Phenotypic and Genetic Characterization of Temperature-Induced Mutagenesis and Mortality in Cupriavidus metallidurans.

Cupriavidus metallidurans strains display a decreased viability when incubated in rich medium at a temperature of 37°C compared to their normal growth temperature of 30°C, a phenomenon coined "temperature-induced mortality and mutagenesis" (TIMM). To scrutinize this aberrant phenotype further, the contributions of specific inducers and protective agents were determined. Different growth media, including lysogeny broth (LB) and Schatz, and components, including casamino acids, in particular amino acids (proline, cysteine, glycine, glutamine, leucine, histidine and phenylalanine) and ammonium, were found to induce TIMM at 37°C. Sorbitol was found to counteract TIMM. Furthermore, although TIMM is well conserved within the C. metallidurans species, multiple and strain-specific TIMM inducers exist. Twenty-nine percent of the TIMM survivors inherited resistance to TIMM. Whole-genome sequencing of two resistant derivatives revealed an important role of an uncharacterized oxidoreductase, indicating putative metabolic poisoning when grown in high-concentration nitrogen-containing media at 37°C.

Limnospira indica PCC8005 growth in photobioreactor: model and simulation of the ISS and ground experiments.

The Arthrospira-B experiment is the first experiment in space ever allowing the online measurements of both oxygen production rate and growth rate of Limnospira indica PCC8005 in batch photobioreactors running on-board ISS. Four bioreactors were integrated in the ISS Biolab facility. Each reactor was composed of two chambers (gas and liquid) separated by a PTFE membrane and was run in batch conditions. Oxygen production was monitored by online measurement of the total pressure increase in the gas chamber. The experiments are composed of several successive batch cultures for each reactor, performed in parallel on ISS and on ground. In this work, a model for the growth of the cyanobacterium Limnospira indica PCC8005 (also known as Arthrospira or spirulina) in these space membrane photobioreactors was proposed and the simulation results obtained are compared to the experimental results gathered in space and on ground. The photobioreactor model was based on a light transfer limitation model, already used to describe and predict the growth and oxygen production in small to large scale ground photobioreactors. It was completed by a model for pH prediction in the liquid phase allowing assessment of the pH increase associated to the bicarbonate consumption for the biomass growth. A membrane gas-liquid transfer model is used to predict the gas pressure increase in the gas chamber. Substrate limitation is considered in the biological model. A quite satisfactory fit was achieved between experimental and simulation results when a suitable mixing of the liquid phase was maintained. The data showed that microgravity has no first order effect on the oxygen production rate of Limnospira indica PCC8005 in a photobioreactor operating in space in zero gravity conditions.

Reactivation of Microbial Strains and Synthetic Communities After a Spaceflight to the International Space Station: Corroborating the Feasibility of Essential Conversions in the MELiSSA Loop.

To sustain human deep space exploration or extra-terrestrial settlements where no resupply from the Earth or other planets is possible, technologies for in situ food production, water, air, and waste recovery need to be developed. The Micro-Ecological Life Support System Alternative (MELiSSA) is such a Regenerative Life Support System (RLSS) and it builds on several bacterial bioprocesses. However, alterations in gravity, temperature, and radiation associated with the space environment can affect survival and functionality of the microorganisms. In this study, representative strains of different carbon and nitrogen metabolisms with application in the MELiSSA were selected for launch and Low Earth Orbit (LEO) exposure. An edible photoautotrophic strain (Arthrospira sp. PCC 8005), a photoheterotrophic strain (Rhodospirillum rubrum S1H), a ureolytic heterotrophic strain (Cupriavidus pinatubonensis 1245), and combinations of C. pinatubonensis 1245 and autotrophic ammonia and nitrite oxidizing strains (Nitrosomonas europaea ATCC19718, Nitrosomonas ureae Nm10, and Nitrobacter winogradskyi Nb255) were sent to the International Space Station (ISS) for 7 days. There, the samples were exposed to 2.8 mGy, a dose 140 times higher than on the Earth, and a temperature of 22°C ± 1°C. On return to the Earth, the cultures were reactivated and their growth and activity were compared with terrestrial controls stored under refrigerated (5°C ± 2°C) or room temperature (22°C ± 1°C and 21°C ± 0°C) conditions. Overall, no difference was observed between terrestrial and ISS samples. Most cultures presented lower cell viability after the test, regardless of the type of exposure, indicating a harsher effect of the storage and sample preparation than the spaceflight itself. Postmission analysis revealed the successful survival and proliferation of all cultures except for Arthrospira, which suffered from the premission depressurization test. These observations validate the possibility of launching, storing, and reactivating bacteria with essential functionalities for microbial bioprocesses in RLSS.