Microbial biomass in the sea: Methods, limitations, and distributions

Transcription

1 Microbial biomass in the sea: Methods, limitations, and distributions Matt Church (MSB 612 / / mjchurch@hawaii.edu) Marine Microplankton Ecology OCN 626 E. F. DeLong

2 The tripartite goals of microbial ecology How many? Biomass At what rate? Metabolism Who s there? Diversity

3 Things to remember about marine microbes 1. Important components of the marine food web. 2. Major drivers of the global carbon cycle. 3. Primary controllers of the global nitrogen cycle.

4 Things we need to know to understand the importance of microbes to ecosystem processes Population size and biomass (biogenic carbon): provides information on energy available to support the food web Growth, production, metabolism: turnover of material through the food web and insight into physiology. Controls on growth and population size

5 Why carbon? Common currency among all organisms. Carbon serves as a proxy for energy transfer through the ecosystem (primary secondary production, respiration). AN = 6 (6P/6N) AW = Oxidation: -4 to +4 Electrons: 1s 2 2s 2 p 2 Isotopes: 11 C, 12 C, 13 C, 14 C Forms: oxides, hydrides, sulfides and halides

6 Major cellular macromolecular constituents All of these serve as potential biomarkers of microbial biomass. Cell constituent % dry weight Protein 55 C, N, S RNA C, N, P Lipopolysaccharides 3 C, P Peptidoglycan 2-4 C, N (cell walls) DNA 3-10 C, N, P

7 Element The Elemental Composition of E. coli % dry Substrate Source Cellular Components C 55 DOC, CO 2 Main constituent of cellular material O 20 O 2, DOM, CO 2 water; O 2 primary electron acceptor Constituent of cell material and cell in aerobic respiration N 10 NH 3, NO 3-, NO 2-, DON, N 2 Constituent of amino acids, nucleic acids, nucleotides, and coenzymes H 8 DOM, H 2 Main constituent of organic compounds and cell water P 3 PO 4 3-, DOP S 1 SO 4, H 2 S, HS, DOM K 1 Potassium salts Mg 0.5 Magnesium salts Ca 0.5 Calcium salts Fe Iron salts, DOM Constituent of nucleic acids, nucleotides, phospholipids, LPS Constituent of cysteine, methionine, glutathione, several coenzymes Main cellular inorganic cation and cofactor for certain enzymes Inorganic cellular cation, cofactor for certain enzymatic reactions Inorganic cellular cation, cofactor for certain enzymes Component of cytochromes and Feproteins; cofactor for many enzymes

8 Primary producers Primary consumers (herbivores) Secondary consumers (carnivores) Tertiary consumers The way biomass is distributed among trophic levels in the food web provides clues to the efficiency of energy transfer through the ecosystem. Note: this is a static depiction-it does not provide information on how fast biomass turns over within each trophic level.

9 The Struggle for Composition % surface ocean Fe Mg Ca S K % cell composition P N C Bacterial biomass relatively enriched in P, N, C, Fe compared to the surface seawater. Energy must be expended to acquire and maintain intracellular concentrations of these elements. Biomass is a key source of these elements.

10 Plant Biomass and Productivity in Marine and Terrestrial Ecosystems Ecosystem Biomass (10 15 g) Net Primary Production (10 15 g year -1 ) Turnover time (years) Marine Land The main punch line: Despite low biomass, plankton biomass turnover is rapid implies rapid cycling of bioelements in the sea.

11 Determining plankton biomass in seawater is complicated Cell densities in seawater are generally low. Cells are often small. Difficult to attribute biomass to specific trophic levels. Confounding influences of non-living material, other organisms, variable sources of nutrients and energy alter some biomarkers but not others, etc.

12 Examples of commonly used methods to estimate microbial biomass in seawater Biomass (mg C L -1 ) ATP Chlorophyll a Total carbon Cell abundance Cell volume Note: none of these are direct measures of biomass (i.e. carbon) except total carbon and it has problems too. Primary producers Primary consumers Secondary consumers Tertiary consumers

13 Size selection is common for evaluating how biomass is partitioned into different trophic levels.

14 ATP as an indicator of biomass All living cells contain ATP ATP degrades rapidly after cell death ATP:C ratio appears well conserved ATP:C ~250:1 (mg : mg) Non-discriminate, includes all living material

15 Particulate carbon Technique: combust (oxidize) organic material and measure resulting CO 2. Need to concentrate cells: typically glass filters (usually ~0.7 µm pore size) or tangential flow (Fukuda et al. 1998) Measurements include variable amounts of detritus (and possibly bacteria). Lee et al. (1995)

16 Euphotic zone typically ~30-75% of particulate material contains ATP.

17 Phytoplankton carbon Phytoplankton carbon determinations are most often derived from measurements of chlorophyll; this requires a conversion factor. Phytoplankton carbon can also be estimated based on cell size and abundances (microscopy and/or flow cytometry). Primary producers

18 Chlorophylls Cyclic tetrapyrole with a magnesium atom chelated in the center of the ring Phytol Chlorophyll c lacks the phytol group

19 Carbon to Chlorophyll Conversions Chlorophyll concentrations can vary as a function of cellular physiology, independent of biomass. Need simultaneous measurements of carbon and chlorophyll. In pure cultures, values typically range from 30 :1 to 100 :1 (mg:mg); most often reported as 50 : 1 Chlorophyll in the near surface ocean can be derived from satellite measurements of ocean color and fluorescence

20 Carbon to Chlorophyll Conversions Chlorophyll concentrations variable depending on environmental history of the cells NAB Eq-Pac Arabian Sea HOT 0 50 Depth (m) How much detritus? Chlorophyll a (ng L -1 ) Particulate Carbon (µmol L -1 ) C: Chl (mg : mg)

21 PC (µmol L -1 ) ATP (ng L -1 ) Depth (m) Chl ATP PC Pigments can vary as a function of light, nutrients, and temperature li mits their utility as proxies for biomass Chlorophyll a (ng L -1 )

22 Commonly used determinations of bacterial biomass (mg C L -1 ) Cell abundance and cell volume (by microscopy and more recently by flow cytometry)

23 Determining Biomass by Microscopy Filter seawater onto polycarbonate filter or concentrate cells by settling (large phytoplankton) Stain cells Visualize cells by light and/or fluorescence microscopy Count cells and measure cell sizes Use biovolume conversion factors to calculate carbon

24 Application of flow cytometry to abundance/cell sizing Provides cell abundances and estimated cell sizes. Ability to distinguish non-pigmented bacteria from bacteria with weak autofluorescence.

25 0 Bacterial abundances in different oceanic ecosystems BATS EQ-PAC (March) EQ-PAC (September) HOT 0 Antarctic Polar Front Ross Sea North Atlantic Bloom Depth (m) Depth (m) Bacterial abundance varies ~2-fold (5 x 10 5 to 1 x 10 6 cells ml -1 ) across very different ocean ecosystems-- except the Ross Sea. 1e+4 1e+5 1e+6 1e+4 1e+5 1e+6 Cell Abundance (cells ml -1 ) Cell Abundance (cells ml -1 )

26 Typical bacterial cell densities in aquatic ecosystems Habitat Cell density (cells ml -1 ) Estuaries >5 x 10 6 Coastal (near shore) 1-5 x 10 6 Open Ocean x 10 6 Deep Sea <0.01 x 10 6

27 Remember that cell abundance carbon biomass Size matters Fagerbakke et al. (1996)