Pfitsch Don W.

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Don W.

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  • Preprint
    Autonomous Microbial Sampler (AMS), a device for the uncontaminated collection of multiple microbial samples from submarine hydrothermal vents and other aquatic environments
    ( 2006-01-11) Taylor, Craig D. ; Doherty, Kenneth W. ; Molyneaux, Stephen J. ; Morrison, Archie T. ; Billings, John D. ; Engstrom, Ivory B. ; Pfitsch, Don W. ; Honjo, Susumu
    An Autonomous Microbial Sampler (AMS) is described that will obtain uncontaminated and exogenous DNA-free microbial samples from most marine, fresh water and hydrothermal ecosystems. Sampling with the AMS may be conducted using manned submersibles, Remotely Operated Vehicles (ROVs), Autonomous Underwater Vehicles (AUVs), or when tethered to a hydrowire during hydrocast operations on research vessels. The modular device consists of a titanium nozzle for sampling in potentially hot environments (>350°C) and fluid-handling components for the collection of six independent filtered or unfiltered samples. An onboard microcomputer permits sampling to be controlled by the investigator, by external devices (e.g., AUV computer), or by internal programming. Temperature, volume pumped and other parameters are recorded during sampling. Complete protection of samples from microbial contamination was observed in tests simulating deployment of the AMS in coastal seawater, where the sampling nozzle was exposed to seawater containing 1x106 cells ml-1 of a red pigmented tracer organism, Serratia marinorubra. Field testing of the AMS at a hydrothermal vent field was successfully undertaken in 2000. Results of DNA destruction studies have revealed that exposure of samples of the Eukaryote Euglena and the bacterium S. marinorubra to 0.5 N sulfuric acid at 23°C for 1 hour was sufficient to remove Polymerase Chain Reaction (PCR) amplifiable DNA. Studies assessing the suitability of hydrogen peroxide as a sterilizing and DNA-destroying agent showed that 20 or 30% hydrogen peroxide sterilized samples of Serratia in 1 hr and destroyed the DNA of Serratia, in 3 hrs, but not 1 or 2 hrs. DNA AWAY™ killed Serratia and destroyed the DNA of both Serratia and the vent microbe (GB-D) of the genus Pyrococcus in 1 hour.