Phytoplankton-Manual

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Phytoplankton
Identification Manual
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Phytoplankton Identification Manual
 National Institute of Oceanography

Disclaimer : The authors are responsible for the contents of this manual
First Edition : March 2004
X.N. Verlencar
Somshekar Desai
National Institute of Oceanography
Dona Paula, Goa - 403 004
Editors
V.K. Dhargalkar
B.S. Ingole
National Institute of Oceanography,

Dona Paula, Goa - 403 004
DTP
Devanand Kavlekar
Bioinformatics Centre,
National Institute of Oceanography, Dona Paula, Goa
Financial Support
Ministry of Environment & Forests, New Delhi

FOREWORD
Since its inception in 1966 the National Institute of Oceanography is involved

in taxonomic classification of marine phytoplankton, zooplankton, benthos and
other flora and fauna under the Project “ Measurement and Mapping of Marine
Resources”. Although the mandate of the project has been diversified with
changing times, the taxonomic identification continues to remain the thrust
area for all biological projects, especially those dealing with baseline studies
on ecobiology and environmental pollution. Visiting post-graduate and post-
doctorate students constantly look for information on taxanomic identification
which is spread over several books and journals.
The project “Survey and Inventerisation of Coastal Biodiversity (West coast)

funded byMinistry of Environment and Forests (MoEF), New Delhi, provided an
opportunity to bring together taxonomic experts from various disciplines. Their
efforts have resulted in preparation of this manual. This manual provides details
of taxonomic classification and description of the concerned organisms /
species. All the figures are well illustrated and detailed identification key is
provided. This should surely guide even a beginner to understand the
identification procedure.
S.R.Shetye
Director. NIO

PREFACE
Marine phytoplankton which constitutes diatoms, dinoflagellates, blue-green

algae, silicoflagellates, cocolithophors etc. contributes about 95% of primary
production in the oceans. On this depends the secondary production (zooplank-
ton) and tertiary production (fish, shellfish, mammals, etc.). Since phytoplankton
serve as a basic food source for animals in the sea their presence in large num-
bers may indicate the abundance of commercially important fish and shellfish
populations.
Coastal waters around India contain diverse groups of phytoplankton. The

biodiversity of these organisms is threatened due to large scale release of do-
mestic and industrial wastes. Hence there is a need to study these organisms in
more details.
The organization of this manual is made with a broad outline to accommo-

date the specific need of our Indian coastal waters. The manual is divided in
chapters which include - method of collection, preservation and identification
procedures. Species chosen for description were selected to provide good repre-
sentation of the commonly important species to give full spectrum of Chaetoceros
to be found in phytoplankton. The details should help the students as well as
researches to be thorough in the method of collection and identification of the
different phytoplanktonic species. We take the responsibility of any inadvertent
errors in this manual.
X. N. Verlecar
S. R. Desai

CONTENTS
1. Introduction
2. Methods of samplings
2.1 Bottle samplers
2.2 Plankton pumps
2.3 Plankton nets
3. Fixation and Preservation
3.1 Lugol’s solution

3.2 Bottling and labeling
4. Preparation for light microscopy

4.1 Acid cleaning
4.2 Specimen mounting
5. Identification of species
6. Bacillariophyceae (Diatoms)
6.1 Structure of the diatom cell
6.2 Gross vegetative structure
6.3 Cell division
6.4 Classification of Diatoms
7. Phyrrophyceae (Dinoflagellates)
8. Micrometry
9. Measurement of Biomass
9.1 Chlorophyll measurements
9.2 Cell counts
9.3 Cell count by drop count method
10. Measurement of productivity
11. Bibliography

1. Introduction
Phytoplankton (‘phyto’ = plant; ‘planktos’ = made to wander) are single celled

marine algae, some of which are capable of movement through the use of flagella
while others drift with currents. These microscopic plants range in size from 1/
1000 of a millimeter to 2 millimeters and float or swim in the upper 100 m of the
ocean, where they are dependent on sunlight for photosynthesis. In addition to
light and oxygen (O2), they require basic simple inorganic chemical nutrients,
such as phosphate (PO4) and nitrate (NO3). They also require carbon in the form
of carbon dioxide (CO2). Some phytoplankton, the diatoms, also require a form of
silicon (silicate, SiO 4 ) because they have a “glass-like” shell.
The marine phytoplankton come in a myriad of shapes, sizes, and forms, some
of them quite beautiful. Some drift on currents while others have an ability to
move around with the aid of flagella (Gymnodinium sanguineum). Some live as
single cells while others form chains or colonies. Marine algae are extremely
important to life on earth—probably the most important living organisms on the
planet. They impact us in at least three ways. First, they appear to be a signifi-
cant factor in controlling atmospheric carbon dioxide (CO2), a green house gas,
which in turn can influence heat retention in the Earth’s atmosphere. Secondly,
the phytoplankton and bacteria are the basis of the marine food web. At this level,
inorganic nutrients like phosphate, nitrate, and carbon dioxide are converted to
larger more complex organic molecules necessary for life. In turn, these micro-
scopic organisms provide the food for the higher trophic levels in the food web or
larger organisms higher in the food web, such as zooplankton, fishes and mam-
mals. For example, bivalve shellfish (oysters, mussels, scallops, clams) almost
exclusively consume phytoplankton for their food.
And lastly, marine algae are important because they can produce a variety of
highly toxic compounds—marine biotoxins. These compounds, some of which
can be released to the surrounding water while others are retained in the phy-
toplankton, can enter the food web and accumulate in fish and shellfish. In most
cases, fish and shellfish do not appear to be affected by these potent com-
pounds, but organisms higher in the food web, such as marine mammals and
humans, can be made ill or even die. It is this very lack of affect on the fish and
shellfish that we consume that makes marine biotoxins so dangerous, since
there is no outward sign that can forewarn the consumer. In virtually all cases,
the marine biotoxins produced by these phytoplankton, can only be detected

through laboratory analysis.
If conditions are right, phytoplankton can sometimes grow and reproduce at such
a high rate that they create dense, highly colored patches in the water. When
this happens, because the growth rate is so high, they deplete necessary nutri-
ents from the water, particularly dissolved oxygen (O2). When this happens fish
can suffocate. This sudden depletion in a small contained area can be a serious
problem in aquaculture since the fish are constrained in pens and cannot escape
into more oxygenated waters.
Algal Blooms: Most of the time, marine waters are characteristically blue or
green and reasonably clear. In the temperate waters of the northern latitudes,
water is seldom as clear as seen in tropical areas, where visibility can exceed
50-75 feet. In temperate waters, the limits of visibility or murkiness is usually the
result of algae in the water. However, in some unusual cases, a single microalgal
species can increase in abundance until they dominate the microscopic plant
community and reach such high concentrations that they discolor the water with
their pigments, these “blooms” of algae are often referred to as a “Red tide”.
Although referred to as “Red tides”, blooms are not only red, but can be brown,
yellow, green, or milky in color. These blooms can be caused by high concentra-
tions of toxic algal species and referred to as a “Harmful Algal Bloom” (abbrevi-
ated as HAB), however non-toxic species can also bloom and harmlessly dis-
color the water. Adverse effects can likewise occur when algal cell concentra-
tions are low and these cells are filtered from the water by shellfish such as
clams, mussels, oysters, scallops, or small fish. Many animals at higher levels
of the marine food chain are impacted by harmful algal blooms. Toxins can be
transferred through successive levels of the food chain, sometimes having lethal
effects.
2. Methods of samplings
There are three methods of sampling of phytoplankton.

1)Bottle samples 2) Plankton pumps 3) Plankton nets
2.1 Bottle Samplers:

Sampling by water sampler is the recommended method (Sournia 1978 p. 33) to
obtain a correct picture of the quantitative composition of the phytoplankton. A
water bottles sample contains all but the rarest organisms in the water mass

sampled and includes the whole size spectrum from the largest entities, like
diatom colonies to the smallest single cells (Tomas, 1997). These are ideal for
quantitative phytoplankton collections as required quantities of water can be col-
lected from the desired depth. Water samples are generally used from vessels,
ships or fish trawlers. Bottle sample method is a simplest method as generally
used for the collection of water samples from any desired depth of shallow sys-
tems like the near shore water, estuaries and mangroves.
2.1.1 Meyers water sampler (Fig 1) : It consists of an ordinary glass or perhaps

bottles of about 1-2 liter capacity and is enclosed with a metal band. It is weighted
below with a lead weight and there are two strong nylon graduated ropes. One
tied to the neck of the bottle and the other to the cork. While operation, the
corked up (closed bottle) is let down to the desired depth where the stopper is
jerked open by a strong pull of the cork rope. Water flows into the bottles and
then the cork rope is released to keep the cork closed. Afterwards, using the
neck rope, the bottle containing the water sample is taken out of the water col-
umns. Up to a depth of only 20m, this type of water samples could be used.
2.1.2 Friedinger’s Water Sampler (Fig 2): It is made of Plexiglas or Perspex with
two hinged covers. While operation, the sampler is sent down in an open state to
the desired depth and can be closed by a drop weight messenger, which falls
down inside on sliding rail and closes the covers and makes the bottle water
tight. By this way, the water together with the planktonic organisms of the speci-
fied column is trapped inside.
2.1.3
for Niskin waterenumeration
phytoplankton sampler (Figfrom
3a, 3b): It is employed
subsurface forvarious
levels to taking water samples
depths. These
bottles are non-metalic, free-flushing sampler recommended for general purpose
water sampling. These samplers can be individually or serially attached on a
hydrocable and activated by messenger, or placed in any kind of Multisampling
System (like G.O., Sea Bird, Falmouth Scientific and Small Multisampler Sys-
tem), and activated by remote or preprogrammed command. The Standard PVC
Niskin type Sampler is made of gray PVC (RAL 7011), spring closure made of
latex tubing with optional stainless steel spring closure, clamp bolts for attach-
ments on a cable and mounting blocks for Multisampling System
attachment. Delivery is made with lanyards for loading on both, cable and Multi
Sampling Systems. All metal parts are made out of special V4A-stainless
steel. Specially made V-LIP seal rings avoid leaking of the sampler. When the
sampler is lowered, the clamp at the lower end and the plug valves are in open
condition, so that, water can pass through the sampler. The sampler is held in
this position by the wire rope. When the messenger is dropped down the rope, it
strikes the release, shutting the valves closed by a locking device. The water
sample of the desired depth so trapped in the bottle can then be pulled up onto
the vessel in a closed condition. For collection water samples from different depths

simultaneously, a series of water samplers are suspended one above the other
from a wire rope and are lowered into the depths in the open state. In this case,
the messenger releases another messenger that was attached to the wire clamp
before lowering. The second messenger closes the next lower sampler releasing
a third messenger and so on.
2.2 Plankton pumps

Plankton pumps are integrating samplers that pump a continuous stream of wa-
ter to the surface and the phytoplankton can then be rapidly concentrated by
continuous filtration. Because the pumps can collect continuously as the tube is
lowered through the water column the samples are integrated from surface to
desired depth. This method has its disadvantages, however e.g., breaking up
colonies, breaking of large Chaetoceros setae, and breaking into pieces long
pinnate cells like Thalassiothrix spp.
2.3 Plankton nets:

Nets permit quantitative studies, since the mesh size will select the type of
phytoplankton collected. Sampling by nets is highly selective, depending on the
mesh size of the gauze, net towing speed, and the species present in the water.
Chaetoceros setae, for instance, may form a fine network inside the gauze and
very small single cells, which in other cases pass through the meshes, are re-
tained. On the other hand, net with very fine meshes (5 or 10 µm) often filter too
little water to provide an adequate diatom sample. The most useful mesh size for
collecting diatoms is 25 µm. Net hauls have the advantage of a simultaneous
collection and concentration of the plankton providing sufficient for species iden-
tification.
A typical plankton net usable in the surface layers is conical in shape and has
the following constituents (Fig 4). A net ring made up of stainless steel and
wrapped and sealed with polythene tubing is present anteriorly. To this, a non-
filtering portion made of a coarse khaki cloth is attached using button and hole
system. The filtering portion is made of monofilament nylon material as described
earlier and is followed by again a non-filtering portion of khaki cloth. To the latter,
a metal net bucket provided with a stop cock is tied with a strong twine.
The determination of the volume of water filtered through any plankton net is
essential for the estimation of the standing crop. The volume of water traversed
by the net is determined as an approximate value by the formula v = r2dΠ.
Where, V, volume of the water filtered by the net; r, radius at the mouth of the net;
d, distance through which the net is towed.
The water collected through the different water samplers is either centrifuged or
passed through fine mesh nylon or filter papers to separate the plankton present
in it. The smaller the sub sample the fewer number of rare species will be ob-

tained. On the other hand, there is no point in concentrating large quantities of a

sample rich in one or a few species. Concentration by settling, centrifugation
and filtration are the most used methods.
Plankton concentration is generally used to over come the damages caused, to

certain groups of phytoplankton especially the setoid diatoms and dinoflagel-
lates, by vacuum filtration, centrifugation. The simple plankton concentrator,
which is quite gentle in its action, consists of a stiff tube (1.2 cm dia: 10cm
height) of Perspex or PVC to the bottom of which a filter is attached. A filter paper
(Whatman No. 42) or membrane filters supported by monofilament nylon netting
which serves as the filter is glued at the bottom of the tube with the aid of ethyl-
ene dichloride. While using the tubes is dipped slowly into a beaker containing
the phytoplankton sample. Through the filter water flows slowly upward into the
tube and is removed with a large pipette. By forcing the tube downward, the rate
of flow through the filter can be increased.
Centrifugation:
With the help of an electrical centrifuge 5-20ml of water sample is centrifuged for
above 10-20 mins at 1500-2000 rpm. The supernatant water is removed by de-
canting. The plankton is precipitated by adding a few drops of 1% Potassium
aluminium sulphate or fixed weak neutralized formalin or Lugol’s solution.
3. Fixation and Preservation
If collected samples kept alive, they should be stored in an ice chest or refrigera-
tor and then only for a few hours. For long term analysis the collected plankton
should be preserved in fixatives and preservative. A very widely used fixative and
preservative for a variety of organisms including plankton is formalin. The com-
mercial formalin is obtained as a 40% (Saturation limit) formaldehyde dissolved
in water. The formalin has to be stored in inert glass or plastic containers and not
in metal containers as the formalin reacts with the latter. The commercial forma-
lin may also contain dissolved impurities such as iron and formic acid which
disintegrate the shells of some planktonic organisms. The acid content of com-
mercial formalin however, may be neutralized, by the addition of excess of cal-
cium carbonates. For preserving net and other phytoplankton sample, 2% neu-
tralized formaldehyde (i.e formalin) may be used.
3.1
atedLugol’s Solution:toItretain
phytoplankton is a good preservative
the flagella especially
and cilia. for flagellated
It consists and and
of 10g iodine cili-
20g potassium iodine dissolved in 200ml of distilled water and 20g of glacial
acetic acid. The solution can be made up a few days ahead and stored in a dark
bottle for convenience. Lugol’s solution is added in a ratio of 1 part to 100 parts
of the seawater sample. For about 250ml of water sample containing
nanophytoplankton about five drops of this preservative is quite sufficient.

3.2 Bottling and Labeling.

Bottling: Storage of phytoplankton especially diatoms in bottles made of soft
glass is preferred. Because surface water is usually under saturated with sili-
cate, storage in bottles of high quality glass like Pyrex which does not release
much silicate or plastic bottles may result in slow solution of delicate frustules
or spines of diatoms. This may happen in plastic bottles in one to a few years.
Further use of glass of very low quality for storage of phytoplankton may result in
precipitates. The bottles are closed by a leak proof cork. After the analysis of
the plankton, contents of the bottles and for permanent storage of plankton, or
wax coating is given around the cork of the bottle after the latter’s closure. This
would help avoiding the loss of formalin by evaporation in the long run.
Labeling: Proper labeling of the collected and bottled plankton samples is essen-
tial. All types of information regarding plankton collection should be written on
the labels so that, the plankton samples can be identified accurately. The label
should contain enough information about the sample collected in order to assure
proper identification of the sample. The label is written with a light coloured water
proof marker or wax pencil.
4. Preparation for light microscopy
Common diatoms can be identified by examination of raw (without acid cleaned)

material in a water mount. Common diatoms such as Chaetoceros spp and
Rhizosolenia spp are identified by their gross morphology and special struc-
tures like Chaetoceros setae and the shape of the Rhizosolenia its and process
(Tomas, 1997). However, this method is not effective for identifying the essential
morphological structures of other genera; for example, the areolation and pro-
cesses of Coscinodiscus and Thalassiosira and the striation and raphe structure
of Navicula and Pesudo-nitzschia (Tomos, 1997). Organic part and cell con-
tents, which obscure the image, have to be removed. Acid cleaning is one of the
methods used to separate diatoms frustules into single valves on which struc-
tures of diatoms are best seen .
4.1 Acid cleaning: Before acid cleaning salt particles associated with the dia-
toms in a test tube should be washed by rinsing and centrifuging in distilled
water. Then test tube with the sample is allowed to dry by removing water.
Adding some hydrochloric acid to the test tube dissolves the calcareous matter
and also loosens any diatoms that may be attached to the debris. After allowing
the test tube with the sample for one or two days, the test tube is well shaken
and the solid matter including the diatoms is allowed to settle at the bottom. The
acid is then decanted off and the sediment is washed by adding water and pour-
ing off again after allowing time for the solids to settle. Finally most of the water
is poured off and concentrated sulphuric acid is added slowly and carefully. Until

red fumes are no longer evolved, small crystals of potassium dichromate are then
added at intervals. The sulphuric-chromic acid mixture is then poured off and
water is added. Acid and dichromate treatment must be repeated until cleaning
is complete if the diatoms are not yet properly cleaned with water.
4.2 Specimen mounting : For, mounting, diatoms are put in a drop of distilled
water on a cover slip that has been smeared with a little Mayer’s egg albumen
which is prepared by mixing 50ml of white of egg with 50ml of glycerin and 1g of
sodium salicylate. After allowing the water to evaporate, the diatoms on the cov-
erslip are thoroughly dried by heating and then using any mounting media like
Canada balsam, Styrax, Hyrax or DPX mount can be done. After cooling the
specimen-mounted slide excess resin is trimmed off by a knife and the prepara-
tion is finally sealed with nail polish or wax.
Glycerin mounting and polyvinyl lactophenol mounting are other methods of mount-
ing diatoms. These are more convenient to mount the diatoms in slides directly
by embedding them in polyvinyl lactophenol.
Canada balsam is ideal for permanent mounts. For longer preservation, diatoms
can also be cleaned and stained with methylene blue and Bengal pink. Subse-
quently they are embedded in Canada balsam in microscopic slides and covered
with cover glasses.
5. Identification of species
Identification of diatoms in water samples is usually best done by using phase

contrast optics, which reveal especially well lightly silicified structures, like deli-
cate Chaetoceros setae, and also the organic chitan threads in Thalassiosiraceae.
(Tomas, 1997).
It is essential to know which side of the diatoms cell is viewed. Intact single cells
with a short pervalvar axis tend to lie up under the coverslip ( Coscinodiscus
radiatus and Pleurosigma sp). Diatoms like Corethron and Rhizosolenia with a
pervalvar axis longer than the cell diameter or the apical axis turn girdle side
upwards. Colony types like Chaetoceros, Fragilariopsis and Thalassiosira) are
normally seen in girdle view in a water mount. Diatoms like Thalassionema,
Asterionellopsis and Pseudo-nitzschia show either valve or girdle side. Cylindri-
cal and discoid diatoms are readily recognized by the general circular outlines in
valves view. When the cells are viewed properly the next step is to look for
special features like setae in Chaetoceraceae, shape of linking processes in
Skelotonema and in unpreserved material, organic threads from the valve in
Thallassiosiraceae.
Frustular elements cleaned of organic material may also be oriented in various

ways in a permanent mount (Tomas, 1997). Flattened valves with a low mantle
will usually be seen in valve view (Some Coscinodiscus spp., most Navicula
spp.), while valves with a high mantle and protuberances may appear in girdle
view (Eucampia and Rhizosolenia ). Lightly silicified bands shaped as those in
Rhizosolenia and Stephanopyxis often lie with girdle side up.
6. Bacillariophyceae (Diatoms)
Diatoms are extremely widespread and occur as the dominant organisms of
many diverse habitants. They are particularly conspicuous in both marine and
freshwater phytoplankton. They are characterized by four main features : 1). The
cell walls are silicified and show characteristic secondary structures. 2). The
photosynthetic pigments include chlorophylls a and c, together with the xantho-
phylls, fucoxanthin. 3). Food storage products include fats and chrysolaminarin.
4). The motile states possess a single pantonematic flagellum.
Although it has been possible to identify a number of structural developments in

the Xanthophyta and Chrysophyta paralleling those in the Chlorophyta, the range
of form has been much more restricted. The reduction in the range of vegetative
types is even more marked in the diatoms, in which only unicellular and colonial
forms can be recognized.
6.1 Structure of the diatom cell:
The diatom cell wall (frustule) consists of two parts, the one fitting over the main
part of the box (Fig. 5A). The outer part (the ‘lid’) is the epitheca and the inner part
is the hypotheca. Each half consists of the main surface, the valve, and the
overlapping connecting bands; the two bands constitute the girdle. The diatom
cell can therefore be viewed from two directions, the girdle view and the valve view
(Fig. 5 B,C). The axis between the middle of the two valves is the long axis (most
diatoms are broader than long) and is called the pervalvar axis, and the one at
right angles to this is the valvar plane.
All diatom frustules have silicified walls, although in Phaeodactylum tricornutum

only one valve is silicified. The walls contain hydrated silica and, since no other
element canifreplace
silicon, and silicon, growth
other elements of diatoms
are present show an
in adequate absolute
amounts, requirement
growth for
is propor-
tional to the silicon concentration. The silicified wall also contains an organic
component, which has been called ‘pectin’ although there is no definitive chemi-
cal evidence for this statement.

The diatom cell wall is not of uniform thickness, and periodic arrangements of
thicker and thinner areas produce a complex series of ‘markings’ on the cell
surface . The general arrangements and symmetry of these ‘markings’ are im-
portant criteria for the division of the Bacillariophyceae into the two orders, the
Centrales and the Pennales. In the former they are arranged with reference to a
central point (Fig. 5E)(this basic picture may be altered when the cell is angular).
In the later, however, the main structural element on the cell surface is a spine,
and the finer secondary structures are arranged as lateral branches (Fig. 5B).
Thus, the value of pinnate forms has a narrow axial area thickened at each end
(polar module), and a dilated central area usually having a median thickening
(central nodule). The axial region sometimes has a slit (raphe)which runs from
one polar nodule to the other; whereas in other species a lighter region of the
axial area gives the superficial appearance of a raphe, and this is called a
pseudoraphe.
The secondary structures of the diatom cell wall represent the fine sculpturing
which occur over much of the value surface. These structures are extremely
variable; their nomenclature is sometimes equally variable and much confusion
has resulted. Examination in the electron microscope has confirmed four basic
kinds of secondary structures : 1. punctae are small perforations of the valve
surface and are frequently arranged in regular lines, or striae, 2. aerolae are
larger, depressed box-like structures , 3.Canaliculi are narrow tubular channels
through the valve wall and 4. costae are ribs formed by the heavy deposition of
silica.
From the
these fourvoluminous
basic kinds literature on the two
of structures, detailed
mainarrangements
wall types areand elaborations
recognized: of
first,
the laminar walls consisting of a single silicified layer with a great variety of
patterns on it (Fig. 5 F), and second , the locular walls consisting of two parallel
layers with a complex reticulum of cross walls between them (Fig.5 G). Ex-
amples of species with lamina walls include Synedra fulgens, Fragilaria construens
and Didymosphenia geminata. Species in which the wall is of the locular type
include Triceratium favus, Coscinodiscus asteromphalus (in both of these spe-
cies the locular nature of the wall can be seen with the light microscope) and
Stephanopyxis palmeriana.
6.2 Gross vegetative structure:
The general shape and superficial appearance of the diatom cell is variable and
many of these gross variations are useful for a preliminary identification of a given
alga. There are six main types of morphological elaborations which appear to be
correlated with the planktonic habit: 1). the flat discoid shape of many centric
diatoms (e.g. Cyclotella comta, Fig.5 D, E); 2). the needle shape of species such

as Rhizosolenia and Synedra; 3). the long, sometimes coiled filaments of some
species of Melosira; 4). the development of elongated bristles (Stephanodiscus)
or horns (Chaetoceros) from the edge of the valves; 5). the stellate colonies of
Asterionella and Tabellaria; and 6). the frequent production of extensive mucilagi-
nous envelopes (e.g. Cyclotella planctonica).
Although, most diatoms are unicellular, a number of colonial species is also

known. There appears to be three basic kinds of colony formation. Firstly, cells
are sometimes joined together by a localized production of mucilage to form
stellate colonies of Asterionella of the filamentous forms of Melosira. Secondly,
some colonies consists of many cells embedded in a common mucilaginous
envelope, and in some genera the envelope has a tubular structure (e.g. some
species of Navicula, Cymbella and Nitzschia) The third type of colony consists
of cells joined by special outgrowth such as the spines of Chaetoceros.
6.3 Cell Division:
Diatoms usually divide at night and the plane of division is always at right angles
to the longitudinal axis, that is, parallel to the valve surfaces. The first indication
that cell division is imminent is that the cell increases in size and the two halves
separate slightly. Mitotic division of the nucleus is followed by fission of the pro-
toplast in a plane parallel to the valve faces. New siliceous valves are then
deposited on the two fresh protoplasmic surfaces. As the connecting bands
develop, the valves of the parent cell separate and the new silica valve becomes
the hypotheca of each daughter cell.
The daughter cell having the original hypotheca of the parent (it is now the epitheca
of the daughter cell) is smaller than the parent cell. Thus, in a population of
diatoms there is normally a progressive decrease in the average cell size (those
species which do not show a progressive decrease in size are usually weakly
silicified and the constancy of size is probably related to the plasticity of the cell
wall).
6.4 Classification of Diatoms:
As pointed out in the beginning the diatoms can be divided into two orders, the
Centrales and the Pennales; this classification being based largely on the sym-
metry and orientation of the secondary structures on the valve surface. 1. Cen-
tric diatoms are non-motile, whereas many species of the pinnate forms exhibit a
gliding movement which is, in some way, dependent on the presence of a raphe.
10

2. Sexual reproduction of the Centrales is oogamous whereas that of the pinnate

species is generally isogamous. 3. although species of the two groups some-
times inhabit the same regions, Centrales are more commonly planktonic and
marine, whereas occurrence of Pennales are less in marine water.
The more detailed classification of diatoms depends almost entirely on the struc-
ture of the siliceous skeleton. The Centrales are divided into three major groups
on the basis of cell shape and are the presence or absence of particular pro-
cesses. Genera such as Coscinodiscus, Cyclotella and Melosira are disc-shaped
with no processes, whereas the valve surfaces of genera such as Biddulphia and
Chaetoceros have various horns. A third group containing genera such as
Rhizosolenia and Corethron also have a complex girdle structure.
The classification of the pinnate diatoms is based largely on extent of develop-

ment of the raphe. Tabellaria and Asterionella are examples of the group of
diatoms which only posses a pseudoraphe. Amongst the other forms it is pos-
sible to identify an increasing tendency for the development of a raphe; the valves
of Eunotia, for example, show the beginnings of raphe development. Achnanthes
and Cocconeis have a raphe on one value only, but most genera (e.g. Navicula,
Bacillaria and Nitzschia) have a raphe on each valve.
Order: Centrales:
Family: Coscinodisceae
Skeletonema costatum (Greville) Cleve (Fig. 6): Valves small, lens shaped with
rounded ends and form long and slender chains with the help of marginal spines;
space between cells larger than cell; dia., 12-15 µm.
Cyclotella meneghiniana Kutzing (Fig. 7): Cell disc shaped with a number of
regularly arranged striations which do not reach center; dia., 17-24 µm.
Cyclotella striata (Kutzing) Grunow (Fig. 8): Cells resemble C. meneghiniana;

with evenly striated border; central area of the cell coarsely punctuate; dia., 15-
30 µm.
Coscinodiscus eccentricus Ehrenberg (Fig. 9): Cell disc shaped; hexagonal
markings seen; areolae of same size (6 in 10 µm) and arranged in tangential
series; margin striated and 18-20 striae in 10µm; dia., 340-104 µm.
11

Family: Actinodisceae
Asterompalus flabellatus (Brebisson) Greville (Fig. 11): Cell slightly convex; valves
slightly ovate; middle sector lines unbranched; 7-8 slightly curved hyaline rays,
of which one is narrower; length 37-63 µm and breadth 32-55 µm.
A. wyvillei Castracane (Fig. 12): Valves rounded with 15 straight hyaline rays, of
which, one is narrower; sector lines branched; dia., 71-74 µm.flattened at ends;
numerous disc-shaped chromatophores; dia., 3-16 µm and length 10-91 µm.
Family: Soleniae
Lauderia annulata Cleve (Fig 13a, b): Cells from straight chain; cells cylindrical
with convex valves; valves with a depression in middle and raised at margin;
adjacent cells touch raised portions; valves with numerous spines and varying
length; dia., 53-83 µm.
Schroederella delicatula (Peragallo) Pavillard (Fig. 14):

Cells cylindrical and form chains; valves with depressions in middle; valve ends
with a crown of spines; a spine like pore canal present at center of each valve;
dia., 14-41µm.
Leptocylindrus danicus Cleve (Fig 15): Cells cylindrical and form chains; valves
flattened at ends; numerous disc-shaped chromatophores; dia., 21-24µm
Rhizosolenia cylindrus Cleve (Fig. 16): Cells cylindrical and valves with fairly
truncated ends; presence of large and bent spines; cell wall hyaline; dia., 21-24
µm.
R. crassispina Schroeder (Fig 17): Cell cylindrical and valves possess truncated
ends; apical processes broadened at base and hair-like afterwards; numerous
disc-shaped chromatophores; dia., 41-54 µm; length 145-278 µm.
Family: Chaetocereae
Chaetoceros lorenzianus Grunow (Fig. 18): Cells from straight chains; apertures
of varying sizes; terminal setae thicker, somewhat shorter than other setae and
run parallel to chain axis; inner setae longer and interlocking; setae four sided;
12

length of cell, 18-61 µm.
C. didymus Ehrenberg (Fig 19): Cells from straight chains; a charasteristic semi-
circular knob like structure present in middle of each valve; distict interlocking of
setae; two plate-like chromatophores present; length of cell, 22-40 µm.
C. diversus Cleve (Fig. 20): Cells form compact and short chains; apertures very
small; setae of some cells thicker, tubular and spinous; other inner setae and
terminal ones hair-like; length of cell, 5-9 µm.
Family: Biddulphieae
Eucampia zoodiacus Ehrenberg (Fig 21): Cells flat united to form spirally twisted
chains with characteristic blunt processes; valves concave in middle so that a
wide aperture between two cells; intercalary bands faint; length of cell, 42-61µm.
Ditylum brightwellii (West) Grunow (Fig 22a, b): Cells prism shaped with three
cornered valver plane; valve margin wavy; a circlet of short spines on valves ends
and a long hollow spine at center of the valve; side of valve measures 42-144 µm.
Biddulphia sinensis Greville (Fig 23): Cells forming short chains, cylindrical and
square to rectangular in girdle view; however, ovate to lanceolate in valvar plane;
presence of two thin blunt horns at corners of valve and two long and thin spines
nearer to horns characterize this species; length of cell 82-215 µm.
B. mobiliensis Bailey (Fig 24): Cell resembles B. sinensis to some extent; cells
moderately squarish with slender horns at corners of valves; length of cell 24-81
µm.
Order: Pinnales
Family: Fragilariodeae
Fragilaria oceanica Cleve (Fig 25): Cells rectangular in girdle view and form com-
pact ribbon-like chain; valves broadly lanceolate with rounded ends; pseudoraphe
narrow linear; transapiacl striae 14 in 10 µm; length of cell, 11-32 µm; breadth 6
µm.
13

Thalassionema nitzschioides Grunow (Fig. 26): Cells form zig-zag chains and
linear-rectangular in girdle view; cells rest at protoplasmic cushions found at
junctions; linear-lanceolate in valve view; marginal striae 12 in 10 µm; length 20-
66 µm; breadth 3m.
Asterionella japonica Cleve (Fig. 27): Cells form spiral colonies; frustules linear,
narrow with parallel sides and knob-like at base; striae not clearly seen; length
43-106 µm; breadth 7-11 µm.
Family: Naviculoideae
Gyrosigma balticum (Ehrenberg) Rabenhorst (Fig 28): Valves linear with curved
and truncated ends; raphe excentric and central area small, oblique; transverse
and longitudinal striae equidistant, 11-12 in 10 µm; length of cell 290-338 µm;
breadth 28-30 µm.
Pleurosigma galapagense Cleve (Fig. 29): Valves very slightly sigmoid; ends
blunt; raphe somewhat sigmoid; transverse striae 18 in 10 µm and oblique striae
15 in 10 µm.
Diploneis weissflogii (A. Schidt) Cleve (Fig. 30): Valves broad and strongly con-
stricted at center; ends fairly rounded; central nodule with horns; transeverse
costae 9 in 10 µm; length 28-58 µm; breadth 10-25 µm (away from constriction)
and 6-15 µm (at constriction).
Navicula longa (Gregory) Ralfs (Fig. 31): Valves long rhombic with fairly pointed
ends; axial area narrow; central area small; striae 9-11 in 10 µm; length 52-56
µm; breadth 10µm.
N. sigma (Kutzing) W. Smith (Fig. 32): Valve linear; somewhat sigmoid in girdle
view and straight in valve view; bulge at center and gradually diminishing in size
towards end; keel punctae 5-6 in 10µm; length, 280-310 µm; 10-11 µm.
N. closterium (Ehrenberg) W. Smith (Fig. 33): Valve spindle shaped in middle;

extremities beak like and slightly curved in opposite directions; striae not visible;
14

length 30-160 µm; breadth 3-7 µm.
7. Pyrrophyceae (Dinoflagellates)
The motile unicellular forms of the dinoflagellates are sometimes important con-
stituents of phytoplankton populations and are only next to diatoms as far as the
phytoplankton biomass is concerned. Although, motile unicells form the bulk of
the class a number of non-motile and multi-cellular types also occur. The di-
noflagellates are unicellular, single or pseudocolonial and show wide variations in
morphology. The size of these organisms ranges from 0.001 to 2 mm; however,
most of the species have a size below 0.2 mm.
The presence of 2 flagella , one encircling the body is located in the transverse
furrow otherwise known as ‘girdle’ or ‘cingulum’ and the other located in the
longitudinal furrow or ‘sulcus’ trailing behind is the characteristic feature of most
of the planktonic dinoflagellates (Fig 34). The body of the cell is covered by an
envelope (cell wall) made of cellulose, pellicle, valves or plates. The plates when
present form the theca and are usually arranged in specific series of taxonomic
importance. The arrangements of plates and plate formulas of certain genera of
dinoflagellates are shown in Figs 34-37. Based on certain characteristics, the
theca may be of five types a) no distinct plates in theca (e.g. Amphidinium,
Noctiluca, Oxyrrhis); b) theca of thin polygonal plates (e.g. Gymnodinium,
Gyrodinium); c) theca of thik plates each with distingshable shape (e.g. Gon-
yaulax, Glenodinium); d) theca of thick plates covered by reticulations (e.g.
Peridinium, Ceratium ) and e) theca of two valves (e.g. Prorocentrum). The highly
ornamental forms are provided with striations, ridges, horns, spines, tentacles
(lists).
The cell has normally a large and fingerprint-like nucleus and two vacuoles.
Among the latter, the larger one is known as ‘pusule’ which is said to help in
phagocytosis. The other vacuole is small and its function is not known. The
chromatophores may or may not be present; if present, they are few, small and
conspicuously coloured (green , yellow, brown or orange). Pigments such as
cholorophyll a. c, β carotene, fucoxanthin, dinoxanthin, peridinin and
diadinoxanthin. The other components of the cell are oil globules, ocelli, eye
spots (stigmata), nematocytes, trichocysts and internal siliceous star-shaped or
net work structure.
Majority of dinoflagellates are autotrophic and a few are holozoic, saprophytic or

phagotrophic. In the autotrophic dinoflagellates, the products of the photosynthe-
sis are starch and lipids. The class Pyrrophyceae comprises two groups, viz.
Desmophyceae and Dinophyceae
Group: Desmophyceae: It is much smaller group and has only two genera, viz.
Prorocentrum and Exuviaella. Both the flagella in these organisms arise from the
15

anterior end of the cell and hence the cingulum and sulcus are absent. The cell
wall is not composed of separate plates unlike in dinophyceans but has only a
longitudinal suture which divides the cell into two valves. The reproduction is by
longitudinal division while the cell is motile. During division, the suture dividing
the two valves separates, so that after fission, each daughter cell retains one
valve from parent.
Order: Prorocentrales
Genus: Prorocentrum : Cells of species generally elongated, oval and armoured;

composed of two opposing longitudinal valves connected by suture and interca-
lary bands; chromatophores small and yellowish-brown.
P. micans Ehrenberg (Fig 38): Cells variously shaped from oval to almost circular
and compressed laterally; apical teeth and protrusions may or may not be present;
however, apical platelet present; valves with poroids (pits) pores, reticulations,
spines or other surface markings; length 34-52 µm, breadth 15-18 µm.
P. rostratum Stein, (Fig 39): Body compressed laterally with a blunt apex; a
finger or rostratum like process present; valves narrow and pointed posteriorly
with an apical tooth on each valve; length, 98 µm; bredth, 18µm.
Genus: Exuviaella Cienkowski: Cells of species oval or subspherical; absence of

anterior projection; two lateral large and brown chromatophores; nucleus poste-
rior.
E. compress Barley and Ostenfeld (Fig 40): Cell oval and not compressed ; each
valve with a smooth tooth anteriorly; two plate-like yellow chromatophores; length
20-25 µm; breadth 18-24 µm.
Group: Dinophyceae: This group differs from Desmophyceae in having a cingu-

lum which divides the cell into an anterior epicone and a posterior hypocone. The
girdle houses a hand-like transverse flagellum which arises through a pore and
causes the cell to spin to some extent on its axis. The sulcus is also present in
this group and it runs from the posterior end of cell part way forwards. The longi-
tudinal flagellum which arises from a pore in the sulcus, runs back and usually
beyond the cell trailing behind in the water.
Order: Dinophysiales
Genus: Dinophysis Ehreneberg: Cells of this species compressed laterally;
epitheca small or rudimentary with oblique set girdle tentacles (lists); upper list
funnel shaped projecting beyond epitheca and strengthened by radial ribs; left
sulcal list not well developed.
16

D. caudata var. pedunculata Schmidt (Fig 41): Hypotheca with distinctive protu-
berances but without posterior sail; length 65-115 µm.
Genus: Phalacroma Stein: Body not much compressed; epitheca is elevated

above transverse lists which are uniformly developed and not conspicuous; chro-
matophores absent.
P. argus Stein (Fig. 42): Body laterally ovate and wider behind girdle; epitheca
and hypotheca rounded; girdle lists ribbed; right sulcal list concave, length 72
µm.
P. cuneus Schutt (Fig 43): Body cuneate laterally; epitheca low and broadly
rounded; hypotheca posteriorly narrowly rounded to subacute; margin of left sul-
cal list slightly sigmoid. 71 µm.
Order: Peridiniales
Noctiluca Suriray: Body kidney or sphere shaped; no girdle and hence epicone
and hypocone not distinct; deep sulcus; short longitudinal flagellum and trans-
verse flagellum represented by a mobile membrane or tooth; well developed ten-
tacle at the posterior end of sulcus.
N. miliaris Suriray (Fig 44): Only one species known under the genus Noctiluca;
characters similar to that of genus; dia., 200-2000 µm.
Genus: Protoperidinium Ehrenberg: Top shaped; presence or absence of antapical

horns; ventral plate (1st apical) of epitheca may be 4, 5 or 6 sided; 2nd intercalary
plate 4, 5 or 6 sided; apical horn affixed or tapering; plate formula 4’, 3a, 7”, 5”’
and 2””.
P. ovatum (Pouchet) Schutt (Fig. 45): Cell slightly compressed; epitheca low,
dome-shaped and tapering sharply into small apical horn; hypotheca also low,
dome-shaped and with two small antapical spines; sulcus subantapical or reach-
ing antapex; five-sided 1st apical and four-sided or five-sided 2nd intercalary; 68-70
µm by 45-60 µm.
P. crassipes Kofoid (Fig 46): Body low and stout; slightly compressed dorsoven-
trally; ventral side rather concave but dorsal convex; apical horn conical abtuse;
antapicals short, stout and close together; antapicals end in a blunt, semi-trun-
cated projection with 2-3 points; right antapical longer than left; 70-85 by 60-72
µm.
Genus: Gonyaulax Diesing: Girdle equatorial and left handed; sulcus indented
and occupies whole venteral area; plate formula 3’ 6”, 6”’, 1p and 1””; chromato-
phores yellow to dark brown.
17

G. polygramma Stein (Fig 47): Body elongated, spindle shaped and swollen mid-
body; both ends projecting into two long horns; length, 135 µm; breadth, 40 µm.
Genus: Ceratium Schrank: Cell dorso-ventrally flattened; girdle left-handed with

lists; epitheca with long hollow apical horn and hypotheca with two hollow
antapicals; chain formation in some species and heteromorphic; chromatophores
numerous yellow; plate formula 4’,5”, 5”’ and 2””.
C. tripos var. atlanticum Ostenfeld (Fig 48): Some what large species; body as
broad as long; epitheca’s left contour slightly convex and right contour strongly
convex; horns strong; apical broader below, larger than the others; antapicals
diverging from one another; antapicals of equal size, bent; 115-130 µm by 20-25
µm.
C. pulchellum B. Schroder (Fig. 49): Robust species; eiptheca with steep left
and very convex right side; apical horn long and strong, slightly wider in middle;
base of hypotheca strongly convex; antapicals short; less stronger than apical;
left curved slightly diverging or parallel to apical; right equal in length or shorter
than left, 130 –140 by 60-65 µm.
C. breve (Ostenfeld and Schmidt) Schroder (Fig. 50): Body with short horns;
epitheca’s right contour strongly convex and left contour steep; hypotheca with
evenly convex base; anatapicals very strong; slightly parallel with apical horn;
75-80 by 60 µm.
C. karstenii Pavillard (Fig. 51): Strong body; right contour of the epitheca convex;
apical horn slightly bent at base; antapical slender; right antapical longer than
left antapical and bent distally towards apical horn; left anatapical at times curved;
90-95 by 25-32 µm.
C. contortum (Gourret) Cleve (Fig. 52): Cell resembles C. karstenii to some ex-
tent; epitheca oblique on right, right contour strongly convex; apical horn twisted,
S shaped; horns slender; antapicals unequal, right longer than left and twisted
towards apical horn; 90-94 by 25-30 µm
8. Micrometry
By micrometry, while viewing through a microscope, the length, breadth and

other details of an organism are measured. The size determination of the phy-
toplankton forms an important aspect, especially, in preparing the report on the
18

occurrence of new species or taxonomic studies for publication.
In micrometry a occular micrometer (graticule) plays an important role. The

ocular micrometer is a circular glass piece which contains a scale of lines which
are engraved or photographically reproduced (Fig. 53). This scale is of 10 mm in
length divided into ten equal divisions. Thus on the scale of the ocular microme-
ter one hundred divisions of 100 µm each.
Calibration: Ocular micrometer is mounted on the diaphragm inside the eyepiece

of the chosen microscope at the focal of the eyelens. On the diaphragm inside
the eyepiece, at which point, the image from the object is also focused, so that,
the two can be viewed simultaneously. Now, not only the object in focus, but
superimposed on the object, the series of lines of the graticule is equally visible.
For the calibration of the graticule, a stage micrometer which is a microscopic
slide of 7.5 x 2.5 cm, on which, has been engraved a scale of 1mm long, divided
into 100 divisions of 10 µm (0.01mm) each (Fig 54). While calibrating, the stage
micrometer is first placed on the stage of the microscope. Then it is focused and
aligned with the ocular micrometer scale. The stage micrometer is then moved
carefully until its zero line is in exact coincidence with that of the ocular meter, in
order to find out how many divisions on the ocular micrometer scale correspond
with a certain number of divisions on the stage micrometer scale. From this, the
value (in µm) of one division of the ocular micrometer under the chosen micro-
scope with fixed objective and eyepiece powers is calculated.
If 30 divisions of the ocular micrometer correspond with 10 divisions of the stage
micrometer
words, thesescale, then these
30 divisions 30 100
occupy divisions are equivalent
µm space of the stagetomicrometer
100 µm. In(asother
one
division occupies 10 µm of the space in the stage micrometer and the total length
of the scale is 1000 µm –1 mm). Thus one ocular micrometer division is equal to
100/30 = 3.3 µm. This calibrated value of the ocular micrometer is of a particular
objective and eyepiece of a microscope.
If size determination of a object is to be done in different objective or eye lens, the

ocular micrometer scale is calibrated for all the combinations of the different
objectives and eyepieces, all value may be tabulated and can be used whenever
it is required.
The size of an individual phytoplankton cell of a species may be determined
using the calibrated ocular micrometer and micrometer as follows. For size
determination, on the stage of the microscope, the specimen for which the size
is to be determined, is now placed instead of the stage micrometer. If the diam-
eter of Cyclotella cell is to be determined the zero of the ocular micrometer is
focused against the edge of the cell and the number of division of the ocular
micrometer occupy the diameter of the cell is found out. Number of calibrated
ocular micrometer divisions multiplied by the corresponding calibrated value would
19

give a diameter of the said cell. For examples, if the graticule divisions are 20
then the diameter of the cell is 20 x 3.3 µm= 66µm.
9. Measurement of Biomass
Assessment of standing crop of phytoplankton in different periods is essential for

any environment as the level of biomass indicates directly or indirectly its fertility
and fishery resources. The biomass may be estimated in various ways.
9.1 Chlorophyll measurements:

This method is chiefly employed to estimate phytoplankton biomass. The most
useful chemical method for determining the total quantity of phytoplankton in
seawater
son et al., is to estimate
1984). This is the amount
a rapid of chlorophyll
method usually
for determining as chlorophyll
phytoplankton a (Par-
density in
a sample involves the extraction and measurement of chlorophyll concentrations.
The amount of pigments as chlorophyll a, b, c and phaeophytin is considered as
a measure of phytoplankton biomass.
After the collection of the sample, it is filtered through a Millipore (Pore size 0.45
µm) or glass fiber (1 µm mesh) filter, and is pumped to dryness (Fig. 55). All
steps should be carried out in the dark to avoid pigment breakdown. The filter
containing the sample is placed in 90% acetone in a plastic vials covered by
aluminium foil and shaken vigorously and gently ground with a homogeniser to
ensure dissolving of the filter (Millipore) before storage in the refrigerator for 20-24
hr. Some recommend, addition of 1 ml of a 1% Magnesium carbonate suspen-
sion on to the filter paper to form a thin bed, which will serve as a precaution
against the development of any acidity and subsequent degradation of pigment in
the extract.
After 20-24 hrs of extraction in the cold and dark, the plastic vial containing filter
paper is brought to room temperature and the volume brought up to the original
level by addition of 90% acetone in a graduated centrifuge tube. The solution is
centrifuged for about 20 minutes at 5000 rpm and the supernatant solution is
considered for the determination of optical density, or transmission percentage
which is mainly with the aid of a flourometer (Parson et al., 1984).
9.2 Cell counts:

The direct estimate of phytoplankton cell density as measures of standing crop
are usually made by this method. The enumeration of nano and net phytoplank-
ton is done by various counting chambers, however, the most commonly used
counting chamber is Sedgwick Rafter cell. The counting cell is filled with the
plankton sample and placed on the mechanical stage of the microscope. Then
the counting cell is left for about half-an-hour for proper sedimentation. The or-
ganisms are then counted from one corner of the counting cell to the other. The
20

Sedgwick Rafter is moved horizontally along the first row of squares and the
organisms in each square of the row are thus counted. The rafter is moved to the
second row and organisms in each square here are counted. (Few transects
may also be counted instead of all the squares. The total number of cells is then
computed by multiplying the number of individuals counted in transects with the
ratio of the whole chamber area to the area of the counted transects.) Replica-
tion of counts of one ml samples is recommended for the statistical treatments.
After counting, the sample is to be returned to the jar containing the whole sample.
The average values are taken into account for calculation. The total number of
phytoplankton present in a liter of water
sample can be calculated using the formula:
N= n x v X 1000
V
Where, N: total number of phytoplankton cells per liter of water filtered;
n: average number of phytoplankton cells in 1 ml of plankton sample.
v: volume of plankton concentrate (ml)
V: volume of total water filtered ( l ).
9.3 Cell counts by drop count method:

The common glass slide mounted with a drop of concentrated phytoplankton
sample in glycerol and covered with cover slip is placed under the microscope
provided with a mechanical stage. The plankton are then counted from the mi-
croscopic field of the left top corner of the slide. Then slide is moved horizontally
along the right side and plankton in each microscopic field are thus counted.
Whenthe
using firstmechanical
microscopic field row
device is finished
of the the
stage. In next
this consecutive
way row ispresent
all the plankton adjusted
in
entire microscopic field are counted. If it is difficult to count all the microscopic
fields, then few microscopic field may be counted.
The total number of cells then calculated by summing the plankton numbers of
all the microscopic fields. If this total number is of one drop of the concentrated
phytoplankton, then total number is in 1 ml of the phytoplankton concentration
has to be calculated. Before calculating this, number of drops which form 1 ml
has to be counted by adding the drops of water into the graduated centrifuge
tube. If one drop of concentrated phytoplankton contains some known number
then cells present in 1 ml can be calculated.
For example if 16 drops forms 1ml, and suppose 50 planktons are counted in one
drop. Then the plankton in 1ml are calculated as follows.
Plankton in 1 ml concentrate = 16 x 5
Plankton per litre = 800 x 1000 ml
= 800000 cells.
21

10. Measurement of Productivity
The basic concept of primary productivity can be summed up in the equation for
carbon fixation by autotropic aerobic algae.
Light
6CO2 + 6H2O → C6H12O6 + 6O2
From this equation it is apparent that a number of methods can be employed to

measure the rate of photosynthesis i.e carbon dioxide uptake, oxygen produc-
tion or the formation of carbon compounds.
According to Litter (1973) and Hoffman and Dawes (1980), of the different meth-
ods: such as labeled carbon (C 14) uptake, oxygen release (oxygen probe), pH
(CO2 uptake), light and dark bottle method (Winkler titration method); the most-
effective method for measuring productivity was C14 uptake.
C14 Method:
Labeled carbon is probably the most extensively used procedure for oceanic
studies of productivity. This method essentially advantageous because it is rela-
tively safe, weak β-emission (0.15 Mev) as well as its long half life (4700yr), so
that storage offers no major problems.
Procedure: The activity per ml of the working solution needed for the different
productivity experiments depends on the production rates expected, duration of
incubation, bottle size, etc. Invariably, 0.2-1 ml of the working solution is used
per bottle containing water sample.
Water samples for which production rates are to be determined are first col-
lected from the specified depths and are transferred to the light and dark bottles
kept in a dark box. Then, a known dose of the working solution is injected rapidly
into the bottles with the help of a graduated hypodermic syringe having a needle
not shorter than 5 cm. The bottles are then incubated for a known period by
suspending them at the respective depths from where the water samples were
taken for experimentation. After the incubation is over, the experimental bottles
are removed from the depths and are stored in a light-free case until the filtration
of water samples is begun. Filtration may be done either on board the ship or in
the laboratory. Aliquots of water samples for filtration are rapidly transferred into a
suitable vacuum filtration apparatus on to a No. 2 membrane filter or Millipore
filter of about 0.5 µ porocity. The vacuum should be applied at about 0.5 atm
which will help avoiding damaging of fragile phytoplankton cells. The filtration
22

should be done in a semidarkend area.
The filters, after their removal from the filtration apparatus are placed onto planchets
which are then kept in a desiccator containing silica gel. Filters obtained from
light and dark bottles are then subjected to counting in a Geiger-Muller counter.
Under constant light source, the rate of production is obtained in mgC/m3/hr by
the following formula:
Where, cpm, counts per minute; the total CO2 is assumed to be constant in
oceanic waters and the value is 90 mg CO 2/l; 1.06, a correction factor for the
isotope discrimination effect and to be used as the 14C incorporation will be slow
compared to 12C; 1000 to convert the value for m3; 12/4 to get the value of C from
CO2 as the molecular weight of CO2, 44 and the atomic weight of C, 12.
Net activity (cpm of light bottle-cpm of dark bottle) X Total CO2 X 1.06 X 1000 X 12/44
Rate of production =
cpm added Hrs of incubation
(photosynthesis)
(mgC/m3/hr)
23

11. Bibliography:
Hoffman, W. E. and C. J. Dawes., 1980. Photosynthetic rates and primary pro-

duction by two Florida benthic red algal species from a salt marsh and a man-
grove community. Bull. Mar. Sci,. 30: 358-364
Littler, M. M. 1973. The productivity of Hawaiin fringing-reef crustose corallinaceae

and an experimental evaluation of production methodology. Limnol. Oceanogr.
18: 946-952.
Parsons, T. R., Y. Maita and C. M. Lalli., 1984. A manual of chemical and bio-
logical methods for seawater analysis. Pergamon Press, New York.
Sournia, A.(ed). 1978. Phytoplankton manual. In “Monographs on Oceanographic

Methodology 6,” pp 337. UNESCO, Paris.
.
Tomas C. R., 1997. Identifying marine phytoplankton. Academic press, Harcourt
Brace and Company, Toronto. Pp. 858
24

Rope
Rope
Lid
Lid
Bottle
Bottle
Frame
Frame
Thick metalbase
Thick metal base
Fig.
Fig. 11
Fig.22
Fig.
25

Fig. 3
26

Fig. 4
Fig. 5
A, B, C, Pinnularia viridis; A, transverse section, B, valve-view, C, girdle view, D, E, Cyclotella

comta; D, girdle view; E, valve view, F, laminar wall, G, locular wall c.n. central nodule, e,
epitheca; g, girdle; h, hypotheca; p.a, pervalvar axis, p.n, polar nodule; r, raphe, v, valve; v.p.
valvar plane.
27

A, B, C, Pinnularia viridis; A, transverse section, B, valve-view, C, girdle view, D, E, Cyclotella

comta; D, girdle view; E, valve view, F, laminar wall, G, locular wall c.n. central nodule, e,
epitheca; g, girdle; h, hypotheca; p.a, pervalvar axis, p.n, polar nodule; r, raphe, v, valve; v.p.
valvar plane.
28

Fig. 7 Fig. 8 Fig. 9

Fig. 6
Fig. 10a Fig. 10b

Fig. 11
Fig. 13a Fig. 13b

Fig. 12
Fig. 16
Fig. 15
Fig. 14
29

Fig. 17 Fig. 19
Fig. 18
Fig. 20 Fig. 21
Fig. 22a
Fig. 22b
Fig. 23 Fig. 24
30

Fig. 25
Fig. 26
Fig. 28 Fig. 29
Fig. 27
Fig. 30
Fig. 31
Fig. 33 Fig. 34
Apica l
Anterior intercalarie s
EPITHECA
Precingulars
Cingulars
Postcingulars
Fig. 32
Posterior intercalaries
HYPOTHECA
Antapi cals
Sulcals
31

Apical plate bar
Suture
Intercalary band
Anter ior view

Fig. 35
Ventral
Nucleus
Sulcus
Transverse flagellum
Flagellar pore
Chloroplast
Sulcus
Longitudinal flagellum
Fig. 36
Lateral
Epicone
Cingulum
Hypocone
Fig. 37
32

Fig. 39
Fig. 40
Fig. 38
Fig. 41
Fig. 42
Fig. 43
Fig. 44
Fig. 45 Fig. 46
33

Fig. 47
Fig. 48
Fig. 49 Fig. 50
Fig. 52
Fig. 51
34

Fig. 53
Fig. 54
Graduated
Upper part
Millipore filter Holding device
Suction bottle
To vaccume pump
Fig. 55
35

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5/21/2018 Phytopla nkton Ma nua l - slide pdf.

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Phytoplankton Identification Manual

 National Institute of Oceanography

First Edition : March 2004

National Institute of Oceanography,

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 2/40

Since its inception in 1966 the National Institute of Oceanography is involved

The project “Survey and Inventerisation of Coastal Biodiversity (West coast)

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 3/40

Marine phytoplankton which constitutes diatoms, dinoflagellates, blue-green

Coastal waters around India contain diverse groups of phytoplankton. The

The organization of this manual is made with a broad outline to accommo-

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 4/40

3. Fixation and Preservation

3.1 Lugol’s solution

4. Preparation for light microscopy

10. Measurement of productivity

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 5/40

Phytoplankton (‘phyto’ = plant; ‘planktos’ = made to wander) are single celled

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 6/40

through laboratory analysis.

There are three methods of sampling of phytoplankton.

2.1 Bottle Samplers:

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 7/40

2.1.1 Meyers water sampler (Fig 1) : It consists of an ordinary glass or perhaps

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 8/40

2.2 Plankton pumps

2.3 Plankton nets:

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 9/40

tained. On the other hand, there is no point in concentrating large quantities of a

Plankton concentration is generally used to over come the damages caused, to

3. Fixation and Preservation

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 10/40

3.2 Bottling and Labeling.

Common diatoms can be identified by examination of raw (without acid cleaned)

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 11/40

Identification of diatoms in water samples is usually best done by using phase

Frustular elements cleaned of organic material may also be oriented in various

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 12/40

Although it has been possible to identify a number of structural developments in

6.1 Structure of the diatom cell:

All diatom frustules have silicified walls, although in Phaeodactylum tricornutum

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 13/40

6.2 Gross vegetative structure:

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Although, most diatoms are unicellular, a number of colonial species is also

6.3 Cell Division:

6.4 Classification of Diatoms:

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2. Sexual reproduction of the Centrales is oogamous whereas that of the pinnate

The classification of the pinnate diatoms is based largely on extent of develop-

Cyclotella striata (Kutzing) Grunow (Fig. 8): Cells resemble C. meneghiniana;

http://slide pdf.c om/re a de r/full/phytopla nkton-ma nua l 16/40

Schroederella delicatula (Peragallo) Pavillard (Fig. 14):

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length of cell, 18-61 µm.

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N. closterium (Ehrenberg) W. Smith (Fig. 33): Valve spindle shaped in middle;