Implications of Microorganisms in Future Crewed Missions to Mars

Crewed deep space missions will likely become a reality in the coming decades. One of the most challenging issues associated with Mars is the lack of resources for sustaining human life. Microbes can be used to help humans settle on Mars. However, they can also be problematic since terrestrial microorganisms carried by astronauts could contaminate the Martian landscape and be mistaken for Martian native life. Organic carbon is one of the resources limited on Mars. Microalgae, capable of producing organic carbon photoautotrophically, can provide various crucial products for human survival, such as oxygen and food. Moreover, due to their ability to accumulate lipids and starch, microalgae, such as Chlamydomonas reinhardtii, can be used as feedstock for heterotrophic bacteria that can produce products required for daily routine, such as plastics. In this work, a two-step system was developed to produce bioplastics by microbial culture. C. reinhardtii was cultured to create biomass, which was processed to produce 16.8 g/100g ± 0.67 of starch as feedstock with a productivity of 2.79 mg/L/h for a Bacillus subtilis culture yielding polyhydroxybutyrate (PHB) biopolymer. A photobioreactor capable of growing large volumes of C. reinhardtii biomass was developed, and different techniques evaluated to estimate the starch content during culture. An optimised protocol for collecting the microalgae cells by combination of coagulation and centrifugation was developed, and then for chemical hydrolysis of the harvested starch into fermentable sugars with acid under high temperatures. The successful cultivation of B. subtilis grown on Chlamydomonas-derived sugars was also demonstrated, producing a maximum of 23% ±1.45 of CDW or 0.18 ± 0.1 g/L of PHB with a productivity of 7.6 mg/L/h. Infrared spectroscopy was used to track the production of starch and PHB in the primary and secondary cultures, respectively. The PHB obtained was successfully used to produce a plastic film with improved flexibility. The use of such PHB in fused deposition modelling 3D printing to create tools was also demonstrated in principle by use of commercially available blends. An improvement to the starch harvesting and hydrolysis stage was attempted with novel genetic tools based on the Golden Gate assembly. An attempt was made to introduce a thermophilic external alpha-amylase with the objective of facilitating the separation and hydrolysis of starch. However, the work could not be completed within the allotted timeframe due to unforeseen challenges. To address the potential negative impact of human-associated microbes leaking out onto the Martian surface, a culture-independent study was conducted at the Mars Desert Research Station (MDRS). Despite its inherent bias, PCR amplification is a highly sensitive method, used here to evaluate the risk of contaminating Mars in future crewed missions to Mars. Microbial DNA extraction, 16s rRNA and ITS sequencing and identification from swab samples of the habitat interior were used to characterise the human-associated microbiome, along with a complementary analysis on soil samples from the immediate vicinity to test for evidence of contamination from the habitat. No contamination was observed, but even if present such habitat-associated microbes in the desert soil would be expected to be present in very low numbers that may have fallen below the detection threshold of this approach.