In situ filtration and preliminary molecular analysis of microbial biomass from the Rainbow hydrothermal plume at 36°15′N on the Mid-Atlantic Ridge

Abstract

Elevated microbial biomass levels play a significant role in altering the chemical composition of hydrothermal plumes. In this work, a procedure was developed for collecting sufficient quantities of microbial biomass from a hydrothermal plume to allow the extraction of DNA suitable for use in molecular analyses of the plume microbial populations. During a geophysical survey along 200 km of the Mid-Atlantic Ridge (MAR), southwest of the Azores Triple Junction, several new sites of hydrothermal activity were identified, including the Rainbow hydrothermal plume, at 36°15′N, the strongest such feature yet reported from the MAR. During the survey, in situ filtration systems were used to collect microbial biomass from the water column onto 293 mm diameter, 0.2 μm-pore membrane filters by pumping large volumes (300–500 l) of seawater through the filters. Filter samples were collected from both the neutrally buoyant portion of the Rainbow plume (at a depth of 2050 m) and from above it in the water column (at 1500 m). Microgram quantities of genomic DNA were extracted and purified from the biomass on these filters. The suitability of the purified DNA for use in molecular analyses was demonstrated by polymerase chain reaction (PCR) amplification of the small subunit ribosomal rRNA (SSU rRNA) gene from the two primary prokaryotic domains, the Bacteria and the Archaea. As indicated by the binding of domain-specific oligonucleotide hybridisation probes to the purified DNA, the plume microbial populations contained a higher proportion of members of the domain Archaea than the populations in the surrounding ambient sea-water.