Analysing the Plankton of Milford Haven
Collection of Samples
Typically, plankton is collected from a small boat pulling a net behind, a method I have always used since the 1980's. Retiring from Dale Fort some years ago, with no access to a boat. it took some months of trial and error to develop a good and consistent method for shore collections. For some years now I have been taking samples approximately at fortnight intervals throughout the year. Winter working from a boat is always difficult anyway and at Dale Fort it was rare to sample outside the late spring and summer.
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My current method is to use a 25 cm diameter plankton net with a 55 micron mesh. This collects a good range of species including the occasional nanoplankton at 10-20 microns. It is probable that some of the larger arrow-worms in the macroplankton manage to avoid becoming trapped but most organisms like tintinnids, dinoflagellates, diatoms, copepods and larvae of the shore invertebrates are collected.
Two samples are made, approximately 10 minutes pulling for each one with care taken when reversing.
The net is pulled along the side of a pontoon or Dale Fort jetty in the western area of Milford Haven. The former structure is removed from the village during autumn and winter, hence the change to the jetty at Dale Fort. The net is kept around a metre below the surface by suspended weights (2 steel spanners) to avoid detritus at the surface. The pull is done from the end of a trekking pole held out from the walkway. This keeps the net away from the edge of the pontoon as well as preventing me from falling into the water.
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To maintain some form of control the trawl is made at the top of the high spring tide which provides maximum depth. This is invariably between 7 - 8am. At this time the zooplankton will still be near the surface and the incoming tide will have swept across the rock and sandy shores bringing early larval forms. As an example, Phoronid larvae upon release from the adult quickly develop and most specimens appearing in the samples are between 6 -10 hours old.
Analysing Samples
The two samples are decanted into separate containers to be transported home and usually within 30 minutes they are being analysed. One is placed in a fridge while the other is being examined. Initially the sample is divided up into several petri dishes and looked at under an inspection microscope x1 - x15 magnification. Any large items, e.g. Gnathia or large copepods, can be moved with a pipette to a separate, small dish. T-light dishes can be ideal. Small samples are pipetted on to glass slides to look under a x10 objective, DIC. Under the coverslip the "square" of sample is systematicly observed starting in the top left corner working down, back and forth until it has been analysed.
Inspection microscope to left of the Olympus BH2. Sony A6500 camera relays image to a 30in 4K screen
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Larger material is photographed either with a Canon 7D ll with 65mm MPE lens or Olympus OM1 on bellows fitted with different objective lenses. Both with twin flash. These are both tethered to a PC for stacking
I only tend to look at live plankton and that means the sample needs to be viewed that day. Tintinnids rarely last more than an hour or two and so the pressure is on. Up to 1000 images plus video sequences are taken, much is for record purposes so that I can go back later and check, especially mis-identification. I rarely stop for ID but make sure I have plenty of photographs, spending the next week going through the pictures.
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Everything is recorded in a spreadsheet including a 5-point scoring system I use for relative abundance. See Data Page
Creating the Photographs
The Porcelain Crab Porcellana platycheles, just over 1mm long. This was taken on the BH2 with DIC at x10. Only a tiny area like the tip of the spine would be in the field of view and so over 40 sections would have to be photographed and then stitched. But to maintain depth of field each photo would be a composite of around 10 stacked images. This meant over 400 images went into creating this photo. The resulting stitch was around 300 megapixels which were then reduced.
This image of the Porcelain Crab was a single composite made from 15 stacked images using the Canon 7D ll. Each of the 15 images is taken at a slightly different focus point. The first is the first place some sharp material can be seen then the fine focus is moved into the specimen just a little to shift the focus to a new point. Take a photo. Then move it again, take a photo and so on until the speciment is completely out of focus. By combining all the sharp pixels using software like Helicon Focus (my preferred software) the final composite has good depth of field but will need some cleaning up.
Video stacking is an extension to the stacking method. I find it is ideal for microscopy, especially if you are in bit of a hurry. The Sony camera has 4K video, meaning each frame is approximately 8MP. Switch the video on with the specimen just out of focus. Then rotate the fine focus, passing through the specimen until it is out of focus. Switch off. This typically makes a 7-8 second burst of video. Drop this into Helicon Focus software which separates each frame into about 200 or so separate TIFF files. Then you render them into a final composite. The main drawback is if a lot of debris or moving bacteria, etc., is going on around the subject and then the composite needs extensive "cleaning". The content aware tool in Lightroom/Photoshop or healing/clone tools in other software will help in irradicating the mess. See Podon.
The marine water flea Podon, created in a video stack
Video clips are also useful to capture rapidly moving creatures, particularly really tiny nanoplankton. Use Helicon Focus to create the separate frames as TIFFs. It is then possible to look at the containing folder with all 200 plus images which you need to go through to find an image that is relatively sharp. That can be moved to a different folder to save. When Helicon is shut down it automatically wipes the video frames.
Single frame from a video clip, nanoplankton highly magnified.
Collecting plankton
Collecting plankton on a high tide at dawn. Looking down the Haven towards the rising sun. The pontoon is in the foreground and the Dale Fort Jetty is just visible, top right. April 2024.
Plankton collections are made from the Dale Fort Jetty (with permission from the FSC) during winter when there is no pontoon. Results are very similar to those taken down in Dale. This view shows a very high spring tide in February, again at dawn.
Helpful Books
More Information on photography and stacking.
Written by me, Julian Cremona, Published by Crowood Press
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Extreme Close-up Photography and Focus Stacking
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Beyond Extreme Close-up Photography
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Photographic Image Enhancement and Workflow
Identification of Plankton
The main books for marine plankton identification :
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Coastal Plankton - photo guide for European Seas by Otto Larink & Wilfried Westheide
Coastal Phytoplankton - photo guide for Northern European Seas by A Kraberg, M Baumann & C Durseen
The useful 1963 book Marine Plankton a practical guide by Newell and Newell is available secondhand through a number of booksellers