I am trying to establish liquid cultures of common seagrass members such as species of Halophila, Halodule and Syringodium. Are these plants respond well to seaweed culture medium as Provasoli's Enriched Seawater medium (which we routinely use for our algal taxa) ? It is appreciated if some one send me the composition of liquid medium which is best for seagrasses.
Culture of plant modules collected in the field
1. Collect plants composed of at least 3 modules (i.e. 1 apical shoot plus 2
shoots and respective internodes and roots) and transport to the laboratory in
seawater. Place the plants in an aerated aquarium until transfer to the culture
media.
2. Prepare one of the following culture media in glass/plastic containers:
i) a monophasic medium of seawater (natural or artificial);
ii) a biphasic medium with natural sediments (preferably sterilized sand) and
seawater;
iii) a biphasic medium with an agar-solidified layer (2% agar w/v with seawater
as solvent) and seawater.
Attention: Plant the seagrass modules in the agar medium immediately before
the complete solidification of the agar, which should be allowed to cool down
first.
3. Distribute the containers with the plants in the three different culture media
per large volume (> 30L) aquaria filled with aerated seawater. Keep the aquaria
in a growth chamber at a light intensity, photoperiod, temperature and salinity
similar to the field conditions, or adjust according to the purpose of the
experiment.
4. Remove randomly without replacement four plants from each media after
the designated time period for the experiment. Determine plant survival and
other significant plant variables (e.g. rhizome elongation rate, shoots and
internodes production), according to the purpose of the experiment.
Culture of seedlings germinated in vitro from seeds
1. Clean the seeds by gently submerge them for 10 min. in a 10% (v:v)
commercial bleach solution made up with autoclaved seawater to avoid
contamination and overgrowth of algae.
2. Place the seeds individually in sterilized assay tubes containing biphasic
media of sandy sediment and seawater (previously autoclaved).
3. Place the assay tubes in a growth chamber at field temperature and light and
optimal germination conditions of salinity usually consisting of hyposaline shock
(e.g. 1 PSU for Zostera noltii, 16 PSU for Cymodocea nodosa), and allow seeds to
germinate, i.e. until the rupture of the seed coat and emergence of the
cotyledon.
4. After the emergence of the first chlorophyll-bearing leaf (seedling stage), transfer the plants to new
sterilized assay tubes containing sterile sandy sediment and full strength seawater. Add a nutritional solution of
macro- and micro-nutrients, PES culture medium (Provasoli 1968), as 1:3 of PES:Seawater.
5. Maintain the seedlings in a growth chamber at field temperature, salinity and light conditions, and transfer
them to bigger culture vessels (Magenta® – G7 300 ml Sigma Co.) when necessary using the same medium and
PES solutions as described above.
Additional information:
Cabaço S, Alexandre A, Santos R (2006) Survival and growth of the seagrass Zostera noltii in different culture media. In: Gambi MC, Borg
JA, Buia MC, Di Carlo G, Pergent-Martini C, Pergent G, Procaccini G (eds) Proceedings of the Mediterranean Seagrass Workshop, Malta
29 May - 4 June 2006. Biologia Marina Mediterranea, 13 (4):24-28.
Alexandre A, Cabaço S, Santos R, Serrão EA (2006) Timing and success of reproductive stages in the seagrass Zostera noltii. Aquatic
Botany 85:219-223.
Cabaço S, Santos R (2010) Reproduction of the eelgrass Zostera marina at the species southern distributional limit in the Eastern
Atlantic. Marine Ecology 31: 300-308.
Provasoli, L (1968) Media and prospects for cultivation of marine algae. In: A. Watanabe and A. Hattori (eds). Cultures and collections of
algae. The Japanese Society of Plant Physiologists, Tokyo. pp. 47–74.
Copy write from-ASSEMBLE-JRA1-Protocol-21.00
What you mean by liquid culutre, cell suspension culture then you refer the paper Subhashini et al., 2014, if you are interested with axenic cultures then refer to Bird et al., 1998; Duarte papers. More than culture medium it is sterilization and controlling of endophytes is the challenge in the seagrass culture.
I seek advice on culturing seagrass modules directly into liquid (single phase) not in biphasic medium. I would like to study effect of certain chemicals on growth and other physical and biochemical attributes. If there are any specific suggestions they are welcome.
Being the rooted flowering aquatic plant seagrasses naturally exposed to biphasic conditions underground root system and water surrounded above ground system. Physiologically too seagrasses absorb nutrients both by roots and leaves, so if you really want to study the effect of certain chemicals on growth and other physical and biochemical attributes you have to study in biphasic condition only or at cellular level by cell culture methods.
But for your interest, there are also options to grow the seeds (Enhalus accoroides, Thalassia etc.) for short durations about 20 days in seawater alone, I will send couple of such papers may be of useful to you.
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