The development of anaerobic enrichment cultures and measurements of the rates of methanogenesis, the turnover of 14C acetate and the degradation of substrates from sediment samples from Bratina Island

Life in ponds and tidal lagoons of the McMurdo Ice Shelf is dominated by benthic cyanobacterial mats. Anaerobic conditions occur beneath these productive mats. A study was conducted to investigate the degradation processes of biogeochemical cycles in the Ice Shelf ecosystem by the heterotrophic microbes that occur below the benthic mats, in particular how these organisms facilitate the degradation of proteins, polysaccharides and other components released into the anaerobic environment. The study was undertaken over 4 years with different investigations each season. In the first season the investigation focussed on i) measurement of methane production rates, ii) turnover of 14C radio labelled acetate and ii) degradation of known substrates added to sediment. Experiments were initiated by transfer of sediment samples to tubes or bottles under a gas stream of N2/CO2. Vessels were sealed with septum stoppers to maintain anaerobic conditions and were incubated at the pond sites. Anaerobic enrichment cultures were attempted to be developed with two types of enrichment media used, “high salt” and “low salt”. The medium were inoculated with sediment from 3 ponds (Salt, Fresh, Skua and Orange ponds) and examined for bacterial growth. Experiments were conducted on sediment gathered from 6 ponds (Skua, Fresh, Brack, Salt, Orange and P70 ponds). In the second season, the seasons work was dominated by sampling a transect from well above the water line of Orange Pond down to about 50cm water depth. Temperature profiles were taken with a thermocouple probe and sediment cores were taken down to the freezing level. Cores were segmented at 2cm intervals. Duplicate segments of six sites along the transect were extracted for nutrient analyses and other segments of the same depth were pooled for measurement of density, porosity, dry weight, organic matter, volatile fatty acids, pH, salinity and for the determination of biological activities under various conditions. The samples were returned to New Zealand for measurements of methane production rates, determination of the terminal carbon flow using the turnover of radio labelled 14C acetate and degradation of known substrates added to the sediment. Enrichment of anaerobic bacteria was also initiated from these sediment. In the third season of study, the study of methanogenesis in Orange pond was the focus. The temperature of the pond was measured throughout the study period. Methanogenesis rates between sediment cores and slurries were compared. The high salt tolerance of the bacterial population in Orange pond was demonstrated when the effect of osmotic pressure on methanogenesis was examined. Salt solutions (10x and 50x multiples of the salt concentration in Orange pond) were added to sediment slurry replicates and methane production was measured. The flux of methane from Orange Pond sediment was investigated in situ by pushing core tubes into the sediment to about 10cm depth and leaving to incubate. Methane flux across the core surface was calculated from the change in methane concentration in the overlying water over the incubation period. Incubation was continued in New Zealand with samples transported from Antarctica. In the fourth and final season, the original six ponds were all sampled for benthic mat, water and sediment and samples were returned to New Zealand for laboratory investigations. Experiments with the sediments were conducted to determine i) the contribution of dimethylsulphide to methane and ii) the rates of sulphate reduction in the sediment system. In addition, a major study on the physiological characteristics of two ‘extremophilic’ anaerobic bacteria isolated from the pond system was completed.