Palmer LTER: Spatial distribution of viruses in the ...
tions (nitrate, phosphate, and silicate) were not significant. Although some significance might be resolved within defined water masses within the database, preliminary findings of high nutrients, low biomass, high assimilation rates, and low carbon-to-nitrogen ratios (Prézelin unpublished data) suggest overall phytoplankton growth was not nutrient-limited but light-limited. One is left asking, however, why distinct phytoplankton communities are evident late in the winter, when successional events would not yet have come into play; whether these early seasonal patterns reflect those left at the end of the last growing season; and whether they will be the prevalent signatures in the upcoming spring/summer season. Our Marine Primary Productivity Group plans to combine the measurements presented here with other LTER datasets to • assess temporal and spatial scales for significant changes in the abundance and diversity of phytoplankton communities within the sampling area, • advance physical-based and optical-based models of primary productivity, and • contribute to ecosystem-based models of trophic interactions within the antarctic food web.
This project was supported by a University of California at Santa Barbara May Company Scholarship to Barbara Sullivan and National Science Foundation grant OCE 93-01322 to Barbara B. Prézelin. Special thanks to B. Boczar, T. Diem, T. Westberry, M. Moline, and the crew of the R/V Polar Duke. (This is LTER contribution number 43.)
References Hofmann, E.E., B.L. Lipphardt, Jr., D.A. Smith, and R.A. Locarnini. 1993. Palmer LTER: Hydrography in the LTER region. Antarctic Journal of the U.S., 28(5), 209-211. Klinck, J.M., and R.C. Smith. 1994. Heat budgets and implications for circulation on the continental shelf west of the Antarctic Peninsula. Abstracts, Proceedings of SCAR Sixth Biology Symposium. Cambridge Press: Scientific Committee on Antarctic Research. [Abstract] Prézelin, B.B., N.P. Boucher, M. Moline, E. Stephens, K. Seydell, and K. Scheppe. 1992. Palmer LTER: Spatial variability in phytoplankton distribution and surface photosynthetic potential within the peninsula grid, November 1991. Antarctic Journal of the U.S., 27(5), 242-245. Stammerjohn, S.E. 1994. Spatial and temporal variability in southern ocean sea ice cover. (Masters of Arts thesis, University of California at Santa Barbara.)
Palmer LTER: Spatial distribution of viruses in the Palmer LTER region ROxANE MARANGER and DAVID F. BIRD, Department of Biology, University of Quebec at Montreal, Montreal,
Quebec H3C 3P8, Canada DAVID M. KARL, School of Ocean and Earth Science and Technology, University of Hawaii, Honolulu, Hawaii 96822
i ruses have been identified as dynamic components in Vseveral aquatic environments including marine and fresh waters (Bergh et al. 1989; Klut and Stockner 1990; Proctor and Fuhrman 1990), marine and freshwater sediments (Paul et al. 1993; Maranger and Bird unpublished data), and polar sea ice (Maranger, Bird, and Juniper in press). High abundances and rapid changes in viral abundance (Bratbak et al. 1990) along with rapid viral decay rates (Heldal and Bratbak 1991) suggest that viruses may play an important role in controlling microbial populations. Viruses are also thought to be involved in carbon transfer within the microbial loop (Bratbak et al. 1992); however, their quantitative role in carbon and nutrient cycling has not been fully established. Viruses have previously been observed in the southern oceans and abundances have been reported for the Drake Passage (D.C. Smith et al. 1992) and for the coastal waters of Paradise Harbor (Bird, Maranger, and Karl 1993). During cruise 94-01 of the R/V Polar Duke (January 1994), we enumerated viruses from surface water samples taken at each station of the Palmer Long-Term Ecosystem Research (LTER) transect lines 300, 400, 500, and 600 (Waters and Smith 1992). Our objective was to determine onshore-to-offshore gradients in viral abundance and to compare these results with other
physical, chemical, and microbiological characteristics of the surface waters. Viruses were counted in different size classes by head capsid diameter [less than 30 nanometers (nm), 30-60 nm, 60-80 nm, greater than 80 nm) to determine changes in the viral community composition between sites. Depth profiles of virus samples were taken at the endpoint stations (nearest to and farthest from shore) of each transect line. Virus samples were fixed with electron microscopy (EM) grade glutaraldehyde (2.5 percent final concentration), stored in polypropylene vials at 4°C, and prepared on board the R/V Polar Duke as soon as sea conditions were calm. Water samples were concentrated using AMICON microconcentrators (10,000 molecular weight cutoff filter) in a micro centrifuge. Viruses from these concentrated water samples were then pelleted directly onto 400-mesh formvar-coated copper EM-grids using an EM-90 rotor in a Beckman airfuge for 30 minutes at 100,000 x g. Once removed, the grids were floated on a drop of filtered (0.02-micrometer) distilled water to remove salts. Grids were stained with uranyl acetate (2 percent final concentration) for 30 minutes. Viruses were counted directly by transmission electron microscopy at high magnification (90,000x). Virus particles were identified on the basis of shape, size, and staining properties. Our criteria for virus
ANTARCTIC JOURNAL - REVIEW 1994 209
identification were quite strict because high abundances of colloidal particles similar in shape and size to virus particles have been reported in the southern oceans (Wells and Goldberg 1994). Viral abundances ranged from 3.2x10 5 to 1.7x10 6 viruses per milliliter (mL') (coefficient of variation of each mean was approximately 30 percent) in the surface waters of the region studied (figure 1). Peaks of viral abundances appeared at stations 40 (nearest to shore), 100, and 120 whereas the lowest abundances observed were at stations 60 and 180. Comparisons of these trends with other physical, chemical, and microbiological variables will be made as they become available. An interesting observation is that the viral population, as classified by head capsid diameter, appeared to be quite variable among sampling stations (figure 2A and 2B). Viruses are largely species-specific and, thus, spatial distribution is influ-
enced by the bacterial community itself. Therefore, a difference in the viral population might imply a difference in the species composition of the bacterial community or a difference in the numerically dominant species of bacterium at different stations. It should not be assumed that all the viruses observed were bacteriophage; it is possible that some were algal viruses. No methods to discern between algal and bacterial viruses in natural systems exist at present, however. Viral abundances were consistently lower in the surface waters of the vertical profiles (figure 3A-D). Low concentrations of viruses in near-surface waters have been observed in Paradise Harbor (Bird et al. 1993) during the austral spring. A surface reduction in virus particles, however, was not observed in the Drake Passage during the austral winter (D.C. Smith et al. 1992). This reduction in virus concentration in the summer might indicate that virus accumulation is inhibited by increased ultraviolet (UV) radiation at the surface (R.C. Smith et al. 1992; Suttle and Chan 1992), resulting in either lower production rates or higher loss rates. U_V radiation may damage viral DNA or proteins, rendering infectious viruses defective, or it may destroy viruses directly by photolysis. The reduced number of viruses observed in the surface waters of the Palmer LTER region could also be due to differences in mixed layer depths between our study region and the Drake Passage. Viruses may be more abundant in the surface waters in areas where there is a deep well-mixed layer. In this case and relative to more stratified waters, the residence time of viruses at the surface will be reduced as a result of mixing and, consequently, so will their exposure to U_V light. A simpler alternative explanation is that viral loss rates are high as a result of rapid sedimentation rates due to bubble scavenging and adhesion to particles. Adsorption to marine snow particles has been shown to be a major factor in controlling viral loss at lower latitudes (Suttle and Chan 1992). A consistent subsurface peak of viruses was observed between 20 and 60 meters in the water column. The peak of viruses appeared closer to the surface at the nearshore stations (figure 3A and 3C) and deeper in the water column at the offshore stations (figure 3B and 3D).
I . 1.8 1.6 1.4 1.2
E
to 0
C) C C
0.8 0.6 0.4 0.2
.0 Cli Cli 1..
>
300 line U 500 line1 L--40° line ---600 line
20 60 100 140 180 220 Station Number
Figure 1. Viral abundances (in millions of virus particles per milliliter) in the surface waters of the transect lines 300, 400, 500, and 600 of the Palmer LTER study region in the southern oceans during January and February 1994. Station number represents distance from shore in kilometers.
• 80 nm
• 80 nm
80
80
60 Cd
>
U) U) Cli C.)
40
,
N
60
40 20
20 40 60 80 100 120 140 160 180 200
0
40 60 80 100 120 140 160 180 200 Station number
Station number
Figure 2. Virus community composition, represented as a percentage of head capsid diameter size classes, for the onshore-to-offshore stations of the (A) 400 transect line and the (B) 600 transect line. ANTARCTIC JOURNAL - REVIEW 1994 210
Viral abundance (106 mr1)
Viral abundance (1O6 m)
0 0.5 1 1.5 2 2.5 3 3.5
0
02 O4 0_R 0 A 1 12
1A 1r]
0 -50 E
E
a. a
a.
a
•ioo -150 -200 -250
Viral abundance (106 m11)
Viral abundance (106 ml-')
0 0.5 1 1.5 2 2.5 3 3.5
E
0 0.5 1 1.5 2 2.5 3
Bratbak, G., M. Heldal, S. Norland, and T.F. Thingstad. 1990. Viruses as partners in spring bloom trophodynamics. Applied and Environmental Microbiology, 56(5), 1400-1405. Bratbak, G., M. Heldal, T.F. Thingstad, B. Riemann, and O.H. Haslund. 1992. Incorporation of viruses into the budget of microbial C-transfer. A first approach. Marine Ecology Progress Series, 83(2/3), 273-280. Heldal, M., and G. Bratbak. 1991. Production and decay of viruses in aquatic environ-
ments. Marine Ecology Progress Series, 72(3), 205-212. Klut, M.E., and J.G. Stockner. 1990. Virus-like particles in an ultra- oligotrophic lake on Vancouver Island, British Columbia.
Canadian Journal of Fisheries and Aquatic Sciences, 47(4), 725-730.
E -C
a. a
Maranger, R., D.F. Bird, and S.K. Juniper. In press. Viral and bacterial dynamics in Arctic sea ice during the spring algal bloom, near Resolute, N.W.T. Marine
Ecology Progress Series.
Paul, Jj, J.B. Rose, S.C. Jiang, C.A. Kellogg, and L. Dickson. 1993. Distribution of viral Figure 3. Profiles of viral abundance in the upper 200 meters of the water column of LTER station abundance in the reef environment of number (A) 500.060, (B) 500.020, (C) 400.040, and (D) 400.200. Key Largo, Florida. Applied and Environmental Microbiology, 59(3), 718-724. We thank the captain and the crew of the R/V Polar Duke, Proctor, L.M., and J. Fuhrman. 1990. Viral mortality of marine bactemembers of the Antarctic Support Associates, and field particria and cyanobacteria. Nature, 343 (6253), 60-62. Smith, D.C., G.F. Steward, F. Azam, and J.T. Hollibaugh. 1992. Virus ipants J. Christian, T. Houlihan, J. Polovina, and S. Vink for and bacteria abundances in the Drake Passage during January and their assistance in sample collection. This research was supAugust 199 1. Antarctic Journal of the U.S., 27(5), 125-127. ported by National Science Foundation grant OPP 91-18439, Smith, R.C., B.B. Prézelin, K.S. Baker, R.R. Bidigare, N.P. Boucher, T. awarded to D.M. Karl, and an NSERC (Canada) operating Coley, D. Karentz, S. Macintyre, H.A. Matlick, D. Menzies, M. grant to D.F. Bird. (This is SOEST contribution number 3635.) Ondrusek, Z. Wan, and K.J. Waters. 1992. Ozone depletion: Ultraviolet radiation and phytoplankton biology in antarctic waters. Science, 255(5047), 952-959. References Suttle, C.A., and F. Chan. 1992. Mechanisms and rates of decay of marine viruses in seawater. Applied and Environmental MicrobiolBergh, 0., K.Y. Bersheim, G. Bratbak, and M. Heldal. 1989. High abunogy, 58(12), 3721-3729. dance of viruses found in aquatic environments. Nature, Waters, K.J., and R.C. Smith. 1992. Palmer LTER: A sampling grid for the 340(6233), 467-468. Palmer LTER program. Antarctic Journal of the U.S., 26(5), 236-238. Bird, D.F., R. Maranger, and D.M. Karl. 1993. Palmer LTER: Aquatic Wells, M.L., and E.D. Goldberg. 1994. The distribution of colloids in virus abundances near the Antarctic Peninsula. Antarctic Journal the North Atlantic and southern oceans. Limnology and Oceanogof the U.S., 28(5), 234-235. raphy, 39(2), 286-302.
ANTARCTIC JOURNAL - REVIEW 1994
211
This project was supported by a University of California at Santa Barbara May Company Scholarship to Barbara Sullivan and National Science Foundation grant OCE 93-01322 to Barbara B. Prézelin. Special thanks to B. Boczar, T. Diem, T. Westberry, M. Moline, and the crew of the R/V Polar Duke. (This is LTER contribution number 43.)
References Hofmann, E.E., B.L. Lipphardt, Jr., D.A. Smith, and R.A. Locarnini. 1993. Palmer LTER: Hydrography in the LTER region. Antarctic Journal of the U.S., 28(5), 209-211. Klinck, J.M., and R.C. Smith. 1994. Heat budgets and implications for circulation on the continental shelf west of the Antarctic Peninsula. Abstracts, Proceedings of SCAR Sixth Biology Symposium. Cambridge Press: Scientific Committee on Antarctic Research. [Abstract] Prézelin, B.B., N.P. Boucher, M. Moline, E. Stephens, K. Seydell, and K. Scheppe. 1992. Palmer LTER: Spatial variability in phytoplankton distribution and surface photosynthetic potential within the peninsula grid, November 1991. Antarctic Journal of the U.S., 27(5), 242-245. Stammerjohn, S.E. 1994. Spatial and temporal variability in southern ocean sea ice cover. (Masters of Arts thesis, University of California at Santa Barbara.)
Palmer LTER: Spatial distribution of viruses in the Palmer LTER region ROxANE MARANGER and DAVID F. BIRD, Department of Biology, University of Quebec at Montreal, Montreal,
Quebec H3C 3P8, Canada DAVID M. KARL, School of Ocean and Earth Science and Technology, University of Hawaii, Honolulu, Hawaii 96822
i ruses have been identified as dynamic components in Vseveral aquatic environments including marine and fresh waters (Bergh et al. 1989; Klut and Stockner 1990; Proctor and Fuhrman 1990), marine and freshwater sediments (Paul et al. 1993; Maranger and Bird unpublished data), and polar sea ice (Maranger, Bird, and Juniper in press). High abundances and rapid changes in viral abundance (Bratbak et al. 1990) along with rapid viral decay rates (Heldal and Bratbak 1991) suggest that viruses may play an important role in controlling microbial populations. Viruses are also thought to be involved in carbon transfer within the microbial loop (Bratbak et al. 1992); however, their quantitative role in carbon and nutrient cycling has not been fully established. Viruses have previously been observed in the southern oceans and abundances have been reported for the Drake Passage (D.C. Smith et al. 1992) and for the coastal waters of Paradise Harbor (Bird, Maranger, and Karl 1993). During cruise 94-01 of the R/V Polar Duke (January 1994), we enumerated viruses from surface water samples taken at each station of the Palmer Long-Term Ecosystem Research (LTER) transect lines 300, 400, 500, and 600 (Waters and Smith 1992). Our objective was to determine onshore-to-offshore gradients in viral abundance and to compare these results with other
physical, chemical, and microbiological characteristics of the surface waters. Viruses were counted in different size classes by head capsid diameter [less than 30 nanometers (nm), 30-60 nm, 60-80 nm, greater than 80 nm) to determine changes in the viral community composition between sites. Depth profiles of virus samples were taken at the endpoint stations (nearest to and farthest from shore) of each transect line. Virus samples were fixed with electron microscopy (EM) grade glutaraldehyde (2.5 percent final concentration), stored in polypropylene vials at 4°C, and prepared on board the R/V Polar Duke as soon as sea conditions were calm. Water samples were concentrated using AMICON microconcentrators (10,000 molecular weight cutoff filter) in a micro centrifuge. Viruses from these concentrated water samples were then pelleted directly onto 400-mesh formvar-coated copper EM-grids using an EM-90 rotor in a Beckman airfuge for 30 minutes at 100,000 x g. Once removed, the grids were floated on a drop of filtered (0.02-micrometer) distilled water to remove salts. Grids were stained with uranyl acetate (2 percent final concentration) for 30 minutes. Viruses were counted directly by transmission electron microscopy at high magnification (90,000x). Virus particles were identified on the basis of shape, size, and staining properties. Our criteria for virus
ANTARCTIC JOURNAL - REVIEW 1994 209
identification were quite strict because high abundances of colloidal particles similar in shape and size to virus particles have been reported in the southern oceans (Wells and Goldberg 1994). Viral abundances ranged from 3.2x10 5 to 1.7x10 6 viruses per milliliter (mL') (coefficient of variation of each mean was approximately 30 percent) in the surface waters of the region studied (figure 1). Peaks of viral abundances appeared at stations 40 (nearest to shore), 100, and 120 whereas the lowest abundances observed were at stations 60 and 180. Comparisons of these trends with other physical, chemical, and microbiological variables will be made as they become available. An interesting observation is that the viral population, as classified by head capsid diameter, appeared to be quite variable among sampling stations (figure 2A and 2B). Viruses are largely species-specific and, thus, spatial distribution is influ-
enced by the bacterial community itself. Therefore, a difference in the viral population might imply a difference in the species composition of the bacterial community or a difference in the numerically dominant species of bacterium at different stations. It should not be assumed that all the viruses observed were bacteriophage; it is possible that some were algal viruses. No methods to discern between algal and bacterial viruses in natural systems exist at present, however. Viral abundances were consistently lower in the surface waters of the vertical profiles (figure 3A-D). Low concentrations of viruses in near-surface waters have been observed in Paradise Harbor (Bird et al. 1993) during the austral spring. A surface reduction in virus particles, however, was not observed in the Drake Passage during the austral winter (D.C. Smith et al. 1992). This reduction in virus concentration in the summer might indicate that virus accumulation is inhibited by increased ultraviolet (UV) radiation at the surface (R.C. Smith et al. 1992; Suttle and Chan 1992), resulting in either lower production rates or higher loss rates. U_V radiation may damage viral DNA or proteins, rendering infectious viruses defective, or it may destroy viruses directly by photolysis. The reduced number of viruses observed in the surface waters of the Palmer LTER region could also be due to differences in mixed layer depths between our study region and the Drake Passage. Viruses may be more abundant in the surface waters in areas where there is a deep well-mixed layer. In this case and relative to more stratified waters, the residence time of viruses at the surface will be reduced as a result of mixing and, consequently, so will their exposure to U_V light. A simpler alternative explanation is that viral loss rates are high as a result of rapid sedimentation rates due to bubble scavenging and adhesion to particles. Adsorption to marine snow particles has been shown to be a major factor in controlling viral loss at lower latitudes (Suttle and Chan 1992). A consistent subsurface peak of viruses was observed between 20 and 60 meters in the water column. The peak of viruses appeared closer to the surface at the nearshore stations (figure 3A and 3C) and deeper in the water column at the offshore stations (figure 3B and 3D).
I . 1.8 1.6 1.4 1.2
E
to 0
C) C C
0.8 0.6 0.4 0.2
.0 Cli Cli 1..
>
300 line U 500 line1 L--40° line ---600 line
20 60 100 140 180 220 Station Number
Figure 1. Viral abundances (in millions of virus particles per milliliter) in the surface waters of the transect lines 300, 400, 500, and 600 of the Palmer LTER study region in the southern oceans during January and February 1994. Station number represents distance from shore in kilometers.
• 80 nm
• 80 nm
80
80
60 Cd
>
U) U) Cli C.)
40
,
N
60
40 20
20 40 60 80 100 120 140 160 180 200
0
40 60 80 100 120 140 160 180 200 Station number
Station number
Figure 2. Virus community composition, represented as a percentage of head capsid diameter size classes, for the onshore-to-offshore stations of the (A) 400 transect line and the (B) 600 transect line. ANTARCTIC JOURNAL - REVIEW 1994 210
Viral abundance (106 mr1)
Viral abundance (1O6 m)
0 0.5 1 1.5 2 2.5 3 3.5
0
02 O4 0_R 0 A 1 12
1A 1r]
0 -50 E
E
a. a
a.
a
•ioo -150 -200 -250
Viral abundance (106 m11)
Viral abundance (106 ml-')
0 0.5 1 1.5 2 2.5 3 3.5
E
0 0.5 1 1.5 2 2.5 3
Bratbak, G., M. Heldal, S. Norland, and T.F. Thingstad. 1990. Viruses as partners in spring bloom trophodynamics. Applied and Environmental Microbiology, 56(5), 1400-1405. Bratbak, G., M. Heldal, T.F. Thingstad, B. Riemann, and O.H. Haslund. 1992. Incorporation of viruses into the budget of microbial C-transfer. A first approach. Marine Ecology Progress Series, 83(2/3), 273-280. Heldal, M., and G. Bratbak. 1991. Production and decay of viruses in aquatic environ-
ments. Marine Ecology Progress Series, 72(3), 205-212. Klut, M.E., and J.G. Stockner. 1990. Virus-like particles in an ultra- oligotrophic lake on Vancouver Island, British Columbia.
Canadian Journal of Fisheries and Aquatic Sciences, 47(4), 725-730.
E -C
a. a
Maranger, R., D.F. Bird, and S.K. Juniper. In press. Viral and bacterial dynamics in Arctic sea ice during the spring algal bloom, near Resolute, N.W.T. Marine
Ecology Progress Series.
Paul, Jj, J.B. Rose, S.C. Jiang, C.A. Kellogg, and L. Dickson. 1993. Distribution of viral Figure 3. Profiles of viral abundance in the upper 200 meters of the water column of LTER station abundance in the reef environment of number (A) 500.060, (B) 500.020, (C) 400.040, and (D) 400.200. Key Largo, Florida. Applied and Environmental Microbiology, 59(3), 718-724. We thank the captain and the crew of the R/V Polar Duke, Proctor, L.M., and J. Fuhrman. 1990. Viral mortality of marine bactemembers of the Antarctic Support Associates, and field particria and cyanobacteria. Nature, 343 (6253), 60-62. Smith, D.C., G.F. Steward, F. Azam, and J.T. Hollibaugh. 1992. Virus ipants J. Christian, T. Houlihan, J. Polovina, and S. Vink for and bacteria abundances in the Drake Passage during January and their assistance in sample collection. This research was supAugust 199 1. Antarctic Journal of the U.S., 27(5), 125-127. ported by National Science Foundation grant OPP 91-18439, Smith, R.C., B.B. Prézelin, K.S. Baker, R.R. Bidigare, N.P. Boucher, T. awarded to D.M. Karl, and an NSERC (Canada) operating Coley, D. Karentz, S. Macintyre, H.A. Matlick, D. Menzies, M. grant to D.F. Bird. (This is SOEST contribution number 3635.) Ondrusek, Z. Wan, and K.J. Waters. 1992. Ozone depletion: Ultraviolet radiation and phytoplankton biology in antarctic waters. Science, 255(5047), 952-959. References Suttle, C.A., and F. Chan. 1992. Mechanisms and rates of decay of marine viruses in seawater. Applied and Environmental MicrobiolBergh, 0., K.Y. Bersheim, G. Bratbak, and M. Heldal. 1989. High abunogy, 58(12), 3721-3729. dance of viruses found in aquatic environments. Nature, Waters, K.J., and R.C. Smith. 1992. Palmer LTER: A sampling grid for the 340(6233), 467-468. Palmer LTER program. Antarctic Journal of the U.S., 26(5), 236-238. Bird, D.F., R. Maranger, and D.M. Karl. 1993. Palmer LTER: Aquatic Wells, M.L., and E.D. Goldberg. 1994. The distribution of colloids in virus abundances near the Antarctic Peninsula. Antarctic Journal the North Atlantic and southern oceans. Limnology and Oceanogof the U.S., 28(5), 234-235. raphy, 39(2), 286-302.
ANTARCTIC JOURNAL - REVIEW 1994
211
