North Pacific Marine Science Organization

PICES Press Vol. 22, No. 2

Microbial Culture Collection at the National Institute for Environmental Studies,Tsukuba, Japanby Masanobu Kawachi and Mary-Hélène NoëlMCC-NIES team (from left to right): Ishimoto (Charales curator), Yumoto (curator, DNA), Mori (curator, cryopreservation), Kawachi (head), Matsui(assistant curator), Noël (curator, projects), Niitsuma (assistant curator, DNA), Sato (curator, flow cytometry), Shimura (Post-Doc, cyanobacteria), Fujii(secretary).

History and characteristics of the MCC-NIES collection

partially supported by the Time Capsule Project conductedby the Ministry of Environment of Japan since 2002.

The Microbial Culture Collection at the National Institutefor Environmental Studies (MCC-NIES), located in
Tsukuba, Japan, was founded as an “environmental studyoriented” culture collection in 1983 when eutrophication oflakes and rivers, and air and water pollution were severe in
Japan. The MCC-NIES started with ca. 250 strains mainlyof red-tide-forming algae (Chattonella antiqua andHeterosigma
akashiwo)
andwater-bloom-forming
cyanobacteria (Microcystis aeruginosa). Although theMCC-NIES is still characterized by such types of strains,the collection now holds almost all eukaryotic lineages anda diversity of cyanobacteria, too. At present the collection
includes 18 phyla, 51 classes, 354 genera, 718 species and2,356 strains.

In 2002, the MCC-NIES was selected as the core repository
for algae in the National BioResource Project (NBRP)
conducted by the Ministry of Education, Culture, Sports,ScienceandTechnologyofJapan(NBRP:
http://www.nbrp.jp). In this framework, more than 200
strains of Microcystis and Anabaena, collected fromrepresentative eutrophic lakes all over Japan, weredeposited by the National Science Museum along withphylogenetically diverse strains of microalgae and protozoa
deposited by the University of Tsukuba. In addition, morethan 300 strains of cyanobacteria and eukaryotic
microalgae maintained at the IAM Collection (Universityof Tokyo) were transferred to the MCC-NIES up until theend of FY 2006, when the IAM Collection was closed.

Most of the MCC-NIES strains have been directly
deposited by researchers, but some have been depositedfrom exchanges with other collections. Around 80% of theMCC-NIES strains were originally sourced from Japan,
giving the collection a high level of specificity.
This is also the only culture collection holding major
endangered macroalgae. The collection, which started exsitu conservation of endangered algae in Japan, has been in
operation since the mid-1990s. In the list of endangered
Japanese wildlife (the red list) compiled by the Ministry ofEnvironment of Japan in 2007, 116 taxa (species andvarieties) of algae are listed as extinct, extinct in the wild,or as endangered in Japan. At present, the MCC-NIES
holds ca. 300 strains of these endangered algae, including
Charales and freshwater red algae. The collection isCharales strains maintained in MCC-NIES.

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North Pacific Marine Science Organization
Since the start of the collection, the Committee forEvaluating Microbial Culture Strains has evaluated theNIES strains upon deposition based on a set of criteria. At
present, the Committee includes nine researchers at NIES
and six supervisors outside NIES. In addition, since 2002
the MCC-NIES has been supervised by the Steering
Committee of the NBRP Algae.

The collection includes:
 Evolutionarily important species such as Mesostigmaviride (NIES-296) and charophytes (NIES-1601);
 Experimental materials that have been well-studied ingenomic, genetic, molecular, and physiological terms,
such as Cyanidioschyzon merolae (NIES-1332/10D),
Chlamydomonas reinhardtii (NIES-2235/C-9), andThermosynechococcus elongatus (NIES-2133/BP-1);
 Ecologically significant species such as Prochlorococcus
marinus (NIES-2086) and Micromonas pusilla (NIES1411);
 Harmful algal species such as Microcystis aeruginosa(NIES-44) and Chattonella marina (NIES-3);
 Commercially useful strains such as Botryococcus
braunii (NIES-836), Porphyridium (NIES-1035) and
Chlorella vulgaris (NIES-227).

Maintenance of strains
About 3/4 of the NIES strains (~ 2,000 strains) aremaintained by subculturing under optimal and/or suboptimal conditions, mostly ranging from 5 to 25°C (37 or
45°C for thermophilic strains) and with a 4 to 50 µmol·m–2·s–1 photon flux density in a 12-h-light:12-h-dark lightregime.

Test tube of Anabaena.

The strains are serially transferred at 10-day to 6-monthintervals. Specially designed software for the collection
allows a flexible gestion of the daily transfers. Maintenanceconditions differ with each algal strain and are individuallyindicated in the catalogue of strains on our web site.
To prevent the loss of strains during maintenance bysubculturing, we conduct weekly growth checks. Once ayear, we also check axenic strains for the absence ofbacteria by using several bacterial check media.
The remaining strains—about 600 including most of thecyanobacterial strains and some of the green and red algal
strains—are cryopreserved only, in the vapor phase ofliquid nitrogen.

A. Thermosynechococcus (NIES-2133), B. Cyanidioschyzon (NIES-1332),
C. Microcystis (NIES-44), D. Chattonella (NIES-3), E. Botryococcus(NIES-836), F. Porphyridium (NIES-1035). Scale bar = 10 μm.

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PICES Press Vol. 22, No. 2
MCC-NIES services
Most of the strains are available for education, research anddevelopment in accordance with the “Agreement fordistribution”. The MCC-NIES also provides genomic
DNA for worldwide distribution.

Packaging for worldwide distribution of culture strains.

Technical advice and follow-up of the distributed strains
are provided to customers. Usual time for delivery of anordered strain is 10 days as most of the strains aremaintained under active growth. For strains kept as
cryopreserved samples, the delay is longer since the MCCNIES first return the culture to active growth before
shipping it.
The MCC-NIES accepts the deposit of strains that areenvironmentally important, as well as those for basic andapplied studies. The collection also accepts the depositionof strain types of cyanobacteria and specimen types of
eukaryotic microalgae as frozen samples.

Upper: Subculture of strains in liquid or agar media. Lower: Cryopreserved strains in liquid nitrogen tanks.

Scientific names and phylogeny

Useful information relating to the DNA barcodes,
photosynthetic pigments, morphology and other relevant
literature are provided for each strain at our web site. The
MCC-NIES collection welcomes joint research andcollaborations; please contact us at mcc@nies.go.jp.
Japanese, English and French are the available languages
for any communication with the collection.

The scientific names of the NIES strains are given
primarily by the depositors. However, we have used DNA
sequencing (mostly of the 18S rRNA gene) to re-evaluate
the strains for which DNA sequence data have not yet beenreported. As a result, we have changed the scientific namesof the misidentified strains, although we have left theirformer names as “Formerly identified as.” We have also
added “Re-identified at NIES by DNA sequencing” in“Identified by” and, if the original scientific names of thesestrains were appropriate, we have simply indicated“Confirmed at NIES by DNA sequencing”, with genenames and accession numbers in “Gene data.” We are still
re-evaluating the remaining strains.

A culture collection is “team work”, each step being
important for the final state of the cultures (from washing,media preparation to sub-culturing, etc.). We take care ofthe culture strains as much as possible and hope to share
our passion for microalgae.

Dr. Masanobu Kawachi (kawach9i@nies.go.jp) is a microalgae taxonomist working at the National Institute for Environmental Studies
based at Tsukuba, Japan. Since 2012, he has been the head of the Microbial Culture Collection. As a Ph.D. student, Masanobu revealed
the food capturing role of the haptonema for the haptophyte Chrysochromulina. He spent 6 months at an Antartica research base, andtravelled worldwide to collect microalgae samples from various environments. With co-workers he is involved in the discovery of a newclass, the Pinguiophyceae, having specific fatty acid content. Taxonomic revision of Chattonella and toxic cyanobacteria are among thefields of research he is working on. He is currently participating in several research projects: on coral symbionts with global warming
concerns, picoplankton diversity with the environmental DNA analysis, and biofuel production from microalgae.
Dr. Mary-Hélène Noël (noel.mary-helene@nies.go.jp) came to the National Institute for Environmental Studies in 1996 as a Post-Doc
right after her Ph.D. graduation from the University of Paris XII, CNRS 386 / E.N.S. With a background in marine biogeochemistry, shespent the first 4 years at the Water and Soil Division of NIES, then started to work on the coccolithophorid life cycle at the Biological
Division and MCC-NIES. Her participation to the culture collection consists of recovering the strains with growth troubles, adapting
strains, and providing advice to customers internationally. She is also involved in research projects on marine picoplankton diversity andlife cycle studies.

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