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A distinctive microbial community has inhabited the wastewater tank within the International Space Station Water Recovery System (WRS) for over 14 years and experienced the stressors associated with the microgravity environment. The WRS generates potable water for the crew from urine distillate, humidity condensate, Sabatier product water, and the occasional off-loading of ground-supplied water. The reservoir for these products, the wastewater tank, does not have a means of microbial control. While samples are occasionally collected for analysis, the time between sample collection and the return to Earth, as well as the lack of preservation, results in a skewed depiction of the microbiome. Routinely observed from these returned samples are high counts (105 – 106 colony forming units per mL) and two prevailing genera, Burkholderia and Ralstonia. The wastewater tank likely contains a more diverse microbiome, as a higher diversity of bacteria and fungus has been noted upstream and downstream of the tank. To characterize the microbial profile of the tank, analysis needs to occur at the time of sample collection. Toward this goal, a method for in situ analysis based on nanopore sequencing was developed. The filter-to-sequencer method evolved from previous work that has been validated onboard the ISS (BEST payload and the BioMole Crew Health Care Systems Facility). The method, including filtration, DNA extraction, purification, amplification, library preparation, and nanopore sequencing will be described. Additionally, data collected with this method from both ISS and terrestrial samples will be detailed. The consumables needed to support in situ analysis of the tank are set to the launch to the ISS in the spring of 2023. This investigation will allow for the first accurate characterization of the microbiome of the tank providing insight for crew health, planetary protection, and has the potential to enable engineering controls for future space station water systems.