Cynobacteria

2. 1. Habitat
2. Morphology
3. Cultivation
4. Reproduction
5. Evolution
6. Genetics
7. Photosynthesis

4. y
Division Cyanophyta
Cyanobacteria ‘formerly known as’
BlueGreen Algae
- Cyano = blue
- Bacteria – acknowledges that they are
more closely related to prokaryotic
bacteria than eukaryotic algae

5.  Introduction to Cyanophyceae:
 It is a primitive group of algae, consists of 150
genera and about 2,500 species. In India, the
division is represented by 98 genera and about 833
species. Members of the class Myxophyceae
(Cyanophyceae) are commonly known as blue
green algae. The name blue green algae is given
because of the presence of a dominant pigment c-
phycocyanin, the blue green pigment.
 In addition, other pigments like chlorophyll a (green),
c-phycoerythrin (red), β-carotene and different
xanthophylls are also present. The members of this
class are the simplest living autotrophic prokaryotes.

6.  సైనోఫిసీ పరిచయం: ఇది ఆల్గే యొక్క ఆదిమ
సమూహం, ఇందులో 150 జాతులు మరియు సుమారు
2,500 జాతులు ఉన్నా యి. భారతదేశంలో, ఈ
విభాగాన్నా 98 జాతులు మరియు 833 జాతులు
సూచిసుున్నా యి. తరగతి మైక్సో ఫిసీ (సైనోఫిసీ)
సభ్యు లను సాధారణంగా నీలం ఆకుపచచ ఆల్గే
అంటారు.
 నీలం ఆకుపచచ ఆల్గే అనే పేరు పెట్టబడంది
ఎందుక్ంటే ఆధిపతు వరణద్రవు ం సి-ఫైక్ససైన్నన్,
నీలం ఆకుపచచ వరణద్రవు ం.
 అరనంగా, క్ల
క్సరోఫిఫి్ ఎ (ద్ీన్), సి-ఫైక్సరిద్ిన్ (ఎరుపు),
β- కెఫిటిన్ మరియు వివిధ జాంతోఫి్ో వంటి ఇతర
వరణద్రవు ం కూడా ఉన్నా యి. ఈ తరగతి సభ్యు లు
సరళమైన జీవన ఆటోద్టోఫిక్ ద్ొకాఫిు ట్లరో.

8.  Important Characteristics of Cyanophyceae:
 1. The individual cells are prokaryotic in nature. The
nucleus is incipient type and they lack membrane bound
organelles.
 2. Both vegetative and reproductive cells are non-
flagellate.
 3. Cell wall is made up of microfibrils and is
differentiated into four (4) layers. The cell wall composed
of mucopeptide, along with carbohydrates, amino acids
and fatty acids.
 4. Locomotion is generally absent, but when occurs, it is
of gliding or jerky type.
 సైనోఫిసీ యొక్క ముఖ్ు మైన లక్షణాలు: విభజన యొక్క ముఖ్ు మైన
లక్షణాలు ద్రంది విధంగా ఉన్నా యి: 1. వు ర ుగత క్ణాలు ద్పక్ృతిలో
ద్ొకాఫిు టిక్. న్యు ర రోయస్ ద్ారంభ రక్ం మరియు వాటిర ొర
క్ట్లటకునా అవయవాలు ల్గవు.
 2. ఏపుగా మరియు పునరుతప తిు క్ణాలు రండూ క్ల
ా
రో లల్గట్ కాన్నవి. 3.
సె్ గోడ మైద్క్సఫైద్ి్ో తో రూొందించబడంది మరియు ఇది

9.  5. The principal pigments are chlorophylls a (green), c-
phycocyanin (blue) and c-phyco- erythrin (red). In
addition, other pigments like β-carotene and different
xanthophylls like myxoxanthin and myxoxanthophyll are
also present.
 6. Membrane bound chromatophore are absent.
Pigments are found embedded in thylakoids.
 7. The reserve foods are cyanophycean starch and
cyanophycean granules (protein).
 5. ద్పధాన వరణద్రవాు లు క్ల
క్సరోఫిఫి్ో ఎ (ఆకుపచచ ), సి-
ఫైక్ససైన్నన్ (నీలం) మరియు సి-ఫైక్స-ఎరిద్ిన్ (ఎరుపు).
అరనంగా, β- కెఫిటిన్ వంటి ఇతర వరణద్రవు ం మరియు
మైకాో క్ో ంతిన్ మరియు మైకాో కాశ ంతోఫి్ వంటి విభినా
శంతోఫి్ో కూడా ఉన్నా యి.
 6. మంద్ేన్ బండ్ ద్క్సమాటోఫోర్ ల్గదు. వరణద్రవు ం
థైలాక్సయిడ్ో లో ొందుపరచబడ ఉంటాయి.
 7. రిజర్్ ఆహారాలు సైనోఫిషియన్ క్ల
సా
ట ర్చ మరియు

10.  8. Many filamentous members possess specialized cells
of disputed function (supposed to be the centre of
N2 fixation) known as heterocysts.
 9. Reproduction takes place by vegetative and asexual
methods. Vegetative reproduction takes place by cell
division, fragmentation etc. Asexual reproduction takes
place by endospores, exospores, akinetes, nannospores
etc.
 10. Sexual reproduction is completely absent. Genetic
recombination is reported in 2 cases.
 8. చాలా మంది ఫిలమంట్స్ సభ్యు లు వివాదాసప ర
ఫంక్షన్ యొక్క ద్పత్యు క్ క్ణాలను క్లిగి ఉంటారు (N2
క్ల
సిరీకక్రణకు కంద్రంగా భావించాలి) హెటెఫిసిక్ల
స్టో అన్న
పిలుసా
ు రు.
 9. ఏపుగా మరియు అలంగిక్ పరధతుల దా్ రా పునరుతప తిు
జరుగుతుంది. వృక్షసంపర పునరుతప తిు క్ణ విభజన,
విచిి నా ం మొరలన వాటి దా్ రా జరుగుతుంది.

11.  Thallus Organisation in Cyanophyceae:
 Plants of this group show much variation in their thallus
organisation.
 The thallus may be of unicellular or colonial forms:
 1. Unicellular Form: In unicellular form, the cells may be
oval or spherical. Common members are Gloeocapsa (Fig.
3.23A), Chroococcus and Synechococcus.
 2. Colonial Form: In most of the members the cells after
division remain attached by their cell wall or remain together
in a common gelatinous matrix, called a colony.
 The colonies may be of two types: a. Non- filamentous,
and b. Filamentous

14. Growth
Forms
Synechococcus Crococcus Cells of Anacystis
Oscillatoria
Spirlina spp.
Anabaena

15. Fossil
Cyanobacteria

16. "Ooid" sand grain containing
Endolithic Cyanobacterium
(CB). Erosion caused by Endolithic Cyanobacteria

18. Structural drawing of the
fine structural features of
a cyanobacterial cell.
(D) DNA fibrils;
(G) gas vesicles; (Gl)
glycogen granules;
(P)plasmalemma; (PB)
polyphosphate
body;
(Ph) polyhedral body;
(Py) phycobilisomes;
(R) ribosomes;
(S) sheat;
(SG) structured granules
(cyanophycin granules);
(W) wall.

19.  a. Non-Filamentous Type: The cells of this type divide
either alternately or in three planes, thereby they form
spherical (Gomphosphaera, Coelosphaerum), cubical
(Eucapsis alpine, Fig. 3.23C), squarish (Merismopedia) or
irregular (Microcystis, Fig. 3.23B) colony.
 b. Filamentous Type: By the repeated cell division in one
plane, single row of cells are formed, known as trichome.
e.g., Oscillatoria (Fig. 3.23D), Spirulina, Arthosporia etc.
The trichome when covered by mucilaginous sheath is
called a filament. The filament may contain single
trichome (Oscillatoria, Lyngbya) or several trichomes
(Hydrocoleus, Microcoleus, Fig. 3.23E).
 The trichomes may be unbranched (Oscillatoria,
Lyngbya), branched (Mastigocladus limilosus, Fig. 3.23J)
and falsely branched (Scytonema, Fig. 3.23K and
Tolypothrix).

20.  ఒక్. న్నన్-ఫిలమంట్స్ రక్ం: ఈ రక్మైన క్ణాలు
ద్పత్యు మాా యంగా ల్గదా మూడు విమాన్నలుగా
విభజసా
ు యి, తదా్ రా అవి గోళాకార (గోంాస్ఫే రా,
క్సలోస్ఫే రం), కూు ిక్్ (యూకాపిో స్ ఆలప న్, Fig. 3.23
సి), సాక ్ రిష్ (మరిస్పో పీడయా) ల్గదా సద్క్మంగా
(మైద్క్ససిసిటస్, ఫిగ్. 3.23 ి)
 కాలనీ. ి. తంతు రక్ం: ఒక్ విమానంలో
పునరావృతమయ్యు క్ణ విభజన దా్ రా, ఒక వరుస
క్ణాలు ఏరప డత్యయి, వీటిన్న ట్రైక్సమ్ అంటారు.
 ఉదా., ఓసిల్గటోరియా (Fig. 3.23D), సిప రులిన్న,
ఆఫిరీస్పప రియా మొరలనవి. ముసిలాజనస్ క్సశంతో
క్పప బడనపుప డు ట్రైక్సమ్ను ఫిలమంట్ అంటారు.
ఫిలమంట్లో సింగి్ ట్రైక్సమ్ (ఓసిల్గటోరియా, లింగాో ు )
ల్గదా అనేక్ ట్రైక్సమ్లు (హైద్డోక్సలియస్, మైద్క్సక్సలియస్,
Fig. 3.23E) ఉండవచ్చచ .
 ట్రైక్సమ్లు ద్రంచ్ చేయబడన్నవి (ఓసిల్గటోరియా,
లింగాో ు ), శఖ్లుగా (మాసిటగోకారో డస్ లిమిలోసస్, Fig.
3.23J) మరియు తపుప గా కొమో లుగా ఉండవచ్చచ

21.  Reproduction in Cyanophyceae:
 The blue green algae (Cyanophyceae) reproduce by both
vegetative and asexual means. Sexual reproduction is
absent.
 The vegetative reproduction performs through fission
(Synechococcus), fragmentation (Oscillatoria,
Cylindrospermum muscicola), hormogonia formation
(Oscillatoria, Nostoc), hormospores (Westiella lanosa),
planococci and Palmelloid stage.
 During asexual reproduction various types of asexual
spores are formed. These are akinetes (Anabaena
sphaerica, Gloeotrichia natans, Calothrix fusca),
endospores (Dermocarpa), exospores (Chamaesiphon)
and nannocyte (Microcystis) (Fig. 3.27).

22.  సైనోఫిసీలో పునరుతప తిు: నీలం ఆకుపచచ ఆల్గే
(సైనోఫిసీ) ఏపుగా మరియు అలంగిక్ మారా
ే ల దా్ రా
పునరుతప తిు చేసుుంది.
 లంగిక్ పునరుతప తిు ల్గదు. ఏపుగా పునరుతప క్ల
తిు విచిి తిు
(సైనెక్సకాక్స్), ద్ాలో ంటేషన్ (ఓసిల్గటోరియా,
సిలింద్డోసెప రుో మ్ మసిో క్సలా), హాఫిో గోన్నయా న్నరాో ణం
(ఓసిల్గటోరియా, నోస్ప
ట క్), హాఫిో స్పప ర్ో (వెసిటఎలరో లానోసా),
క్ల
ా
రో నోక్సర మరియు ామలా
రో యిడ్ రశల దా్ రా
జరుగుతుంది.
 అలంగిక్ పునరుతప తిు సమయంలో వివిధ రకాల అలంగిక్
బీజాంశలు ఏరప డత్యయి. అవి అరన్నటెస్ (అన్నబెన్న
స్ఫప రికా, క్ల
గోరోట్రద్టిచియా న్నటాన్ో , క్లోద్ిక్ో ఫుసాక ),
ఎండోస్పప ర్ో (డెఫిో కారాప ), ఎక్సో స్పప ర్ో (చమసిఫోన్)
మరియు న్ననోసైట్ (మైద్క్ససిసిటస్) (Fig. 3.27).

24. • cyanobacteria can be found in almost every conceivable
environment, from oceans to fresh water to bare rock to soil.
• They can occur as planktonic cells or form phototrophic biofilms in
fresh water and marine environments,
• they occur in damp soil, or even temporarily moistened rocks
indeserts.
• A few are endosymbionts in lichens, plants, various protists,
or sponges and provide energy for the host.
• Some live in the fur of sloths, providing a form of camouflage.
• Aquatic cyanobacteria are probably best known for the
extensive and highly visible blooms that can form in both
freshwater and the marine environment and can have the
appearance of blue-
green paint or scum.
Habitat

25. Hot Spring at Yellowstone Park:
The dark color is due to the presence of
Cyanobacteria.
Limestone deposit at Yellowstone Park:
The localized areas of green are due to
the presence of Cyanobacteria

26. A schematic outline of the acquisition, reduction,
and loss of genomes and compartments during
evolution. Black arrows indicate evolutionary
pathways; white arrows indicate endosymbiotic
events in the host cell.
Endosymbiotic event 1 occurred at the origin of
eukaryotes. The proteobacterial endosymbiont
gave rise to mitochondria (the smaller organelles in
the bottom part of the diagram).
Endosymbiotic event 2 occurred at the origin of
plastid-containing cells.
Endosymbiotic event 3 represents the secondary
and higher-order endosymbioses giving rise to
numerous algal phyla, as well as apicomplexans
(such as Plasmodium), which have residual plastids,
and to trypanosomes, which have no plastid at all.
Black, filled circles indicate nuclei or nucleomorphs;
ellipses within organelles indicate bacterially
derived genomes, which may be reduced or lost
completely.
More than one kind of host cell and of
endosymbiont is involved in the secondary, and in
the higher-order, symbioses. The genome of the
Archaebacterium is not represented in the diagram.

27. n
• Old 3.5 billion years
• Dominated as biogenic reefs
• During Proterozoic – Age of Bacteria
(2.5 bya – 750 mya) they were wide spread
• Then multicellularity took over
• Cyanobacteria were first algae!

28. a
Production
• For BGA production dig a small pit of 6x3x9 feet size in the soil and lay
down a polythene sheet in the pit to check to percolation of water. Large
galvanized steel tray containing soil can also be used for this purpose.
Now 10 Kg soil, 200 gm of super phosphate, 6 litre water and 100 gm BGA
dry flakes containing wooden dust or mother culture of BGA are added
into the prepared pit. If found any pest in the pit, spray melathion
solution ( 1 ml melathion in one litre water) to destroy pests.
• If green algae and diatoms are found in pit, use 0.05% CuSO4 solution
which will kill the green algae. After 12-15 days, a thick layer of BGA will
be found floating on the water surface of the pit. You can easily
collect/harvest the BGA directly from the pit or let the pit dry after water
evaporation and take out dry flakes of BGA and fill it in small polypack
(100-200 gm) for sale.
• By this simple method BGA culture/incoulum can be prepared for use in
the paddy fields. The same methods may be used in the paddy fields for
the large scale productions of BGA culture.

29. CULTIVATION STOCK
CULTURE
•The stock culture for
maintenance of laboratory
culture, 2- 3 mL of a 3 weeks
old cyanobacterial stock
culture was used as
inoculum in 50 mL of
autoclaved BG 11 medium in
150 mL Erlenmeyer flasks.
•The cultivation was carried
out at 20 ±2°C, under
continuous illumination of
8gmol/m2 by cool
fluorescence lamps.
•The stock cultures were
maintained for 20-30 days.

30. CULTIVATION SHAKE
CULTURE
Aliquots of 50 mL from the stationary phase stock cultures were used to inoculate
500 mL of autoclaved BG11 medium in 1.5 liter Fehrnbach flasks. These samples
were cultivated at 20 ±20°C, under continuous illumination of 8gmol/m2 by
cool fluorescence lamps. The cyanobacterial cultures were harvested after 4-6
weeks.
The cells were separated from the medium by centrifugation (4000 rpm/ 10 min/
100C) followed by filtration with filter paper. The biomasses were lyophilized and
stored at -20°C until use while cultivation media were concentrated to 1/10 (v/v) by
rotary evaporation in vacuum at 400°C and extracted immediately with EtOAc
solvent.

31. The large scale cultivation was carried out in a 45 liter-
glass fermentor. The fermentor was cleaned by
distilled water and 70% isopropanol before use. At the
beginning, the fermentor was filled with 15 L of
medium and after 1- 2 hours 1.5 L of growing culture
(after 20 days of cultivation in three Fehrnbach flasks)
was added. Afterwards, every day 5 L of medium were
added into the fermentor until 35 L of medium were
reached. The cultures were illuminated continuously
with banks of cool white fluorescent tubes of
8gmol/m2 and incubated at temperature of 26°C to
28°C adjusted using a heater. The pH-value of the large
scale culture was adjusted to 7.4-8.5 using CO2
supplementation.
The biomass was collected by centrifugation at 6500 rpm in a
refrigerated continuous-flow centrifuge and lyophilized, then
stored at -20°C.

32. Composition :
Magnesium Sulphate
(MgSO4 .7H2O)
0.025 g
Calcium chloride 0.05 g
Sodium chloride 0.20 g
Dipotassium hydrogen
phosphate
0.35 g
A5 trace elements stock
solution
1.0 ml
Glass-distilled water 1,000 ml
General purpose media for cyanobacteria (blue green algae) :
Allen and Arnon's Medium (modified):
This medium is generally used for nitrogen-fixing cyanobacteria. If
0.20 g of potassium nitrate is added, the
medium supports the growth of many non-
nitrogen-fixing cyanobacteria.

33. Life cycle of
Cyanobacteri
a

35. Heterocysts
are the sites
for
Nitrogen
fixation.

36. Akinetes are asexual propagules which
derived from the vegetative cells. It
protects in unfavourable condition this
aids in asexual reproduction

37. Provides a buffering
microenvironment.

38. Pilli :
secretion

39. Storage products
1. Phosphate Storage:
•Polyphosphate bodies contain metals, mostly potassium,
calcium, and magnesium. Polyphosphate bodies in
heterocysts of Anabaena showed an increased content of S
and Mo and a reduction in the Ca content. The fact that
polyphosphate bodies can take up heavy metals led to the
hypothesis that polyphosphate bodies may be important in
heavy-metal accumulation.
•The enzyme involved in the synthesis of polyphosphates in
cyanobacteria is thought to be polyphosphate synthetase
(polyphosphate kinase), which catalyses the formation of
polyphosphate from ATP. No polyphosphate is required as a
primer but the enzyme requires magnesium.Polyphosphates
are broken down by alkaline polyphosphatase.

40. Storage products
2. Cyanophycin
•Nitrogen can be stored in cyanobacteria in the
form of electron-dense granules, called structured
granules, containing cyanophycin granule
polypeptide (CGP) .
•CGP consists of a simple polypeptide, composed of
arginine and aspartic acid in a 1 : 1 molar ratio and
is called multi-L-arginil-poly(L-aspartic acid), or
cyanophycin.
•Cyanophycin is unique to cyanobacteria ,but is not
found in all species.

41. 3. Phycobilin pigments:
•Phycobilisomes are composed of light-harvesting
phycobilin pigments that transfer absorbed light to
photosystem II reaction centres.
• Phycocyanin is sometimes regarded as a nitrogen
storage compound.
•It is rapidly degraded during N starvation and the
apoprotein synthesis is specifically repressed.
• However, kinetic analyses have shown that when N
is available, phycocyanin is always synthesised after
the formation of cyanophycin and when N is limiting,
phycocyanin is degraded after CGP.
Storage products

42. s
The compartmentalization of the cyanobacterial cell:The thylakoid membrane,the internal
membrane system that separates the cytoplasm from the lumen and that is present in
virtually all cyanobacteria,contains both photosynthetic and respiratory electron transport
chains. These electron transport chains intersect,and in part utilize the same components in
the membrane. Note that oxygenic photosynthesis (conversion of CO2 and water to sugars
using the energy from light) essentially is the reverse of respiration (conversion of sugars to
CO2 and water releasing energy). The cytoplasmic membrane,separating the cytoplasm from
the periplasm,contains a respiratory electron transport chain but not photosynthetic
complexes in most cyanobacteria. Therefore,in most cyanobacteria,photosynthetic
electron transport occurs solely in thylakoids, whereas respiratory electron flow takes place in
both the thylakoid and cytoplasmic membrane systems.

43. C-Phycocyanins
Absorb Green-Yellow
Light
(615-620A).
Allophycocyanins
Absorb
Orange-Red (650-
670A)
C- Phycoerythrin
Absorbs Green Light
(495-570)

44. (PS II) uses light energy to split water and to reduce the PQ pool. Electrons are
transported from the PQ pool to the cytochrome b6f complex and from there to a
soluble electron carrier on the luminal side of the thylakoid membrane. In
cyanobacteria this soluble carrier may be plastocyanin or cytochrome
c553,depending on the species and on the availability of copper (plastocyanin is a
copper containing enzyme). Either of these soluble one-electron carriers can reduce
the oxidized PS I reaction centre chlorophyll,P700 . This oxidized form of the reaction
centre chlorophyll is formed by a light-induced transfer of an electron from PS I to
ferredoxin (Fd) and eventually to NADP. Reduced NADP can be used for CO2 fixation.
Photosynthetic electron transfer leads to a proton gradient across the thylakoid
membrane. In PS II,protons are released into the lumen upon water splitting,and
protons formed upon plastoquinol oxidation by the cytochrome b6f complex are
released into the lumen as well. The proton gradient across the thylakoid membrane
is used for ATP synthesis by the ATP synthase in the thylakoid; this ATP may be
applied for CO2 fixation and for other cell processes.

46. REGULATION OF
PHOTOSYNTHESIS
• If light is abundant,the photosynthetic electron
transport chain has a much higher capacity of
electron flow than has the respiratory chain,
• but at very low light intensity or in darkness
respiratory rates are higher than those of
photosynthesis.
Phycobilisomes which are attached to the Surface of the
Thylakoids . Phycobilisomes contain Accessory
Pigments for Photosynthesis . These are Water Soluble and
are stabilized by bonds to Proteins.

47. PS I activity is abundant relative to that
of the cytochrome b6f complex

48. n

49. Conditions affect
on blooms
A juicy, yummy microbial
mat, full of cyanobacterial
goodness from
Yellowstone Park.

50. Nutrient Availability: Nutrients are a limiting factor for cyanobacteria populations. As long as the
correct nutrients are in excess, they can grow until some other factor, often light or temperature,
becomes limiting.
Competition: Ability to adapt to the environment is a big factors determining whether a bloom will
form. Many blue-greens are less edible, have gas vacuoles that help them float, can sequester
nutrients at the sediment water interface, or can fix dissolved nitrogen, any of which can give them
a competitive advantage over other algae and lead to bloom formation.
Light Intensity: Since cyanobacteria are phytoplankton, light is important and different species
thrive under different light intensities. If light is not extinguished by particles or color in the water, a
bloom is more likely. Many blue-greens thrive under low light, and so may be favored unless light is
nearly absent (such as in some high particulate reservoir systems).
Mixing: Mixing allows nutrients to be more evenly distributed and affects other aspects of water
quality that in turn affect algal abundance and composition. Mixing can also move algae to depths
with less light, limiting growth and survival. In general, blue-greens do better wtih less mixing
(Cylindrospermopsis is one taxon that seems to do well in mixed systems, though).
Temperature: Surface water temperatures consistently above 28 degrees Celsius (82 degrees
Fahrenheit) encourage blue-green blooms, although blooms may still occur in late fall (October,
November) in the Northern U.S.
Species: The above factors influence different species very differently, because each species or
taxon has a unique way of dealing with their environment. There are generalizations that apply to
blooms and blue-green dominance, but ther are exceptions in most cases. Algal bloom formation is
a complicated ecological process.
Toxicity: Not all blue-greens are toxic, so while risk may be higher during a bloom, high biomass
does not necessarily result in toxicity. Also, although many toxin producing algae produce taste and
odor compounds, the presence or absence of geosmin or MIB is not a predictor of the presence of
toxins.

51. The pH and moisture of soil and
population of cyanobacteria in
four seasons of the year

52. Effects of cyanobacteria on plant and
soil. Analysis was performed with
independent Samples t-test

53. Algalization in paddy
field
(1) Increase in soil pores with having filamentous structure and
production of adhesive substances.
(2) Excretion of growth-promoting substances such as hormones
(auxin, gibberellin), vitamins, amino acids (Roger and Reynaud
1982, Rodriguez et al. 2006).
(3) Increase in water- holding capacity through their jelly structure
(Roger and Reynaud 1982).
(4) Increase in soil biomass after their death and decomposition.
(5) Decrease in soil salinity.
(6) Preventing weeds growth.
(7) Increase in soil phosphate by excretion of organic acids (Wilson
2006)

54. Rice growth with
Cyanobacteria

55. Mishra and Pabbi (2004)
Effect of cyanobacterial
biofertilizer inoculation on rice
grain yield at a farmer’s field

56. for production at farmers’ level is not popular among
the
farming community.
The main limitations of this technology are:
•due to open air nature of production it can be produced for only a
limited period in a year (3-4 months in summer; Production has to be
stopped during rainy and winter season),
•high level of contamination due to open type of production,
•slow production rate,
• low population density and hence need for heavy inoculum
per hectare.

57. 1. The individual unit in the polyhouses can be of either
RCC, brick and mortar, or even polythene lined pits in the
ground. The algae are grown individually as species, by
inoculating separate tanks with laboratory grown pure
cultures, so as to ensure the presence of each required
strain in the final product.
2. Once fully grown, the culture is harvested, mixed with
the carrier material, presoaked overnight in water and
multani mitti (in 1:1 ratio) and sun dried. The dried
material is ground and packed in suitable size polythene
bags, sealed and stored for future use.
3. The final product contains 10,000 to 1,00,000 units or
propagules per gm of carrier material and, therefore,
500 g material is sufficient to inoculate one acre of rice
growing area.
Mishra and Pabbi (2004)

58. Luxuriant growth of Cyanobacteria
in a paddy field

59. Types of
Photobioreactor
• Tubular Reactors
–Horizontal
–Vertical
• Flat panel reactors
• Vertical column reactors
• Bubble column reactors
• Air lift reactors
• Stirred tank photobioreactors
• Immobilized bioreactors

60. Among the proposed
photobioreactors, tubular
photobioreactor is one of the most
suitable types for outdoor mass
cultures. Most outdoor tubular
photobioreactors are
usually constructed with either
glass or plastic tube and their
cultures are re-circulated either with
pump or preferably with airlift
system. They can be in form of
horizontal / serpentine, vertical near
horizontal, conical, inclined
photobioreactor.
Aeration and mixing of the cultures in
tubular photobioreactors are usually

61. Tubular photobioreactor are very suitable for outdoor
mass cultures of algae since they have large
illumination surface area. Tubular photobioreactors
consist of straight, or looped
arranged in
coiled
various ways for
transparent
maximizing
designed
capture. Properly
photobioreactors completely isolate the
tubing
sunlight
tubular
culture from potentially contaminating
environments, hence, allowing extended
external
duration
monoalgal culture.
photoinhibition is very common in outdoor tubular
photobioreactors .When a tubular photobioreactor is
scaled up by increasing the diameter of
the illumination surface to volume ratio
tubes,
would
decrease. On the other hand, the length of the tube
can be kept as short as possible while a tubular
photobioreactor is scaled up by increasing the
diameter of the tubes. In this case, the cells at the
lower part of the tube will not receive enough light
for cell growth (due to light shading effect) unless
there is a good mixing system.

62. • Prospects
Large illumination surface
area, suitable for outdoor
cultures, fairly good biomass
productivities, relatively
cheap.
• Limitations
Gradients of pH, dissolved
oxygen and CO2 along the
tubes, fouling, some degree
of wall growth, requires large
land space.

63. Vertical Bioreactor Bioreactor

64. Industrial size
Photobioreacto
r

65. Seaweed
cultivation

66.  Seaweeds are macrophytic algae, a primitive type of
plants lacking true roots, stems and leaves.
 Most seaweeds belong to one of three divisions - the
Chlorophyta (green algae), the Phaeophyta (brown
algae) and the Rhodophyta (red algae).
 There are about 900 species of green seaweed, 4000
red species and 1500 brown species found in nature.
 The green seaweeds Enteromorpha, Ulva, Caulerpa and
Codium are utilized exclusively as source of food.
Khan and Satam(2003)

67. 1) the Single Rope Floating Raft
(SRFR) technique developed by
CSMCRI is suitable for
culturing seaweeds in wide area
and greater depth.
2) A long polypropylene rope of 10
mm diameter is attached to 2
wooden stakes with 2 synthetic
fiber anchor cables and kept
afloat with synthetic floats.
3) The length of the cable is twice
the depth of the sea (3 to 4 m).
Each raft is kept afloat by
means of 25-30 floats. The
cultivation rope (1 m long x 6 m
diameter polypropylene) is
hung with the floating rope.
4) A stone is attached to the
lower end of the cultivation
rope to keep it in a vertical
position.
5) Generally 10 fragments of
Gracilaria edulis are inserted
on each rope. The distance
between two rafts is kept at 2

68. Chinese people provide fertilizer to
seaweeds
Hungry fish and animals cause
damage to seaweeds

70. INTRODUTION
A biofertilizer is a substance which contains living
microorganisms, when applied to seed, plant surfaces,
or soil, colonizes the rhizosphere or the interior of the
plant and promotes growth by increasing the supply or
availability of primary nutrients to the host plant.
Bio-fertilizers add nutrients
processes of nitrogen
through the natural
fixation, solubilizing
phosphorus, and stimulating plant growth through the
synthesis of growth-promoting substances.

71. What is Bio fertilizer?
Biofertilizers are natural fertilizers that are microbial
inoculants of bacteria, algae and fungi (separately or
in combination).
which may help biological nitrogen fixation for the
benefit of plants.
They help build up the soil micro-flora and there by
the soil health.
Biofertilizer also include organic fertilizers(manure,
etc.)
Use of bio-fertilizer is recommended for improving the
soil fertility in organic farming

72. TYPES OF BIOFERTILIZERS
Bacterial
Fungal
Algal
Aquatic fern
Earthworms
VAM fungi

73. Bacteria:
Symbiotic nitrogen fixers.
Rhizobium, Azospirillum spp Free living
nitrogen fixers.
Azotobacter, Klebsiella etc.,
Algal biofertilizers:
BGA in association with Azolla
Anabena, Nostoc, Ocillatoria
Phosphate solubilising bacteria:
Pseudomonas, Bacillus megaterium
Fungal biofertilizer
VAM
Earthworms

74. Bacterial biofertilizers
The live cells of bacteria used as a biofertilizers
These microbes contains unique gene called as
Nif-Gene which make them capable of fixing
nitrogen.
The nitrogen fixing bacteria work under two conditions,
Symbiotically
Free living bacteria (non-symbiotic).
The symbiotic bacteria make an association with crop plants
through forming nodules in their roots.
The free living bacteria do not form any association but live
freely and fix atmospheric nitrogen.

75. Symbiotic nitrogen fixers.
Most important symbiotic Nitrogen fixing bacteria is
Rhizobium and Azospirillum.
Rhizobium:
Rhizobium lives in the root hairs of the legumes by
forming nodules
Plant root supply essential minerals and newly synthesized
substance to the bacteria
The name Rhizobium was established by Frank in 1889.
This genus has seven distinct species based on "Cross
Inoculation Group Concept".

76.  More than twenty cross-inoculations groups have been
established.
 A new classification has been established for Rhizobium.
 That is 'slow growing rhizobia' known as Bradyrhizobium
and the other group is 'fast growing rhizobia' called
Rhizobium.
 Rhizobium can fix 50-300 kg/ha
Rhizobium

77. Azospirillum:
It mainly present in cereal plants.
inhabits both root cells as well as surrounding of roots
forming symbiotic relation and increasing nitrogen fixing
potential of the cereal plant.
Azospirillum is recognized as a dominant soil microbe
nitrogen in the range of 20- 40 kg/ha in the rhizosphere in
non-leguminous plants such as cereals, millets, Oilseeds,
cotton etc.
Considerable quantity of nitrogen fertilizer up to 25-30 %
can be saved by the use of Azospirillum inoculant.
These species have been commercially exploited for the use
as nitrogen supplying Bio-Fertilizers.

78. Free living bacteria
Large number of free living or non -symbiotic bacteria (does
not form nodules but makes association by living in the
rhizosphere) present in soil.
Commonly used free living bacteria are
Azotobacter Klebsiella
it will not associated with plant.
Azotobacter is a biofertilizer which provides the required
amount of nitrogen to the plant from the soil.

79. Azotobactor
Azotobactor is a heterotrophic free living nitrogen fixing
bacteria present in alkaline and neutral soils.
Azotobactor is the most commonly occurring species in
arable soils of India.
Apart from its ability to fix atmospheric nitrogen in soils, it
can also synthesize growth promoting substances such as
auxins and gibberellins and also to some extent the
vitamins.

80. Many strains of Azotobactor also exhibit fungicidal
properties against certain species of fungus.
Response of Azotobactor has been seen in rice, maize,
cotton, sugarcane, pearl millet, vegetable and some
plantation crops.
It improves seed germination and plant growth.
Azotobacter is heaviest breathing organism
and requires a large amount of organic
carbon for its growth.

81. Mass production
isolated bacterial cultures were subculture in to nutrient
broth
The cultures were grown under shaking condition at 30±2°C
The culture incubated until it reaches maximum cell
population of 10¹º to 10¹¹
Under optimum condition this population level could be
attained within 4-5 days for Rhizobium 5-7 days for
Azospirillum and 6-7 days for Azotobacter.
The culture obtained in the flask is called Starter culture
For large scale production , inoculum from starter culture is
transferred in to large flasks / fermentor and grown until
required level of cell count is reached

82. prepare appropriate media for specific to bacterial
inoculant in required quantity
Inoculated with specific bacterial strain for aseptic
condition Incubated at 30±2ºC for 5-7 days in rotary
shaker
Observe growth of the culture and estimate the
population
( starter culture)
The above the media is prepared in large quantities in

83. Sterilized and cooled well
Media in a fermentor is inoculated with the log phase of culture
grown in large flask (usually 1-2 % of inoculum is sufficient)
cells are grown in fermentor by providing aeration & continuous
stirring
Broth is checked for the population of inoculated organisms
Cells are harvested with the population load of 109 cells/ml

84. Carrier material
the use of ideal carrier material is necessary for the
production of god quality of biofertilizer
Peat soil, lignite, vermiculture, charcoal, press mud,
farmyard manure and soil mixture are used as a carrier
materials
Neutralized peat soil/lignite are found to be better carrier
materials
Ideal carrier material should be
Cheaper in cost
Locally available
High organic matter content
No toxic chemical
Water holding capacity of more than 50%
Easy to process

85. Preparation of inoculants packet
Neutralized and sterilized carrier material is spread in a
clean, dry, sterile metallic or plastic
Bacterial culture drawn from the fermentor is added to the
sterilized carrier and mixed well by manual or mechanical
mixer
Inoculants are packed in a polythene bags sealed with
electric sealer

86. Specification of the polythene bags
 Polythene bags should be of low density grade
 Thickness of bag should be around 50-75 micron
 Packet should be marked with the
 Name of the manufacture
 Name of the product
 Strain number
 The crops to which recommended
 Method of inoculation
 Date of manufacture
 Batch number
 Date of expiry
 Price
 Full address
 storage instruction

87. Vesicular Arbuscular Mycorrhiza
(VAM)
The term mycorrhiza was taken from Greek language
meaning
'fungus root'.term was coined by Frank in 1885
The mycorrhiza is a mutualistic association between fungal
mycelia and plant roots.
VAM is an endotrophic (live inside) mycorrhiza formed by
aseptated phycomycetous fungi.
VAM help in nutrient transfer mainly of phosphorus, zinc
and sulfur.

88. Mycorrhizae is the symbiotic association between plant
roots and soil fungus of the 7 types of mycorrhizae,
VAM plays a great role in inducing plant growth.
VAM are symbiotic entophytic soil fungi, which colonize
the roots of approximately 80% plants.
The VAM hyphae also help is retaining moisture around the
root zone of plants
It increases the resistance to root borne or soil borne
pathogens and Nematodes.

89. They also mobilize different nutrients like Cu(copper),
K(potassium), Al(aluminum), Mn(manganese), Fe
(iron)and Mg (magnesium) from the soil to the plant roots.
They posses vesicles (sac like structure) for storage of
nutrients and arbuscular for funneling them into root
system.
Morphology
External hyphae
Arbuscles
Vesicles
External hyphae vesicles
Arbuscles

90. Mechanism of Action
The VAM forms an association with plant roots.
It penetrates in the root cortex and
spreads aroundthe roots of the plant.
As the name indicates, they posses sac like structure called
vesicules which stores phosphorus as phospholipids.
The other structure called arbuscule helps
bringing the distant nutrients to the vesicules
and root.

91. Mass production
VAM spores isolated
Spores mixed with sterilized soil
Soil filled in pots
Host plant transplanted in pots
Kept 3-4 months in green house

92. Soil in the pot along with roots of host
plant is macerated
Dried till it attains 5% moisture
Dried soil inoculants used for field
application

93. Uses of V
AM
Enhances the feeding areas of the plant root is as the
hyphae spreads around the roots.
Mobilizes the nutrients from distantance to root. Stores the
nutrients (sp. phosphorus).
Removes the toxic chemicals (example : phenolics) which
otherwise hinder nutrient availability.
Provide protection against other fungi and nematodes
It increase growth rate in plants (citrus, maize, wheat, etc.)
It reduces sensitivity of crop towards
high level of salts and heavy metals

94. Algae as a biofertilizer
Another group of free living nitrogen fixers are cyanobacteria.
Commonly called as Blue green algae.
More than 100 species of BGA can fix nitrogen. Nitrogen
fixation takes place in specialized cells called ‘Heterocyst’
BGA very common in rice field.
Unlike Azotobacter BGA are not inhibited by the presence of
chemical fertilizers.
No chemical fertilizers added, inoculation of the algae can
result in 10-14% increase in crop yields.

95. They are easy to produce
Usually they are mass produced in cement tanks filled with
fresh water.
Not require any processing Quite and cheap
Cost of 10kg may be Rs.30-40 only
Beneficial in certain crops like vegetables, cotton,
sugarcane.
Eg. of some algal biofertilizers are
Anabena
Nostoc
Oscillatoria

96. Azolla is a tiny fresh water fern common in ponds, ditches and
rice fields.
It has been used as a biofertilizer for a rice in all major rice
growing countries including India, Thailand, Korea, Philippines,
Brazil and West Africa.
The nitrogen fixing work is accomplished by the symbiotic
relationship between the fern and BGA, Anabena azollae.
In addition to nitrogen the decomposed Azolla also provides K,
P, Zn and Fe to the crop
Azolla as a bio fertilizer

97. Azolla biomass gets doubled within 5-7 days by vegetative
methods.
fix 40-80 kg nitrogen / ha / year. good manure for flooded
rice.
Increase of crop yield up to 15-20% has been observed while
fertilizing the rice with Azolla
Hybrids are growing faster Tolerant to heat and cold Fix 4-
5% more nitrogen

98. Bio - fertilizers application methods
There are three ways of using these N-fixing/P.S.M. bacteria.
Seed treatment
Root dipping Soil applications

99. Seed Treatment
Seed treatment is a most common method adopted for all
types of inoculant. The seed treatment is effective and
economic.
Seed treatment with Rhizobium, Azotobacter, Azospirillum
along with P.S.M.
seed treatment can be done with any of two or more
bacteria.
no side effect.
important things has the seeds must be coated first with
Rhizobium or Azotobacter or Azospirillum when each seeds
get a layer of above bacteria then the P.S.M. inoculant has to
be treated on outer layer of the seeds.

100. This method will provide maximum number of population
of each bacteria required for better results.
Mixing the any of two bacteria and the treatment of seed
will not provide maximum number of bacteria of
individuals.

101. Root dipping
Application of Azospirillum with the paddy/vegetable plants
this method is needed.
The required quantity of Azospirillum has to be mixed with
5-10 ltr of water at one corner of the field and all the plants
have to kept for minimum ½ an hour before sowing .

102. Soil application
P.S.M. has to be used as a soil application use 2 kgs of
P.S.M. per acre. Mix P.S.M. with 400 to 600 kgs of
Cowdung along with ½ bag of rock phosphate if available.
The mixture of P.S.M., Cowdung and rock phosphate have
to be kept under any tree shade or celling for over night and
maintain 50% moisture.
Use the mixture as a soil application in rows or during
leveling of soil.

103. Precautions
Store biofertilizer packets in cool and dry place away from
direct sunlight and heat.
Use right combination of biofertilizers
Rhizobium is crop specific, so use in specified crop Do not
mix with chemicals
Use the packet before expiry, only on the specified crop, by
the recommended method.

104. Advantage of biofertilizers
 Renewable source of nutrients Sustain soil health
 Supplement chemical fertilizers. Replace 25-30%
chemical fertilizers Increase the grain yields by 10-
40%.
 Decompose plant residues, and stabilize C:N ratio
of soil Improve texture, structure and water holding
capacity of soil No adverse effect on plant growth
and soil fertility.

105. Stimulates plant growth by secreting growth hormones.
Secrete fungistatic and antibiotic like substances
Solubilize and mobilize nutrients
Eco-friendly, non-pollutants and cost effective method

106. Disadvantages
Biofertilizers require special care for long-term storage
because they are alive.
must be used before their expiry date.
If other microorganisms contaminate the carrier medium or
if growers use the wrong strain, they are not as effective.
Biofertilizers lose their effectiveness if the soil is too hot or
dry.