Comparison of Niskin vs. in situ approaches for analysis of gene expression in deep Mediterranean Sea water samples

Author Posting. © The Author(s), 2014. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Deep Sea Research Part II: Topical Studies in Oceanography 129 (2016): 213-222, doi:10.1016/j.dsr2.2014.10.020.Obtaining an accurate picture of microbial processes occurring in situ is essential for our understanding of marine biogeochemical cycles of global importance. Water samples are typically collected at depth and returned to the sea surface for processing and downstream experiments. Metatranscriptome analysis is one powerful approach for investigating metabolic activities of microorganisms in their habitat and which can be informative for determining responses of microbiota to disturbances such as the Deepwater Horizon oil spill. For studies of microbial processes occurring in the deep sea, however, sample handling, pressure, and other changes during sample recovery can subject microorganisms to physiological changes that alter the expression profile of labile messenger RNA. Here we report a comparison of gene expression profiles for whole microbial communities in a bathypelagic water column sample collected in the Eastern Mediterranean Sea using Niskin bottle sample collection and a new water column sampler for studies of marine microbial ecology, the Microbial Sampler – In Situ Incubation Device (MS-SID). For some taxa, gene expression profiles from samples collected and preserved 33 in situ were significantly different from potentially more stressful Niskin sampling and 34 preservation on deck. Some categories of transcribed genes also appear to be affected by sample 35 handling more than others. This suggests that for future studies of marine microbial ecology, 36 particularly targeting deep sea samples, an in situ sample collection and preservation approach 37 should be considered.This research was funded by NSF OCE-1061774 to VE and CT, NSF DBI-0424599 to CT and NSF OCE-0849578 to VE and colleague J. Bernhard. Cruise participation was partially supported by Deutsche Forschungsgemeinschaft (DFG) grant STO414/10-1 to T. Stoeck

Autonomous Microbial Sampler (AMS), a device for the uncontaminated collection of multiple microbial samples from submarine hydrothermal vents and other aquatic environments

Author Posting. © Elsevier B.V., 2006. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Deep Sea Research Part I: Oceanographic Research Papers 53 (2006): 894-916, doi:10.1016/j.dsr.2006.01.009.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.This work was supported by a DOC/NOAA Small Business Innovative Research Award, Contract No. 50-DKNA-9-90116 awarded to McLane Research Laboratories, Inc. and (via subcontract) to the Woods Hole Oceanographic Institution. Some of the microbial testing work was also supported by the National Science Foundation, Grant No. IBN-0131557 and the Woods Hole Oceanographic Inst. Deep Ocean Exploration Institute Grant No. 25051131