This Guide covers sample collection, preservation, examination and identification.

Sample collection

The first thing to consider is how you collect your specimens. Basically, there are two main growth strategies for algae, either to grow on a surface (periphytic), or to float in the water (planktonic), and the collection technique must take account of this. (Some algae grow epiphytically, and there are also algae which grow endophytically - within cells of higher plants).

Plankton

Plankton samples are most conveniently taken with a net (mesh size about 50 micrometres (µm.)). This concentrates the sample, but at the cost of possibly losing some of the very small plankters (microplankton). The method is qualitative, or at best, semi-quantitative (as it is hard to know how much water has actually passed through your net). The net will also collect all the zooplankton, which will start to graze your algae. Adding preservative will kill the zooplankton, but there are disadvantages in this for algal identification. Sieves may be used to strain out the larger animals, but the smaller grazers (eg. rotifers) will remain, and will graze the microplankton.

A water sample of known volume can be taken and allowed to settle, to concentrate it. This will take a long time, and preservative will need to be added. Samples can also be filtered through membrane filters which are then examined directly under the microscope. The filter and sample may need special treatment to clear it for microscopical examination.

Periphyton

"Periphyton" actually refers to all organisms growing attached to a surface, and includes bacteria, algae, protozoa, and microscopic multicellular animals. I am restricting this discussion to algae. Sampling technique differs according to the surface in question, which varies from rocks (epilithic) to mud (epipelic), as well as vegetation (epiphytic growth).

Soft sediments can be scraped with a spoon or other implement to sample the growth, while rocks can be scraped with a blade, or more conveniently a fingernail, and stones can be lifted from the water and brushed into a container using an old paintbrush or toothbrush. This can be used to obtain a qualitative estimate of species coverage, as a known area can be harvested, but the vigorous brushing can make identification of some species tricky. For best viewing, picking with the fingers is best.

Samples should be collected into a container half filled with a much larger volume of water than the sample: densely packed samples will soon start to decompose.

Alternative sampling techniques

Periphyton can be sampled on artificial surfaces which you place in the water for a period of time. The most useful surface is a microscope slide, as you can then examine your catch directly on the microscope. Apparatus can be devised for holding the slides, either attached to a float or a brick (for example). A reasonable growth of periphyton will be found aftr a week or so, but the algae will be quite densely packed on the slide. I have found that exposing the slides in a pond for about 24 hours then incubating them in filtered pond water for 2-3 weeks gives well-spaced young algal plants for identification (see the pages on Stigeoclonium identification). If possible incubate in a vessel floating in the pond. Colonisation in running water is usually slower.

Preservation (or not)

For best results, look at the samples while they are still fresh, as all preservatives have their disadvantages. Lugol's Iodine is widely used and appears to work well with diatoms and some green algae, so long as it is not used in excess (enough is added to turn the sample a "pale straw colour", about 0.3 ml per 100ml sample, but the amount needed varies with the amount of algal material in the sample). Some filamentous Zygnematacean algae, however, break up readily in Lugol's; also the iodine reacts with starch, showing up pyrenoids (a useful diagnostic test), but altering the colour which causes problems with some algae, e.g. cyanophyes. I have used 4% formalin to avoid these problems, but it is extremely toxic and air extraction must be used at all times with treated samples. It also disrupts the structure of some algae. Glutaraldehyde is reported to be better, but is also toxic. For a discussion of preservatives see this site.

Not:

Fresh plankton samples need the most immediate attention as grazing zooplankton can decimate your sample. Periphyton samples do not suffer the same problem, because the algae are larger and there tend to be few grazers anyway, though Chironomid larvae can chomp through filamentous algae quite alarmingly. Periphyton samples will keep quite happily in a fridge for several weeks or longer, provided the algae are not too crowded.

I have recently examined filtered phytoplankton sampled which were stored in a dark fridge for more than 10 months, and found that some algae, especially desmids, had survived, as had some zooplankton.

Examination

Set your microscope up for Köhler illumination *

Spread your sample thinly on a slide and place a coverslip on top. Scan using the X10 objective, then switch to a higher power when you find something interesting. Modern microscopes are fitted with parfocal lenses: they should be more or less in focus when you swing a different objective in (unless someone has fitted a lens from a different make or model). Focus up and down to view the object's upper surface, circumference, and (sometimes) lower surface. The 3-dimensional nature of algal cells can make them difficult to view (and photograph sucessfully), but shutting down the condenser iris a little can help by giving a greater depth of field.

Identification

I routinely use the following keys:

• EJ Cox, 1996: Identification of Freshwater Diatoms from Live Material; Chapman and Hall, London, ISBN 0 412 49380 2
• DM John, BA Whitton and AJ Brook, 2002: The Freshwater Algal Flora of the British Isles, Cambridge University Press, Cambridge. ISBN 0 521 77051 3
• EM Lind and AJ Brook, 1980: Desmids of the English Lake District; Freshwater Biological Asociation, Ambleside. ISBN 0 900386 40 1
• GW Prescott,1970: How to Know the Freshwater Algae; Wm C Brown, Dubuque.

Prescott was my only key for a long time and I still find it useful for a quick guide to possible ID, but it is out of date and many of the names have been changed. Similarly Lind and Brook is useful mainly for its portability. The best current key is John, Whitton and Brook, with current nomenclature and a fairly extensive coverage of British algae. However, it does not cover identification of diatoms. Cox is my choice here, especially as it allows you to identify fresh samples (without extensive treatment and mounting), but the book is currently out of print (taken over by Kluwer publishers).

Learning to use a key is a bit of a slog. You need to familiarise yourself with the terminology and look for the key characteristics in your unknown sample. The illustrations are a good guide to an identification, but you may need to plough through a key as well to confirm your ID. I hope the photographs on these pages help.

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