Explorations in BioLuminescence
From fireflies to stinky squids... the future is bright
Isolation of luminous bacteria from fish or squid
P. phosphoreum is one of the commonest spoilage organisms of fish such as cod. It is not known to cause disease, but there are reports of people being startled by glowing fish fingers in the fridge!
To isolate glowing bacteria from fish, obtain a freshly-caught seafish or squid. It is very important that the fish has not been frozen or washed in fresh water. It is also better if the fish has not been stored on ice. Place the fish in a container with 3% NaCl solution. The liquid should be deep enough to come half way up the fish.
Cover the container and store the fish for 24 hours at about 12–15 °C. Note: if this temperature is difficult to achieve, place the fish in a fridge at about 4 °C for 48–72 hours.
After incubation, take the container with the fish into a dark room. When your eyes have been adopted to dark, light spots will be visible on the skin of the fish. Use a sterile toothpick or disposable sterile loop or needle to aseptically transfer the brightest spots onto sterile fish enrichment agar plates. Tip: some people find it useful to use a dim red lamp (e.g., a photographic safety lamp) for this step. Turn the fish away from the lamp so that the glowing colonies are in the shade and therefore visible.
Transfer the cultures to new agar plates every second day if you are incubating them at temperatures around 12 °C or once a week if you are storing them in a fridge. P. phosphoreum will grow at 4 °C; V. fischeri will not. By selecting the brightest colonies when inoculating, it should be possible to isolate a pure culture.
Fish enrichment medium
- Boil 250 g fish meat in 1 l of water.
- Add 30 g NaCl and sieve to remove solids.
- Add 10 g peptone, 10 ml glycerol and 1 g yeast extract.
- Adjust the pH to 7.
- Autoclave at 121 °C for 15–20 minutes.
For a solid medium add 15–20 g of agar to every litre of broth.
Safety note Several species of Vibrio are pathogenic. The chances of inadvertently isolating pathogens in this procedure can be reduced by using at least 3% salt solution and incubating fish and plates at no more than 15 °C. Human pathogens are unlikely to grow under such conditions.