Planetary protection - Approaching uncultivable microorganisms

With ESA's MiDiv project [Rettberg, P., Fritze, D., Verbarg, S., Nellen, J., Horneck, G., Stackebrandt, E., Kminek, G. Final report ESA project MiDiv, ESA contract number 17538/03/NL/VS] the exobiology group at the German Aerospace Centre (DLR) in Cologne started to investigate the microbial diversity found on spacecrafts and in assembly halls used by Europe [Rettberg, P., Nellen, J., Horneck, G., Fritze, D., Verbarg, S., Stackebrandt, E., Kminek, G. Determination of the microbial diversity of spacecraft assembly facilities: first results of the ESA project MiDiv. Adv. Space Res., 2005]. So far this examination was limited to cultivable microorganisms, probably excluding the majority of organisms present. To approach the microbial diversity of uncultivable microorganisms, new methods, have to be implemented into the analysing process. Therefore, we describe in this paper the adaptation of an existing protocol (surface sampling with swabs) to be used with a method based on the detection of DNA-fragments by polymerase chain reaction (PCR). Our data indicate that it is feasible to adapt the standardized sample taking process to be used with the PCR-method. While using a classical swabbing/cultivation approach for the detection of microorganisms on a surface, two impairing factors have to be accounted for. First, not all bacteria present on a surface will be taken up by the swab and second, not all the bacteria taken up will be released again from the swab. This will lead to a diminished overall estimation of the microorganisms present on a surface. Our studies further suggested that the estimated data had to be corrected by a factor of 2-3 to correspond with the actual numbers of spores spotted.