Secondary Mission - Laboratory Analysis of Specimens

The biology mission at MDRS was focused on laboratory analysis of both the 09 Feb 02 and 10 Feb 02 samples and the exploration of new territory by the EVA team.

Secondary biology mission goals were met today through laboratory analysis of the toe different sorts of samples returned from the EVAs of 09FEB02 and 10FEB02. Way Point 18 apparent endolithic organisms (green layer just subsurface of exposed sunside rock surface) were used to develop a technique for extracting endolithic cells from the rock substrate. Way Point 23 apparent sublithic (AKA hypolithic) organisms (green coating at junction of exposed rock surface and buried rock surface) was used to develop techniques for removing lithic microorganisms from hard rock surfaces that cannot be easily fragmented.

In order to prepare the endolithic sample for analysis, subsamples of about 10 grams from each was placed into microfuge tubes. In order to get the sample from the rock substrate, hard steel snips were used to chip small fragments from the region possessing the green layer under the rock surface. These chips were further pulverized using a flat surface plier. The chips and finer materials were collected into virgin ziplock bags. By hand sifting, a course fragment partition was separated from a finer fragment partition. The finer fragments were then placed into a 10ml centifuge tube and hand settled into an upper courser layer and the extrafine materials on bottom – the course layer was poured off and placed into a separate centrifuge tube. The finest layer was then used to recreate the sample protocol established yesterday for friable hypoliths. (e.g., subsample was solubilized in about 1.0 ml of sterile H2O, vortexed for 30 seconds, and microfuged for about 1.0 minute. The microfuged samples were then decanted, saving the relatively cleared liquid above the pellet into a separate microfuge tube. The remaining pellet in each case was then used to subsample the surface of each pellet by taking a small portion of the pellet surface and solubilize same in a separate microfuge tube using a very small amount of sterile water (just enough to create a very thick susupension of the pellet surface material). The samples were then subjected to various microscopic analyses. In each case, where possible under the conditions of the laboratory (handheld digital camera focusing through the monocular eyepiece), a digital photographic record was made. Each dry sub sample was used to visualize dry sample using the dissection (gross specimen) microscope. Each liquid aliquot (decanted liquid and resuspended pellet) was viewed at each magnification of the Olympus scope as a dried slide smear (no slip cover), and as a wet mount. Wet mount slides were further subjected to oil immersion microscopy. All microscopic mounts were viewed under bright and dark (fluorescent – none, red, and yellow filtered) field analysis.

One subsample of each of the endolith and sublithic organism was used to create a 2% glutaraldehyde preserved sample and placed under refrigeration. One such subsample was saved for analysis in the tertiary mission (organophosphorus hydrolase activity).

The microscopy of the endolithic sample showed clear evidence of cellular organisms (cellular inclusions, ovoid, pigmented). Certain of the cells were very small and at least certain of them appeared to be motile (directional movement in excess of movement due to Brownian motion). Such cells were observed in both the decanted liquid and in the resuspended pellet. There were also crystalline structures. There were apparent colonial (end to end, segmented or branched) structures of much larger cells. All motile cells appeared to be individual. None of the cells appeared to fluoresce (brightly, at least) under any filter scenario (at least in a fashion or intensity consistent with the control fluorescent beads).

The microscopy of the sublithic sample also showed clear evidence of cellular organisms (cellular inclusions, ovoid, pigmented). There were very few individual cells as opposed to other samples imaged to date. There were apparent colonial (end to end, segmented or branched) structures of average sized cells. No motile cells appeared. Certain of the colonial structures appeared to fluoresce using a yellow filter UV. The flouresence might be mere background, however it was differentiated between green, yellow and orange colors. None of the cells appeared to fluoresce under any filter scenario in a fashion or intensity consistent with the control fluorescent beads).

In conclusion, with further analysis still to do by way of confirmation, it would appear that we have devised a way to extract and image microorganism associated with both endolithic and tightly bound to hard rock sublithic surface microorganisms. The microscopy of the endolithic sample showed clear evidence of a substantial green-colored cellular organism. Such cells were observed in both the decanted liquid and in the resuspended pellet.

Future work will include both the tertiary analysis and starting to do the lithic organisms survey.

Primary Mission - Further Biological Exploration

Today’s 11FEB02 EVA team visited several sites starting with the Lower Blue Hills area northwest of the hab, and finishing close to the Coal Mine wash area. Vegetation was plentiful in the Lower Blue Hills area and became sparser the further northwest we went. In the Lower Blue Hills area there were few rocks to examine for lithic organisms. However, there were mollusk fossils like those found yesterday in this location. Other landscapes further north and closer to the Coal Mine wash area contained many varieties of rocks that did not appear to have growth on any location about the rocks. We then descended to Waypoint 35 and found large rocks covered in a desert varnish fashion, as described yesterday. There may be endolithic growth below the desert varnish, and a sample was taken for lab analysis. Lichens were plentiful in this area, including a large white variety not seen in other locations. Frozen water was found at Waypoint 36, and a sample was taken. This is the only water found so far in our area and will be microscopically examined. Also, a green clay-like sample was taken in the same area for comparison with suspected green algae/cyanobacteria we are seeing in secondary mission analyses. See the other report section for Waypoint locations and descriptions.