08uj-2026-01-31_08_25_06-s41598-023-49680-3.pdf
08uj-2026-01-31_08_25_06-s41598-023-49680-3.pdf
OPEN Effects of altered gravity on growth and morphology in Wolffia globosa implications for bioregenerative life support systems and space-based agriculture
Understanding the response of plants to varied gravitational conditions is vital for developing effective food production in space bioregenerative life support systems. This study examines the impact of altered gravity conditions on the growth and morphological responses of Wolffia globosa (commonly known as "water lentils" or "duckweed"), assessing its potential as a space crop. Although an experiment testing the effect of simulated microgravity on Wolffia globosa has been previously conducted, for the first time, we investigated the effect of multiple gravity levels on the growth and morphological traits of Wolffia globosa plants. The plant responses to simulated microgravity, simulated partial gravity (Moon), and hypergravity environments were evaluated using random positioning machines and the large-diameter centrifuge. As hypothesized, we observed a slight reaction to different gravitational levels in the growth and morphological traits of Wolffia globosa. The relative growth rates of plants subjected to simulated microgravity and partial gravity were reduced when compared to those in other gravity levels. The morphological analysis revealed differences in plant dimensions and frond length-to-width ratios under diverse gravity conditions. Our findings showed that Wolffia globosa is responsive to gravitational changes, with its growth and morphological adaptations being slightly influenced by varying gravitational environments. As for other crop species, growth was reduced by the microgravity conditions; however, relative growth rate remained substantial at zero point three three per day. In conclusion, this study underscores the potential of Wolffia globosa as a space crop and its adaptability to diverse gravitational conditions, contributing to the development of sustainable food production and bioregenerative life support systems for future space exploration missions.
Wolffia globosa, commonly known as "water lentils" is a member of the Wolffoideae subfamily within the Lemnaceae family, shares similar biological traits with its Lemnoideae relatives: it is the smallest flowering plant and shows the fastest growth rate in the plant kingdom and lacks a pseudo root system. Due to its rapid growth rate, high protein content, and nutritional value, it has emerged as a promising candidate for sustainable food production, particularly in agriculturally challenging regions. This unique plant has recently garnered attention also as a potential bioregenerative life support system candidate, offering traits that align with closed-loop, resource-efficient systems like those adopted for the ESA MELISSA Loop. Nevertheless, as for other candidate space crops, to assess the suitability of Wolffia's in space cultivation and its ability to recycle resources efficiently, further laboratory tests investigating growth, nutrition, and genetic responses under extreme conditions are fundamental.
Studying the effects of various stimuli, including different gravity environments, radiations, and their interactions, on the growth and development of plants can be challenging and costly. Space limitations within onboard test facilities, such as the International Space Station or orbiting vectors, often restrict the number of replicates, making studies under these conditions even more demanding. However, cost-effective alternatives exist in the form of facilities that simulate microgravity and partial gravity levels by continuously altering the gravitational vector. These facilities offer higher replicate numbers and serve as robust testbeds for experiments involving different gravity levels.
Hypergravity, characterized by gravitational forces greater than Earth's standard one G, is a significant factor in space exploration, affecting humans and plant life. Hypergravity is most prominent during the maneuvers of take-off and landing phases of spacecraft. These brief yet intense episodes of increased gravitational force can influence various aspects of plant biology and growth. Understanding how plants respond to hypergravity is essential for optimizing their cultivation in space environments and for Earth's agriculture. Furthermore, it provides valuable insights into the mechanisms plants employ to withstand and adapt to extreme gravitational conditions.
Earlier studies under simulated microgravity have shown stable anatomical morphology in Wolfia plants, possible effects of hypergravity on plant growth and reproduction have not yet been investigated. Testing plant reactions of potential space crops under varying gravity levels, including those experienced during take-off, landing, or in partial gravities, is reported as crucial. Consequently, exploring the adaptability of Wolfia plants under different gravitational conditions significantly contributes to our understanding of their potential.
Recent genome sequencing of Wolffia australiana has shed light on the gravity-sensing mechanisms and photomorphogenesis in Wolffia plants. Due to the loss of gravity sensing genes in the sister species Wolffia australiana, we also hypothesized for Wolffia globosa a lack of the gravity sensing mechanisms, resulting in a reduced effect of the different gravity levels on the growth and morphological characteristics.
More specifically, we aim to study the effects of altered gravity conditions on the growth and morphology of Wolfia globosa. Leveraging the capabilities of random positioning machines and large-diameter centrifuges, we simulated microgravity, partial gravity (Moon), and hypergravity conditions to comprehensively explore the impact of varying gravitational environments on this plant species. These machines allow us to generate and study a range of gravity conditions, thereby elucidating Wolffia globosa's responses to these diverse gravity environments and assessing its potential utilization in space-based food production systems.
Materials and methods Plant material and cultivation
Materials and methods Plant material and cultivation
Plants of Wolffia globosa (nine nine one zero) have been provided by Prof. Klaus Apperoth from the Department of Plant Physiology of the University of Jena, Germany. Upon receive, plant material was surface sterilized with zero point three percent bleach/water solution for five minutes. After fourteen days from disinfection, plants were subcultured for thirty days in N-medium under axenic conditions. Therefore, plant material has been transferred under a laminar flow hood in sets of six-well plates, each filled with five milliliters of N-medium solution and sealed with micropore tape. Before the experimental run, plants were acclimatized for twenty-four hours at thirty degrees Celsius. After acclimatization, an average of two hundred four fronds were transferred in each well of a six-multiwell plate previously filled with five milliliters of N-medium and zero point eight percent of Agar to achieve a semi-solid substrate. The experiment was conducted at thirty degrees Celsius average temperature with a photoperiod of sixteen/eight hours light/dark and a total Photon Flux Density of seventy-two point three seven plus or minus ten point three zero micromole per meter squared per second for one hundred sixty-eight hours. Carbon dioxide concentration has been monitored throughout the experiment, resulting in an average concentration of
Experimental hardware has been developed to hold two multiwell plates in a setup that minimizes the gravity gradient between the upper and lower multiwell plates. Furthermore, the experimental hardware had to be designed to fit in most test facilities, setting the lower constraints to an overall dimension of fifteen by fifteen by fifteen centimeters of the smallest Random Positioning Machine available. Considering these constraints, we designed and developed the experimental hardware using the free available software SketchUp. Due to the constraints described before, multiwell plates had to be stacked vertically. This setup minimizes the distance of the two centers of mass of both multiwells so that the difference in acceleration can be neglected. The experimental hardware has been equipped with LED white light to ensure proper plant development.
Furthermore, three centimeter five volt fans have been added to the experimental hardware to increase uniformity between the temperature inside and outside the experimental hardware. The prototype of the experimental hardware has been built in plywood. Plywood has been laser cut, and parts have been glued in place with vinyl glue. The template for laser cutting can be found in supplementary material Appendix A. Nevertheless, the experimental hardware can be three D printed in any plastic material, and information for the three D model can be found on the directory: GitHub.
The inner dimension of each experimental hardware is designed to fit a multiwell plate of standard dimensions twelve point seven by eight point five by two point two centimeters. From the multiwell lid to the lighting system, a fixed distance of two point zero centimeters has been set up to guarantee optimal light intensity for plant growth. The experimental hardware's design and dimensions are reported below.