Microorganisms living under high pressures and/or high temperatures hold vital clues for defining the extent of Earth’s biosphere and for exploring the emergence of life itself. Not only does a majority of the modern prokaryotic biosphere thrive in the piezosphere, but high-pressure habitable niches were likely even more widespread on early Earth and throughout the solar system. To date, high-pressure microbial sampling and growth has been limited by expense, expertise and customized equipment needs. To expand the scope of high-pressure microbial research and to grow the research community that can access and investigate high-pressure subsurface samples, the Deep Carbon Observatory (DCO) supported the aquisition of a suite of custom 50mL Pressurized Underwater Sample Handler (PUSH50). The PUSH50 instrumentation is the cornerstone of the emerging DCO high-pressure biology facility that will be known as Piezophile Research Instrumentation for Microbial Exploration (PRIME).
The design specifications for the PUSH50 were defined during the DCO Bioreactor Workshop in August 2013. Twenty industry and academic scientists from six nations discussed and proposed improved instrumentation to facilitate high-pressure biological and geochemical investigations. Workshop attendees outlined design specifications for a new generation of high-pressure and high-temperature vessels to sample subsurface fluids as well as to transport these high-pressure samples between laboratories in airline-certified pressure vessels. A broader community of interested scientists commented on “transporter” specifications and design elements following the workshop. This feedback was used to fine-tune the final specifications in a request for proposals sent to leading companies in the high-pressure equipment industry. In late 2014, following proposal review, the system proposed by Top Industrie (Vaux Le Pénil, France) was selected and technical specifications and procurement details were finalized between DCO representatives. The combination sampler/transport vessels, now known as PUSH50s, were delivered to Drs. Daniel and Rogers in September 2015.
Key attributes of the PUSH50 are its capability to both retrieve and transport biological fluid samples under high pressure, eliminating sample decompression that characterizes other methodologies. The PUSH50 includes a 50mL sterilizable PEEK variable volume, floating piston reactor housed in a high-pressure vessel rated for 100 MPa (1 kbar) and 160°C, and is certified for airline travel. Additionally, a full deployment package includes a CTD-rosette compatible skid, a manual pressure generator, pressure compensator and recording pressure sensor.  With DCO instrumentation support, two PRIME instrumentation locations—each home to a full deployment package and five samplers—are being established in the laboratories of Isabelle Daniel (Laboratoire de Géologie de Lyon, Université de Lyon, France) and Karyn Rogers (Rensselaer Polytechnic Institute, USA).
In Winter 2015/16, the PUSH50s are being tested in the DCO PRIME satellite laboratories of Daniel and Rogers. The PUSH50 is compatible with SeaBird carousels  and is currently being outfitted for deployment on submersible vehicles including the DSV Alvin and ROV Jason II, similar to the Seewald Isobaric Sampler  and the Japanese WHATS II sampler . Upcoming modifications include an ICL-compatible thermocouple, a sampling nozzle, and adapter for manipulator arm use. The DCO PRIME facility will also develop training videos and user protocols. A three-member PRIME Standing Committee will be established to ensure that the instrumentation is appropriately managed and maintained. Prospective users will submit a short proposal that will be assessed, approved and scheduled by the PRIME Standing Committee. Users will be expected to pay a relatively small access fee to cover insurance, maintenance and shipping costs. Final reports based on user experiences will ensure that the PUSH50s are optimized for community needs. Once their testing is complete, the PUSH50s, along with accompanying deployment equipment, will be available to the broader DCO community.
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