Phytoplankton Bacterial Interactions and their influence on Biogeochemical cycles

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1 Phytoplankton Bacterial Interactions and their influence on Biogeochemical cycles Outline Intro: the role of plankton and bacteria in the ocean. The importance of microscales Biogeochemical strategies and ectohydrolic enzymes Biochemical diversity 2 1

2 Plankton blooms and the biological pump 2

3 Ecosystem pathways in the ocean High turnover and lysis rate (50% of PP) Couples primary productivity to microbial ecosystems Mechanisms of death? Turnover times Terrestrial 16 years Oceanic <2-14 days Valiela (1995) Marine Ecological Processes 3

4 Bacteria are everywhere! (more curious when absent) Tracy Mincer, 2011 Bates et al, 2003 HP Grossart, 2011 A bacteria-eye view of the ocean s sunlit layer. Seawater is carbon-based matter continuum, a gel of tangled polymers with embedded strings, sheets, and bundles of fibrils and particles, including living organisms, as hotspots. Bacteria acting on marine snow or algae can control sedimentation and primary productivity, diverse micro-niches can support high bacterial diversity. Azam, The Marine Microscale 8 4

5 Technological advances allow new understanding: Flow cytometry Multiple displacement amplification Next-gen sequencing NanoSIMS (Secondary Ion Mass Spectrometry) Fundamentally new context for ecology and biogeochemistry Better understanding better predictive powers 9 The Marine Microscale Bacterial adaptations utilizing organic matter Hunting: motility and chemotaxis Gathering: different systems for patchy nutrients Eating: Ecotoenzymes (proteases, lipases, nucleases, phosphatases, glucosidase). 10 5

6 The start of enzyme studies The study of ectohydrolytic enzymes in the sea first in in the early 80s implementing fluorogenic methyumbelliferylbased substrates Variable activity over time most likely variable species composition in the community, which in turn will influence how OM is processed and degraded Martenez et al., Variability of cell specific enzyme activity Measured ecohydrolytic enzymes in 44 Isolates (including attached and free) Variation at the order of magnitude for all enzymes measured Many specialists, no super bug 12 6

7 Attached vs free-living enzyme activity α-gluc β-gluc Per volume Illustrated that higher activities are not due to elevated bacterial abundance on aggregates, but rather due to higher cell-specific activity on aggregates. 13 protease Karner, M., and G. J. Herndl Attached vs free-living enzyme activity α-gluc β-gluc Per cell Found marine snow harbored bacterial ectohyrolitic activity 2.5 to 10 times that of ambient water. 14 protease Karner, M., and G. J. Herndl

8 Attached vs. free-living enzyme activity Bacterial abundance in (A) Phytoplankton aggregates and (B) the surrounding water after a simulated bloom. Particles were highly active sites ( hotspots ) of intense enzymatic activities compared to bulk seawater populations. 15 Martinez et al., 1996 Increased activity not necessarily high growth Higher enzyme activities not correlated with higher bacterial growth (called secondary production) Indicates that particles are sites of rapid remineralization that is not coupled with growth. 16 Karner, M., and G. J. Herndl

9 Particle decomposition paradox Aggregation may be triggered by some bacterial-diatom interaction Grossart et al., 2004 Aggregates are sites of intense remineralization and release of DOM Marine snow aggregates show high ectohyrolytic enzyme activity, but low carbon demand Suggests that hydrolysis and uptake are only loosely coupled 17 9

10 Ectoenzymes and biogeochemistry: a diatom s dilemma 19 Fate of Diatom Silica Dissolution: Production (D:P) ratio is 0.6 So 60% of diatom silica comes from dissolved frustules! 20 10

11 Bacterial-mediate regeneration of diatom silica C. fusimormis T. weisflogii Little to no silica dissolution seen in the bacteria-free sample their activity was the rate-limiting step in BSi dissolution Colonizing bacteria showed high growth rates and high protease activities to the diatom microenvironment 21 Bidle and Azam, 1999 Bacterial-mediated Si cycling is highly variable 22 Diatom Experiment Turnover rates C. fusimormis T. weisflogii All bacterial isolates accelerated silica dissolution, but individual rates varied by ~300% Isolates that displayed enhanced colonization and protease activities were the most effective at regenerating silicon Bidle and Azam,

12 Effect of bacterial protease inhibition an inhibitor cocktail, consisting of antibiotics and protease inhibitors, successfully abolished regeneration of diatom POC and BSi 23 Effect of bacterial protease inhibition Bidle and Azam (2001) observed a strong correlation between protease activity and Si regeneration cell wall-associated protein classes (e.g., frustulins and silaffins) found (not shown) direct addition of Pronase E caused the release of 14 C and BSi from uniformly 14 C-labeled diatoms 24 12

13 Natural ecosystem example Bidle et al examined whether proteolytic hydrolysis and production of attached bacteria were correlated with silica dissolution parameters during a bloom Integrated protease activities were strongly correlated (r2 = 0.91, p < 0.001) Dissolution influenced by the physiological state of the diatom cells. Integrated pdis = dissolution integrated over depth BSi dissolution rate (Vdis) 25 Most were characterized by high diatom biomass with minimal depth variation in the specific BSi dissolution rate (V dis ) and D:P ratio Differences between BSi dissolution parameters for high vs. low biomass stations might represent a bloom to post-bloom transition

14 Effectiveness of protease inhibitor treatments on Si cycling Demonstrates a direct role for bacteria as principle BSi regeneration mechanism during natural diatom blooms selectively inhibiting bacterial ectoprotease activity via antibiotic and protease inhibitor (leupeptin) treatment and measuring V dis over 24 h period (compared to control samples). 27 Bloom vs Post-bloom trends in the global data 2 major groups: (1) High 1 - D:P rations (meaning they have low BSi dissolution) (2) Low 1 - D:P rations (meaning they have extensive BSi dissolution). These process influence Si cycling which influence bloom dynamics

15 This research brought up fundamentally important issues about bacterial ectoproteases and their in situ activity 1) How is their activity regulated? And what how might this affect biogeochemical cycling? 2) How diverse are they? 29 Different environments, different dissolution rates Biological Si and C org preservation in different open ocean systems There is dramatic preservation beneath the cold Southern Ocean (Si:C ratios can reach 20-60) the continental margins, have a Si:C ratio of 0.6 which is indicative of tighter coupling. In both regions, the Si:C ratios of freshly produced diatoms are

16 Interplay between temperature and Si:C regeneration Shows a relationship between particulate organic carbon (POC) utilization and biogenic silica (BSi) dissolution from diatoms incubating at different temperatures 31 Bidle s research provides a framework to interpret spatial differences in Si and C preservation in different oceanic systems, based on temperature Selective stripping of C org and decoupling of Si and C preservation is enhanced in permanently cold water. Systems experiencing seasonal temperature shifts may display greater variability in BSiO 2 /C org coupling Warmer temperatures would enhance POC hydrolysis and expose diatom frustules to elevated dissolution

17 Temperature response in different oceans Sub-optimal performance at in situ temperatures Differential response of ectoproteases (LAPase) and glucosidases (BGase P) in subtropical, equatorial, and polar region Possibly finding latitudinal trends in processing and utilization of C and N. Strongly suggest distinct bacterial phenotypes with distinct isozymes. 33 Influence on nitrogen sources on ectoprotease activity Christian and Karl (1998) found that general nitrogen availability was a poor predictor of LAPase (ectoprotease) activity. the addition of either ammonia (NH4) or Glycine/Proline did not alter the LAPase activity from the control incubations. The addition of specific amino acids, especially histidine and phenyalanine, noticeable repressed LAPase activity. perhaps indicating that high ectoprotease activity by marine bacteria is a sign of auxotrophy 34 17

18 Summary Natural bacterial assemblages can exert different biological pressures on organic matter Activity depends on different bacteria present Free-living and particle attached may have very different ectohydrolytic enzyme activities Ectohydrolytic enzymes can mechanistically control ocean biogeochemistry Such as the cycling of Bsi and C in the open ocean and distribution of Si in sediments in ocean basins Environment (temperature, nitrogen, etc) affects ectohydrolytic enzyme activity 35 18