## 5 Conclusions  
Spacecraft may utilize controlled-release silver technology in potable water systems. Imparted silver ions will not be removed from the potable water effluent. Until this study, there was no existing evaluation of ionic silver levels on the space crop growth, plant nutrient content, and root zone microbiome. As a result, there were no established guidelines for its use in the space crop production setting. Currently, plant chambers such as the Advanced Plant Habitat on the International Space Station recycle their own transpired water but still require potable water to prime their water tanks and plumbing, and for occasional water volume replenishment. From this work, we determined that in a substrate-based system such as arcillite, lettuce plant growth, nutrient content, microbial diversity, and abundance are not affected by irrigation water with as high as 500 ppb ionic silver. Lettuce plants exposed to silver-treated water showed reduced growth for shoots at 31 ppb silver and severely stunted growth at 125 and 500 ppb when grown in a hydroponics system. Leaves from the hydroponic system showed the accumulation of silver and a reduced uptake of other essential elements as the silver concentration increased. It was observed that the number of microbes associated with roots and water from the growth trays of the hydroponic system increased as the concentration of silver increased. The types of microbes found on the roots and in the trays were similar but present in different relative amounts. This suggests that ionic silver exerted selective pressure on the irrigation water and the lettuce root microbiome in the hydroponics system. Future space crop production systems that lack cation-exchange capacity will need to remove silver to below 31 ppb to ensure plant health and adequate biomass for crop production. However, this biocide effect on water systems by silver does not preclude microbial resistance from occurring. In fact, subinhibitory amounts of silver over time can lead to greater microbial resistance to silver and the buildup of microbial biofilms, and this is an area for further investigation. **Supplementary Materials:** The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/microorganisms12030515/s1](https://www.mdpi.com/article/10.3390/microorganisms12030515/s1)]([https://www.mdpi.com/article/10.3390/microorganisms12030515/s1](https://www.mdpi.com/article/10.3390/microorganisms12030515/s1)), Supplementary Table S1: Metaphlan3 abundance counts for 126 samples. Supplementary Table S2: Enriched microbial genetic pathways and normalized counts for silver treatments in biological triplicate. Supplementary Table S3: Fold change and \(p\)-value for enriched microbial genetic pathways in arcillite samples. **Author Contributions:** Conceptualization, A.O. and R.W. ; methodology, L.S. and A.O. ; formal analysis, B.C., A.O. and A.D. ([PERSON]); investigation, T.I., A.D. ([PERSON]), L.S., B.C., A.D. ([PERSON]), W.L., C.S. and C.K. ; resources, A.O. ; data curation, A.O. and A.D. ([PERSON]); writing--original draft preparation, A.O. and L.S. ; writing--review and editing, A.O., L.S., R.W., A.D. ([PERSON]), B.L. and T.S. ; visualization, A.O. and B.L. ; supervision, A.O., R.W. and L.S. ; project administration, A.O. and L.S. ; funding acquisition, A.O. and R.W. All authors have read and agreed to the published version of the manuscript. **Funding:** This research was made possible by Center Innovation Funding (CIF) from NASA's Science and Technology Mission Directorate (STMD) to Kennedy Space Center (KSC) Exploration and Research Technology (ER&T) Division. **Data Availability Statement:** Whole genome metagenomics data are available at NASA's Genelab. **Acknowledgments:** We would like to thank our KSC resource officers, [PERSON] and [PERSON], for ensuring access to funds to complete this work.