Visualising symbiotic bacteria: a practical application of fluorescence in situ hybridisation (FISH)

Many marine microbiologists use fluorescence in situ hybridisation (FISH) to identify and visualise specific bacteria. Cells are permeabilized to allow probes to enter, once inside the cell the probes target specific complementary nucleic acid sequences and become hybridized to them. The probes are labelled with fluorescence which can then be analysed using epifluroscence, laser scanning microscopy or flow cytometry. Typically when researchers use this technique to study symbiotic bacteria the tissue in question is paraffin or frozen sectioned; the former technique involves lengthy preparation by trained and skilled individuals whilst the later often compromises resolution. Authors of the current paper developed a FISH procedure which allows entire gill filaments to be used. The preparation of which simply requires careful dissection techniques and the ability to follow an off the shelf FISH kit, further more using this technique a single gill filament would correspond to the same area as 80 paraffin or frozen sections.

The authors of this paper were specifically interested in the symbiosis between chemosynthetic bacteria and two species of deep-sea mussels. The technique they developed using a whole mounted gill filament allowed them to visualise the the spatial distribution of thiotrophic and methanotrophic bacteria for which they had developed specific probes for. They found that in both species the majority of the lateral surface contained bacteriocytes for both thiotrophic and methanotrophic bacteria; however cilary cells did not contain bacteriocytes. In one species this was visualised by dark band-like zones which did not fluoresce which were the ciliary junctions, in another dark non-fluorescing oval areas were seen which corresponded to ciliary tufts on the gill. Authors predict that using this technique in addition to confocal laser-scanning a three dimensional analysis could also take place.

I feel the authors of this paper have taken the logical step forward in science in order to use the FISH technique to its full potential which has allowed them to gather more information about the specific microbial community they were interested in. Visualisation of the spatial distribution of microbial communities provides information on both community structure as well as possible insights into ecological function. It therefore makes perfect sense to examine larger areas at any one time, not to mention the amount of time it saves compared to looking at tiny 10µm thick sections. I agree with the authors in their prediction that this technique could be used for other species with symbiotic bacterial communities and possibly even for multi-layered tissues if the correct permeability conditions were optimized.

Fujinoki, M., Koito, T., Fujiwara, Y., Kawato, M., Tada, Y., Hamasaki, K., Jimbo, M., et al. (2012). Whole-mount fluorescence in situ hybridization to visualize symbiotic bacteria in the gills of deep‑sea mussels. Aquatic Biology, 14(2), 135–140.

http://www.int-res.com/abstracts/ab/v14/n2/p135-140/