Classification of algal divisions

Modern classifications of the Eukaryotes, as discussed in lecture and in Campbell, have begun to address the real relationships among the many groups of algae and other eukaryotes. Based on this work, it is apparent that the “algae” as a whole are definitely not monophyletic. However, the different groups of algae can be classified on the basis of a number of characteristics. Color has been an important means of classifying algae, and gives many groups their names. However, other characteristics, such as type of photosynthetic food reserve, flagella type, cell wall structure and composition, and life history, have been important in further distinguishing the algal divisions. The major divisions and their characteristics are given in the table at the end of this exercise. We will cover the following groups of algae:

Chlorophyta and Charophyta (within the modern grouping “plants”)
Euglenozoa (or Euglenophyta)
Phaeophyta, Chrysophyta, and Bacillariophyta (within the Strameopiles)
Dinoflagellata (within the Alveolata)

(A) Photosynthetic pigments and chloroplasts

The photosynthetic pigments are located in chloroplasts. The pigment common to all algae is chlorophyll a. Other types of chlorophyll are present such as chlorophyll b, c, and d. Chlorophyll a is the site of photosynthetic light reactions. The other chlorophylls transfer their light energy to chlorophyll a and are therefore called accessory pigments. Accessory pigments absorb light at a different wavelength than chlorophyll a, usually at slightly shorter wavelengths and transfer the energy via fluorescence. In addition to some of the chlorophyll pigments, other groups of pigments are considered accessory, and include carotenoid pigments (usually yellow and orange in color), xanthophylls (yellow-green and brown colors), and phycobilins (either bluish or red in color). The production of accessory pigments can often obscure the underlying green chlorophyll pigment. In addition, individual species may vary in the production of accessory pigments depending upon the depth at which they live or their stage of growth. Note the different colors of green, brown, and red algae.

(1) Chloroplast shapes
Prepare whole mounts and examine the live material of Chlorella (or Ulothrix), Spirogyra, Oedogonium, and Cladophora under your compound microscope. Notice the variation in the shape of the chloroplast. The most common appearance in the green algae is called a parietal chloroplast, which means the chloroplast lies against the cell wall. It may or may not surround the entire cell, sometimes it is located only on one side. Which alga(e) have parietal chloroplasts? What do you notice about the shape of the chloroplast in Spirogyra? Can you guess how this alga got its name? When the chloroplast becomes heavily divided it appears net-like or reticulate. Which alga(e) exhibit this type of chloroplast? Can theorize on the evolutionary significance of various chloroplast shapes? Now examine the live material of a red alga under your compound microscope. What shape are the chloroplasts? When many small chloroplasts are found in a single cell, we call them discoid, nevertheless, they are often found along the cell wall in the parietal position. Most higher plants have discoid chloroplasts.

(2) Colors
Compare the colors of chloroplasts observed above with that of a diatom and a thin section of a kelp (Fucus or Laminaria). Compare the colors with the chart on accessory pigments. What colors can be attributed to which pigment names? What are the advantages of the various accessory pigment combinations?

Demonstrate the presence of chlorophyll a within the brown and red algae by taking a piece of each one and dipping it in boiling water to extract the accessory pigment. What color does the water become? What color remains in the algae tissue? Now place the tissue in a beaker of cold acetone. What color is the solvent and algal tissue after extraction? What does this procedure demonstrate about the solubility and chemical nature of the two types of pigments?

(B) Food reserves

Re-examine the slide of Spirogyra. Do you notice the small dots embedded in the chloroplast? These are called pyrenoids and are the site of starch storage in the green algae. Starch will stain blue-black in the presence of iodine. Place a drop of IKI (iodine stain) on the side of your cover slip and using a tissue on the opposite side, draw the stain through the slide mount. Watch the pyrenoids during this process. What color do they appear? Try this test with other green algae observed above. Where are the pyrenoids located?

Try the test with a red alga. The red algae have a storage product similar to starch, but the bonding in the glucose molecule chains is different. The color reaction is usually brown. Did you observe any pyrenoids?

Finally, observe the diatoms. Try the iodine stain. Diatoms store oil droplets as their primary food reserve. If these diatoms have been growing under optimal conditions, you should be able to observe the oil droplets disperse throughout the cell.

(C) Flagella

Locomotion in algae is largely based on the action of flagella. The figure below illustrates the wide variety of flagella present in the algae. The primary distinctions used for classification are the number of flagella, their location on the cell, and their morphology. Two major types of flagella are recognized: the smooth or acronematic and the hairy or pleuronematic types. The smooth flagella generally moves by whiplash motion and the hairy flagella moves by a pulling motion.

(1) Live Mounts
Make live mounts of Euglena, Chlamydomonas, Gonium, and Volvox. Observe them under the microscope while they are active. They usually move quite rapidly and because you will need to observe them under high power, your field of view is limited. Now add some methyl cellulose or glycerine to slow them down. You will need to adjust your microscope carefully and perhaps find a cell that is "stuck" to the cover slip.

(a) Chlamydomonas and Gonium
Chlamydomonas is a unicellular green alga with two equal flagella. Observe how they move in a whiplash fashion. Gonium is a colonial alga with 8 to 16 Chlamydomonas-type cells united together. Is there any coordination to the movement of the flagella? Describe how this colony moves through the water.

(b) Volvox
It is difficult to observe the flagella on the individual cells in Volvox, but you should be able to see the slow movement of the colony if you haven't squashed it too hard under your coverslip.

(c) Euglena
Euglena has two flagella, however, only one is readily visible. The other flagellum is short and does not extend beyond the cell body. How do the flagella in Euglena move? Euglena can lose its flagella under certain environmental conditions and will move through the substrate by an amoeboid-like action called metaboly. Can you observe such motion in your material? If you have difficulty observing flagella in your live preparations, observe the prepared slides that have been specially stained for flagella. Unfortunately, you will miss seeing them in action.

(d) Diatoms
Not all algae have flagellar stages in their life history. For example, red algae do not have flagella and the reproductive spores are distributed solely by wave and currents. Diatoms are another group of algae that lack flagella, yet are able to move over substrates by secreting mucilage through pores in their cell wall. Make a preparation of pennate diatoms. Do not place too much water on the slide and you may want to include some fine mud as a substrate on which these diatoms can move. The diatoms are noticeable by their golden-brown color. Observe their movement. On intertidal mud flats, diatoms often move to the surface of the mud at low tide turning a brown mud flat a bright golden color.

(D) Cell Walls, material and construction

Like the higher plants, many algae have cellulose as part of their cell wall. The cell wall is constructed of microfibrils of cellulose with the microfibrils intertwined and criss-crossed in layers, like a sheet of plywood. Cell walls are difficult to observe in algae as they are clear and rather thin compared to cell walls in plants. Stains are used to distinguish cell walls and a few prepared slides may be available for demonstration on the front desk

In addition to cellulosic cell walls, algae have a number of other compounds used in cell wall construction. Euglena and Chlamydomonas have proteinaceous materials in their cell walls. The cell wall material in Euglena is arranged in a spiral pattern around the cell and is called a pellicle. This flexible wall allows it to move freely in the bottom mud of ponds and lakes.

(1) Silica
Another compound found in algal cell walls is silica (SO2). Diatoms are the primary algae exhibiting this type of construction and their cell wall is called a frustule. The cell wall is quite resistant to decay and in some parts of the world (such as Lompoc, California), large deposits of marine diatoms are mined for diatomaceous earth. This material is used as an ingredient in many commercial products such as polishes, paint removers, detergents, and insulation. It is also used as a filtering agent for pool filters and beer clarification. Obtain some diatomaceous earth and examine it under your compound microscope. A frustule is composed of two halves that fit together much like a petri dish. Much of the material will be broken, but you should be able to observe the ornate patterns of the cell walls. Unlike the pennate (or elongate) cells you observed above, most of the planktonic marine diatoms are circular and are called centric diatoms. The ornamentation is used as a taxonomic tool in the identification of the diatoms. The cell wall in living diatoms is often obscured by the protoplasm and the cells must be cleared in nitric acid before identification.

(2) Calcium carbonate
Calcium carbonate (CaCO3) is found in the cell walls of many groups of algae, including Chrysophyta, Chlorophyta, and especially Rhodophyta. Calcium carbonate is often a by-product of photosynthesis and the crystals are deposited in the cell wall matrix. In the seaweeds, the presence of calcium carbonate is thought to have evolved as a defense against herbivores. Examine the calcareous algae available in the laboratory. In the red algae, the calcareous members are found in two forms; those which are erect and jointed are called articulate, and those which grow flat over the surface of rocks are called crustose. Examine both types under your dissecting microscope. You should be able to see small pores in the surface of the crustose algae, which are the location of the reproductive structures. In the articulate algae notice the filaments at the joints of each segment that hold the segments together. Place a small amount of 10% HCl on the articulate alga and wait for 5 minutes (be careful with HCl, it is an acid and can cause skin irritation. If you get any in your eyes, wash out immediately). The calcium carbonate should dissolve and you will be able to see the underlying filamentous structure of the alga.

(3) Polysaccharides
Many algae also have polysaccharides in their cell walls that are important commercial products, notably algin, carrageenan, and agar. You have probably come in contact with one of these products today if you brushed your teeth, put on make-up, ate some ice cream, drank chocolate milk, or had a cheese product. These chemicals are used as stabilizers in a variety of industrial and food related products and have become common in our diet and household products. See the demonstration of materials that use these polysaccharides. Algin comes from kelp and its production is a major industry in California, especially in southern California between Santa Barbara and San Diego. Carrageenan and agar come from red seaweeds. You can extract agar from local seaweeds, but it is not of high quality in terms of gelling strength.