Hostname: page-component-cd9895bd7-q99xh Total loading time: 0 Render date: 2024-12-28T18:15:53.008Z Has data issue: false hasContentIssue false

A molecular method to assess bioburden embedded within silicon-based resins used on modern spacecraft materials

Published online by Cambridge University Press:  16 February 2012

Christina N. Stam*
Affiliation:
Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA
James Bruckner
Affiliation:
Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA
J. Andy Spry
Affiliation:
Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA
Kasthuri Venkateswaran
Affiliation:
Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA
Myron T. La Duc
Affiliation:
Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA

Abstract

Current assessments of bioburden embedded in spacecraft materials are based on work performed in the Viking era (1970s), and the ability to culture organisms extracted from such materials. To circumvent the limitations of such approaches, DNA-based techniques were evaluated alongside established culturing techniques to determine the recovery and survival of bacterial spores encapsulated in spacecraft-qualified polymer materials. Varying concentrations of Bacillus pumilus SAFR-032 spores were completely embedded in silicone epoxy. An organic dimethylacetamide-based solvent was used to digest the epoxy and spore recovery was evaluated via gyrB-targeted qPCR, direct agar plating, most probably number analysis, and microscopy. Although full-strength solvent was shown to inhibit the germination and/or outgrowth of spores, dilution in excess of 100-fold allowed recovery with no significant decrease in cultivability. Similarly, qPCR (quantitative PCR) detection sensitivities as low as ∼103 CFU ml−1 were achieved upon removal of inhibitory substances associated with the epoxy and/or solvent. These detection and enumeration methods show promise for use in assessing the embedded bioburden of spacecraft hardware.

Type
Research Article
Copyright
Copyright © Cambridge University Press 2012

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Baker, S.S., Rugh, C.L. & Kamalay, J.C. (1990). Biotechniques 9, 268272.Google Scholar
Berhards, J., Weitzel, B., Werner, M., Rimpler, M. & Georgii, A. (1992). Histochem. 98, 145154.CrossRefGoogle Scholar
Bizzini, A., Jaton, K., Romo, D., Bille, J., Prod'hom, G. & Greub, G. (2010). J. Clin. Microbiol. 48, 15491554.CrossRefGoogle Scholar
Brewer, W.A., Paik, W.W., Smith, C.D., Robillard, C.L. & Green, R.H. (1972). Orig. Life Evol. Biosph. 3, 198205.CrossRefGoogle Scholar
Cano, R., Poinar, H. & Poinar, G.O. Jr. (1992). Med. Sci. Res. 20, 249251.Google Scholar
Dickinson, D.N., La Duc, M.T., Haskins, W.E., Gornushkin, I., Winefordner, J.D., Powell, D.H. & Venkateswaran, K. (2004). Appl. Environ. Microbiol. 70, 475482.CrossRefGoogle Scholar
Driks, A. (1999). Microbiol. Mol. Biol. Rev. 63, 120.CrossRefGoogle Scholar
Gilbert, M.T., Cuccui, J., White, W., Lynnerup, N., Titball, R.W., Cooper, A. & Prentice, M.B. (2004). Microbiology 150, 341354.CrossRefGoogle Scholar
Kempf, M.J., Chen, F., Kern, R. & Venkateswaran, K. (2005). Astrobiology 5, 391405.CrossRefGoogle Scholar
La Duc, M.T., Nicholson, W., Kern, R. & Venkateswaran, K. (2003). Environ. Microbiol. 5, 977985.CrossRefGoogle Scholar
La Duc, M.T., Osman, S. & Venkateswaran, K. (2009b). J. Rapid Meth. Auto. Microbiol. 17, 350368.CrossRefGoogle Scholar
La Duc, M.T., Osman, S., Vaishampayan, P., Piceno, Y., Andersen, G., Spry, J.A. & Venkateswaran, K. (2009a). Appl. Environ. Microbiol. 75, 65596567.CrossRefGoogle Scholar
Lampel, K.A., Orlandi, P.A. & Kornegay, L. (2000). Appl. Environ. Microbiol. 66, 45394542.CrossRefGoogle Scholar
Muller, S. & Nebe-von-Caron, G. (2010). FEMS Microbiol. Rev. 34, 554587.CrossRefGoogle Scholar
Nicholson, W. & Setlow, P. (1990). in Molecular Biological Methods for Bacillus, pp. 391450. John Wiley and Sons Inc., New York, NY.Google Scholar
NRC. (2006). Preventing the Forward Contamination of Mars, Committee on Preventing the Forward Contamination of Mars, National Research Council, National Academies Press, Washington, DC.Google Scholar
Olsson-Francis, K., de la Torre, R. & Cockell, C.S. (2010). Appl. Environ. Microbiol. 76, 21152121.CrossRefGoogle Scholar
Puleo, J.R., Fields, N.D., Bergstrom, S.L., Oxborrow, G.S., Stabekis, P.D. & Koukol, R. (1977). Appl. Environ. Microbiol. 33, 379384.CrossRefGoogle Scholar
Raghunathan, A., Ferguson, H.R. Jr., Bornarth, C.J., Song, W., Driscoll, M. & Lasken, R.S. (2005). Appl Environ Microbiol. 71, 33423347.CrossRefGoogle Scholar
Rowe, R., Todd, R. & Waide, J. (1977). Appl. Environ. Microbiol. 33, 675680.CrossRefGoogle Scholar
Satomi, M., La Duc, M.T. & Venkateswaran, K. (2006). Int. J. Syst. Evol. Microbiol. 56, 17351740.CrossRefGoogle Scholar
Schaeffer, P., Millet, J. & Aubert, J. (1965). Proc. Natl. Acad. Sci. U.S.A 54, 704711.CrossRefGoogle Scholar
Setlow, P. (2007). Trends Microbiol. 15, 172180.CrossRefGoogle Scholar
Vasin, V.B. & Trofimov, V.I. (1995). Adv. Space Res. 15, 273276.CrossRefGoogle Scholar
Venkateswaran, K., Chung, S., Allton, J. & Kern, R. (2004). Astrobiology 4, 377390.Google Scholar
Venkateswaran, K., Hattori, N., La Duc, M.T. & Kern, R. (2003). J. Microbiol. Methods 52, 367377.CrossRefGoogle Scholar
Venkateswaran, K., Satomi, M., Chung, S., Kern, R.G., Koukol, R., Basic, C. & White, D. (2001). Syst. Appl. Microbiol. 24, 311320.CrossRefGoogle Scholar
Wang, H. & Edwards, K.J. (2009). Geomicrobiol. J. 26, 463469.CrossRefGoogle Scholar
Wang, Y., Hammes, F., De Roy, K., Verstraete, W. & Boon, N. (2010). Trends Biotechnol. 28, 416424.CrossRefGoogle Scholar
Ward, D.M., Weller, R. & Bateson, M.M. (1990). Nature 345, 6365.CrossRefGoogle Scholar
Zeitlin, C., Guetersloh, S.B., Heilbronn, L.H. & Miller, J. (2006). Nucl. Instrum. Methods Phys. Res. B, Beam Interact. Mat. At. (Netherlands) 252, 308318.CrossRefGoogle Scholar
Zitterkopf, N. L. (2008). LabMedicine 39, 623625.Google Scholar