Wade H. Jeffrey

AMBIENT SOLAR RADIATION-INDUCED PHOTODAMAGE IN MARINE BACTERIOPLANKTON

Abstract— There has been much recent concern about the effects of increased UV radiation at certain locations on the earths surface. There have been extensive studies of ultraviolet radiation effects on phytoplankton and primary production, yet the effects of UVB upon bacterioplankton have been largely overlooked. Bacteria play a central role in the cycling of nutrients and energy flow to higher trophic levels, serving as both mineralizers and secondary producers that are consumed by higher organisms. We have begun to investigate the induction of DNA photodamage by UVB in marine planktonic communities using a highly specific radioimmunoassay to measure cyclobutane pyrimidine dimers in samples collected from the northern Gulf of Mexico. DNA damage in the bacterioplankton size‐fraction (< 0.8 μ.m) was greater than in the larger eukaryotic size fraction (>0.8 μm <120 μm) in 9 of 10 samples. Diel patterns of dimer accumulation and repair were observed in surface waters over a 48 h period in the bacterioplankton size fraction and in the larger eukaryotic plankton size fraction. Depth profiles of DNA damage in the bacterioplankton size fraction appear to be dependent on surface water mixing. Damage was greatest in surface waters, decreased with depth and could be detected to 10 m in calm seas. No net accumulation of damage was observed in moderate seas, even at the surface. Solar radiation was found to inhibit significantly both 3H‐thymidine and 14C‐leucine incorporation. Ultraviolet B was responsible for approximately half of the total inhibition of 3H‐thymidine incorporation, UVA contributing the other half of the inhibition. The vast majority of 14C‐leucine incorporation inhibition was due to UVB, suggesting that protein synthesis is less affected by UVA. The results demonstrate that direct measures of DNA damage can be made of indigenous planktonic communities and that bacterioplankton are highly susceptible to UVB damage and may serve as a more sensitive indicator of UVR stress than other microorganisms.

Estimation of Biologically Damaging UV Levels in Marine Surface Waters with DNA and Viral Dosimeters

Abstract We have surveyed the biologically harmful radiation penetrating the water column along a transect in the western Gulf of Mexico using dosimeters consisting of intact viruses or naked calf-thymus DNA (ctDNA). The indigenous marine bacteriophage PWH3a-P1, which lytically infects the heterotrophic bacterium Vibrio natriegens (strain PWH3a), displayed decay rates for infectivity approaching 1.0 h−1 in surface waters when deployed in a seawater-based dosimeter. The accumulation of pyrimidine dimers in ctDNA dosimeters provided a strong correlation to these results, with pyrimidine dimers representing more than 0.3% (up to ca 3800 dimers Mb−1 DNA) of the total DNA in dosimeters exposed to sea surface levels of solar radiation. The results demonstrate a strong correlation between the dimer formation in the DNA dosimeters, the decay rates of viral infectivity and the penetration of UVB radiation into the water column. The decay of viral infectivity attenuated with depth in a manner similar to the decay of solar radiation and was still significant at 10 m in offshore oligotrophic water and at dimer frequencies less than 0.1% (ca 200–300 dimers Mb−1 DNA).

Sunlight-induced DNA Damage in Marine Micro-organisms Collected Along a Latitudinal Gradient from 70°N to 68°S

We examined ultraviolet radiation (UVR)‐induced DNA damage in marine micro‐organisms collected from surface seawater along a latitudinal transect in the Central Pacific Ocean from 70°N to 68°S. Samples were collected predawn and incubated under ambient UVR in transparent incubators at in situ temperatures until late afternoon at which time they were filtered into primarily bacterioplankton and eukaryotic fractions. Cyclobutane pyrimidine dimers (CPDs) and (6‐4) photoproducts [(6‐4)PDs] were quantified in DNA extracts using radioimmunoassays. UVB was lowest in the polar regions and highest near the equator and correlations between UVB and DNA damage were observed. The eukaryotic fraction showed significant CPDs across the entire transect; (6‐4)PDs were detected only in the tropics. The bacterial fraction showed no accumulation of (6‐4)PDs at any latitude, although residual (6‐4)PDs were observed. Bacterial cell volumes were greatest in the sub‐Arctic and northern temperate latitudes and lower in the tropics and southern hemisphere, a unique observation that parallels Bergmann’s rule. A strong negative correlation was observed between cell volume and CPDs. The environmental impact of solar UVR on marine micro‐organisms in the open ocean is complex and our results suggest that several factors such as DNA repair, cell size, temperature, salinity, nutrients and species composition are important in determining relative sensitivity.

Development and Application of a Novel Immunoassay for Measuring Oxidative DNA Damage in the Environment

Abstract We developed a facile, cost-effective competitive binding assay for the analysis of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) in DNA, using a polyclonal rabbit antiserum raised against an 8-oxodGuo hapten coupled to bovine serum albumin and radiolabeled synthetic ligand containing multiple 8-oxodGuo residues. This radioimmunoassay (RIA) displays a high affinity for 8-oxodGuo in DNA, with a detection limit of ∼1 adduct in 105 bases of DNA. 8-oxodGuo standards for RIA were quantified by high-performance liquid chromatography and electrochemical detection in DNA diluted in methylene blue and exposed to visible light. As an initial application we quantified 8-oxodGuo in dosimeters deployed at increasing depths in the Southern Ocean during the austral spring of the 1998 field season or at the surface at Palmer Station, Antarctica, throughout the 1999 field season. Cyclobutane pyrimidine dimers (CPD) were quantified using an established RIA. We found that the frequency of both photoproducts decreased with depth. However, CPD induction was attenuated at a faster rate than 8-oxodGuo, correlating with the differential attenuation of solar ultraviolet wavelengths in the water column. CPD induction was closely related with ultraviolet-B radiation (UVB) attenuation, whereas the lower attenuation of 8-oxodGuo suggests that oxidative damage is more closely related to ultraviolet-A radiation (UVA) irradiance. The ratio of 8-oxodGuou200a:u200aCPD was also found to covary with changes in stratospheric ozone concentrations at Palmer Station. These data demonstrate the usefulness of these assays for environmental photobiology and the potential for their use in studying the relative impacts of UVB versus UVA, including ozone depletion events.

Biogeography and environmental genomics of the Roseobacter -affiliated pelagic CHAB-I-5 lineage

The identification and functional characterization of microbial communities remains a prevailing topic in microbial oceanography as information on environmentally relevant pelagic prokaryotes is still limited. The Roseobacter group, an abundant lineage of marine Alphaproteobacteria, can constitute large proportions of the bacterioplankton. Roseobacters also occur associated with eukaryotic organisms and possess streamlined as well as larger genomes from 2.2 to >5u2005Mpb. Here, we show that one pelagic cluster of this group, CHAB-I-5, occurs globally from tropical to polar regions and accounts for up to 22% of the active North Sea bacterioplankton in the summer. The first sequenced genome of a CHAB-I-5 organism comprises 3.6u2005Mbp and exhibits features of an oligotrophic lifestyle. In a metatranscriptome of North Sea surface waters, 98% of the encoded genes were present, and genes encoding various ABC transporters, glutamate synthase and CO oxidation were particularly upregulated. Phylogenetic gene content analyses of 41 genomes of the Roseobacter group revealed a unique cluster of pelagic organisms distinct from other lineages of this group, highlighting the adaptation to life in nutrient-depleted environments.

Measurement of UVB-induced DNA damage in marine planktonic communities

Publisher Summary Ultraviolet radiation (UVR) has been recognized for many years as a potential stress for organisms in a variety of environments. Decrease in phytoplankton production may result in a decline in bacterial production, which may be compounded by direct UVB effects on bacterioplankton. UVR may directly affect viruses, bacteria, phytoplankton, or zooplankton via direct DNA damage and reduced rates of production. The effects of UVR on marine bacterioplankton are investigated by using radiolabeled precursor molecules such as 3 H thymidine, and 3 H- or 14 C-leucine. It has been shown that bacterioplankton experience significant amounts of DNA damage (CPDs) in surface waters, often twice the amount of larger eukaryotic cells. Bacterioplankton accumulate DNA damage over a solar day. DNA damage extend to depths of 10 m or more in calm waters but the amount of damage may be significantly altered by surface water mixing events. Radioimmunoassay (RIA) technique is used for the measurement of specific DNA photoproducts in the DNA of UV-irradiated cells. DNA damage results are reported per unit (megabase) DNA, and are therefore independent of the concentration of DNA present in the original sample or the amount of DNA assayed. The sensitivity of the RIA is determined by the affinity of the antibody and specific activity of the radiolabeled antigen (probe).