Culture Collection Yerseke (CCY)

A collection of microalgae and cyanobacteria, for strain

selection, growth optimization, and product maximization

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www.intesusal.eu 1

Lucas J. Stal

Department of Marine Microbiology

Royal Netherlands Institute for Sea Research

Professor of Marine Microbiology

University of Amsterdam

lucas.stal@nioz.nl

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Mission Statement

The Culture Collection Yerseke (CCY) is a non-profit facility that is specialized

in the cultivation of (marine) phototrophic microorganisms. The collection

comprises unicellular algae (green algae and diatoms) and cyanobacteria. The

collection now also extends to anoxygenic phototrophs and other bacteria that

live in marine phototrophic communities such as microbial mats and biofilms or

planktonic assemblages. CCY was established in 1997 by the Department of

Marine Microbiology of the Netherlands Institute of Ecology (NIOO-KNAW),

now the Royal Netherlands Institute for Sea Research (NIOZ), in Yerseke.

CCY provides a platform for maintaining novel strains of microorganisms that

have been isolated in the course of scientific research projects. These strains

are maintained alive as cultures or cryo-preserved. CCY provides expertise on

isolation, cultivation, and identification of novel strains and on their cryo-

preservation. The collection is available to academic and commercial parties,

except when agreements about specific strains prevent this.

CCY Collection Media How to Order Projects News Contact

CCY.NIOZ.NL

Mission Statement

The Culture Collection Yerseke (CCY) is a non-profit facility that is specialized in

the cultivation of (marine) phototrophic microorganisms. The collection

comprises unicellular algae (green algae and diatoms) and cyanobacteria. The

collection now also extends to anoxygenic phototrophs and other bacteria that

live in marine phototrophic communities such as microbial mats and biofilms or

planktonic assemblages. CCY was established in 1997 by the Department of

Marine Microbiology of the Netherlands Institute of Ecology (NIOO-KNAW), now

the Royal Netherlands Institute for Sea Research (NIOZ), in Yerseke. CCY

provides a platform for maintaining novel strains of microorganisms that have

been isolated in the course of scientific research projects. These strains are

maintained alive as cultures or cryo-preserved. CCY provides expertise on

isolation, cultivation, and identification of novel strains and on their cryo-

preservation. The collection is available to academic and commercial parties,

except when agreements about specific strains prevent this.

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Culture collections are not stamp or coin

collections

Why collect cultures?

Cultures are genetic repositories

Isolation, purification and cultivation

Characterization and identification

Optimization of growth media and cultivation conditions

Screening for bioactives and other valuable compounds

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Collection of ~700 microorganisms:

mainly photoautotrophs

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Prorocentrum lima Green algae

Tetraselmis striata Phaeodactylum tricornutum

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CO2

O2

Fatty acid,

sugars,

proteins..

Pediastrum CCY0386

Photoautotrophs

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Cryopreservation

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Cryopreservation

Necessary because:

• Guarantee the stability of the genetic composition

• To avoid human errors (e.g. mislabeling and contamination)

• Decrease maintenance costs and labor-intensive activities (e.g. sub-

culturing, medium preparation, glassware cleaning)

• Allows the collection of many more cultures

• Needs minimum storage space

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Controlling the rates of cooling:

• Controlled-cooling canister as Mr. Frosty (approx. -1C per min -1)

• Controlled Rate Freezer (-XC per min -1)

Mr. Frosty

Isopropanol resevoir

Cryovial holder

Cryomed1010

Controlled rate freezer

Cryopreservation

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plasmolysis Intracellular ice

Cryopreservation: cooling rate

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CryoProtective Agents (CPAs):

• Penetrating Agents (MeOH,

DMSO, Glycerol)

• Lower the intracellular water

freezing-point

• Decrease plasmolysis

Cryopreservation

Phormidium

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Lipid droplets in Phaeodactylum

stained with Bodipy. The chloroplast

fluoresces red.

Lipid droplets in Phaeodactylum

stained with Nile Red

Lipids

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408 strains of the Culture Collection Yerseke (CCY) were tested

The BIODIPY test revealed 220 positive strains (in which green

fluorescence was detected), of which 165 showed strong

fluorescence or many fluorescing lipid bodies.

CCY0216 (Nitzschia sp., diatom) CCY0224 (Surirella brebisonii, diatom) CCY0305 (Odontella sp., diatom)

CCY0399 (Navicula sp., diatom) CCY0348 (Phormidium sp., cyanobacteria) CCY9611 (Gloeothece sp. cyanobacteria)

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100% N 50% N 10% N 0% N

BODIPY labeling of CCY0033 after nitrogen starvation

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CCY0033

CCY0438

CCY0913

+glycerol

+glycerol

+glycerol

Lipids droplets in Phaeodactylum CCY0033

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The cells are almost completely filled with two lipid droplets

Effect of glycerol on lipid accumulation

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6/5/2015 InteSusAl - confidential 20

Culture fed with glycerol (50 g/l) Culture did not receive glycerol

Effect of glycerol on cell size

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Growth condition Cell size (μm)

14⁰C-100%N 4.7

14⁰C, 100%N + 50g/l glycerol 5.3

23⁰C, 100%N 4.4

23⁰C, 100%N + 50 g/l glycerol 5.2

23⁰C, 10%N 4.4

23⁰C, 10%N + 50g/l glycerol 5.4

Cells 13% (14oC), 18% (23oC, 100% N) or 23% (23oC, 10% N) bigger when fed with glycerol

Lipid content (% of dry weight)

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Growth condition % lipids C:N Pigments

pg/cell

POC

mmol/l

PON

mmol/l

14⁰C, 100%N 7.1 9 0.51 23.9 2.68

14⁰C, 100%N + 50g/l glycerol 28.8 25 0.36 72.1 2.87

23⁰C, 100%N 11.4 8 0.63 15.3 2.03

23⁰C, 100%N + 50 g/l glycerol 33.0 23 0.42 79.5 3.43

23⁰C, 10%N 12.2 13 0.45 18.1 1.44

23⁰C, 10%N + 50g/l glycerol 47.5 78 0.15 66.5 0.86

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The Marine Microbiome Applied (MaMA)

‘Where Industry meets Academia’

An Industry Conference back-to-back to an Open Science

Meeting on the biotechnical application of marine microbes

organized by MaCuMBA

Berlin-Adlershof

27-30 June 2016

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www.macumbaproject.eu

ANNOUNCING: