Diversity of Microbial Eukaryotes Along the West Antarctic Peninsula in Austral Spring.

During a cruise from October to November 2019, along the West Antarctic Peninsula, between 64.32 and 68.37°S, we assessed the diversity and composition of the active microbial eukaryotic community within three size fractions: micro- (> 20 µm), nano- (20-5 µm), and pico-size fractions (5-0.2 µm). The communities and the environmental parameters displayed latitudinal gradients, and we observed a strong similarity in the microbial eukaryotic communities as well as the environmental parameters between the sub-surface and the deep chlorophyll maximum (DCM) depths. Chlorophyll concentrations were low, and the mixed layer was shallow for most of the 17 stations sampled. The richness of the microplankton was higher in Marguerite Bay (our southernmost stations), compared to more northern stations, while the diversity for the nano- and pico-plankton was relatively stable across latitude. The microplankton communities were dominated by autotrophs, mostly diatoms, while mixotrophs (phototrophs-consuming bacteria and kleptoplastidic ciliates, mostly alveolates, and cryptophytes) were the most abundant and active members of the nano- and picoplankton communities. While phototrophy was the dominant trophic mode, heterotrophy (mixotrophy, phagotrophy, and parasitism) tended to increase southward. The samples from Marguerite Bay showed a distinct community with a high diversity of nanoplankton predators, including spirotrich ciliates, and dinoflagellates, while cryptophytes were observed elsewhere. Some lineages were significantly related-either positively or negatively-to ice coverage (e.g., positive for Pelagophyceae, negative for Spirotrichea) and temperature (e.g., positive for Cryptophyceae, negative for Spirotrichea). This suggests that climate changes will have a strong impact on the microbial eukaryotic community.

Greenhouse gases and biogeochemical diel fluctuations in a high-altitude wetland.

The landscapes of high-altitude wetland ecosystems are characterized by different kinds of aquatic sites, including ponds holding conspicuous microbial life. Here, we examined a representative pond of the wetland landscape for dynamics of greenhouse gases, and their association with other relevant biogeochemical conditions including diel shifts of microbial communities' structure and activity over two consecutive days. Satellite image analysis indicates that the area of ponds cover 238 of 381.3 Ha (i.e., 62.4%), representing a significant landscape in this wetland. Solar radiation, wind velocity and temperature varied daily and between the days sampled, influencing the biogeochemical dynamics in the pond, shifting the pond reservoir of inorganic versus dissolved organic nitrogen/phosphorus bioavailability, between day 1 and day 2. Day 2 was characterized by high dissolved organic nitrogen/phosphorus and N2O accumulation. CH4 presented a positive excess showing maxima at hours of high radiation during both days. The microbial community in the sediment was diverse and enriched in keystone active groups potentially related with GHG recycling including bacteria and archaea, such as Cyanobacteria, Verrucomicrobia, Rhodobacterales and Nanoarchaeaota (Woesearchaeia). Archaea account for the microbial community composition changes between both days and for the secondary productivity in the water measured during day 2. The results indicate that an intense recycling of organic matter occurs in the pond systems and that the activity of the microbial community is correlated with the availability of nutrients. Together, the above results indicate a net sink of CO2 and N2O, which has also been reported for other natural and artificial ponds. Overall, our two-day fluctuation study in a representative pond of a high-altitude wetland aquatic landscape indicates the need to explore in more detail the short-term besides the long-term biogeochemical variability in arid ecosystems of the Andes plateau, where wetlands are hotspots of life currently under high anthropogenic pressure.

Aquatic Thermal Reservoirs of Microbial Life in a Remote and Extreme High Andean Hydrothermal System.

Hydrothermal systems are ideal to understand how microbial communities cope with challenging conditions. Lirima, our study site, is a polyextreme, high-altitude, hydrothermal ecosystem located in the Chilean Andean highlands. Herein, we analyze the benthic communities of three nearby springs in a gradient of temperature (42-72 °C represented by stations P42, P53, and P72) and pH, and we characterize their microbial diversity by using bacteria 16S rRNA (V4) gene metabarcoding and 16S rRNA gene clone libraries (bacteria and archaea). Bacterial clone libraries of P42 and P53 springs showed that the community composition was mainly represented by phototrophic bacteria (Chlorobia, 3%, Cyanobacteria 3%, at P42; Chlorobia 5%, and Chloroflexi 5% at P53), Firmicutes (32% at P42 and 43% at P53) and Gammaproteobacteria (13% at P42 and 29% at P53). Furthermore, bacterial communities that were analyzed by 16S rRNA gene metabarcoding were characterized by an overall predominance of Chloroflexi in springs with lower temperatures (33% at P42), followed by Firmicutes in hotter springs (50% at P72). The archaeal diversity of P42 and P53 were represented by taxa belonging to Crenarchaeota, Diapherotrites, Nanoarchaeota, Hadesarchaeota, Thaumarchaeota, and Euryarchaeota. The microbial diversity of the Lirima hydrothermal system is represented by groups from deep branches of the tree of life, suggesting this ecosystem as a reservoir of primitive life and a key system to study the processes that shaped the evolution of the biosphere.

Oceanic Microplankton Do Not Adhere to the Latitudinal Diversity Gradient.

A latitudinal biodiversity gradient has captivated ecologists for years, and has become a widely recognized pattern in biogeography, manifest as an increase in biodiversity from the poles to the tropics. Oceanographers have attempted to discern whether these distribution patterns are shared with marine biota, and a lively debate has emerged concerning the global distribution of microbes. Limitations in sampling resolution for such large-scale assessments have often prohibited definitive conclusions. We evaluated microbial planktonic communities along a ~ 15,400-km Pacific Ocean transect with DNA from samples acquired every 2 degrees of latitude within a 3-month period between late August and early November 2003. Next-generation sequencing targeting the Bacteria, Archaea, and Eukarya yielded ~ 10.8 million high-quality sequences. Beta-analysis revealed geographic patterns of microbial communities, primarily the Bacteria and Archaea domains. None of the domains exhibited a unimodal pattern of alpha-diversity with respect to latitude. Bacteria communities increased in richness from Arctic to Antarctic waters, whereas Archaea and Eukarya communities showed no latitudinal or polar trends. Based on our analyses, environmental factors related to latitude thought to influence various macrofauna may not define microplankton diversity patterns of richness in the global ocean.

Bacterial Survival under Extreme UV Radiation: A Comparative Proteomics Study of Rhodobacter sp., Isolated from High Altitude Wetlands in Chile.

Salar de Huasco, defined as a polyextreme environment, is a high altitude saline wetland in the Chilean Altiplano (3800 m.a.s.l.), permanently exposed to the highest solar radiation doses registered in the world. We present here the first comparative proteomics study of a photoheterotrophic bacterium, Rhodobacter sp., isolated from this remote and hostile habitat. We developed an innovative experimental approach using different sources of radiation (in situ sunlight and UVB lamps), cut-off filters (Mylar, Lee filters) and a high-throughput, label-free quantitative proteomics method to comprehensively analyze the effect of seven spectral bands on protein regulation. A hierarchical cluster analysis of 40 common proteins revealed that all conditions containing the most damaging UVB radiation induced similar pattern of protein regulation compared with UVA and visible light spectral bands. Moreover, it appeared that the cellular adaptation of Rhodobacter sp. to osmotic stress encountered in the hypersaline environment from which it was originally isolated, might further a higher resistance to damaging UV radiation. Indeed, proteins involved in the synthesis and transport of key osmoprotectants, such as glycine betaine and inositol, were found in very high abundance under UV radiation compared to the dark control, suggesting the function of osmolytes as efficient reactive oxygen scavengers. Our study also revealed a RecA-independent response and a tightly regulated network of protein quality control involving proteases and chaperones to selectively degrade misfolded and/or damaged proteins.

Microbial Activity Response to Solar Radiation across Contrasting Environmental Conditions in Salar de Huasco, Northern Chilean Altiplano.

In high altitude environments, extreme levels of solar radiation and important differences of ionic concentrations over narrow spatial scales may modulate microbial activity. In Salar de Huasco, a high-altitude wetland in the Andean mountains, the high diversity of microbial communities has been characterized and associated with strong environmental variability. Communities that differed in light history and environmental conditions, such as nutrient concentrations and salinity from different spatial locations, were assessed for bacterial secondary production (BSP, 3H-leucine incorporation) response from short-term exposures to solar radiation. We sampled during austral spring seven stations categorized as: (a) source stations, with recently emerged groundwater (no-previous solar exposure); (b) stream running water stations; (c) stations connected to source waters but far downstream from source points; and (d) isolated ponds disconnected from ground sources or streams with a longer isolation and solar exposure history. Very high values of 0.25 µE m-2 s-1, 72 W m-2 and 12 W m-2 were measured for PAR, UVA, and UVB incident solar radiation, respectively. The environmental factors measured formed two groups of stations reflected by principal component analyses (near to groundwater sources and isolated systems) where isolated ponds had the highest BSP and microbial abundance (35 microalgae taxa, picoeukaryotes, nanoflagellates, and bacteria) plus higher salinities and PO43- concentrations. BSP short-term response (4 h) to solar radiation was measured by 3H-leucine incorporation under four different solar conditions: full sun, no UVB, PAR, and dark. Microbial communities established in waters with the longest surface exposure (e.g., isolated ponds) had the lowest BSP response to solar radiation treatments, and thus were likely best adapted to solar radiation exposure contrary to ground source waters. These results support our light history (solar exposure) hypothesis where the more isolated the community is from ground water sources, the better adapted it is to solar radiation. We suggest that factors other than solar radiation (e.g., salinity, PO43-, NO3-) are also important in determining microbial productivity in heterogeneous environments such as the Salar de Huasco.

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 >5 Mpb. 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 3.6 Mbp 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.

Photochemical changes in water accommodated fractions of MC252 and surrogate oil created during solar exposure as determined by FT-ICR MS.

To determine effects of photochemical weathering of petroleum, surrogate and Macondo (MC252) crude oils were exposed to solar radiation during the formation of Water Accommodated Fractions (WAFs) in sterile seawater. Samples were incubated in either unfiltered sunlight, with ultraviolet radiation blocked (Photosynthetically Active Radiation [PAR] only), or in darkness. WAFs were collected at two time points over the course of a week. Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) analyses of water soluble species formed during exposure to sunlight were compared for the different treatments. Photochemical alterations resulted in differences in compound class distributions. In general, surrogate oil was photo-oxidized across a wider carbon number range compared to MC252. While photochemical differences were observed between MC252 and surrogate oils, microbial production in seawater responded similarly to both WAFs from both types of oils with the majority of the inhibition resulting from oil exposure to visible light.

Polycyclic aromatic hydrocarbon concentrations across the Florida Panhandle continental shelf and slope after the BP MC 252 well failure.

The Florida Panhandle continental shelf environment was exposed to oil from the BP oil well failure in the Gulf of Mexico during 2010. Floating mats of oil were documented by satellite, but the distribution of dissolved components of the oil in this region was unknown. Shipek® grab samples of sediments were taken during repeated cruises between June 2010 and June 2012 to test for selected polycyclic aromatic hydrocarbons (PAHs) as indicators of this contamination. Sediments were collected as composite samples, extracted using standard techniques, and PAHs were quantified by GC/MS-SIM. PAHs in samples from the continental slope in May 2011 were highest near to the failed well site and were reduced in samples taken one year later. PAHs from continental shelf sediments during the spill (June 2010) ranged from 10 to 165 ng g(-1). Subsequent cruises yielded variable and reduced amounts of PAHs across the shelf. The data suggest that PAHs were distributed widely across the shelf, and their subsequent loss to background levels suggests these compounds were of oil spill origin. PAH half-life estimates by regression were 70-122 days for slope and 201 days for shelf stations.

The significance of the Dewar valence photoisomer as a UV radiation-induced DNA photoproduct in marine microbial communities.

Induction of pyrimidine dimers in DNA by solar UV radiation has drastic effects on microorganisms. To better define the nature of these DNA photoproducts in marine bacterioplankton and eukaryotes, a study was performed during a cruise along a latitudinal transect in the Pacific Ocean. The frequency of all possible cyclobutane pyrimidine dimers, pyrimidine (6-4) pyrimidone photoproducts (64PPs) and their related Dewar valence isomers (DEWs) was determined by high-performance liquid chromatography-mass spectrometry. Studied samples were bacterioplankton and eukaryotic fractions isolated from sea water either collected before sunrise or exposed to ambient sunlight from sunrise to sunset. Isolated DNA dosimeters were also exposed to daily sunlight for comparison purposes. A first major result was the observation in all samples of large amounts of DEWs, a class of photoproducts rarely considered outside photochemical studies. Evidence was obtained for a major role of UVA in the formation of these photoisomerization products of 64PPs. Considerations on the ratio between the different classes of photoproducts in basal and induced DNA damage suggests that photoenzymatic repair (PER) is an important DNA repair mechanism used by marine microorganisms occupying surface seawater in the open ocean. This result emphasizes the biological role of DEWs which are very poor substrate for PER.

Latitudinal gradients in degradation of marine dissolved organic carbon.

Heterotrophic microbial communities cycle nearly half of net primary productivity in the ocean, and play a particularly important role in transformations of dissolved organic carbon (DOC). The specific means by which these communities mediate the transformations of organic carbon are largely unknown, since the vast majority of marine bacteria have not been isolated in culture, and most measurements of DOC degradation rates have focused on uptake and metabolism of either bulk DOC or of simple model compounds (e.g. specific amino acids or sugars). Genomic investigations provide information about the potential capabilities of organisms and communities but not the extent to which such potential is expressed. We tested directly the capabilities of heterotrophic microbial communities in surface ocean waters at 32 stations spanning latitudes from 76°S to 79°N to hydrolyze a range of high molecular weight organic substrates and thereby initiate organic matter degradation. These data demonstrate the existence of a latitudinal gradient in the range of complex substrates available to heterotrophic microbial communities, paralleling the global gradient in bacterial species richness. As changing climate increasingly affects the marine environment, changes in the spectrum of substrates accessible by microbial communities may lead to shifts in the location and rate at which marine DOC is respired. Since the inventory of DOC in the ocean is comparable in magnitude to the atmospheric CO(2) reservoir, such a change could profoundly affect the global carbon cycle.

Bacterial growth response to photoactive quinones.

Quinones are known producers of reactive oxygen species (ROS) that may be toxic in natural aquatic environments. In this study, the effects of parent quinones and their photodegradation products on bacterial growth were determined, and photochemical ROS formation rates were measured. Using (3)H-leucine incorporation to measure growth of the bacterium Pseudomonas aeruginosa and natural seawater bacterioplankton, growth inhibition was observed when samples were exposed to dichlone, chloranil and sodium anthraquinone-2-sulfonate (AQ2S). For seawater, compared with other quinones tested, dichlone showed the greatest toxicity in the dark, and AQ2S toxicity was greatest during simultaneous exposure to sunlight. Photodegraded chloranil and dichlone showed decreased toxicity compared with nonirradiated samples. For P. aeruginosa, AQ2S and its photodegradation products showed the greatest toxicity during simultaneous exposure to sunlight. Chloranil photodegradation products showed reduced toxicity compared with the parent compound during simultaneous exposure to sunlight. Dichlone was the only compound to show any toxicity to P. aeruginosa in the dark, and its photodegradation products were more toxic than the parent compound. Based on the results of dark and light controlled experiments measuring bacterial growth and estimated ROS production rates, ROS alone does not account for relative differences in toxicity between these quinones.

Temperature and solar radiation interactions on 3H-leucine incorporation by bacterioplankton in a subtropical estuary.

Although the effects of UV radiation are thought to be temperature independent, the photoinhibition of aquatic bacteria may be temperature dependent owing to enzymatic repair kinetics, an important consideration for climate change analyses. We examined the interactions between temperature and solar radiation in water samples collected from the Blackwater River, Pensacola Bay, and the coastal Gulf of Mexico (Florida) in July 2008. Subsamples were incubated in the dark for 20 h at either the in situ temperature, +5 degrees C from in situ or -5 degrees C from in situ after which they were amended with (3)H-leucine and irradiated in full sunlight at their respective temperatures and compared to samples incubated simultaneously in the dark. Temperature and light significantly affected (3)H-leucine incorporation at all locations and interactive effects between temperature and sunlight were found for Pensacola Bay and the Gulf. Generally, warmer waters reduced photoinhibition. The -5 degrees C treatment was always significantly more inhibited than the +5 degrees C treatment, but the in situ temperature and +5 degrees C and -5 degrees C treatments were not always significantly different. Photoinhibition reduction at warmer temperatures suggests specific effects on photobiology not observed in general cellular activity may be important in determining interactive ecosystem effects of climate change.

Sunlight-induced DNA damage in marine micro-organisms collected along a latitudinal gradient from 70 degrees N to 68 degrees 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 degrees N to 68 degrees 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.

Ultraviolet radiation in the rhône river lenses of low salinity and in marine waters of the northwestern mediterranean sea: attenuation and effects on bacterial activities and net community production.

The high content in nutrients of freshwater outflows induces highly productive and buoyant plumes spreading over marine waters (MW). As a consequence, the growth of organisms developing in these low-salinity waters (LSW) might be potentially affected by UV-R (280-400 nm). This study investigated the penetration of UV-R and its impact on net community production (NCP) and bacterial protein (B(PROT)S) and DNA (B(DNA)S) synthesis in mesotrophic-LSW formed from the Rhône River and in oligotrophic MW of the Northwestern Mediterranean Sea (Gulf of Lions) in May 2006. High concentrations of chlorophyll a (up to 8 microg L(-1)) measured in the LSW (<37.8 psu, 0-10 m) were the main factor influencing the diffuse attenuation coefficients (K(d)) of both UV-R and photosynthetically active radiation (PAR). The mean ratio of the K(d) measured between the LSW and the MW increased with wavelength from 2.4 at 305 nm to 2.9 at 380 nm and 3.1 for PAR indicating more similarity in the UV region. NCP was severely inhibited by UV-R at the surface of the LSW, whereas no effect was measured in the surrounding MW. In contrast, B(PROT)S and B(DNA)S were affected deeper by UV-R in the MW (up to 8 m depth) compared to the LSW where inhibition was only observed at the surface. Differences in response of bacteria in LSW and MW are largely explained by differences in UV-R transparency; however, transplant experiments indicate that bacterial assemblages from the MW were also more sensitive to UV-R than those present in the LSW. We also observed that higher activity of bacteria after nutrient additions increased their sensitivity to UV-R during the day, but favored their recovery during the night incubation period for both LSW and MW. Results suggest that riverine and nutrient inputs may alter the effects of UV-R on microbial activity by attenuating the UV-R penetration and by modifying the physiology of bacteria.

Effect of stratospheric ozone depletion and enhanced ultraviolet radiation on marine bacteria at Palmer Station, Antarctica in the early austral spring.

We investigated the interactions between ozone-depleted air masses and subsequent changes in UVB on marine bacterial abundance and production at Palmer Station, Antarctica from September to November 1999. During periods of low total column ozone (TCO), bacterial cell concentrations declined by 57%. Photoinhibition of bacterial [(3)H]-leucine (Leu) and [(3)H]-thymidine (TdR) incorporation due to UVB was greatest during periods of low TCO in September and early October. During diel ( approximately 28 h) exposure experiments, light treatment samples exhibited >75-100% inhibition of TdR incorporation by mid-afternoon. Leu incorporation exhibited maximum inhibition (50-100%) at sunset and early evening hours. Leu and TdR incorporation in light treatment samples did not exhibit recovery during subsequent periods of darkness. Bacterial Leu and TdR incorporation rates were inversely related to Setlow Dose during a period of recovery from low TCO. These data further suggested a threshold exposure below which bacterial Leu and TdR incorporation recovered rapidly. Recovery of bacterial production after acute Setlow Dose exposures lagged recovery of TCO and was linearly related to TCO measured 2 days previously. This lag in recovery may have resulted from the energetically expensive repair of UVR-induced DNA damage acquired during periods of low TCO.

Autoclave method for rapid preparation of bacterial PCR-template DNA.

An autoclave method for preparing bacterial DNA for PCR template is presented, it eliminates the use of detergents, organic solvents, and mechanical cellular disruption approaches, thereby significantly reducing processing time and costs while increasing reproducibility. Bacteria are lysed by rapid heating and depressurization in an autoclave. The lysate, cleared by microcentrifugation, was either used directly in the PCR reaction, or concentrated by ultrafiltration. This approach was compared with seven established methods of DNA template preparation from four bacterial sources which included boiling Triton X-100 and SDS, bead beating, lysozyme/proteinase K, and CTAB lysis method components. Bacteria examined were Enterococcus and Escherichia coli, a natural marine bacterial community and an Antarctic cyanobacterial-mat. DNAs were tested for their suitability as PCR templates by repetitive element random amplified polymorphic DNA (RAPD) and denaturing gradient gel electrophoresis (DGGE) analysis. The autoclave method produced PCR amplifiable template comparable or superior to the other methods, with greater reproducibility, much shorter processing time, and at a significantly lower cost.

Estimation of biologically damaging UV levels in marine surface waters with DNA and viral dosimeters.

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).

Development and application of a novel immunoassay for measuring oxidative DNA damage in the environment.

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 approximately 1 adduct in 10(5) 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-oxodGuo: CPD 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.

Seasonal fluctuation of DNA photodamage in marine plankton assemblages at Palmer Station, Antarctica.

Ultraviolet radiation-induced DNA damage frequencies were measured in DNA dosimeters and natural plankton communities during the austral spring at Palmer Station, Antarctica, during the 1999-2000 field season. We found that the fluence of solar ultraviolet-B radiation (UV-B) at the earth's surface correlated with stratospheric ozone concentrations, with significant ozone depletion observed because of "ozone hole" conditions. To verify the interdependence of ozone depletion and DNA damage in natural microbial communities, seawater was collected daily or weekly from Arthur Harbor at Palmer Station, Antarctica, throughout "ozone season," exposed to ambient sunlight between 0600 and 1800 h and fractionated using membrane filtration to separate phytoplankton and bacterioplankton populations. DNA from these fractions was isolated and DNA damage measured using radioimmunoassay. Under low-ozone conditions cyclobutane dimer concentrations in bacterioplankton and phytoplankton communities were maximal. DNA damage measured in dosimeters correlated closely with ozone concentrations and UV-B fluence. Our studies offer further support to the theory that stratospheric deozonation is detrimental to marine planktonic organisms in the Southern Ocean.