Identity, Abundance, and Reactivation Kinetics of Thermophilic Fermentative Endospores in Cold Marine Sediment and Seawater

Marta Volpi, Bente Aa Lomstein, Andreas Sichert, Hans Røy, Bo B Jørgensen, Kasper U Kjeldsen

Cold marine sediments harbor endospores of fermentative and sulfate-reducing, thermophilic bacteria. These dormant populations of endospores are believed to accumulate in the seabed via passive dispersal by ocean currents followed by sedimentation from the water column. However, the magnitude of this process is poorly understood because the endospores present in seawater were so far not identified, and only the abundance of thermophilic sulfate-reducing endospores in the seabed has been quantified. We investigated the distribution of thermophilic fermentative endospores (TFEs) in water column and sediment of Aarhus Bay, Denmark, to test the role of suspended dispersal and determine the rate of endospore deposition and the endospore abundance in the sediment. We furthermore aimed to determine the time course of reactivation of the germinating TFEs. TFEs were induced to germinate and grow by incubating pasteurized sediment and water samples anaerobically at 50°C. We observed a sudden release of the endospore component dipicolinic acid immediately upon incubation suggesting fast endospore reactivation in response to heating. Volatile fatty acids (VFAs) and H2 began to accumulate exponentially after 3.5 h of incubation showing that reactivation was followed by a short phase of outgrowth before germinated cells began to divide. Thermophilic fermenters were mainly present in the sediment as endospores because the rate of VFA accumulation was identical in pasteurized and non-pasteurized samples. Germinating TFEs were identified taxonomically by reverse transcription, PCR amplification and sequencing of 16S rRNA. The water column and sediment shared the same phylotypes, thereby confirming the potential for seawater dispersal. The abundance of TFEs was estimated by most probable number enumeration, rates of VFA production, and released amounts of dipicolinic acid during germination. The surface sediment contained ∼10(5)-10(6) inducible TFEs cm(-3). TFEs thus outnumber thermophilic sulfate-reducing endospores by an order of magnitude. The abundance of cultivable TFEs decreased exponentially with sediment depth with a half-life of 350 years. We estimate that 6 × 10(9) anaerobic thermophilic endospores are deposited on the seafloor per m(2) per year in Aarhus Bay, and that these thermophiles represent >10% of the total endospore community in the surface sediment.