Quantification of encapsulated bioburden in spacecraft polymer materials by cultivation-dependent and molecular methods

PLoS One. 2014 Apr 15;9(4):e94265. doi: 10.1371/journal.pone.0094265. eCollection 2014.

Abstract

Bioburden encapsulated in spacecraft polymers (such as adhesives and coatings) poses a potential risk to jeopardize scientific exploration of other celestial bodies. This is particularly critical for spacecraft components intended for hard landing. So far, it remained unclear if polymers are indeed a source of microbial contamination. In addition, data with respect to survival of microbes during the embedding/polymerization process are sparse. In this study we developed testing strategies to quantitatively examine encapsulated bioburden in five different polymers used frequently and in large quantities on spaceflight hardware. As quantitative extraction of the bioburden from polymerized (solid) materials did not prove feasible, contaminants were extracted from uncured precursors. Cultivation-based analyses revealed <0.1-2.5 colony forming units (cfu) per cm3 polymer, whereas quantitative PCR-based detection of contaminants indicated considerably higher values, despite low DNA extraction efficiency. Results obtained from this approach reflect the most conservative proxy for encapsulated bioburden, as they give the maximum bioburden of the polymers irrespective of any additional physical and chemical stress occurring during polymerization. To address the latter issue, we deployed an embedding model to elucidate and monitor the physiological status of embedded Bacillus safensis spores in a cured polymer. Staining approaches using AlexaFluor succinimidyl ester 488 (AF488), propidium monoazide (PMA), CTC (5-cyano-2,3-diotolyl tetrazolium chloride) demonstrated that embedded spores retained integrity, germination and cultivation ability even after polymerization of the adhesive Scotch-Weld 2216 B/A. Using the methods presented here, we were able to estimate the worst case contribution of encapsulated bioburden in different polymers to the bioburden of spacecraft. We demonstrated that spores were not affected by polymerization processes. Besides Planetary Protection considerations, our results could prove useful for the manufacturing of food packaging, pharmacy industry and implant technology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacillus / genetics
  • Bacillus / isolation & purification*
  • Bacillus / physiology
  • Microbial Viability
  • Polymers*
  • Solvents / chemistry
  • Spacecraft*
  • Spores, Bacterial / physiology

Substances

  • Polymers
  • Solvents

Grants and funding

This study has been funded by the European Space Agency (ESA) in the context of the project ‘Evaluation of Encapsulated Bioburden on Flight Hardware’ (contract no. AO/1-6079/09/NL/CP). This publication was supported by the German research foundation (DFG) within the funding programme Open Access Publishing. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.