate under a laminar-flow hood. Two of the beakers were inoculated with P. aeruginosa, which is tolerant of a wide range of environmental conditions, but is not known to be halotolerant. After two weeks, a halite precipitate developed in each beaker, with crystals ranging in size from a few μm to 2-3 mm across. Thick sections were made from the halite crystals, which contained abundant single-phase (all liquid) and a few two-phase (liquid and gas) fluid inclusions. Bacterial cells were present in inclusions in halite precipitated from the inoculated beakers, but not present in halite from the sterile control beakers. The bacteria were present in less than 10 percent of the fluid inclusions, in populations ranging from one to several dozen cells in each inclusion. Entrapment of cells occurred prior to complete desiccation of the saline solution, and the cells did not indicate any preference for attachment to halite crystals. The mechanism of entrapment, therefore, is unclear. The cells exhibited similar pseudo-Brownian motion both in the inclusions and in the hypersaline solution, in contrast to the run-and-tumble motion of cells in fresh water. The cells could have expired, or the high density of the saline solution could be inhibiting the flagellum-driven motion of the cells. The bacteria fluoresced brightly (UV illumination) in fresh water, in the saline solution, and in fluid inclusions. The bright fluorescence could indicate that the bacteria survived entrapment as fluorescence was greatly reduced after sterilization by autoclaving. A single inclusion containing cells was imaged once and again three days later. Exact cell counts were difficult to obtain, but the cell population appeared to increase from the first to the second image. The results of this study have a wide-range of implications for studies of molecular paleontology and fluid inclusions. Cells living and possibly reproducing within inclusions could respire gases and affect the chemistry of the fluid. Fluid inclusions also could preserve large unstable biomolecules, such as DNA, long after the cells expire.