This is a supplemental file to a published research article in Gravitational and Space Research.
Citation: McKaig, J., Caro, T., Hyer, A., Talburt, E., Verma, S., Cui, K., Boguraev, A., Heit, M., Johnson, A., Johnson, E., Jong, A., Shepard, B., Stankiewiz, J., Tran, N., & Rask, J. (2019). A High-Altitude Balloon Platform for Space Life Sciences Education, Gravitational and Space Research, 7(1), 62-69. doi: https://doi.org/10.2478/gsr-2019-0007
Sampling devices that actively draw air are efficient in the sample volume they can collect compared to passive sampling instruments, but this efficiency is proportional to ambient air pressure. This experiment examines the diminishing returns of active sampling as a HAB payload rises. We used an aerosol black carbon detector (ABCD) and emphasize that any active sampling system can be used for similar experiments.
Our team used an ABCD that was developed, manufactured, and donated to us by Lawrence Berkeley National Labs (Caubel et al., 2018). The ABCD uses a rotary vane pump to draw air and a spectrophotometric assay to quantify black carbon concentration. The ABCD drew air for the entire duration of the flight, and its efficiency dramatically decreased as the ambient air pressure dropped. Future teams are encouraged to prototype or acquire active sampling devices to determine the minimum ambient pressure required to robustly sample aerosols, and how these performance decreases affect the fidelity of gathered results.
The SHAB flights of the ABCD demonstrated successful validation of active air intake systems for sampling aerosols at high altitude. The figure shows the measurements taken by the ABCD payload during the SHAB-2 flight.It should be noted that, per the ABCD manufacturer’s instructions, black carbon measurements taken when the flow rate is below 200 ccm are not reliable. Because the flow rate fell below 200 ccm quickly, the black carbon concentrations are not reliable above an altitude of 1000m. Although there is a dramatic spike in the black carbon concentration data (E), this spike corresponds to a flow-rate below 100ccm (B). Additionally, the negative spike in attenuation (D) is also within the suboptimal flow rate region of time.
The purpose of this experiment was to determine how a freshwater tardigrade species performs with a combination of environmental stressors: low pressures, low temperatures, and moderate radiation levels. Hypsibius dujardini was chosen because the species is well-characterized and relatively easy to culture. Approximately 15 hours before launch, 100 tardigrades and 10 exuvias (with enclosed eggs) were collected, washed, and injected into an OptiCell Cell Culture Chamber (Cat#: 392-0603) containing 4 mL of fresh Chalkey’s media and 20 µl of chlorococcum algae. At launch, the OptiCell was secured to the side of the payload. Upon payload retrieval, samples were removed and examined via light microscopy. Live, deceased, intact, and destroyed organisms/eggs were enumerated.
Approximately six hours after retrieval of the SHAB-2 payload, the OptiCell was recovered, analyzed and examined. We suppose that, because the payload reached a maximum altitude of 20421.6m with a minimum temperature of -18°C, only 7 of the original 100 tardigrades were fully intact, with some appearing to become malformed and distorted as compared their original morphology. All intact tardigrades were deceased, with microscopy examples of deceased tardigrades in the Figre. Closer examination of debris, which was not present in the OptiCell before flight, revealed that it consisted of tardigrade limbs and segments, including squished (A) and headless (D) tardigrades. We hypothesize that the tardigrades experienced rapid depressurization while onboard the ascending HAB payload. As a result of the rapid loss of pressure, gaseous nitrogen (N2) and oxygen (O2) likely bubbled out of the tardigrades’ coelomic fluid, causing rapid expansion of their interior volume and whole-body rupture. None of the eggs injected into the OptiCell appeared to be viable, as observed over the course of a one week post-flight egg-monitoring study.
It is important to note that these results were observed without a ground control condition. Thus, we cannot be confident that our results are due to the experimental conditions. We encourage future teams to validate these results.
The goal of this experiment was to assess the survivorship of various bacteria in the upper troposphere and lower stratosphere. Biological Sample Trays (BSTs) outfitted with aluminum coupons containing microbial samples were carried aloft and exposed to the ambient conditions for a desired duration. Coupons with identical cell concentrations were held in a sterile biosafety cabinet as a ground control.
Prior to flight, aluminum coupons were autoclaved, allowed to cool and dry, and inoculated with 10 µL of microbial culture in quadruplicate. Our methods used cultures of Pseudomonas syringae (B728A) grown to a concentration of 109 CFU/mL and Paenibacillus xerothermodurans grown to a concentration of 105 CFU/mL, enumerated by the MPN method. After flight, the coupons were aseptically removed from the payload. Cells were washed from the coupons using 20 µL of sterile water, which was pipetted up and down until cells no longer adhered to coupon surface. P. syringae was plated onto King’s Hard Agar and P. xerothermodurans was plated onto R2A minimal media. Viable cells were enumerated by the MPN method.
This experiment was designed to study the possible association between black carbon aerosols and microbial biomass. Two Biological Sample Trays (BSTs) were outfitted with filter paper-covered coupons for sampling of microbial cells. As the balloon rose, the onboard flight computer controlled the actuation of the BSTs to the exterior of the payload at predetermined altitudes (e.g. 35,000ft, 90,000ft). Biological material was collected as the BSTs rose by impact of the laminar flow of air against the filter paper. After 900 seconds, the BSTs retracted to the interior of the payload and shielded the paper from further ambient exposure. Upon payload recovery, the filter paper was removed and placed in a sterile, conical tube along with 5 mL of sterile water, and the tubes were vortexed. Aliquots of 1 mL were plated onto plates of King’s Agar (nutrient rich) and R2A minimal media. Viable cells were enumerated by the most probable number (MPN) method.
We note that the BSTs provided likely do not allow for optimal air flow through the filters as the coupon is a solid sheet of acrylic. We encourage future teams to optimize this design so as to maximize the volume of air that can be sampled by laminar flow.