Investigating Interactions between Lignin and Marine

Microbes

Connor StonesiferMentor: Natasha McDonald

C.D.O.M• CDOM= large factor in regulating the

underwater light field– Absorbs light

• Problem: We don’t really know what it’s comprised of!

• Problem: We don’t really know how microbes interact with it!

• Clue: Lignin is a constituent of CDOM

Lignin• Phenolic organic polymer

– Aromatic and a constituent of CDOM

• Derived from cell walls of woody vascular plants (terrestrial origin)

• 9 standard phenol monomers, no standard composition

• Found in all natural waters (including open ocean)

Experimental Questions

• Can lignin persist in a marine microbial environment?

• Can lignin be used a nutrient source by marine microbes in oligotrophic waters?

• How does lignin’s molecular complexity affect its ability to be degraded by marine microbes?

• Are the by-products of lignin degradation more or less labile than the by-products of the metabolism of simpler nutrients?

Experiment 2 Set Up

Control: 70/30 Mix Filtered and Unfiltered Seawater

GNP: + Glucose (10uM C)

+ NH4Cl (1 uM)+ K2HPO4 (0.1 uM)

LNP: + Lignin+ NH4Cl (1 uM)+ K2HPO4 (0.1 uM)

Lignin: + Lignin (10 uM carbon)

Experiment 2 SamplingControl

GNP

Lignin

LNP

CDOM Lignin Concentrations (T0, 10 days, and 30 days)Bacterial Abundance DOC FISH probes DNAEEMS

Experiment 2 Results: Plot of Control/GNP

Plot 2: Lignin and LNP1

ligmean = 0.5542; ligstd = 0.1290; lnpmean = 0.5808; lnpstd = 0.1199

Changing Absorption Spectra of Lignin Over Time: T4 (45 hours)

Changing Absorption Spectra of Lignin Over Time : T5 (62 Hours)

Changing Absorption Spectra of Lignin Over Time : T7 (86 Hours)

Changing Absorption Spectra of Lignin Over Time : T8 (93 Hours)

Changing Absorption Spectra of Lignin Over Time: T9 (110 Hours)

What do we know now?

• We now know lignin is being removed by some process from the absorbance spectrum

Now it’s time to look at what bacteria are doing...

16

0 2 4 6 8 10 120.0E+00

2.0E+05

4.0E+05

6.0E+05

8.0E+05

1.0E+06

1.2E+06

1.4E+06

1.6E+06

CDOM14-2 (Lignin Experiment 2) Bacterioplankton Counts

C1C2L1L2LNP1LNP2GNP1GNP2

Time (Days)

Bact

eria

/mL

Standard error bars represent standard deviation (N=2)

GNP showed ~8x increase

LNP showed ~3-3.5x increase

Lignin showed ~2.5x increase

Controls showed ~1.5x increase

Conclusion

• Our preliminary studies strongly suggest that marine bacteria can use lignin as a nutrient source.– Lignin therefore may be a carbon source for

bacteria in oligotrophic waters.

• Nitrogen and phosphorus are not an important nutrient amendment for bacterial metabolism of lignin, a complex nutrient substrate.

• Future analysis should examine the by-products of lignin metabolism, the bacterial communities involved, and the mechanism for degradation

THANK YOU!