Quantification of anaerobic thermophilic endospores in marine sediment by microcalorimetry, and its use in bioprospecting for gas and oil

Abstract

Endospores of anaerobic thermophilic bacteria (thermospores) are ubiquitous in cold marine sediments. These misplaced thermophiles likely originate from warm environments and are delivered to the seafloor by passive dispersal through the water column. The few studies of the abundance of thermospores that exist have only quantified a subset of the anaerobic metabolic types possibly present and the data density has been too low to address patterns of dispersal. Here, we introduce isothermal microcalorimetry as a fast tool to quantify the abundance of thermospores in environmental samples. We developed and tested the method on samples from the water column and sediment of Aarhus Bay, Denmark. The thermospores were stimulated to germinate and grow by heating the samples to 50°C inside an isothermal microcalorimeter and the metabolic heat production was followed across the exponential growth phase. The number of germinated thermospores was then calculated by dividing the total heat production rate by the cell specific heat production rate, empirically determined in parallel experiments. The abundance of thermospores in the water column of Aarhus Bay was 1.2 × 103−3.6 × 103 cells × mL−1 seawater. The abundance of thermospores in sediment in Aarhus Bay was 1.1 × 106−6.1 × 106 cells × mL−1 sediment and their half-life was ∼70 yr. As application example we used isothermal microcalorimetry to test if thermospores were aggregated at a pockmark in Limfjorden (Denmark) with seepage of thermogenic gas and oil.