W.H. JEFFREY, Center for Environmental Diagnostics and Bioremediation, University of West Florida, Pensacola, Florida 32514
R.V. MILLER, Department of Microbiology, Oklahoma State University, Stillwater, Oklahoma 74078
D.L. MITCHELL, M.D. Anderson Cancer Center, Smithville, Texas 78957
There is now strong evidence that ultraviolet radiation (UVR) is increasing over certain locations on the Earth's surface. Of primary concern has been the annual pattern of ozone depletion over Antarctica and the southern oceans where ozone levels have declined as much as 74 percent compared to pre-ozone-hole events. Reduction of ozone concentration selectively limits stratospheric adsorption of ultraviolet-B (UV-B) radiation [280-320 nanometers (nm)], resulting in higher irradiance on the Earth's surface. As a result, studies of the impact of natural UVR on marine microorganisms have received much attention. The impact of increased UV-B due to ozone depletion on phytoplankton and primary production has attracted extensive interest. The effects of UV-B on bacteria, in contrast, have been largely overlooked. It is apparent from previous studies in the southern ocean and elsewhere that bacteria play a vital role in mineralization of nutrients and provide a trophic link to higher organisms. The objectives of our study have been to identify the effects of UVR and ozone depletion on bacterioplankton in the southern ocean. Our approach has been to combine state-of-the-art molecular approaches with more traditional microbial ecology methodologies. We have examined the extent of DNA damage in bacterioplankton resulting from UVR and as a function of ozone depletion with the ultimate goal of estimating the effect of the stress on carbon fluxes through bacterial assemblages.
Cyclobutane pyrimidine dimers (CPDs) are one of the unique photoproducts created by UV-B and these DNA lesions may be identified using radioimmunoassays (Mitchell 1996). If these photolesions are not repaired, they may affect bacterial DNA and mRNA synthesis resulting in gene mutations, altered physiological activities, or lethality. Because they are induced by UV-B, quantification of these photoproducts is a direct means by which UVR effects may be monitored.
Samples were collected during two research cruises aboard the R/V Polar Duke in the Gerlache Strait (approximately 64°20'S 62°00'W) between 12 October and 5 November 1995 and 1 and 25 October 1996. By limiting the ship's travel, we also minimized variability in results due to changing water masses while experiencing significant fluctuations in column ozone concentrations. Conditions were very different between the two cruises. The 1995 cruise was characterized by very heavy ice, whereas in 1996, ice was minimal at the beginning of the cruise but formed as the cruise progressed. Air temperatures were significantly colder in 1996, and the much heavier cloud cover and snow in 1996 reduced the quality of light compared to the 1995 cruise. Water temperatures were approximately -1.5°C in 1996. In 1995, they averaged approximately -0.6°C. In contrast, production was much greater in 1966: microbial biomass was approximately an order of magnitude greater than in 1995.
Our primary objectives were to determine the distribution of UV-B-induced DNA damage as a function of depth in the water column, time of day (diel studies), and how these may change as a function of ozone conditions and sea state (i.e., surface-water mixing). Results from representative depth profiles are presented herein. Depth profiles of damage were determined by collecting water at discrete depths at sunrise and again near sunset. The bacterioplankton fraction was separated from the larger organisms and concentrated by filtration onto 0.2-micrometer (m) pore filters. DNA damage was determined upon return to our laboratories (Jeffrey et al. 1996). Potential damage with depth was estimated by deploying DNA dosimeters (solutions of calf thymus DNA in quartz tubes; Jeffrey et al. 1996) at discrete depths during sunlight hours. By comparing DNA damage collected from in situ dosimeters with water column bacterioplankton depth samples, we have been able to identify the role of mixing in distribution of UV-B effects.
On calm days, DNA damage was maximal at the surface, decreased with depth, and was detectable to approximately 20 meters (m) when skies were clear and ozone concentrations low (figure 1). In contrast, on days with significant wind-driven mixing, a very different pattern was observed (figure 2). Cyclobutane dimers (figure 2 A ) in the bacterioplankton population did not significantly increase in the water column during the day. Dosimeters held at fixed depth in the water column on that day demonstrate that in the absence of mixing, damage is maximal at the surface and decreases with depth as would be predicted by light-attenuation profiles. These results suggest that when mixing occurs, planktonic cells do not remain at shallow depths, where damage may occur, long enough for significant damage induction. Instead, they spend the majority of the day at deeper depths where damage is minimal, but where repair processes (both photoreactivation and light-independent repair) continue.Our data suggest that the extent of UV-B effects (e.g., DNA damage) may not be predictable from profiles of UVR attenuation in the water column. In the majority of instances, we observed marked differences in the amount of damage in the water column samples compared to in situ incubations. Similar results were also obtained in the Gulf of Mexico (Jeffrey et al. 1996). Although DNA damage may occur in surface waters to significant depths (e.g., >20 m) during calm seas, high winds and surface-water mixing found in high-energy environments such as the southern oceans may result in reduced impact of UVR. These results demonstrate the difficulty in predicting in situ UVR effects based on physical measurements. Further, although these effects have been observed for DNA damage in bacterioplankton, other reports have suggested that surface-water mixing may intensify the distribution of phytoplankton production inhibition (Cullen et al. 1994). These results suggest that different trophic levels and processes may be affected differently by changes in the physical environment, further complicating predictive modeling of UVR effects.
This work was supported by National Science Foundation grant number OPP 94-19037. We thank Peter Aas, Melissa Booth, Richard Coffin, Ross Downer, Sonya Holder, LeAnna Hutchinson, Cheryl Kelley, Maille Lyons, Erin McKee, Dean Pakulski, and Steven Ripp for sample collection aboard the R/V Polar Duke cruises.
Cullen, J.J., P.J. Neale, R.F. Davis, and D.R.S. Lean. 1994. Ultraviolet radiation, vertical mixing, and primary productivity in the Antarctic. EOS, Transactions of the American Geophysical Union , 75, 200.
Jeffrey, W.H., R.J. Pledger, P. Aas, S. Hager, R.B. Coffin, R. Von Haven, and D.L. Mitchell. 1996. Diel and depth profiles of DNA photodamage in bacterioplankton exposed to ambient solar ultraviolet radiation. Marine Ecology Progress Series, 137, 293-304.
Mitchell, D.L. 1996. Radioimmunoassay of DNA damaged by ultraviolet light. In G. Pfeifer (Ed.), Technologies for detection of DNA damage and mutations . New York: Plenum Publishing.