Presented by
M. Tresh
MBRC/LIBFISH Aqua Component Group
1. INTRODUCTION
The first level of the marine food chain is constituted by micro-algae, which are the main feed for most of zooplankton. Phytoplankton is used in a hatchery as basic live feed for early stages of fish larvae. After a few days, larvae are switched to zooplankton (Rotifers, Daphnia sp. and Artemia sp.). It is however essential to produce various types of phytoplankton continuously in order to nourish different zooplankton as well, which are major components of fish and shrimp larvae diets before they can be fed on artificial pellets.
The use of pure algae cultures to feed fish larvae is necessary in order to avoid poisonous species and parasites. Pure cultures of some species of algae are therefore produced in large quantities to cover the requirements of hatcheries.
2. WORK AT MBRC
2.1 Initial situation
During the previous months members of the LIBFISH Aqua Working Group have been cultivating phytoplankton inside 1,000 I fiberglass tanks. Further to microscopic examination, it appeared that this ‘green water’ contained several species of algae.
2.2 Requirements
Pure culture of phytoplankton is needed for the pilot hatchery programme being planned at the MBRC.
2.3 Isolation of Chlorella sp.
Two litres of green water from the 1,000 litre fiberglass tank were transferred to the laboratory. The culture has been filtered with 50μ, plankton net. Then, 10ml of this filtered green water was introduced within a 1 litre bottle and mixed up with cultivation media at room temperature. The bottle was kept in sunlight.
After one week, the culture was microscopically examined and it was found that most of the algae present were of Chlorella sp., e.g. C. nitzchia, C. navicula, and C. punnularia.
The cultivation medium was then transferred into one 2 litre bottle and one 3 litre bottle. The medium was maintained at room temperature in natural sunlight and subjected to forced aeration for one week, during which a good growth was observed.
Serial dilution cultures were done to isolate Chlorella sp. Four dilution steps with five parallels had to be done for each sample, as shown in the accompanying figure.
Figure VIII/1. Isolation of Chlorella sp.
The steps of dilution are thus:
A 10ml-sample is taken and 1ml of it is mixed with 9ml media.
---> first dilution
From the last test tube 1ml is inoculated to each of five test tubes.
---> 2nd dilution 10-1
This step is repeated to produce the next dilution,
---> 3rd dilution 10-2
and the final dilution.
---> 4th dilution 10-3
All serial dilution cultures were incubated at room temperature with aeration and fluorescent light for one week and were then examined microscopically.
2.4 Results
In the first dilution all test tubes contain mixed culture.
In the second dilution, one test tube contained pure Chlorella sp.
In the third dilution, four test tubes contained pure Chlorella sp.
In the fourth dilution, three test tubes were with pure Chlorella sp.
3. CONCLUSION
The exercise described above allowed the recovery of pure Chlorella sp. strain from a mixed culture.
Pure strains of phytoplankton should be kept for long periods in a special lab, annexed to the hatchery. Whenever necessary, they can be used to produce large cultures to cover the needs of the hatchery.
