## 3 Results
### Yeast Growth  
At the time the initial studies were designed and initiated in 2011, one priority was to determine whether microgravity influenced overall cell growth. Although a simple outcome measure, growth was an important consideration for yeast as a potential infectious risk in spaceflight, but also provided insight into yeast adaptation to the microgravity environment. Embedded within the growth studies was a plan to allow the yeast to grow in microgravity for as long as possible, in spite of the requirement for a sealed growth chamber to ensure crew safety in the presence of an opportunistic pathogen. As such, the FPA bioreactors were prepared with two concentrations of yeast; a high inoculum (High, 1 \(\times\) 10\({}^{5}\) cells/mL) to ensure growth, and a low inoculum (Low, 100 cells/mL) to allow maximum replicative potential before stationary phase was reached. The protocol was also designed such that yeast growth was activated early (Early) in the mission to minimize any potential storage stress on the cells while maintained in water-induced stasis. In addition, comparable samples were stored and activated late (Late) in the mission so that cells could also be returned live for post-mission analyses. This design also allowed some comparison between cells activated Early and Late in the mission to evaluate the effects of storage in the microgravity environment. As seen in Figure 3A, cell growth in most of the samples peaked at \(\sim\)1 \(\times\) 10\({}^{6}\) cells/mL, although the Low inoculum samples activated Early in the mission (Early Low) grew to a slightly higher density and demonstrated a minor advantage to cells grown in microgravity (Early Low, * \(p\) = 0.02). It is worth noting that in Micro-6, cells that were scheduled to be returned live and stored at 4 \({}^{\circ}\)C actually experienced a 3-day transport temperature of nearly 25 \({}^{\circ}\)C (Figure 2). The cell growth data in Figure 3A for viable samples (\"Viable Low' and \"Viable High') suggest that the cells indeed continued to grow. The higher level of growth in the flight samples as compared to terrestrial controls likely reflects the elevated temperature during transport. There was not a mechanism to track temperatures in real-time in order to subject terrestrial controls samples to conditions comparable to that experienced by the flight samples. Interestingly, the additional growth of the viable samples in both microgravity and terrestrial conditions indicates that the culture environment was capable of supporting at least 1-2 additional doublings, thus other factors must have been contributing to the apparent growth cap (1 \(\times\) 10\({}^{6}\) cells/mL) observed in the other Micro-6 samples (Figure 3A). For clarity, the on-orbit growth period was sufficiently long for the Low inoculum samples to reach a density of 6 \(\times\) 10\({}^{7}\) cells/mL and the High inoculum samples to reach a density of >1 \(\times\) 10\({}^{8}\) cells/mL, using generation times established in ground-based studies. As seen in Table 2, generation times were calculated, factoring in the differences in inoculum size, and found to be similar between flight samples and the terrestrial controls. Interestingly, samples with the High cell inoculum had much longer generation times (300+ min/generation as compared to 150+ min/generation), suggesting the cell density of 1 \(\times\) 10\({}^{6}\) cells/mL was reached and the culture likely went into stationary phase. A kinetic analysis of cell growth was not performed due to resource constraints and safety precautions, so temporal aspects of the yeast growth dynamics cannot be concluded from these studies.