Negative staining methode

NEGATIVE STAINING ::

principle: 

Negative staining use acidic dye like nigrosine, congo red. Acidic dyes have negatively charged chromogen which will not bind (react) with bacterial cells because of similar acidic chargr on the surface of bacterial cells . The stain does not stain the bacteria because of ionic repulsion. 

1. Natural shape and size of the cell is not disorted because the heat fixation is not required during the staining.

1. It is possible to observe those bacteria which are difficult to stain , such as spirochetes

1. requirements:

• Young culture of E. coli
• Nigrosin staining solution

PROCEDURE :

1.- place a small drop of nigrosine at the end of the slide. for culture on solid media, add a loopful of distilled water and emulsify a small amount of the culture in the Nigrosine-water drop. for broth cultures, mix full of the culture into the drop of Nigro Sine. do not spread the drop or let dry.

2. Using the end edge of another slide, spread the Drop out to produce a Samear varying from opaque black to gray. the angle of spreading slide will determine the thickness of the smear.

3. Let the smear air dry and examine ( do not  fix the slide)

RESULT:

● organism are observed as transparent objects against the dark background .

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Information of Spirogyra

 Spirogyra:

Kingdom : plant

Subkingdom- : Cryptogams
Division: Thallophyta
Sub-division Algae
Class: Chlorophyta
Sub-class: Chlorophyceae
Cohort: Zygnematales                                               (Conjugales)
Family: Zygnemataceae
Genus: Spirogyra

Occurrence :- 

Spirogyra is found in the form of bright green free floating masses in freshwater reservoirs, ponds, rivers, lakes and slow flowing streams.  Spirogyra is found floating on the surface of water.  The thallus of spirogyra covered by mucilage therefore the filaments appear slimy & smooth.  Hence spirogyra is also known as “Pond silk" or “water silk”. Genus includes about 289 species and out of these 94 have been reported from India.

Morphology: - 

The plant body is called as thallus.  The thallus is in the form of silky, hair-like filament. The spirogyra is multicellular, unbranched, cylindrical & filamentous.  These filaments are long, cylindrical, free-floating and slimy. As all its cells are similar in structure, the thallus is termed as homothallus.  The cells are arranged end to end, one above the other in a single row. All the cells in the filaments are similar and usually much longer than broad. The filament cannot be differentiated into apex or base, but some Species have basal cell with which the filament is attached to the rock.  This basal cell is known as hold fast.  During day time, O2 evolved due to photosynthesis fills the interstices of the filaments and keep them floating on the surface of water.

Cell - structure: 

The cells are cylindrical. Their length is more than the width.  It consist of two parts the cell wall and the protoplast.
The structure of cell wall is similar in all the cells of the filament.  It is thin & made up of 2 layers.  The inner layer is made up of Cellulose while the outer layer is formed of pectin.  The outer part of pectose layer is converted into pectin, which dissolves in water to form a mucilaginous sheath. This mucilage sheath is considered as a third layer of cell wall.  Due to this reason the spirogyra filaments are slippery. Normally the size of the cell is about 0.25 mm.
Protoplast: It is differentiated into plasma membrane, cytoplasm and nucleus.  The plasma membrane is thin and living. The central portion of the cell is occupied by a large central vacuole because of that cytoplasm is divided into two parts.  The cytoplasm which is found attached with cell wall is known as ectoplasm and the one which surrounds a centrally located nucleus is known as endoplasm.   Fine cytoplasmic strands keep ectoplasm & endoplasm interconnected.  A big central vacuole surrounds the endoplasm & the nucleus. The ectoplasm surrounding the vacuole is called primordial utricle. The nucleus is kept suspended within endoplasm by means of cytoplasmic strands. 
The nuclear possesses 1 to 2 nucleoli. The cell possesses all the organelles. 
The chloroplast are vary in numbers from 1 to 16.  Each chloroplast is an elongated, green, flat, ribbon like band which is spirally arranged from one end of cell to the other end.  The twisting of the spiral shaped chloroplast is on the left side.  The margin of the chloroplast may be uniform, wavy or serrated.  .  A series of small, rounded, spherical bodies situated at equidistance in the green band of chloroplast.  These structures are called as pyrenoids.  These pyrenoids store protein in the centre and starch in the surrounding region.

Reproduction: 

Two types of reproduction are found in spirogyra viz.
l) Vegetative   (2) Sexual.

Vegetative reproduction
i) Fragmentation
ii) Aplanospores
iii) Akinete

i) Fragmentatio: 

It takes place normally in favourable conditions.  In favourable conditions, some of the intercalary (in between) cells are destroyed. The result is the fragmentation of the filament into small pieces.  Each piece or fragment survives and when conditions are favourable, each fragment will grow in length by cell division, attaining the full length of the filament.  Thus as many fragments as many filaments are formed. 
a) Mechanical injury:
 the rapid flow of water, biting of an aquatic animals or the animals which come to drink water may cause fragmentation. 
b) By sudden changes in the habitat factors such as temperature or acidity of water the middle lamellae of the end walls become gelatinous.  Consequently a turgor difference arises between adjacent cells, causing one cell to bulge in to the other. The strain on the junctions between the cells finally reaches the breaking point.
c) In some species with replicate end walls the rings of one cell get evaginated forcing apart the cells.
d) In some species formation of H- shaped piece (H0colligata) in the end walls of adjacent cells also causes fragmentation. The H-piece slips off. Consequently the two cells come apart.

Aplanospores :-

 S. aplanospora (Randhava-1938) reproduces by aplanospores. Aplanospores formed singly in the vegetative cells.

iii) Akinete :- 

S. farlowii (Transeau-1915) reproduces by formation of thick lamellated walled akinets.


Sexual reproduction:
 spirogyra reproduces sexually by conjugation. In Spirogyra isogamous type of sexual reproduction is seen.  The gametes are similar in size and shape The gametes of spirogyra are aplanogametes (non-.motile).  When conditions are unfavourable, spirogyra reproduces sexually.

There are two types of Conjugation.  (1) Scalariform  (2) Lateral.

Scalariform conjugation:

 This is seen in the beginning of summer season.  During this time two filaments lie closer & remain parallel in such a way that their cells face each other.  Both the filaments remain covered by the same mucilage covering.  The cells face each other & by the touch stimulus, they are stimulated to form small protuberances.  Both the opposite protuberances grow & meet each other.  By the action of enzymes, catalase & cellulase, their contact walls dissolve forming a continuous tubular passage, called conjugation tube.  Such tubes are formed successively between each pair of the cells of the 2 filament.  They appear like a ladder or staircase.  Hence the name Scalariform conjugation.
At this time, the protoplasm of each cell of both the filaments shrinks & forms a gamete.  The cells are referred as gametangia.  The gametes of one filament become active and they pass through the conjugation tube by amoeboid movement and meet the inactive gametes of the other filaments.  The active gametes are known as male gametes.  While the inactive gametes are known as female gametes Thus the fusion of male & female gametes takes place This Is termed as conjugation.  As a result of conjugation, diploid cell, called zygote is formed.  The zygote is covered by a protective layer and it is now known as zygospore.  The cell if both the filaments as well as conjugation tubes disintegrate, liberating the zygospores.  The zygospores sink at the bottom & remain dormant for some period

Spirogyra—Transfer of one aplanogamete to other aplanogametangia A. Two filaments lying parallel to each other, B. Formation of papillate outgrowths to form conjugation tube, C. Ladder-like (Scalariform) conjugation. One aplanogamete moving into the aplanogametangia of the other filament, D. Zygospores formed in one filament and empty aplanogametangia of the other filament

Lateral -conjugation:

 In some species of spirogyra, lateral conjugation is found. A conjugation tube is formed on the lateral sides of a adjacent cells of the same filament: It is termed as lateral conjugation.  Lateral conjugation is of the two types.

(I) Indirect: In this type of lateral conjugation, small protuberance develops from each of the adjacent cells on the same lateral side. The two protuberances then meet each other and as the growth of the transverse wall does not keep space with the growth of protuberances, a sort of lateral conjugation tube is formed between 2 adjacent cells.
Now, the protoplasm of these cells shrinks and forms the gametes. The active gamete i.e. male gamete now passes through the lateral conjugation tube by amoeboid movement & meets the inactive i.e. female gamete. Both these gametes fuse with each other, forming a diploid zygote. The zygote then gets covered by a protective layer and is termed as zygospore.  Male gamete producing cells & female gamete producing tells may be found alternatively or in alternative pair in the filament.

(ii) Direct: Prof. O.P. Ayenger noticed that in spirogyra jogensis this type of lateral conjugation occurs.  No conjugation tube is formed in this type of lateral conjugation. First the protoplasm of cells shrinks & forms the gametes. Now the active i e male gamete directly pushes the partition wall and it gets perforated. The male gamete enters through the perforation and fuses with the inactive or male gamete forming a diploid zygote.  The zygote later on changes into zygospore by enveloping itself into a protective sheath.

Zygospore and its germination: 

Zygopspore is elliptical or spherical body covered by 3 layered protective walls.  It is diploid and hence corresponds to sporophytic generation.  The cells of the filament are destroyed and the zygospores are liberated and sink at the bottom. Here they remain in dormant condition for a period of 4 to 6 months.  The chloroplast gets destroyed.  Starch is converted into oil.  3It therefore appears grayish or yellowish.  When the favourable season approaches, the zygospore germinates.  This time the outer two layers break off and the inner cellulose layer grows as germ tube.  Its diploid nucleus undergoes the reduction division forming 4 haploid nuclei.  3-of these haploid nuclei, degenerate and the one remaining shift into germ tube.  Now by transverse division the germ tube divide into two cells.  A proximal cell helps in attachment to the substratum while the apical cell divides and redivdes forming the filament of spirogyra.

Parthenogenesis: 

Sometimes when conditions are, not very favourable, the filaments are arranged adjacent to each other and forms gametes but conjugation tubes-are not formed.  So, the gametes, cannot fuse at such time, the gamete itself gets covered by a protective layer and forms a structure similar to zygospore.  But it is haploid and it is known as azygospore.   When conditions become favourable, the azygospore germinates without reduction division into new filament.  This is known as parthenogenesis as parthenogenesis.

Life cycle of Spirogyra: 

Gametophytic stage is the principal stage in the life cycle of spirogyra.  It is haploid and produces haploid gametes.  By conjugation a diploid zygote is formed.  Being diploid, it represents a sporophytic stage. This is a very recessive and dormant stage.  In favourable conditions, it germinates by meiosis forming again a gametophytic stage.  Thus, in the life cycle gametophytic stage alternates with a sporophytic stage and so alternation of generation is found.

Information of Mucor

Mucor:

Kingdom: Plantae

Sub - kingdom: Cryptogams

Division: Thallophyta

Sub -Division : Fungi

Class : Phycomycetes

Sub - class: Zygomycetes

Cohort Mucorales

Family: Mucoraceae

Genus: Mucor

Occurance: 

Mucor is a very common fungus which can be seen on decaying organic materials such as fruits, prickles old wet shoes etc.  Mucor stolonifer grow on damp bread while Mucor mucedo grow on cow dung or horse dung.  In the laboratory it can be cultured by keeping moist bread in the bell jar for the period of two to three days.  During this time bread gets covered by fine cottony white filaments which are nothing but hyphae of mucor forming mycelium.  This genus is optimum temperature and moisture are two factors essential for the rapid growth of mucor.

Thallus of mucor is known as mycelium which is white delicate highly branched, unicellular and achlorophylous.  Hypahal wall is made up of fungus cellulose while cytoplasm is granular and vacuolated with many nuclei.  Reserve food is in the form of sugar, glycogen and droplets of lipids.  Hyphae forming mycelium are tubular and aseptate but at the time of formation of reproductive organs the hyphae become septate.  In beginning mycelium spreads on substratum.  Some hyphae are spread on the substratum and some enter the substratum and absorb nutritive substances.  Sporangiphores are essential for the asexual reproduction and they arise solitary in genus mucor.

Reproduction: 

Mucor shows three types of reproduction 

(1) Vegetative reproduction, 

( 2) Asexual reproduction and 

(3) Sexual reproduction.

(1) Vegetative Reproduction (Fragmentation):

 Mucor filaments may break down by mechanical injury and such fragments develop into a new mucor plant.  This method of reproduction is known as "reproduction by fragmentation method".

(a) Reproduction by stolons:  

In this mode of reproduction mycelium develops some aerial hyphae which bend down and touch the substratum.  Later on it gives out rhizoidal hyphae and sporangiophore.  Thus, gradually it covers the entire region of substratum.  Stolon breaks and results into two separate plants.  Such type of reproduction is met with only Rhizopus genus.

(2)Asexual Reproduction: 

In mucor asexual reproduction takes place by Aplanospores, Chlamedospores and oidiospores.

(a) By aplanospore: 

This type of asexual reproduction is very common and such spores develop in humid environment.  When substratum is nutritive in nature.

 

(i) Development of Sporangium and spores: From the thallus of mucor, some aerial hyphae develop which are known as spornagiophores.  The apical region becomes rounded in which several nuclei; cytoplasm and food particles enter in.  This round dome shape region is known as sporangium which appears white in the beginning but later on due to development of spores, it appears black.  Wall of the sporangium is thin and spiny.  The peripheral cytoplasm which touches the wall of sporangium is dense and shows more nuclei while inner cytoplasm is thin and with few nuclei.  Dome shape structure develops in the center with thin cytoplasm.  Peripheral and inner cytoplasm separated by a layer of small vacuoles.  As a result dome shaped columela and spore producing region get well defined.  Now, peripheral protoplast shows cleavage furrows and divides the protoplasm into several small coenocytic groups.  Each group gets covered by a wall and then it is known as spore.  Each spore is minute and coenocytic i.e. multinucleate.

(ii) Dehiscence of sporangium and dispersal of spores: During development of sporangium, columela and sporangiophore receive more thin protoplast in liquid form and hence it exerts more pressure on columela.  As a result sporangium dehisces and spores are dispersed.  After the dehiscence, remnants of sporangial wall appears as rugged collar.  Mature spores are very minute, light in weight and are dispersed by the wind.  In favourable condition, they germinate on the substratum and produce new mycelium.

(b) By Chlamydospores, Oidiospores and torula stage: When mucor is immersed in sugar solution, its mycelium gets septate and becomes multicellular.  Some of the cells swells up become enlarged in size and become thick walled.  These cells are as chlamydospores.  These chlamydospore behave as resting spores.

Some times, the septate mycelium gets fragmented. Each small fragment consists of thin walled cells called oidial cells.  These cells shows budding like yeast cell in sugar solution.  This stage is referred as yeast stage of mucor. This stage shows anaerobic respiration and produces alcohol in sugar solution.  Hence, this stage is named as anaerobic stage or alcoholic stage or alcoholic fermentation stage of mucor.

Torula can be defined as a small swollen region.  During immersion of mucor in sugar solution, hyphae become septate and form small swollen regions as chlamydospores.  This stage is therefore called torula condition.

(2) Sexual Reproduction:

 Mucor shows isogamous type of reproduction.  Some species are homothallic while some are heterothallic.  In heterothallic species hyphae are similar externally while physiologically they are differ from each other.  Dumbel shaped lateral branches develop from heterothallic filaments as shown in the diagram.  Apical region is called progametangium which shows development of a septum.  Terminal portion is known as gametangium and remaining portion is called suspensor.  Gametangium shows denser granular protoplast with many nuclei while suspensor shows vacuolated protoplast with few nuclei.  As gametangia are similar in appearance, instead of referring them as male and female gametangia, they are referred as +ve strain and -ve strain gametangia.  These gametangia possess gametes which are referred as coenogemets or aplanogametes.  When two gametangia meet and unite with each other, the intervening walls between them dissolve.  At this time, +ve strains and -ve strains nuclei fuse in pairs.  The combined protoplast first results into a cell called zygote which increases in size, its wall becomes thick, black and warty and it is now known as zygospore.  This zygospore is isodiametric having a diameter 70-80 µ.  Here, glycogen and lipid droplets are seen as reserve food material.  After the detachment from suspensor, zygospore passes 6-9 months in resting phase.

   

• Germination of Zygospore:
  
  

The zygospore germinates under suitable conditions.  The zygospore protoplast absorbs water and swells up.  As a result, outer wall splits open and a small, stout vertical hypha develops form the inner wall which is known as promycellium or germ tube which grows to a limited extent.  Now, apical end swells and assumes round shape which develops a transverse wall and thus sporangium develops which includes several nuclei and protoplast.  Nuclei undergo meiosis and resultant haploid daughter nuclei are of +ve and -ve strain.  Each nucleus shows protoplast and results into a round haploid spore which is referred as aplanospores.  It may be of +ve or -ve stain.  On germination, it results into +ve stain or -ve stain filaments.  On maturity, wall of sporangium breaks open.  Aplanospores are light in weight and hence, dispersed by the wind.  On getting suitable substratum and condition, they germinate.  As there are two types of aplanospores (+ve and -ve) +ve and -ve strain mycelium are produced.

Parthenogenesis: Some times, the gametangia fails to fuse.  Each such gamete functions as parthenospore which develops into a thick walled structure called azygospore.  If conditions are favourable, it germinates and results into a mycelium without any meiotic division.

Mucor express gametophyte phase and sporophytic phase by mycelium and by zygospore respectively.  They alternate with each other.  Here, haploid mycelium (n) is the main phase and while zygospore (2n) is the subsidiary phase of life cycle.

Lipid and their forms

Lipid

•Lipids are fatty acid of alcohols and related
substance which are in soluble in water but soluble
in number of no polar organic solvents such as
ether, chloroform and benzene.
•Lipids are basically made of carbon, hydrogen, and
sometimes oxygen in which the content of oxygen is
always small as compared to hydrogen and carbon.
•Many lipids contain small amount of additional
element like phosphorus, nitrogen and sulphur.
•This lipid include fats, oils, waxes and related
compound.
•The basic components of all lipids are fatty acids.

 1.Simple lipid

• These are ester of fatty acids with various alcohols e.g
glycerol in neutral fats and cytyl alcohol in waxes.
• a. Neutral fat or triglycerides
• These are formed by esterification of three molecule of
fatty acid with one molecules of trihydric alcohol,
glycerol.
• If two number of fatty acids attached to a glycerol, the
ester is called diglyceride, or monoglyceride if there is
only one molecules of fatty acid attached to a glycerol
molecules.
• Animal triglycerides contain a higher proportion
of long chained saturated fatty acids such as
palmitic and stearic acid, resulting higher
melting points.
• Thus at room temp, they are semisolid or solid .
• Most plant triglyceride contain a larger
proportion of short chained unsaturated fatty
acids like oleic, linoleic or linolenic acids.
• They have low melting points and are liquids at
room temp.

B. Waxes

• They are fatty acid esters of long chain
monohydric alcohols like cytyl, ceryl or mericyl.
• Waxes are chemically immobile as they do not
have double bonds in their hydrocarbon chains
and are highly insoluble in water.
• Plant waxes occur in cuticle on the leaf surface.
• In animal, cutaneous gland are known to secrete
wax lanolin for forming a protective water soluble
coating on animal hair.
• Bees wax is formed from palmitic acid C16H32O2,
and mericyl alcohol C30H61OH or ear wax is
secreted by coetaneous gland for lubricating ear
drum.

• C. Cutin

• It is complex lipid produced by cross esterification and
polymerisation of hydroxy fatty acid as well as other
fatty acids or without esterification by alcohol other
than glycerol.
• Cutin occurs in the aerial epidermal cell walls a wells as
a separate layer of cuticle on the outside of these
epidermal cells.
• Cuticle has 50-90% cutin.

• D. Suberin

• It is a mixture of fatty material having condensation
products of glycerol and phenolic acid or its derivatives.
• It occurs in the walls of cork cells and endodermal cell
making the cell wall strong and impregnable.

2. Compound lipids

• These are esters of fatty acids and alcohol with
additional compounds such as phosphoric acid,
sugars, proteins ect.
• These are classified
• Phospholipids
• They are triglyceride lipid where one fatty acid is
replaced by phosphoric acid which is often linked to
additional nitrogenous group like choline in lecithin,
ethanolamine in cephalin, serine or inositol.
• Phospholipids carry both hydrophobic nonpolar and
hydrophilic polar groups.
• In sphingophospholipid the phospholipid
contain amino alcohol sphingosine.
• Sphingomyelins contain an additional
phosphate attached to choline.
• In sphingomyelin, amino group of the
sphingosine backbone is linked to a fatty acid
by an amide bond.
• On hydrolysis, sphingomyelins yield fatty acid,
phosphoric acid, choline and a complex amino
alcohol sphingosine.
• They are found in large quantities in brain and
nerve tissue, especially in the myelin sheath of
the nerves

• Glycolipids

• They are sugar containing lipid and consist of high
molecular weight fatty acid, sphingosine and sugar
residue galactose.
• They occur in nerve membrane, adrenals, kidney,
leucocytes and retina.
• Individual cerebrosides are differentiated by the
type of fatty acids in the molecule such as kerasin
containing the saturated lignoceric acid, cerebron
containing the cerebronic acid, nervon containing
the nervonic acid.

• • 4. Hydrocarbons

• These are substances which do not have any
structural relation to fatty acids but yet are grouped
with lipid only because of their similar solubility
properties.
• They include carotenoids, vit-A, E, K
• Carotenoids is related to lipid. They are fatty acid
like carbon chain carrying carbon at each end.
• They have general formula C40H56 .
• These compound serve as pigment in both plant
and animal matter.
• These are two classes of carotenoids viz., carotenes,
xanthophylls.
• These contain N acetylneuraminic acid, fatty acids,
sphingosine and three molecules of hexose (glucose
and galactose.
• Gangliosides occur in grey matter.

• Lipoproteins
• Protein molecules conjugated with triglycerides, 

cholesterol or phospholipids are called lipoprotein.
• Two important groups of lipoprotein are low density
lipoprotein and high density lipoprotein.
• It present in blood, milk and egg yolk.

3. Steroids

• The steroid are generally considered along with
lipid, although they differ from them in
chemical structure.
• All steroids have in common the basic ring
structure which possess 17-carbon nucleus of
four fused hydrocarbon ring , cyclopentano
perhydro phenenthrene.
• The steroid include such as substance as
cholesterol and other sterols, the sex hormones
and the hormones of the adrenal cortex.

• Sterols

• Steroid with 8-10 carbon atom in side chain at
position 17 and an alcoholic hydroxyl group at
position 3 are classed as sterols e.g cholesterol,
stigmasterol, campesterol, sitosterol, ergosterol.
• Cholesteol is the common sterol found in many
animals, human being and some plants.
• It occurs in both free and combined form when it is
esterified with fatty acid.
• It is precursor of steroid hormone like
progesterone, estradiol, testosterone, aldosterone,
cortisol, ecdysone.

• 4. Hydrocarbons

• These are substances which do not have any
structural relation to fatty acids but yet are grouped
with lipid only because of their similar solubility
properties.
• They include carotenoids, vit-A, E, K
• Carotenoids is related to lipid. They are fatty acid
like carbon chain carrying carbon at each end.
• They have general formula C40H56 .
• These compound serve as pigment in both plant
and animal matter.
• These are two classes of carotenoids viz., carotenes,
xanthophylls.

Information of zygnema

Zygnema:

Taxonomic Position:

Division: Chlorophyta

Class: Chlorophyceae

Order: Zygnemales

Family: Zygnemaceae

Genus: Zygnema

 Habitat:

Zygnema is common green algae having about a hundred species. All are aquatic except Z.terrestre, Randh. They are found in considerable quantities in quiet fresh water of ponds, lakes and also on slow flowing streams as yellowish green free floating masses on the surface of the water.

 Habit:

 It is a simple, fine, unbranched thread or a filament consisting of several cylindrical, rectangular cells arranged one above the other in a row. 

 Cell Structure: 

1. Each cell contains a small mass of protoplast surrounded by a cell wall.

2. The cell wall is two layered – inner cellulosic and outer pectic, covered my mucilage sheath

3. The protoplast has a cell membrane, cytoplasm, nucleus and two stellate or star shaped chloroplasts. 

4. The chloroplasts are embedded in the cytoplasm.

5. Each chloroplast consists of a central body with a single pyrenoid at the center.

6. A number of delicate strands radiate out from the center of the body of the chloroplast and extend towards the plasma membrane.

7. A single centrally situated nucleus lies embedded in the middle of the broad strand of cytoplasm connecting the two chloroplasts.

 

 Reproduction in Zygnema:

 Zygnema shows vegetative, asexual and sexual modes of reproduction

Vegetative Reproduction – It is shown by fragmentation. The filament breaks into short segments of living cells or individual cells. These may show cell division or simply grow into new Zygnema filaments.

Asexual Reproduction – It is accomplished by Akinets or thick walled Aplanospores.

Akinetes – An Akinets is a thick walled, resting, vegetative cell. It may be formed singly, in rows of two or three or in later stages as long chains.

Aplanospores – They are produced singly in the vegetative cells. They may be round or ovoid in form and have thick, variously colored walls. However, according to Randhawa (1938) the mature Aplanospore has dark greenish blue colored wall. The shape of Aplanospore varies from oval to pyramidal or barrel shaped.

Sexual Reproduction – Zygnema shows scalariform and lateral form of conjugation. 

(a) Scalariform Conjugation:

It is similar to that of Spirogyra. At the onset, the filaments come to lie side-by-side in pairs. Small dome shaped protuberances grow towards each other from the opposite pairs of cells. The protuberances elongate and finally meet. At the point of contact, the intervening walls dissolve. With this the two outgrowths form a conjugation tube. This open passage is known as the conjugation canal. Most species of Zygnema are usually isogamous (no visible difference between the gametes and the conjugating filaments). Now the protoplasts of the conjugating cells form gametes, singly. The protoplast or the gamete, which functions as the male gamete shrinks from the parent cell wall, migrates through its respective conjugation canal and fuses with its partner in the opposite cell, which can be termed as female gamete. The movement of the male gamete is often amoeboid. This fusion leads to the formation of the zygospore.

(b) Lateral Conjugation :

 It is commonly found in Z.gangeticum and Z.heydrichii. In this case the adjoining cells of the same filament give out tube like protuberances on either side of the septum. When the protuberances finally meet, the separating septum breaks at the point of contact and a communication link is established between the two adjoining cells. The contents of one cell migrate into the other where the fusion of two protoplasts takes place to form a zygospore.

Zygospore – The fusion cell formed by the union of protoplasmic masses of the two conjugating strands is called a zygospore. It contains four chloroplasts and a single diploid nucleus. The zygospore secretes a wall around it which gradually thickens. This wall is differentiated into three layers:

1. Exospore – Thin, hyaline outer layer which is cellulosic or pectic in nature.

2. Mesospore – Thick middle layer made up of cellulose.

3. Endospore – Thin delicate innermost layer made of cellulose.

The Zygospores sink to the bottom of the pond after disintegration of the walls of the female cells or conjugation tubes. Upon release the zygospores enter a resting period. Two out of the four chloroplasts, disintegrate. The Zygospore is now capable of resisting unfavorable conditions. It is filled with starch and oil. It germinates during favorable conditions.

Germination of Zygospore – The zygospore is released after the disintegration of the walls and comes to rest at the bottom of the pond. At the beginning of the next growing season the zygospore germinates to form a new plant. Prior to germination, two of the four chloroplasts disintegrate. The diploid nucleus undergoes meiosis to from four haploid nucleii. Three of these disintegrate and only one remains. This is the functional haploid nucleus. It divides and redivides to form daughter cells. In this way a new filament is formed.

Information of Nostoc alagae

Nostoc:Taxonomic position:

Division : Cyanophyta

Class : Cyanophyceae

Order : Nostocales

Family : Nostocaceae

Genus : Nostoc

Habitat:

Nostoc is a filamentous form of both terrestrial & aquatic habitats. The colonies may be free floating or attached 

Terrestrial sp. grows on damp soil & forms leathery or rubbery sheet, it is also common in the arctic and alpine meadows.

Aquatic sp. occur are found as free floating thalli in sunny pools ,ponds and lakes or lying at the bottom attached to submerged vegetation .some species favor running water ,especially fast flowing mountain streams .attached to stones along the stream  beds. Nostoc also occurs in symbiotic association with fungi to form lichens.

 Habit:

Nostoc filaments grow in large colonies as closely packed trichomes embedded in a firm matrix of gelatinous material.

The mucilaginous lump of colony may occur free floating or attached.

Each trichome is of varying length and is made of many bead like cells.

It is enclosed by its own mucilaginous sheath.

Thus Nostoc colonies appear as yellowish of bluish green mass of jelly. Several colonies are enclosed within a gelatinous mass forming a macroscopic compound colony.

 Morphology:

Structure of the trichome:

1) Each trichome is composed of numerous rounded or oval cells.

2) The cells are joined closely from end to end into a trichome resembling a string of beads.

3) At frequent intervals along the trichomes, colorless empty looking, barrel shaped cells called heterocysts are found 

4) Heterocysts are slightly larger and have thicker walls than other vegetative cells.

5) Heterocyst is mostly intercalary but sometimes terminal in position.

6) They occur singly but sometimes in series.

7) Each intercalary heterocyst has two polar pores through which cytoplasmic connections are maintained with adjacent cells.

8) Under certain conditions some vegetative cells may become enlarged stored with food materials and greatly thickened to become resting bodies called akinets.

9) Each cell of the trichome consists of cell wall surrounding the protoplast.

10) The protoplast can be vaguely differentiated into outer pigmented cytoplasmic region called chromoplasm.

11) The chromoplasm contains colourless granules of myxophycean or cynophycean starch and cyano granules of protein mature.

Reproduction:-

Nostoc reproduces entirely vegetatively by following methods:-

1) Colony fragmentation: - the Nostoc colony, as it gets larger breaks up into smaller pieces due to storms or other disturbances. Each of these grows up to the size of parent colony.

2) Hormogone formation: - this vegetative method is common in Nostoc. The trichome ruptures at junction of heterocyst and vegetative cell, which is the weakest link in the chain. In this way smaller segments of living cells called “Hormogonia” (sing. Hormogone) become isolate. they plip out of the enclosing gelatinous matrix and establishes new colonies by division 

3) Akinets formation (): under certain condition any cell or some of the cell of the trichome becomes enlarged and each secrets a thick, highly resistant wall around it. They get filled with reserved food materials such specially modified cells are called akinets or resting pores Akinets are well adopted to survive during unfavorable conditions like water storage or unsuitable temperature. With the return of favorable season, each akinets germinates to form a new filament of Nostoc.

4) Other methods of reproduction like heterocyst germinate or endospore formation are also found.