Isolation of novel extreme-tolerant cyanobacteria from a rock-dwelling microbial community by using exposure to low Earth orbit

Abstract

Many cyanobacteria are known to tolerate environmental extremes. Motivated by an interest in selecting cyanobacteria for applications in space, we launched rocks from a limestone cliff in Beer, Devon, United Kingdom, containing an epilithic and endolithic rock-dwelling community of cyanobacteria into low Earth orbit (LEO) at a height of approximately 300 kilometers. The community was exposed for 10 days to isolate cyanobacteria that can survive exposure to the extreme radiation and desiccating conditions associated with space. Culture-independent (16S rRNA) and culture-dependent methods showed that the cyanobacterial community was composed of Pleurocapsales, Oscillatoriales, and Chroococcales. A single cyanobacterium, a previously uncharacterized extremophile, was isolated after exposure to LEO. We were able to isolate the cyanobacterium from the limestone cliff after exposing the rock-dwelling community to desiccation and vacuum (0.7 x 10(-3) kPa) in the laboratory. The ability of the organism to survive the conditions in space may be linked to the formation of dense colonies. These experiments show how extreme environmental conditions, including space, can be used to select for novel microorganisms. Furthermore, it improves our knowledge of environmental tolerances of extremophilic rock-dwelling cyanobacteria.

FIG. 1.

(A) Map of the United Kingdom showing the location of the sample site, Beer, Devon, United Kingdom. (Reprinted with permission of The Open University, Milton Keynes, United Kingdom.) (B) The rocks were collected from the upper greensand layer of the limestone cliffs, which is covered with a homogeneous epilithic covering of cyanobacteria.

FIG. 2.

Evaluation of the representation of the bacterial clones obtained from the limestone rock from Beer by rarefaction analysis. The appearance of new OTU strains was plotted as a function of the number of clones, which were analyzed randomly. The rarefaction curves were generated using FastGroupII (49). The curve was calculated with sequence similarity cutoff values of 97.0% (species level).

FIG. 3.

16S rRNA gene phylogenetic tree of the cyanobacterial community of the limestone rocks. The neighbor-joining tree was constructed using an approximately 700-nucleotide region of the 16S rRNA gene, which was amplified with cyanobacterium-specific primers (31). The phylogenetic tree was based on 92 sequences from the clone library, 9 sequences from the isolates, and 56 cyanobacterial sequences from GenBank, which were cut to the same length as the clone sequences. The phylotypes contained the following number of sequences: phylotype A, 33; phylotype B, 2; phylotype C, 10; phylotype D, 3; phylotype E, 13; phylotype F, 2; phylotype G, 9; phylotype H, 5; phylotype I, 9; phylotype J, 12; phylotype K, 1; phylotype L, 1; phylotype M, 1; and phylotype N, 1. The phylotypes are in boldface, and OU_20 is underlined. The scale bar corresponds to 0.02 changes per nucleotide. The percentage of bootstrap replicates (1,000 replicates) resulting in the same cluster is given near the respective nodes for bootstrap values higher than 80%.

FIG. 4.

TEM images of the cyanobacterium OU_20 that survived exposure to low Earth orbit.