Doug Bartlett

Center for Marine Biotechnology and Biomedicine

Marine Biology Research Division

Scripps Institution of Oceanography

University of California, San Diego

Deep-Sea High-Pressure Microbiology:

Fundamental Science and Biotechnological

Possibilities

Intro slide

Perhaps location of sub dives and

lander deployments?

Chris - do you have such a

image?

Lutz and Falkowski. 2012.

Science 336:301.

Deepsea Challenger

Manned Submersible

QuickTime™ and aH.264 decompressor

are needed to see this picture.

Gigantism in the

New Britain Trench

TMAO

TMAO

Glycine

GPC

DMG

s-Inos

Challenger Deep Amphipods have high TMAO,

other Methylamines & Scyllo-Inositol

Shallow-water shrimp

glycine CD amphipods

TMAO

Scyllo-inositol

Dimethylglycine

Glycerophospho-

choline

Paul Yancey

Culturing Deep-sea Microbes Plastic bulbs

Pressure vessels bags

120 100 80 60 40 20 0 0

20

40

60

80

100

120

Temperature (°C)

Desulfovibrio profundus 500-1

Photobacterium profundum SS9 Shewanella violacea DSS12

Moritella sp. PE36 Shewanella benthica F1A

Psychromonas sp. CNPT3

Shewanella benthica DB6705

Shewanella benthica DB21MT-2

Colwellia hadaliensis BNL1

Colwellia MT41

Methanococcus jannaschii

Methanococcus thermolithotrophicus

Thermococcus barophilus Pyrococcus abyssi GE5

Pre

ssu

re (

MP

a)

Shewanella benthica KT99

Carnobacterium AT7

Marinitoga piezophila KA3

Desulfovibrio hydrothermalis AM13

Thioprofundum lithotrophica 106

Methanopyrus kandleri 116

Piezobacter thermophilus 108 Dermacoccus abyssi

Characteristics of Cultured Deep-sea Microbes

Pyrococcus CH1

120 100 80 60 40 20 0 0

20

40

60

80

100

120

Temperature (°C)

Desulfovibrio profundus 500-1

Photobacterium profundum SS9 Shewanella violacea DSS12

Moritella sp. PE36 Shewanella benthica F1A

Psychromonas sp. CNPT3

Shewanella benthica DB6705

Shewanella benthica DB21MT-2

Colwellia hadaliensis BNL1

Colwellia MT41

Methanococcus jannaschii

Methanococcus thermolithotrophicus

Thermococcus barophilus Pyrococcus abyssi GE5

Pre

ssu

re (

MP

a)

Shewanella benthica KT99

Carnobacterium AT7

Marinitoga piezophila KA3

Desulfovibrio hydrothermalis AM13

Thioprofundum lithotrophica 106

Methanopyrus kandleri 116

Piezobacter thermophilus 108 Dermacoccus abyssi

Characteristics of Cultured Deep-sea Microbes

Pyrococcus CH1

Adaptive Laboratory Evolution of E. coli to

Growth at High Pressure

Single colony isolates

Increased Unsaturated Fatty

Acid Levels in Evolved Strain

QuickTime™ and aH.264 decompressor

are needed to see this picture.

Pressure Tolerance Range of

Thermococcus Strain 175 at 90ºC

1.00E+00

1.00E+01

1.00E+02

1.00E+03

1.00E+04

1.00E+05

1.00E+06

1.00E+07

1.00E+08

0 50 100 150

Gro

wth

(C

ell

s/m

L)

Pressure (MPa)

48hrs

24 hrs

Initial

Final

Formate Driven H2 Production

From: IMG

3728

40

Total OrthologClusters in 20genomes

Ortholog Clustersin Piezophilegenomes

Membrane transporters 5

Membrane proteins 3

Transcriptional Regulators 4

Universal stress protein 2

Flagellin 1

Ortholog Clusters in

Piezophile/Piezotolerant Genomes

Deep-Ocean Microbial Communities are Distinct

% Similarity

Deep

Marine

Shallow

Marine

Assessing microbial activity using “click” chemistry

K. Barry Sharpless EdU + DAPI

HPG

DAPI

HPG + DAPI

Farooq Azam

HPG – Protein synthesis EdU – DNA synthesis

Protein synthesis (HPG) reflects pressure adaptation

P. profundum SS9 – moderately piezophilic

S. benthica KT99 – obligately piezophilic

0.1 MPa 27 MPa 90 MPa

0.1 MPa 27 MPa 90 MPa

Single-Cell Genomics

[Image courtesy of Susanne Ruemmele (Rutgers University)]

The New View of Diversity Within the Archaea and

Bacteria

Rinke et al. Nature 499:431 (2013)

# cells examined

# species

(Operational

Taxonomic

Units)

Challenger Deep Sorted Cells

(>20,000 cells sorted)

Sediment

Amphipod

exoskeleton

Amphipod gut

Amphipod

seawater

Cyanobacteria

Sediment

# cells examined

# species

(Operational

Taxonomic

Units)

Challenger Deep Sorted Cells

(>20,000 cells sorted)

Sediment

Amphipod

exoskeleton

Amphipod gut

Amphipod

seawater Amphipod

seawater

Psychromonas

Deep-Sea High-Pressure

Microbiology

• The opportunities to collect ultra-deep samples remain rare, but new technologies make this more feasible than ever.

• Piezophily requires extensive reengineering of the cell.

• Deep-sea microial communities are distinct and diverse.

Biotechnology Considerations

• Recalcitrant organic hydrolysis

• Bioremediation

• Organic osmolytes

• Secondary metabolites

• Novel enzyme mechanisms at low volume change

• Contained expression

Thank You

James Cameron and the entire Deepsea Challenge Expedition team

Marine Biological Laboratory: Julie Huber Laboratory

JAMSTEC: Ken Takai Laboratory

Whitman College: Paul Yancey

J. Craig Venter Research Institute: Roger Lasken, Mark Novotny,

Bartlett Laboratory: Roger Chastain, Rosa Léon Zayas, Emiley Eloe,

Logan Peoples, Dr. Priya Narasingarao