Bioluminescence investigations

Bioluminescence Investigations

What

We are investigating life that gives off light!!

already we have dinoflagellates (without flagella!) and Photobacterium phosphorum !
(hoping for some glowing mycelia, but wonder if it is simply phosphorescence there...)

Who is involved?

It all started with 2 students from Nyon, who wanted to do their 'travail de maturité' with some bioluminscent bacteria, to make a mosaic!

• Inès Nerodenkova
• Luca DN

They ordered Vibrio fischeri from a French biotech company in a special kit.

while a bit of light was visible in cultures, in comparison to controls, and determined by unbiased dark-adapted participants, it was not very bright, and no good pictures were obtained of it...

Advice was requested from

• Siouxsie Wiles - Australian researcher, who recommended Photobacterium phosphorum, as brighter.

Help was also obtained from

• Hackuarium members (Rachel, Luc H)

• more members are getting involved (Esther) and
• new members may be coming on board (Christelle) for more fun!

esp, as Rachel also ordered some bioluminescent Dinoflagellates from a company in the states.

Recent results in the new BioLab in Ecublens

The strain obtained by the students from Nyon last year actually isn't V. fischeri, but the recommended Photobacterium phosphorum

• pcr with 16S primers 671 and 672...
  

671: primer name UV_27F Sequence: AGR GTT YGA TYM TGG CTC AG
672: primer name UV_907R Sequence: CCG TCA ATT CMT TTR AGT TT

For additional documentation, 27F and 907R are in this table:
https://www.nature.com/articles/nprot.2014.067/tables/1
(there are references of other classic papers to the right of this table)

after PCR (with Q5 pol to allow proofreading and avoid errors), band purification (Qiagen columns) and
then, sequencing (light-seq from GATC, just about 5chf/result!) ,
we obtained very clear results!
File:36AJ98 77018090 77018090.pdf

blast showed about 92% identity over the 16S rRNA region sequenced vs. V. fischeri, so it seems clear that the french co also thought the P. phos was brighter, but didn't want to waste their kit boxes! :)

• Christelle got some good pictures from plates, but new experiments with more salt are being run now!

• This is one of the best pics of the dinoflagellates (about a half hour after their 'twilight' - they are strict in terms of circadian rhythms) so far...

Will add a pic of these dino cells soon!!

What's Next

The challenge is to get their real color in images, and to make them bright...
extra salt is seemingly very important!
but maybe not malic acid... The second test for this is ongoing!
Clearly, temperature is a big factor in getting colonies bright! After being in the frig, we see more!!
However, the brightest colonies do not always streak out well on fresh plates. Maybe the aldehydes in the light reaction are toxic, and this is part of the reason for quorum effect regulation - to make sure there are some other bugs that will not be bright, and remain able to grow!??
more results to come soon!