INTRODUCTION

INTRODUCTION

PART – I

An organism is always in the state of perfect balance with the environment.

The environment refers to various conditions surrounding an organism which

directly or indirectly influence the life and development of the organism and its

population. In an ecosystem a basic functional unit called “Organism” interact

with the environment and other chain of organisms various physico chemical and

climatic conditions.

Water is important natural resource for the survival of living beings. Water

reservoirs, lakes, dams, ponds, rivers serve as natural habitat and help to maintain

ecological balance of different kinds microbes in natural habitats in water. In

water bodies various organism act together and allows continuous recycling of

each chemical element available in the system. When this stops for some

reason, pollution results and pollution pushes the environment out of balance and

the scheme of nature is that it will react to reestablish the balance Dugan (1974)

In biological literature, this nine letter dirty word pollution has several

connotations; to biologists, it can presage the development of conditions adverse

to the continued existence of certain types of microbes inhabiting in a body water.

In Inland water study of lakes, ponds or dams usually seem to receive

nutrients from bottom sediments. Interaction between sediments and over water

mass usually govern the productivity potential of water body. Human beings by

their anthropogenic activity are making fresh water as dumping grounds for

receiving solid and liquid waste from nearby human settlement. Eighty percent of

water supply of cities find its way to drainage system as domestic and industrial

waste. Most of the precipitation take place during rainy season which contributes

substantially to the surface flow. During this period of time a heavy inflow of water

results in to the natural aquatic systems and result in to exposing benthic substrata

in to main chain.

Lakes are locked up systems and basin soil plays a predominant role in

determining water quality. In tropical reservoirs phosphate level in water usually

govern ecology of lakes. Usually there is a quick recycling and rapid turnover of

nutrients in lakes, Ehrligh (1960); Abbot (1967). Plankton by virtue of their

drifting habit and short turnover period, constitute the major link in the trophic

structure and events in the reservoir ecosystem. A rich plankton community with

well marked seral succession is hallmark of Indian reservoirs.

In water bodies, it is the myriad of diverse organism acting in convert that

allows continuous recycling of each chemical element available in the system.

When this stops for some reasons, pollution results, which pushes the

environment out of balance, and the scheme of nature is that it will react of

reestablish the balance (Dugan, 1974).

Microbes, though they are intricate in their detail offer examples of changes

in molecular structures, metabolic pathways or structural elements which are often

taken as full time monitors of this process. As pollutional stress is increased on

the system, a number of ambient species are usually eliminated or reduced in

number while the other native tolerant species become even move dominant thus

widening the range of individual per species (Cairns and Lanza, 1972). It is then

likely that any possible permutation or combination of nutrient factors may be

involved in eliciting microbial growth in water, depending upon other environmental

influence. And, eutrophic water have one of the most important problems in the

progressive enrichment of water with nutrient concomitant with mass production of

microbes, increased water productivity and other undesirable biotic changes.

Such a progressive deterioration of water is regarded as eutrophication Cairns and

Lanza (1972).

The term “Water bloom” is widely understood to refer to the accumulation of

planktonic algae and microbes at the surface of lakes and reservoirs. Although

the constituent algae and microbes are themselves microscopic in size, the

intensity of scums and rapidity with which they develop are such as to impress the

most casual observer; blooms can form and disperse again within a matter of

hours, Reynolds and Walsby (1975). The great sometimes cataclysmic

productivity phenomenon is world ide and described by various synonyms. The

flowering, breaking or working of water, Hutchinson (1967); Wasserbluthe, flos

aque Whipple (1899); Pea soup Nemerow (1974); ‘Tsvetenie Vody’ the Russian

equivalent; Topachevski (1968). In temperate regions, blooms may develop most

frequently during calm weather in summer and autumn; in tropics they can form

bloomsat almost any time of the year, Reynolds and Walsby (1975). In some

countries additional algal and microbial bloom is welcomed since directly or

indirectly it may be beneficial to freshwater fisheries, Prowse (1964). Algae from

blooms mainly Spirulina SP are also consumed directly by man in Chad, Pirie

(1969).

In western countries and in India, however, algal blooms are usually

regarded as being objectionable. They are plentifully expensive nuisance in water

supplies where the algae present problems of filtration, and many species taint

water with unpleasant taste and adours. Blooms also spoil fishing and water

sports in recreational lakes. Some bloom forming algae are toxic and have

been implicated in many instances of fish kills, Gorham (1964); Chaeko and

Ganapati (1949) ; Venkatraman et al., (1957) ; Boyd et al., (1975) ; Barica (1975)

and remain fatal to domestic animals Gorham (1964).

Growing concern over problems caused by water blooms is one of the

factors which has prompted a great deal of research in to biology of algae and

microbes. The tropholytic zone has a steady supply of free carbon dioxide, which

reacts with carbonates to produce bicarbonates. This results in an increase of

bicarbonates towards the bottom.

Similarly, due to the increase in the hydrogen ions, the pH drops rapidly.

Thus the increase in total alkalinity, specific conductivity and CO2 decrease in pH

value occur towards the bottom layers and these Laxers acts as useful indicators

of productivity. Blue-green algae from the main stay of plankton community in

vast majority of the manmade lakes are studied. The overwhelming presence of

Microcystis aeruginosa in Indian reservois is remarkable. The productive water

of the Gangetic plains, Deccan Plateau, South Tamil Nadu and Orrisa invariably

have good standing crops of Microcystis. A common feature of all these

reservoirs is the bright sunshine, isothermal water column, klinogracde oxygen

curve and an extensive Eatchment area, draining calcium high from forested or

cultivated lands. The species are almost omnipresent in the southern peninsula,

except in the reservoir of Karnataka and Kerala which tend to be oligotrophic and

have poor plankton count with desmids and other green algae as the main

constituents. Reservoirs of Rajasthan receiving scanty rainfall and poor flushing

rate favour macrophytes and despite being productive they do not harbour blooms

of Microcystis. The oligotrophic lakes of the North-East have a desmid dominant

plankton community. Most of the reservoirs have three plankton pulse coinciding

with the post – monsoon (Sep to Nov.), winter (Dec to Feb) and summer (March to

May) seasons. The monsoon (Jun, August) flushing disturbs and often dislodges

the standing crop of plankton. When using obiotic measures alone, a pulse after

a storm for instance, may not be detected. This is because storm pulse is

relatively quick a phenomenon and an observer may not be present to witness the

disturbance occurring. However, this same disturbance would be directly

reflected in the biotic community and therefore detectable at a later date through

use of an appropriate biomonitoring programme. When studying the ebb and flow

of life of life in lakes, it is natural to know what kinds of organism are present, how

the importance of each organism fluctuate in relation to that of others and how

much living matter is produced in a given time, while other pertinent questions

which follow regarding abundance of some organism and not others ‘ what

controls the succession of dominant species ‘ what determines the fertility of lakes.

Some of the more recent observational studies centered round the sequence of

events leading up to bloom, formation, and there has been much speculation on

the factors promoting the growth of bloom forming populations.

It is impossible to understand the ecology of microbes in a lake without the

knowledge of physico – chemical properties of water. A comprehensive

biomonitoring process involves both physicochemical and biological approach and

gives the exact status of the aquatic ecosystem. Lund (1967) established the fact

that year round seasonal studies of water body is an important parameter for

understanding any lake. The best way to understand life in water is to consider

the forms that occur in them studies separately and such a consideration will give

a better picture of plankton distribution and periodicity of biosystem and

organisms. Planktonic microbes requirement is different from order to order and

these groups tend to have different ecological preferences.

Aquatic ecology of Microbes

Major factors that influence the aquatic life could be light, temperature, and

chemical composition of water. Planktons growth of microbes in lake may

additionally be subjected to the direct effect of water movement, chemical

inhabitant, anthoropogenic activity and many myriad factors. Plankton provides a

relevant and a convenient point of focus for research into the mechanism of

eutrophication and for development of measure of control its adverse impacts on

ecosystem.

The term “Plankton” refers to those minute aquatic forms which are non

motile or insufficiently motile to overcome the transport by currents and living

suspended in open or pelagic water. The planktonic plants are called

Phytoplankton and planktonic animals are called Zooplankton. (APHA -1985)

PHYSICO-CHEMICAL FACTORS

Reservoir, lakes and ponds are aquatic ecosystem, their biotope reveals

that they have certain characteristic feature of its own Chemical characteristic of

the water of a reservoir vary with its age, Jhingran (1977) and lakes distributed

even with in a small geographic area remain limnologically different, Gulati (1976) ;

Barica (1978). Different lakes show several pattern and shift in eutrophic levels

siltation in mansoon months from anthropogenic sources also is important for

degradation of logic system in India. As a result a considerable part of standing

crop of biotic communities at upper trophic level gets disturbed. Physico-chemical

factors like lights depth, O2, C2, PH, alkalinity and mineral concentration are

important major parameters of study which influence, the life in water, Micheal and

Sharma (1988).

Basin also play an important role in determining the chemical water quality

through soil – water interface. In reservoirs, nutrient input from allochthonous

sources often determine the water quality, nutrient regime and basic production

potential.

Most of the Indian reservoirs are characterized by low levels of Phosphate

and Nitrates. Phosphates usually range from 0.1 mg/L in reservoirs. 4 to 13 mg/l

phosphate was found in Manasarovar in Madhya Pradesh Low Nitrate Nitrogen in

Indian lake do not usually control productivity. In Kankaria and Naroda lakes in

Ahmedabad, phosphate was present either very low or absent, though both the

lakes recorded eutrophic conditions, Uttarwar (1980).

Rapid turn over of nutrients and quick recycling of various nutrients in water

lakes place. Hedge and Philips (1958) showed that 90 % of the phosphorous was

taken up by phytoplankton within 20 minutes. Abbot (1967) reported rapid turn

over of nutrients in lakes. Phosphates and Nitrates form the part of total dissolved

solid and reflects chemical condition of the reservoirs Khan and Zutschi (1978)

Suggested that temperature governs productivity in water. Konopka and Brock

(1978); Philipose (1960), recorded blue green algal dominance with the rise of

water temperature. There is a clear cut evidence from literature of interaction

existed between temperature and low light intensity from photosynthesis of natural

population and occurrence of effective photosynthesis in lake water. This

represents true adaptation to changing irradiance and was reported by Roger

(1978). Physico Chemical data of more than 100 reservoirs in the country

concluded that morphometric, edaphic and water quality parameters can not be

used as benchmark to predict organic productivity in the lake. However, each

reservoir ecology is determined by a variety of factors and may define the tropical

status of the lake. Bacterial decomposition of organic matter may reflect rate and

consumption of oxygen. Increased in DO content defines high rate of

photosynthesis through out the water column. Almost all productive reservoirs of

the country irrespective of their geographic location depict a klinograde Oxygen

curve.

In lake there is a system of vertical stratification of oxygen along with other

chemical substances and PH. Parameters like pH, CO2, alkalinity and specific

conductivity react to this situation. In upper photolytic zones constant

photosynthesis takes place as a result free CO2 is released.

Free CO2 react with carbonate and produce bicarbonates. These

bicarbonates get deposited on to the bottom and increase Hydrogen Ion

concentrations and in this processes pH drops towards acidic scale. These

reactions together with specific conductivity and CO2 decrease pH values in the

bottom layers and act as useful indicator of productivity.

Higher rainfall in monsoon brings about heavy discharges of water in the

reservoirs and retard macrophyte community growth. In the lack of suitable

available substratum grown of periphyton is discouraged. On the other hand,

planktons because of their drifting habit and short life span proliferate to such an

extent that every suitable environment niche is utilized this Indian reservoirs and

lakes maintain a succession of plankton community. The parameters like bright

sunshine, isothermal water column, klinograde oxygen, curve, extensive

catchments drain enrich earth metals like Ca, mg from forest and cultivated land

thus a complex phenomenon occur in rained catchments reservoirs and lakes.

Any loss of organic matter from healthy organism before they are

consumed is a loss from food chain. It seems unlikely that the organization of the

each of any organism can be such as to prevent entirely the escape of organism

substances formed within them into the surrounding medium. The terms

“extracellular release” or “excretion” are used to describe the loss of soluble

organic compounds by healthy cells of microorganisms. The definition attempts

to separate “true” extracellular substances from organic compounds released by

moribund organisms or liberated during cell lysis Nalewajko, 1977.

Extracellular release appears to be a widespread phenomenon in

microorganisms. The production of great variety of extracellular substances by

various organism is now well established. It is also clear that such substance

often play important roles in growth and physiology of organisms, as well as

aquatic food chains and ecosystems in general.

There is little disagreement that dead micro-organism and their residue

form further contribution along the food chain and contribute appreciable amount

to the non-living organic pool in water. Through degradation organic matter from

these sources probably eventually becomes part of the “dissolved” organic matter,

(Sharp 1975), which are readily utilized by bacteria, Wright, (1970). The release

of extracellular substance often referred as liberation, or excretion.

CHAPTER II

REVIEW OF LITERATURE

Any possible permutation or combination of nutrient factors may be involved

in eliciting algal growth in waters depending upon other environmental influences

and eutrophic water have one of the most important problems in progressive

enrichment of water with nutrient concomitant with mass production of algae,

increased water productivity and other undesirable biotic changes. Such a

progressive deterioration of water is regarded as eutrophication (Cairns and Lanza

1972). When for fiscal or logistic reasons, excess nutrients can not be eliminated

from freshwater lakes, certain species of phytoplankton grow to excess. The term

“Water bloom” is widely understood to refer to the accumulation of planktonic

algae at the surface of lakes and reservoirs. Although the constituent algae are

themselves microscopic in size, the intensity of scums and rapidity with which they

develop are such as to impress the most causal observer. Blooms can form and

disperse again within a matter of hours (Reynolds and Walsby 1975).

Algae are plentifully expensive nuisance in water supplies where algae

present problems of filtration and many species taint the water with unpleasant

taste and odours. Blooms also spoil fishing and water sports in recreational

lakes.

When studying the ebb and flow of life in lakes, it is natural to know what

kinds of organisms are present, how the importance of each organism fluctuates in

relation to that of others and how much living matter is produced in a given time,

while other pertinent questions which follows regarding abundance of some

organisms and not others, what controls the succession of dominant species ;

what determines the fertility of lakes. Some of the more recent observational

studies centered round the sequence of events leading upto bloom formation and

there has been much speculation on the factors promoting the growth of bloom

forming populations. The sequence of algae in aquatic environment is well

documented ; the general spring bloom of diatoms, the early summer growth of

green algae and blue green algal blooms in lake summer (Hutchison, 1967). Some

attempts in literature have also been made to give more general comments on

ecology of individual genera and species. While some species remain planktonic

thought out the winter others, may be largely or entirely restricted to the bottom of

a lake.

The various genera may differ also in their behaviors and the time of

maximum population density near the surface of a lake. Most important of

freshwater plankton are probably the members of Bacillariophyceae or diatoms,

and most significant diatom being centric diatom. Among many features of

planktonic algae requiring investigation, a few may be stressed. It is clear that a

variety of changes in the environment may tend to favour the growth of planktonic

forms.

Fresh water resources are the integral parts of mans’ life. These resources

have received mans’ attraction and attention for various activities like fisheries,

irrigation, navigation, transportation, recreation and daily consumption. In past

years organization like IBP, IUCN, UNESCO have devoted their attention on

various aspects of these fresh water resources and their ecology.

F.A. Forel (1841-1912) of Switzerland is considered as father of limnology

for his work on Swiss lake and published three volumes entitled. “Le Lemnase”,

on Lake Geneva. During 1888-1909 he studied physic chemical, biological

characteristics of lake. In 1901 his first book on limnology was published under

the heading of “Handbunch der seankund allagenmiene”. British researchers

extensively worked on fresh water fauna, Hamilton (1822); Day (1973) and Braun

(1849) algal flora

In Asia sewage ponds have been known from centuries but data on their

performance is available for last fifty years now. In rural areas waste stabilization

ponds are made first of its kinds was a pond for Madras University Campus,

Tamilnadu built in 1957 Jhingaran (1982). In the beginning of nineteenth century,

Uturney, Prasad, Bhutia, Arora, Bond, Anderson et al. Brehm, Tewell, Hauer,

Kiefer and Donner described the fresh water species in the Indian Sub Continent

(cf.Michael 1980). Prasad studied the seasonal variation of pond organisms and

therefore was perhaps the first limnologival work in India, Nasar et al., (1982) ;

Patil (1476) Ganpati and Sreenivasan (1968, 1976) worked on almost all

reservoirs in South India for fishery and waste water treatment. Ganpati (1959)

has published a review on ecology of tropical water. Sreenivasan carried out

immense limnological investigation in South India in 1972, 1976, 1977 covered

and various aspects of limnology and biological productivity on man made and

natural lakes in South India.

Hydrobiological study in relation to various aspects of Rajasthan was

carried out by Jakhar et al., (1990) and Hazarika (1994) studied water bodies of

Assam. Michal (1969) has worked on various aspects of limnology of a natural fish

pond in Barrackpore. Ganpati (1955) carried out systematic observation on

ecology of tropical water. Uttarwar (1980) surveyed two lakes Kankaria and

Naroda lakes in Ahmedabad to define of the trophic status of the lakes. Important

contribution of India freshwater was made by Zafar (1967, 1968 a.b) ; Verma

(1967, 1969); Vashisht (1968) ; Vashisht and Dhir (1970) ; Kaul (1977) ; Vass

(1970) ; Munawar (1970) ; Sreenivasan (1970, 1976) ; Unni (1983, 1985) ; Adoni

(1985 ) ; Seenaya and Zafar (1979) ; Awtramani (1980) ; Sehgal (1983) ; Jayadevi

(1985) ; Rao et al., (1995) ; Chandrashekhar and Kodarkar (1994) ; Kodarkar

(1994) ; Jayachandra et al., (1995) ; Choubey (1997). Many researchers have

carried out studies on physico chemical and biological characteristics of river and

dam. Trivedi and Goel (1986) ; Saxena (1990); Patil and Sanbag (1993) ; Shaikh

and Yeragi (2004) ; Kodarkar (1994) ; Kulkarni and Rao et al., (2002) ; Jakher and

Rawat (2003) ; Jain and Seethapati (1996) ; Reddy et al.,(1994) are few to

mention in the literature.

Notable contribution in limnology and algal ecology was made by following

authors Das and Srivastva (1956, 1959); Chakraborthy and Singh (1959) ; George

(1966) ; Gulati (1964) ; Khan and Siddiqui (1966, 1971); Kant and Kachroo

(1971) ; Tandon and Singh (1972) ; Bayade and Verma (1985); Singh and

Mahajan (1987) ; Malik and Bose (1987) ; Gosh and George (1989). Notable work

on eutrophication of lakes was reported by Hasler (1947) ; Valbentyne (1957) ;

Vollenweider (1968) ; Palmer (1969) ; Morgan (1972) ; Zutshi et al., (1973) ; Kaul

(1977) ; Madhusudan et all., (1984) ; Goel et al., (1985) ; Davis (1986) ; Trivedi

(1988) ; Parshbey (2003) ; Sharma et al ., (2009); Kumawat and Javale (2003) ;

Kumar Hegde (2005) ; Yeole and Patil (2005) ; Chavhan et al., (2006).

Various hydrobiological aspects of water bodies was carried out by Munwar

(1970); Chandrashekhar and Kondarkar (1994) ; Roy and Singh Rajni (1999) ;

Singh (2002); Lande (2004) ; and Malik et al., (2004); Zafar (1964) ; Aboo and

Mancel (1967) ; Kunt and Kachroo (1975) ; Shashikant and Kachroo (1977) ;

Himagauri et al., (1987) ; Zafar (1991) ; Pandey et al., (1992) ; Kadam (2000) ;

Rao and Shrivastava (2002) ; Jakher and Rawat (2003) ; Patil and Talmale

(2004) ; Pande and Varma (2004) Lal (1996) carried out studies on effect of mass

bathing on water quality in Pushkar lake. Jana and Sarkar (1982) ; and Islam

(1990) reported effect of high alkalinity and greater productivity in lakes, they also

reported effect of high alkalinity and greater productivity in lakes, they reported

higher hardness in correlation with bicarbonate in aquatic systems.

Roger (1979) suggested occurrence of increased algal sporulation as

responsible for higher pH counts. Planktonic abundance and coexistence of

species depend upon individual cell size and was reported by Tsimul Skaya and

Galazachve (1978). Sreenivasan (1969) reported presence of high nitrogen and

organic carbon content in Amaravathy as significant with context to its productivity.

Wood and Gibson (1973) concluded Lough Neagh as world’s most eutrophic basin

where P remained as critical nutrient. Parr and Smith (1976) on Lough Neagh

recognized both P and N being important for prolonged algal growth. Bishwas

and Cadler (1955) recognized importance of bottom zone as vital for inhibiting

active decomposers.

In Gujarat, Ganpati and Pathak (1972) reported results on Sayaji Sarovar

and Ajwa reservoir, Baroda;Pandey and Kaul (1976) on Lalpari lake, Rajkot;

Gandhi (1964) surveyed diatoms of Ahmedabad and Gupte (1961) studied

seasonal variation in Chandola and Malkhani Talav in Ahmedabad and North

Gujrat. Uttarwar 1980 worked on Kankaria and Naroda Lakes in afremdalaxd.

Low pH and low concentration of DO usually favour growth of Myxophyceae

Rao (1953), reported higher DO record lower growth of blue green algae in

summer months. With high temperature in summer months blue green are

favored, Zafar (1967). Singh (1960) attributed CO2 in water as responsible for

production of blue greens. They reported favoring blue greens with higher O2 and

CO2 values. Microcystis aeruginosa usually form permanent blooms in polluted

water when temperature, sunshine, CO2, Phosphate, albuminoid ammonia was on

higher scale in domestically polluted water. Ganpati (1943) recorded still higher

number of Spirelina nordstedtii along with presence of few Phormidium fragile.

Flagellates

Various investigators have reported existence of a close relationship

between organism and water bodies. Lakes in general have lower CO2 and large

lakes recorded higher values of flagellates and CO2 during summer months. DO

effect presence or absence of warm water. Seenaya (1971) reported existence

higher O2 and higher blooms. Zafar (1958) reported the importance of Carbon and

flagllates in water than availability of nitrogen. Water temperature is another

factor which effect the presence of flagellates.

Myxophyceae

This is a ubiquitous group found in water with various specific factors like

pH temperature, O2, CO2, Nitrate, Phosphate, Organic matter control the growth

of mycrophycads in tropical water. Sreenivasan (1970) stated that organic matter

plays a deciding role in the formation of blooms. When nitrates and phosphate

are low Mycrophyceae algae increase, Munwar (1970). Munwar considered

calcium as important factor for the growth of algae. However, there are other

reports of presence of higher calcium and lower Myxophycean algae. Blue green

are observed more in number in small ponds alongwith presence of Spirulina and

Phormidium.

There is generally a slight liberation of such substances from healthy cell of

various species of chlorophyceae, whereas, in growing cultures of species of

Myxophyceae a considerable proportion of the total organic matter synthesized

regularly appears in a soluble form in the medium (Fogg and Wastlake, 1952).

Even extracellular products form only a small part of the total material synthesized

by an alga it is nevertheless possible that the substance concerned may be

biological active at low concentrations in their microenvironments. Oligotrophic

waters are characterized by higher percent of extracellular release (PER)

(Nalewajko, 1974). It is important to note, however, that the amounts of carbon

excreted and the rates of excretion are far more useful parameters. A significant

fraction of the carbon and nitrogen fixed by blue – green algae is subsequently

excreted into the environment. Saunders (1972) showed that phytophankton

generally release higher molecular weight organic substances rather than simple

sugars, amino acids, and other simple substances. Growth hormones and vitamins

have been reported as constituents of cell exudated of many algae, but the reports

are limited. Bentley 1958, reported auxin like substances in Oscillatoria and other

algae. He also detected water soluble ether insoluble auxins or auxin precursors.

Zutshi and Vass (1978) reported Dal lake water as alkaline, slightly buffered

and remained at a higher trophic status. Such marked alkalinity is brought about

by planktonic algae is also documented by Talling (1976) and alkalinity and high

planktons count was reported by Russo (1978) Selected aspects of hydrobiology

of Pulicat lake was studied by Chako et al., 1953) and Chilka lake was studied by

Ramananthan et al., (1964). Sreenivasana (1977/78) reviewed preliminary

studies of two narrow deep reservoirs which hold poor nutrient conditions.

Results of Suraj Kund, U.P. was reported by Sarkar and Rai (1964) and that of

Govindsagar reservoir, Himachal Pradesh was reported by Sarkar et al (1977).

Bohr (1975) showed presence of a close relationship between temperature, pH

and dissolved oxygen in two Jodhpur (Rajasthan) reservoirs. Bhargawa and Alum

(1979) reported existence of seasonal succession of diatoms in relation to salinity

and alkalinity in Sambhar lake (Rajasthan). Sreenivasan (1969) found nitrate and

phosphorus lacking or occurring only in traces in South India reservoirs.

Phytoplankton

Phytoplankton taxa was observed during sometime of the year or the other.

Oscillatoria Microcystis, Chlorococus, Merimosopedia, Nostoc, were observed in

lake water. Desmids like Scenedesmus, Tetraedron, Crucigenia, Cosmarium

were seen in the lake water. List of species found in during study period is given in

separate table. However the present work was not centered on algae studies,

therefore a small list in enclosed. Summer blue green algae dominance was noted

in this lake water, during winter desmids were recorded. A probable list of algae

was recorded in the table.

Sr. No. Name of phytoplanktons

1. Anabaena

2. Chorella

3. Closterium

4. Diatoms

5. Euglena

6. Hydrodictyon

7. Microcystis

8. Merismopedia

9. Spirogyra

10. Scendesmus

11. Volvox

12. Vaucheria

13. Zygnema

CHAPTER III

MATERIALS AND METHODS

Map and Chart

DESCRIPATION OF LOCATION AND STUDY SITE

Geographically this place is located 22 km of Pandharkawada Yavatmal

towards South (Maharashtra). This place is located on 79.1 latitude 945

longitude. This area witnesses as nearly as 52 rainy days with nearly 100 mm

rainfall. This place is nearly 35 M.S.L. and climate is dry and hot. The lake is

spread on 10 areas of land with 25 to 30 feet deep. This is a perennial lake.

Population of village is around 5500. Every year temple organized fain in the

month of November of Kartik Shudha full moon also this temple is visited by

pilgrims on every full moon day. Nearly 1,50,000 people visit the site and

celebrate the fair for 5 days. On every full moon days thousand of people visit

the lake village. This lake has no attested history. There is no inlet and out let

end water in mainly from rainy source. People wash their cloths regularly on the

banks of lake and animal always wade through water. Thus there is a big load of

pollution as pollutants reach the water. There are aquatic weeds present in the

water. This water always gives poltray coloured appearance. Regular sampling

station were decided from four corner of the lake. Study is restricted to following

method described in APHA (1998).

Selection of sampling of the site was decided based on preliminary studies

of Khateshwar lake. To understand pollutional status of the lake study a few

sports were fixed and sample were drawn at 4 week interval from fixed stations

from June 2007 to December 2007. A year was chosen as a basic unit for study.

Different study spots were fixed on preliminary studies. After investigation and

after intensive study few sampling station study sport were dropped as it was

found less important. Methods described below was strictly followed from

examination of water and waste water (APHA, 1998).

All Chemical and reagents and glasses used during this piece of

work was of high of purity grade.

TABLE No. 3.1

Methods adopted for water analysis

Sr.

No.

Parameters Methods Tolerance Limit

1. Turbidity

(Water

Transparency

)

Turbidity Tube Method 10

2. Water

temperature

Temperature Sensitive

Probe

400

3. pH Electrometric Method 5.5 to 9.0

4. Free CO2 Titrimetric Method

5. DO Winkler’s iodometric Method 6.0

6. Salinity Titrimetric Method 250-1000 (Inland

water)

7. Total Alkalinity Titrimetric Method 200

8. Total

Hardness

Titrimetric Method 300

9. Nitrates Spectrophotometric Method 45-100

(Inland Water)

All values are expressed in mg.l -1 except for Turbidity, Temperature and pH.

Temperature:

Temperature effect chemical, biological reactions of organism present in

water. Rise in water temperature speed up chemical reaction in water and reduce

the solubility of gases and impart unpleasant odour to water. Water temperature

range from 7 to 11 0 C gives a pleasant and refreshing taste to water. At higher

temperature with less dissolved gases water becomes tasteless and do not

quench the trust, Trivedi and Goel (1986).

With higher water temperature organisms metabolic activities increase and

they require more and more Oxygen, however, with rise in temperature Oxygen

solubility decreases. Organisms remain sensitive to higher temperature, Day

(1994) reported that the disease resistance decrease in fishes with rise in

temperature.

Ambient water temperature was measured by using good grade mercury

weir thermometer. Usually subsurface samples were reported by drawing water

samples with the help of Ruttners water sampler. Temperature was recorded

immediately on the spot.

Transparency:

For noting transperancy in the lake Secchi disc was used. With the help of

shaded disc and graduated rope secchi disc values were recorded at the lake on

the spot usually in the morning hours and are expressed in centimeters.

Turbidity of water body is caused by various particles including planktonic

organisms. Particle size more than 10 micro meter caused turbidity and it is an

expression of optical density in which the light is scattered by the particles.

Present in water, NEERI (1987). The light is essential for carrying out

photosynthesis and is called as photic zone. Turbidity gives a measurement of

active photosynthetic zone.

TDS (Total Dissolved Solids):

TDS reflects presence of dissolved solids mainly composed of various ions

like carbonate, bicarbonate, chloride, phosphate, sodium, potassium, magnesium,

iron, like substances. In potable water dissolved solids should not cross 100

mg/lit. TDS hold an indirect effect on organisms. 100 ml of water is evaporated

on petridish on water bath at usually 60-700 c temperature. The salts were

collected on pre-evaporated dish and dishes were cooled in dessicators.

Diference in weight of evaporating dishes was noted as TDS and is expressed in

mg/L.-1

TDS = __(A-B) _ x 100

V

Where A = Final weight of dish and evaporated salt. (g)

B = Initial weight of the disc.

V = Volume of sample taken (ml)

pH :

pH is –ve logarithm of Hydrogen ion concentration. It ranges from 0-14

scale where 7 remains neutral. This value usually depend on the concentration

of CO2, carbonate and bicarbonate equilibrium. In biological system pH

equilibrium remain changed. Effect of pH on various chemical, biological

activities in water remains always crucial, NEERI (1987) the statue of water

bodies. DO test form the basis for noting biological oxygen Demand (BOD) and

BOD remains a benchmark for defining pollution and measuring biological waste

present in water.

In the present analysis water analysis kit was used to note concentration of

Dissolved Oxygen in water and results expressed in mg/lit. APHA (1989)

Total Alkalinity:

Alkalinity of water is defined as its acid neutralizing capacity. Alkalinity is a

measure of amount of strong acid needed to lower the pH of sample to 8.3, which

gives free alkalinity (Phenolphthalein alkalinity) and at pH 4.5 gives total alkalinity.

Total alkalinity is the sum of hydroxides, carbonates and bicarbonates. Alkalinity

was measured by titration method following Adoni (1985) ; APHA (1998).

Requirements:

Sulphuric acid (0.02 N) Phenolphthalein indicator, methyl orange indictor,

titration assembly.

Proceedure:

1) Take 50 ml of sample in Erylenmere Flask and add two drops of

phenolphthalein indicator.

2) Slight pink colour appears, titrate the sample with sulphuric acid to the

colorless end point and note the reading as ‘P (ml of titrant used for

phenolphthalein alkalinity)

3) Add two drops of methyl orange in the same flask and continue to titrate

further till the colour changes from yellow to orange Note this reading as

‘t’ (total ml of the titrant used for both titrations)

Calculations:

ml of titrant ‘P’ x 100

Phenolphthalein alkalinity (mg/l) =

ml of sample

ml of titrant ‘t’ x 100

Total alkalinity (mg/l) =

ml of sample

Hardness:

Ca, Mg, cations impart hardness to water. Total hardness of water show

the sum total of alkaline metal cations present in it. Hardness caused by

carbonates, bicarbonate is temporary while sulphate, chloride, calcium,

magnesium, impart permanent hardness to water. Geological strata of

catchment area generally record hardness to water where as water hardness play

an important role in distribution of aquatic biota. Hardness of drinking water can

be classified as follows.

Soft - 0 to 60 ml/L

Medium - 60 to 120 mg/L

Hard - 120 to 180 mg/L

Very Hard - more than 180 mg/L

To determine hardness EDTA is used as titrant and Eriochrome Black T as

indicator at pH about 12-0, Mg ++ precipitate and Ca ++ ions remain in the solution.

Requirements:

Standard EDTA titrant (0.01m), Eriochrome Black T indicator, Ammonia

buffer solution, titration assembly, inhibitor solution (Sol. Hydroxyl amine

Hydrochloride)

Procedure:

(a) Total hardness :

1. Take 50 ml of the sample in a flask, add 1 ml of Ammonia buffer and a

pinch of inhibitor.

2. Add 5 drops of Eriochrome Black T indicator. The colour of the sample

turns wine red.

3. Titrate the sample against EDTA solution until a clear blue colour

appears.

4. Note the readings and calculate the total hardness.

Calculation:

ml of titrant used (EDTA) x 100

Total hardness (mg/l) as CaCO3 =

ml of sample

Salinity:

In natural water chloride is invariably present Chlorides are mixed in aquatic

ecosystem by dissolution of deposits, discharges, drainage, sewage, and domestic

sewage sources. High chlorides can damage agricultural crop. Domestic

excretes record high quantities of sodium chlorides and they serve as indicator of

pollution.

Chloride level as high as 250 mg/l is safe for human consumption, a level

above this imparts a salty taste to the portable water.

Requirements:

Silver nitrate titrant (0.02N), (AgNO3), Potassium chromate (K2CrO4) as

indicator, titration assembly.

Procedure:

1. Take 50 ml sample in a flask and add 5 drops of potassium chromate

indicator. This imparts yellow colour to the sample.

2. Titrate with standard silver nitrate solution until brick red end point is

obtained.

3. Note the reading.

Calculations:

ml of titrant used x N x 35-45 x 1000

Salinity (mg/l) =

Vol. of the sample in ml

N = Normality of titrant : (0.02 N)

Nitrate (NO3-N) :

In water bodies, nitrogen is present in various forms and in highly oxidisible,

interconvertible compound, it is found as nitrate, nitrite, ammonia and organic

nitrogen. Significant sources of Nitrogen are fertilizers, vegetables, domestic and

industrial effluents. High amount of nitrate is indicative of pollution. Nitrates

exceeding 40 mg/L cause methonoglobemia or blue body syndrome APHA (1985)

and cattle mortality. Nitrites are present as intermediate for during nitrification

and denitrification reaction and is converted in to nitrate or ammonia. Presence

of nitrate in water indicates organic pollution.

Requirements

Spectrophotometers, phenoldisulphonic acid, potassium hydroxide solution,

flask, hot air oven.

Producer:

1. Evaporate 25 ml. sample overnight in a hot air oven at 500 C.

2. Dissolve the residue in 0.5 ml of phenol disulphonic acid.

3. Add 5 ml of distilled water and 1.5 ml of KOH solution. Stirr it, till yellow

colour develops.

4. Read the absorbance a 410 nm on a spectrophotometer using distilled

water bank.

5. Find out the value of Nitrates with the help of standard curve.

Nitrite (NO2-N):

Under acid i.e condition (pH 2 to 2.5) nitrite ions (NO2-N) as nitrous acid

react with sulphonilic acid forming diazonium salt that combines with

naphthylamine hydrochloride to form pinkish red azodye. The resultant optical

density (OD) is directly proportional to the concentration of nitrite present in the

sample.

Requirements:

Spectrophotometer, EDTA. Sulphanilic acid and naphthylamine

hydrochloride.

Procedure:

1. In 50 ml of filtered sample add 1 ml each of EDTA. Sulphanilic acid and

naphthylamine hydrochloride solution.

2. Appearance of wine red colour indicates presence of nitrites. Measure

the absorbance at 520 nm on spectrophotometer.

3. Read the concentration of NO2 from the standard graph.

Biological Oxygen Demand:

BOD is an amount of oxygen utilized by microorganism in stabilizing the

organic matter. Amount of organic matter degraded aerobically is an average

which provides a basis which is proportionate to demand for oxygen. BOD

generally forms a qualitative quickly degradable organic substance index.

BOD is measured by incubating samples at 200 C for 5 days by using

aerated water and dilute sewage, APHA (1998) and is expressed in mgl -1

Biological Study:

Plants and animals usually swimming or suspended non motile or motile or

brought by transport of currents are defined as phytoplanktons or zooplankton.

They are microscopic or small in size when found in water bodies. Phytoplankton

is microscopic unicellular, colonial or filamentous form mostly are autotrophic.

Phytoplanktons are grazed by zooplanktons usually reflect various seasonal,

successional pattern and serve as indicator to assess quality of water. Planktons

support heterotrophic community and are usually the members of chlorophyceae,

cyanophyceae, bacillariophyceae and freshwater forms. They fix inorganic

carbon and build up organic matter to serve as primary producers in aquatic

ecosystem.

Usually a unit of sample is sedimented to analyze photoplankton quality and

quantity. Plankton nets are usually used in algal research. However for

nanoplanktons study sedimentation process is used. For zooplanktons bigger

amount of sample is sedimented. Samples thus concentrated are then subjected

to qualitative analysis of algal forms by using monographs and keys.

Preservation of these planktons is usually carried out by using lugol for

phytoplanktons.

A liter of water sample was collected every month separately for is the

analysis. Phytoplanktons were counted in 1 ml. sample by Sedgewick-Rafter cell

method and identified following Fritsch (1975); Desikacharya (1975).

CHAHPTER IV

OBSERVATION AND RESULTS

Data was collected following procedure described in earlier chapter. Data

was collected from four corners of lake.

Physico-chemical parameter were studied at the same time phytoplanktons

data was also obtained. Result illustrated and represented and described in this

chapter.

Physical appearance

Turbidity Temperature

Odour

pH

Carbonate

Bicarbonate

Dissolved oxygen

Chlorides/Salinity

Nitrate –N

Nitrate – N

Transparency

BOD

Temperature:

Temperature was ranging between 21 to 30 C during July to September

temperature do not vary. Temperature decreased in December to 190 C and

from February it recorded higher values.

Physical appearance

Turbidity:

Water remains turbid to slightly turbid throughout the year during study period.

Odour:

Lake recorded dirty smell and in summer it imported dirty to highly dirty smell.

pH:

pH of the lake was between 8 to 9 in December it was on lower side where

summer recorded 9 as highest value, however monsoon pH values declined.

Alkalinity:

Total alkalinity both CO3, HCO3 resulted in higher concentrating during

summer.

Dissolved Oxygen:

Dissolved Oxygen recorded higher values in summer and in September DO

values were less DO was noted higher in March. Summer recorded well

oxygenated situation in epilemnetic zones; DO was lower in Oct.

Salinity:

These values fluctuated. In winter months chlorides showed higher values

also during the month of May / June chlorides recorded higher values.

Nitrate – N:

This was towards higher side during winter. However, lower nitrate values

recorded in summer.

Water Transparency:

Transparency results recorded that water remained clear with more photic

zone during summer, however water transparency was lower than in summer are

during rest of the years.

BIOLOGICAL OXYGEN DEMAND (BOD):

BOD values recorded the existence of higher biological degradation in the

lake throughout the year. BOD scale was towards lower side during early

months of summer, however no specific seasonal trend was noted for this

parameter.

Total Hardness

The water was hard during winter and lake hardness values were erratic

during rest of the moth.

Nitrate – N was lower during winter and recorded higher values in summer.

Phytoplankton

Oscillatoria among blue green algae diatom sand Chlorococcales, flagellate

were observed. Microcystis recorded its absence in July and August. It showed it

stability by forming blooms from December. In the lack of Microcystis Oscillatoria

was seen in July and also recorded in summer Chrococcas showed its presence.

Blue green algae Merismopedia, Nostoc was also recorded few in number.

Flagellate This group was present during on monsoon, Diatoms recorded in

monsoon and after.

Desmids was recorded by several species of Scenedesmus other Dersmids

like Tetraedron Crucigania Cosmarium were recorded in water.

List of species found during study period is given. Various tax recorded

some time or the other during study period is represented in the table. Efforts are

made to identify these taxa to it’s taxonomic status, however the study was not

pertained to taxonomy of algae, therefore generic data is presented.

Periodic data of Khateshware lake(Physio-chemical parameters of water).

Expressed in mg/L.

TABLE NO. 4.1

↓ →Temp

.Transparenc

y pH DOHCO

3 CO3

Salinity

BOD

Total Hardne

ssNitrite

s NitratesJUN-05 30 6.1 9 8.5 374 89 610 24.1 153 0.478 0.021JULY-

05 29 8.5 9 7.8 149 121 600 19.1 162 0.34 0.023AUG-

05 29 14 8.8 6.5 234 34 480 21.2 167 0.121 0.022SEPT-

05 29 15.1 8.7 6.3 174 21 490 18.3 180 0.426 0.02OCT-05 28 12.5 8.6 4.2 265 31 487 21.9 183 0.521 0.034NOV-

05 26 11.3 8.6 7.9 245 30 360 20.7 181 0.302 0.042DEC-

05 19 11.2 8.5 9.5 306 45 387 22.3 120 0.162 0.034JAN-06 20 11 8.5 5.4 428 70 430 20.1 117 0.064 0.031FEB-06 21 11.2 8.6 9.8 445 40 511 18.7 121 0.079 0.029MAR-

06 24 10 8.9 11 307 50 470 21.3 122 0.102 0.027APR-06 27 8.1 8.9 7.8 336 43 530 16.7 37 0.097 0.025MAY-

06 31 6.2 9 8.2 532U.N.

D 521 21.3 48 0.764 0.022

Periodic data of Khateshware lake(Physio-chemical parameters of water).

Expressed in mg/L from Jun-2005-May-2006.

Main graph

PHYTOPLANKTON

Phytoplankton is chlorophyll bearing suspended microscopic organisms

consisting of algae with representative from all major taxonomic kinds. The

majority of members belong to Chlorophyceae. Cyanophyceae and

Bacillariophyceae hold unique ability to fix inorganic carbon to build organic matter

through primary production makes these study a subject of prime importance.

The quality and quantity of phytoplankton and their seasonal successional patterns

have been successfully utilized to assess the quality of water and its capacity to

sustain heterotropic communities.

QUALITATIVE AND QUANTITATIVE ANALYSIS

Among the several methods known for the collection of plankton the use of

plankton net is most common even for algal work which however should be

abandoned because many minute (e.g. nonoplankton mainly diatoms) pass

through the pores of the net resulting in under estimations of the population

density. Therefore, the direct concentration of the phytoplankton of the water

sample by sedimentation (and occasionally by centrifugation) is a prerequisite for

accurate qualitative analysis.

Chemical Alkalinity:

Alkalinity measurements were carried out at the lake sites following pheno

phthelein for carbonates and methyl orange indicator and result expressed as CO3

or HCO3 in mg 1 –1.

Physical appearance :

Odour:

Odour is noted at the lake site and odour and colour noted.

TDS total dissolved soil :

Total dissolved solid were calculated by evaporating fixed amount of water

in crucibles on hot water bath at 1000C and difference in weight is calculated.

COLLECTION AND PRESERVATION OF SAMPLE

From the predetermined stations along the lakes the samples were

collected every month for analysis of different parameters in the following

manners. The sampling was always done during early hours of morning at all

stations. Samples collected were accompanied by sampling data remainder and

a tag (appendix) for sampling data and station. All sample were collected in

triplicate and a survey was carried for 12 months. Always subsurface sample

were collected.

For general parameters :

(Alkalinity Hardness, Salinity)

5 liter of sample wasllected in wide mouth transparent polythene containers

previously treated by Rodine as described by Schwoerbel (1970). No chemical

preservative was added and was immediately stored at 40 C.

For Dissolved Oxygen (DO)

The samples were collected in 300 ml bottles using APHA type. DO

samples assembly. DO was fixed on the spot, adding alkali-azide iodide with

manganese sulphate solution.

For phytoplankton analysis

One liter of sample was collected for phytoplankton analysis and it was

preserved at the lake site with Lugols Solution following 1958, but Lugols with

acetic acid (Habro and Willen, 1977).

Method for determination of different parameter.

After bringing the fixed sample the sample were allowed to sediment for 5

to 6 days and the sediments are resuspended in 10ml (Ragothaman, 1975) of

buffered formoline (UNESCO1s, 1974, Harbro and Willen, 1977).

The relative number of genera in the Cyanophyceae, Bacillariophyceae and

Chlorophyceae were determined for each sample.

In present investigation work on natural lake in Khateshwar village is

selected for assessment. The village has population of 5500 residents, but

certain at regular intervals the lake is under stress when pilgrims visit to site and

camp at several days at several time beside this, the visitor wash cloths and cattle

feed on lake sides addition of domestic waste and visitor activity is an common

phenomenon for addition of nutrients in Khateshwar lake. Lake required full

assessment. Study of plankton of lake and study of forest flora from surrounding

environment can define status of lake. The research was carried out in following

lines.

1. The study is novel to its kind because this lake is not attempted by any

researcher earlier.

2. Seasonal study of plankton was carried out.

3. Chemical study was carried out.

4. Data collected is presented and assessed, will the help of available with

scientific literature.

Table No. 3.1

Detail of physico-chemical analysis adopted for this work

Test Method and Reference

Temperature

Transparency

Ambient water temperature noted on

spot transparency by sechhi Disc.

Mercury weir

thermometer

pH Ambient water pH and in Laboratory pH meter

Alkalinity Standard sulphuric acid phenolpthlein

and methyl orange APHA 1998

Titration

Chlorides Argentometric method APHA, 1998 Titration

Total Na EDTA titration APHA, 1998 Titration

Hardness

Dissolved

Oxygen; BOD

Wrinker Azide titration APHA 1998 Titration

Nitrite – N

Nitrote – N

Following APHA 1998 Spectro colour

meter at 520 nm

DISCUSSION

General discussion on water hydromicrobialogy of Khateshwar lake.

Aquatic ecosystems are prone to innumerable social economic problems,

these freshwater systems are over utilized in rural India, as against water

conservation carried out in advanced countries. India fall in tropical area where

excesses nutrient addition is uncontrolled, through there are huge number of

legislations, but these are never taken care of for creating balanced ecosystem

water is scares and human activity is growing. Much of work related to

ecophysiology of aquatic bodies is carried out with respect to bloom formation is

reported. In India major algal forms which grow to excesses are Microcystis,

Merismopedia, Nostoc, Chroococcus, Rivulatria. Bloom- formation in eutrophic

fresh water is an overt expression of multiple interacting factors and bloom formed

like Microcystic is momentarily favored by effects of these factors. In a particular

period of the year, the whole gamut of factors so harmoniously synchronise their

optima that lake water are blasted with overgrowth of Microcystis.

Environmental parameters which trigger the bloom formation can roughly

be divided into two classes ; course adjusters and fine adjusters, light intensity and

macronutrients are those among course adjusters and temperature and release of

certain micronutrients from bottom mud and bacterial activities, oxygen status of

the water are those among fine adjusters which trigger the bloom formation.

Some workers like Keating (1977) has included extra cellular matabolites also

amongst the fine adjusters.

Under this discussion results obtained from physico-chemical analysis of

environmental parameter of the lake will be discussed along with biological

species, then the results obtained from the vitro studies of the lake microbes will

be discussed. These discussion may lead to some concluding remarks for

biocontrol of the bloom forming species.

This method is novel of its kind to correlate environmental and laboratory

outcomes. Ecological perception with respect to study of hydrobiology and

control of native tolerant species with the help of microorganisms is totally novel of

its kind for this area. Present attempt will serve a useful tool for the researches to

come. In the present discussion interpolation of these result is carefully done

and fruitful attempt are made in the following lines.

LIGHT PENETRATION

Solar radiation directly effects growth of micro-organisms in water.

Berlinsko and Mann (1978); besides heterotrophic and photoheterotrophic mode of

nutrition for microorganisms is common in natural water. Photoauto trophy

remains in active mode for blue green algae and various forms of life.

Transparency factor depends upon the turbidity and organic compounds present in

water. In the present study summer recorded higher euphoric zone and it was

almost double than it was in winter. High wind induced turbidity or inflow usually

govern like penetration in water. A combination of factors like light intensity and

day length in summer, water temperature, reduced circulation may cause plankton

succession (Hickman, 1974). It is known that smaller algae reproduce themselves

at a quicker rate (Golterman, 1975). Similarly, spelling and Blum (1974), reported

unicellular algae being favored over filamentous ones during winter and low light

intensity.

Summer usually record high crowding of microorganisms in euphotic zone

in this lake

TEMPERATURE

Temperature of water generally govern productinity of the lake ;

temperature hold great effect on water microbiology. With temperature gases

dissolve and metabolic processes also increase. Zutsi and Khan (1978) ;

Konokpa and Brooke (1978) reported a direct bearing of water and production of

plankton oxidation of organic matter goes higher during summer. In the presnt

survey of Khateshwar lake water temperature followed air temperature and

recorded winter minima and summer maxima. Biological system usually hold a

good tolerance for temperature range. Reynolds (1971) reported higher water

blooms only at onset of summer, during winter algal range of the water recorded

various forms of algae, particularly green algae was found during winter. Thus it

is noted from the results that the lake supported typical tropical summer flora and

the growth of the biota remained increasing. With the temperature rise number of

species decreased during summer therefore this lake is defined eutrophic. In the

present instance higher crop of phytoplankton was recorded. Besides

combination of factors that allows one species of algae to hold dominance over

others, light could be one among them. Plants need light for their photosynthetic

requirements. Physiological adaptation of low light intensity is recognized in

many algae. Similar results were reported by Round, 1964; sreenivasan 1964;

and also by Reynolds and Walsby 1975.

IONIC COMPOSiTION AND pH

In soft water Lake Hutchinson (1967); algae and microorganism remained

higher in number, besides calcium, bicarbonate, ions and pH buffering availability

of free CO2 is important Reynolds and Walsby (1975), In the present study pH

remained higher and lake water was within a well buffered situation. Thus the

results obtained for this parameter were well within the range for Indian lake water.

Bicarbonate and carbonate result are usually governed by soil composition. In

higher temperature carbonate may be precipitated out in the form of bicarbonates

Hutchinson (1957);

Presence of higher carbonate during rainy season and also during winter,

may be due to result of higher run off and higher biological activity in lakes. King

(1970) reported that low alkalinity and high pH microorganisms remain carbon

limited.

Lee et al., (1978), in contrast to Kuentzel (1969) suggested that except

under atypical highly fertile condition carbon rarely limits total algal biomass. It

would appear that carbon limitation of algal growth remains distance possibility in

a productive lake Lange (1973) unless there are adequate reserves provided by

the bicarbonate buffering system. Lakes which are well buffered by high

concentration of bicarbonate low pH value should be favorable to the growth of

blue-green algae) (Reynolds and walshy (1975). Results obtained during present

study can be correlatedpositively with there results.

In a natural water body, carbonate bicarbonates parameters define

biological activities and enzymatic activities balance, however this parameter

never remain limiting to trigger the over growth of biota and this parameter remain

soil dependent.

Under different pH, alkalinity conditions few species may be favored and

few may be eliminated, however then there is surplus growth and eutrophication.

The parameter understudy remained insignificant. In lake Khateshwar summer

flora recorded eutrophophic conditions.

Salinity

Besides soil water and related characters there are two other factors which

are known to effect ionic composition of natural waters in semi arid and arid

regions. Some effect of net water loss from lakes due to evaporation, climatic

factors including air temperature and wind velocity precipitation and role of surface

run off effect the relative water loss, which results in gain of total dissolved solids.

It was expected that small lakes would be more saline and more eutrophic

than large ones as is the case in larger areas which provide homogeneous

characters Rawson (1955), however, in contrast, Barica (1978) showed that

neither salinity nor chlorophyll depend upon morphometric characteristics of the

lakes. It is relevant to quote here Jhingran (1977) and Gulati (1976) who believed

that chemical characteristic of the water of a reservoir change with its age. chloride

values for this lake was erratic with higher records for may and June. Jhingran

(1977) reported that chemical characteristics of water of a reservoir vary with its

age and lakes distributed with in a small geographical area remain limnologically

different.

Rhode (1949) Barica (1978) considered most of the inland waters to be

bicarbonate type ; they characterized other types as regional or “local pecularities”

influenced by geochemical and climatic conditions tending to counteract and

cancel the normal tendency of fresh water to develop into a bicarbonate – Ca type.

However, no such peculiarity was seen in the lakes under study. In the present

investigation the range of salinity obtained was in confirmity with chloride value

reported for Jaipur lakes Sharma (1978) Higher values of chlorides are due in no

small measure to the nature of soil in this part. This lake supported various

seasonal phytoplankton.

Dissolved Oxygen (D.O.)

In the presennt instance higher summer DO values recorded well

oxygenated situation in the lake, which directly reflects photosynthetic activity

carried out by the phytoplankton. In lake subsurface winter DO was slightly

lower. This record metabolically active situation in the lake. pH and minerals

and nutrients remained quite available in the lake in dissolved form and

phytoplankton was represented and by various species. Oxygen contents do not

dissolve in water to a great concentration in the lake higher DO result maximum in

January is in a conformity with several Indian authors like Banarkar et. at., (2005)

on Chandrawati tank. Khatawakar et. al., (2002). Sakhare and Joshi (2003),

availability of O2 in dissolved form in natural water usually support zooplanktons

growth.

BOD

BOD represents biological activity, available DO, bacteria and organic

matter composition. BOD usually follow two cycles, where carbonaceous phase

decompose to produce CO2 and H2O and nitrification phase which follow nitrate

nitrate path way the action of nitrozo-monas and nitro-bactar. A steady phase

unchangel phase of BOD was recorded from the study from Khateshwar lake

water explain constant decomposition and supply throughout the season during

study period

Nitrogen Nitrate

The forms of nitrogen that are generally available for aquatic plant growth

are nitrate and ammonia. Normally algae require ten times as much nitrogen for

growth as phosphorus, natural water contain at least this relative quantity of

available nitrogen over phosphorus Lee et al, (1978). Blue greens frequently

become dominant in lakes at about the same time that concentration of the

nutrients reach their seasonal minima Lund (1965) Hutchinson, (1967). Nicholls

(1976) has observed abundant growth of blue green in water where inorganic

nitrogen and phosphorus concentrations were less than detectable levels.

The present lake recorded higher nitrates during winter and lower nitrites

during the same period of time. This represents that nitrite to nitrate was active

during winter and nitrate degradation was seen in summer. Nitrate reflect

eutrophi condition of water with a domestic source of pollution organic matter

decomposition is defined during onset of hot season in the present lake, which is

also reflected from BOD values.

PHYTOPLANKTON

The problem of the relationship between bacteria and algae is especially

relevant with the emphasis of aquatic ecology today. As to nature of this

relationship there is no simple formula. Some bacteria may benefit from algae

and vice versa. They may also adversely affect one another by secretions of

substance with antibiotic activity.

Ecology of phytoplankton in polluted water bodies and the usefulness of

different phytoplankton species and groups serve as indicators of pollution Status

of water have been studied in depth by a number of workers since the emergence

of the concept of “Biological indicators pollution”. In the present instance the lake

was represented by several groups. of algae Microsystic recorded higher during

July and August ; when Microsystis was absent Chrococcus, Merimopedia,

Oscillatoria were recorded. The lake was also represented by desmids and

diatoms.

The lake understudy was represented with presence of pollutional grade

species like, Anebena, Microcystis, Merismopedia and Scendesmus.

Discussion – Phytoplankton Counts.

Thus it is clearly seen that association between blue-green algae and

bacteria appear ubiquitous in aquatic ecosystems and are often more pronaunced

during over growth of algae.

Changes in temperature, salinity conditions can cause fluctuations in

bacteria and algal members, which may have nothing to do with bacteria algae

relationship itself (Hieper, 1975).

Several authors on the relationship between algae and bacteria tend to

support that healthy algae were not subject to degradation by bacteria or fungi –

organic compound assimilated by bacteria. “May be one of the factors leading to

algal bloom in lakes and ponds especially when growth is not limited by the supply

of phosphorus or other inorganic elements”. This intimate relationship must have

been a reason among others that Microcystic which grows in lake understudy. The

description of Microzones induced by bacteria associated with bluegreen algae

indicaters how the algae might benefit, but may of the mechanics of nutrients and

gaseous exchange remain uncleared.

PART – II

INTRODUCTION

Accelerated eutrophication of natural water bodies is at present, a very

important problem calling for possibly a speedy solution. The possibility of

slowing down eutrophication process or of its control, closely depend on obtaining

a knowledge on dynamics of algal growth and its nutritional requirements. An

individual can survive, grow and reproduce in a changing environment. So long as

the changes are not beyond limits, Genetically determined potentiality for the

morphological and physiological adaptation, algae may respond differently to

different substances added to it for this reson it is desirable to carry out

experiments in the laboratory For the biocontrol of it. In the present experiment

fungi was isolated from lake Khateshwar and biological control with the help of

using bacterial strain was adopted for biocontral applications.

Genus Bacillus is well known for antibiotic producer, antifugal activity of

Bacillus was observed against plant pathogenic fungal including Penicillium sp.,

Rizopus sp ; Candida sp., Fusarium sp., on the potato dextrose agar by Agar Well

Diffusion Method. The author was benefited to work in Department of

Biotechnology , Swami Ramanand Tirth University, Nanded (MS) and strains were

isolated from Lake Khateshwar under study.

Fungal diseases can have major constraints on crop production and in

conventional agriculture and environmental studies ; chemical fungicides are

routinely used to provide disease control. However, as these chemicals are often

toxic and potentially harmful to man and the environment, alternative methods for

control are needed. Biological control is potential alternative approach to

chemical treatment and there are number of fungal biocontrol products

commercially available, including some that are registered as biopesticides.

These are likely to increase in coming years.

The search for new safe, broad spectrum antifungal antibodies with greater

potency has been progressing slowly Gupic et al., (2002). One reasion for the

slow progress compared to antibacterial is that, like mammalian cells, fungi are

eukaryotes and therefore agents that inhibit protein. RNA or DNA biosynthesis in

fungi have greater potential for toxicity to the host as well (Georgo – Papadacou

and Ticazz, (1994). The other reason is that until recently, the incidence of life

threatening fungal infections was perceived as being too low to warrant aggressive

research by the pharmaceutical companies (Georgo Papaddicou and Ticazz,

1996).

2. REVIEW OF LITERATURE

A compound produced by Bacillus pumilus (MSH) that inhibits Mucoraceae

and Aspergillus species is described. Fungicidal activity was demonstrated by

lawn spotting and by diffusion through 0.45 um millipore membranes placed on 5

% sheep blood agar, nutrient agar, trypticase soy agar and muller- Hington agar,

followed by spore incubation correlated with the zone of haemolysis produced by

B. pumius (MSH). The active compound inhibited mucor and Aspergillus spore

germination and aborted elongating hyphae, presumably by inducing a cell wall

lesion. Antifungal activity was stable in agar for a minimum of 8 days, resistant to

pronase degradation, and partially inactivated by chloroform exposure and at pH

5.6. its molecular mass was determined by diffusion through dialysis membrane to

500 – 3000 da. Attempts at further isolation of the compound have proven

unsuccessful to date. Alternaria disease are among the most common diseases of

many plant in the world Rocoa (2001). They affect primarily the leaves, stems,

flowers and fruits of annual plants especially vegetables Basim (1997).

The over use of chemical pesticides has caused soil pollution and harmful

effects of human beings. Accordingly biological control of soil borne disease has

been attracting attention. Many reports or reviews in this area have already

appeared Bernal (2001). Biological controlled agent are potential alternatives for

the chemical fungicides presently used in the agriculture to fight plant diseases.

Bacillus spp., strains are an example of a promising safe fungal biological control

agents. This work, describes that IB. subtilis B2, B. licheniformis B40, B. subtilis

mB2.9, B.subtils + B2.2, B. licheniformis MB40, B. licheniformis + B40.2, which

showed in vitro antibiotic activity against Cornea et al., (2003) were subjected to

pot test to investigated their ability to protect plant against fungal disease.

A Bacillus strain, denoted as Py-1, was isolated from vascular bundle of

cotton. Biochemical, physiological and loss DNA sequence analysis proved that

it should belong to Bacillus subtilis. The Py-1 strain showed strong ability against

many common plant fungal pathogens in vitro. The antibiotics produced by this

strain were stable in neutral and basic conditions, and not sensitive to high

temperature from the culture broth of Py-1 strain. Five antifungal compounds

were isolated by acidic precipitation, methanol extraction, gel filtration and reverse

phase HPLC. Advanced identification was performed by mass spectrometry and

nuclear magnetic resonance spectroscopy. These five antifungal compounds

were proved to be the isomers of iturin A : A2, A3, A4, A6 and A7. In fast atom

bombardment mass spectrometry / mass spectrometry collision induced

dissociation spectra, fragmentation ion from two prior linear acylium ions were

observed, and the prior ion, Try-Asn-Gin-Pro-Asn-Scr-BAA-Asn-Co, was first

reported.

Fusarium wilt causes huge economical losses in a wide variety of crops

(Inouel et al., (2002). The pathogen, Fusarium oxysporum, infects plants through

the roots by direct penetration or wounds, colonized the vascular tissue and

causes plant death Simons et al., (1998). Chemical soil fumigation is the main

treatment of Fusarium wilt. Broad spectrum biocides / particularly methyl

bromide, can be used to fumigate the soil, but they cause serious environmental

damage Fravel et al., (2003).

Recently, scientists have paid attention to biological methods of defense

against plant diseases. Control of pathogens by antagonistic microorganisms or

their antibiotic products is not considered a viable diseases control technology

(Han et al., 2005). A Bacillus strain with ineffective ability against the Fusarium

wilt pathogen was isolated from the vascular tissue of a cotton Fusarium wilt

resistant strain and named Py-1.

3. MATERIALS AND METHODS

Collection of samples

Soil from deferent region of Khateshwar lake was collected for isolation.

Water sample were also collected from the sites.

Enrichment

The isolates obtained from starch agar plates were enriched in pure form

into starch broth tubes and the tubes were subjected to growth at 400 C for 24-48

hrs. Serial subcultures and transfers were made to maintain the cultures of

Bacillus species thus obtained.

Isolation

Mix 1 g of soil samples collected from lake in 10 ml of sterile distilled water

tube. Inoculate 0.1 ml from mixture in to the sterile starch agar plate and sterile

starch broth tube at pH 8, 10 and 12.

Preparation of starch agar

Starch agar medium was prepared as follows starch 20g, Peptone 5 g, Beef

extract 3 g, Agar agar 15 g, Distilled water 1000 ml. Medium was autoclaved at

1210 C for 15-20 min. pH of the medium was adjusted with 0.1 N Na OH or 0.1 N

HCl.

Preparation of starch broth

Starch broth content was as follows starch 20g, Peptone 5 g, Beef extract 3

g Distilled water 1000 ml.

Medium was autoclaved at 1210 C for 15-20 min. pH of the medium was

adjusted with 0.1 N Na OH or 0.1 N HCl.

Inoculated plates and tubes of starch agar and starch broth respectively

were incubated at 400 C for 24-48 hours. After incubation period different species

of Bacillus were obtained and their activity was checked on starch agar plates.

Identification

Morphological and cultural characters were observed by performing certain

biochemical tests. These isolates were labeled as LB1, LB2, LB3.

Isolation of fungi

The four species of fungi were also isolated as Penicillium sp., Rhizopus

sp., Candida spand Fusarium sp.

Extraction

Starch agar broth of LB1 pH 8, LB2 pH 10 and LB 3 pH 12 were configured

by taking the appropriate amount of the sample in each sterile Eppendorf tube of

the centrifuge. These tube were centrifuged for 10000 rpm of 20 min.

The supernatant was collected into three sterile screw cap tube. This

supernatant were used for determining antifungal activity against plant pathogenic

fungi by agar diffusion assay method.

Bacterial culture

Starch broth of pH 8, pH 10 and pH 12 were inoculated by loopful of LB1,

LB2 and LB3, respectively, which was also used for determining antifungal activity

against isolated fungi using agar well diffusion method.

Assay using extracts and whole bacterial cultures.

The sterile potato dextrose agar (PDA) was distributed in sterile petriplates.

The fungal suspension of Penicillium, Rizopus, Candida and Fusarium was

prepared separately in sterile saline tube. These suspensions were eventually

spread with the help of spreader on medium then petridish was kept at room

temperature for 30 min after that a well was prepared with the help of borer. The

wells were separately filled with either bacterial cultures or cell extracts as

mentioned. Plates were incubates at room temperature for 2-3 days and

observed zone of inhibition. The results were recorded and expressed in terms of

zone of inhibition of pathogen i.e. fungi in millimeter.

Percent Relative Activity.

For percent relative activity of the isolated fungi considered standard zones

of inhibition against Gresiofulvin zone with A. Solani as a test organism was

considered standard.

Gresiofulvin antifungal was discovered to be produced by P. Gresiofulvin

and now a days by several species of Penicillium. It is highly toxic to powdery

mildew of beans and downy mildew of cucumbers. It is also used to control

Alternaria sanlani in tomato, Scerotinia fructigena in apple and Botrylies cinerea

in lettuce.

Per cent relative activity of LB1, LB2, and LB3 were calculated by

compairing with standard zones of inhibition with Gresiofulvin against A. Solani

on PDA. The standard solution was prepared by dissolving 100 mg of antifungal

compound in 1 ml of ethanol. This solution (0.1 ml) was added in the agar wells

and the zones of inhibition obtained were considered as standard.

4. RESULTS AND DISCUSSION

Table 4.1 shows colony characteristic of LB1, LB2, LB3 after 24 hr

incubation on starch agar.

Table 4.1 Morphological and colony characteristics of LB1, LB2 and LB3.

Colonies/Colony

Character

LB1 LB2 LB3

Size 1 mm 1 mm 1mm

Shape Circular Circular Circular

Colour White Dirty White Pale Yellow

Elevation Convex Convex Convex

Margin Entire Entire Entire

Opacity Opaque Opaque Opaque

Consistency Sticky Sticky Sticky

Surface Smooth Smooth Smooth

Gram’s nature Gram Positive Gram Positive Gram Positive

Morphology Short rods Short rods Short rods

Strain LB1, LB2, LB3 exhibited amylase test positive.

Table 4.2 antifungal activity of gresiofivom against A. solani of PDA.

Antifungal pH (8.0) pH (10) pH (12)

Gresiofulvin 16 mm 12mm 08mm

The date in Table 4.2 shows that antifungal activity of resiofulvin against I

A. solani of PDA. It is consider as a standard table for calculating percentage

relative activity.

Data in the Table revealed that different species of Bacillus show the

antifungal activity against plant pathogenic fungi at its respective pH. It was seen

that as indicated in the Table 4.3 crude extract of LB1, LB2 and LB3 showed

significant result of zone of inhibitant against Candida sp. And Rhizopus species

as compared to the other fungal species.

Table 4.3 Antifungal activity of Bacillusi sp. Against different target fungi at

pH 8.

Bacterial sp Inhibition zone diameter (mm) *

Penicilium

sp

Rhizopus sp Candida sp. Fusarium sp.

LB1 7 8.5 8.5 7

LB2 5.5 7 8 6

LB3 7 7 5.5 5.5

(*This activity was performed by using agar well diffusion technique.)

Percentage relative activity

When compared with per cent relative activity of standard resiofulvin

against Alternaria solani per cent relative activity of Bacillus sp. given in table 4.4.

and Fig. 4.5. it reveals that per cent relative activity of LB1, LB2 and LB3 was

maximum of significant against the Candida and Rhizopus species at pH 8.

Table 4.4. per cent relative activity of centrifuged extract of the isolated at pH

8.

Bacterial sp Per cent relative activity at pH 8*

Penicilium

sp

Rhizopus sp Candida sp. Fusarium sp.

LB1 36.66 42.44 42.44 36.38

LB2 32.22 336.38 42.44 32.33

LB3 36.44 36.44 32.22 32.22

(* Per cent relative activity of the Bacillus at pH 8 calculated by using the standard

Gresiofulvin against A. solani on potato dextrose agar.

Per cent relative activity of Bacillus sp. against pathogenic Fungi

From the Table 4.3 and Table 4.4 and Fig. 4.5 it is revealed that the crude

extract of LB1, LB2, and LB 3 show the moderate antifungal activity at pH 8 as

compared to other fungal strains.

While under the similar pH i.e. 9 pH conditions but centrifuged extract of

LB1, LB2, and LB3 showed in Table 4.4 that strain LB1, LB2 and LB3 shows

higher than crude extract the antifungal activity against Penicillium sp. and

Rhizopus sp. but all the LB1, LB2 and LB 3 shows moderate antifungal activity

against the Candida sp. and Fusarium sp. at pH 8.

Effect of Bacillus sp. against pathogenic fungi at pH 8.

Bacterial sp Inhibition zone diameter (mm)*

Penicilium

sp

Rhizopus sp Candida sp. Fusarium sp.

LB1 5 4 5 4

LB2 4 5 5 3

LB3 3 5 3 4

(*This activity was performed by using agar well diffusion technique.)

Percent Relative Activity

When compared with per cent relative activity of standard. Gresiofulvin

against Alternaria Solani per cent relative activity of Bacillus sp. it is calculated as

shown in the Tables showed that per cent relative activity of LB1, LB2 and LB3 is

maximum/moderate against the Candida sp. and Rhizopus sp.

Per cent relative activity of Bacillus sp. at pH 8.

Bacterial sp Per cent relative activity at pH 8

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 31.11 26.66 31.11 26.25

LB2 25.66 32.44 32.66 22.22

LB3 21.22 31.33 15.88 21.22

(*Per cent relative activity of the Bacillus at pH 8 (centrifuged extract) calculated

by using the standard Gresiofulvin against A. Solani on potato dextrose agar. )

Per

cent relative activity of Bacillum sp. against pathogenic fungal at pH 8

Table 4.9 crude extract of LB1, LB2 and LB3 showed significant result

against the plant pathogenic fungi but it is more effective against candida sp. and

Rhisopus sp. at pH 10.

Table 4.9 effect of Bacillus sp. against plant pathogenic fungi

(centrifuged extract) at pH 10.

Bacterial sp Inhibition zone diameters (mm)*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 5 6 5 5

LB2 7 5 4 7

LB3 5 6 5 7

(*This activity was performed by using agar well diffusion technique.)

Per cent Relative Activity

When compared with per cent relative activity of standard Gresiofulvin

against Alternaria Soloni per cent relative activity of Bacillus sp. it is was shown

in Tables and reveled that per cent relative activity of LB1, LB2 and LB3 is

moderate against or significant against the Rhizopus sp and then Fusarium sp.

Per cent relative activity of Bacillus sp. (centrifuged extract)

at pH 10.

Bacterial sp Per cent relative activity at pH 8*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 45.15 52.84 45.15 44.15

LB2 52.33 45.55 36.66 52.88

LB3 45.55 45.88 36.66 54.44

(*Per cent relative activity of the Bacillus at pH 10 (Bacterial culture) calculated by

using the standard Gresiofulvin against A. Solani on potato dextrose agar.

Per cent relative activity of Bacillum sp. against pathogenic fungel

(centrifuged extract) at pH 10

From Table it was seen that the LB2 and LB3 was showing maximum zone

of inhibition and maximum per cent relative activity, it shows that LB2 and LB3

show to produce antifungal activity against Rhizopus and Fusarium sp.

Results show that the centrifuged extract of LB1, LB2 and LB3 show

increased level of antifungal activity than that crude extract at pH 10 it reveals that

the LB1, LB2 and LB3 show the increased level of antifungal activity against the

Rhizopus sp. and than the Fusarium sp.

Effect of Bacillus sp. against pathogenic fungi at pH 10.

Bacterial sp Inhavition zone diameters (mm)*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 4 5 5 4

LB2 5 6 4 5

LB3 4 5 4 5

(*This activity was performed by using agar well diffusion technique.)

Per cent Relative Activity

When compared with per cent relative activity of standard Gresiofulvin

against Alternaria solani per cent relative activity of Bacillus sp. is calculated as

shows in Table 4. 13

Per cent relative activity of Bacillus at pH 10.

Bacterial sp Per cent relative activity at pH 8*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 30.76 38.46 38.46 30.76

LB2 38.46 38.46 30.76 38.46

LB3 30.76 38.46 30.76 38.46

(*Per cent relative activity of the Bacillus at pH 10 (Bacterial culture) calculated by

using the standard Gresiofulvin against A. Solani on potato dextrose agar.)

Per cent relative activity of Bacillum sp. against pathogenic fungo at pH 10

From above table per cent relative activity of bacteria against given fungal

species can be calculated, it showed that the per cent relative activity and zone of

inhibition increased with the Rhizopus sp. and than Fusarium sp. at pH 10 as

compared with the other species of fungi.

It was observed that as indicated in the Tables that is crude extract of the

LB1, LB2 and LB3 showed significant zone of inhibition against all the plant

pathogenic fungi but the antifungal activity against the Candida and Fusarium

sp.revorded significant zone of inhibition at pH 12.

Effect of Bacillus sp. against pathogenic fungi

(centrifuged extract) at pH 12.

Bacterial sp Inhavition zone diameters (mm)*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 3 2 3 3

LB2 Nil 3 2 Nil

LB3 2 3 3 2

(*This activity was performed by using agar well diffusion technique.)

Per cent Relative Activity

When compared with per cent relative activity of standard Gresiofulvin

against Alternaria solani per cent relative activity of Bacillus sp. is calculated and

recorded in Table 4. 16

Per cent Relative Activity of Bacillus centrifuged extract) at pH 12.

Bacterial sp Per cent relative activity at pH 8*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 28 20 28 28

LB2 Nil 28 20 Nil

LB3 20 28 28 20

(*Per cent relative activity of the Bacillus at pH 12 (Bacterial culture) calculated by

using the standard. Gresiofulvin against A. Solani on potato dextrose agar.)

From above it revealed that the inhibition zone in per cent relative activity, it

showed that Candida sp. and Fusarium sp. shows maximum inhibition zone and

Rhizopus sp. did not show any significant activity.

Per cent relative activity of Bacillum sp. against pathogenic fungel at pH 12

Above results showed that the centrifuged extract of LB1, LB2 and LB3

show the antifungal activity at pH 12 revealed that the LB1, LB2 and LB3 showed

increased antifungal activity against the Rhzopus sp.

Effect of Bacillus sp. against pathogenic fungi at pH 12.

Bacterial sp Inhibition zone diameters (mm)*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 2 2 2 2

LB2 Nil 1 1 1

LB3 1 1 Nil 2

(*This activity was performed by using agar well diffusion technique.)

AT pH 8 and pH 9 these results show crude extract of LB1, LB2, LB3

antifungal activity. But the activity is less as compared with other pH activities.

Per cent Relative Activity

The per cent relative activity compared with per cent relative activity of

standard Gresiofulvin against Alternaria solani per cent relative activity of

Bacillus sp. is calculated. It revealed that it showed antifungal activity against

Rhizopus sp.

Table 4.19 Per cent relative activity of Bacillus sp. at pH 12.

Bacterial sp Per cent relative activity at pH 8*

Penicilium sp Rhizopus sp Candida sp. Fusarium sp.

LB1 20 20 20 20

LB2 Nil 10 10 10

LB3 10 10 Nil 20

(*Per cent relative activity of the Bacillus at pH 12 (Bacterial culture) calculated by

using the standard. Gresiofulvin against A. Solani on potato dextrose agar.)

Per cent relative activity of Bacillum sp. against pathogenic fungal

(centrifuged extract) at pH 12

Above results show antifungal activity of LB1, LB2, and LB3. Activity is

very less as compare to pH8 and pH10.

5 CONCLUSON

From the persent study, it is concluded that Organisins isolated from

Khateshwar lake at different pH i.e. pH8, pH 10 and pH 12 was capable of

producing antifungal compounds against pathogenic fungi.

On the basis of results obtained in the present investigation, it is concluded

that strains LB1, LB2, LB3 showed maximum antifungal activity against Penicillium

sp. Rhizopus sp., Candida sp. and Fusarium sp. at pH 10. Accordingly, results

of strains LB1, LB2 and LB3 showed less activity against Penicillium sp.,

Rhizopus sp., Candida sp. and Fusarium sp. at pH 12.

Fungal disease can have major constrains on the crop production and in

conventional agriculture and in environmental studies chemical fungicides are

routinely applied to provide disease control. However, these chemicals are often

toxic and potentially harmful to man and the environment. Thus the use of

microorganisms for biological process has become an effective alternative to

control pathogen.

SUMMARY OF EUTROPHICATION AT KHATESHWAR LAKE AND

CONCLUDING REMARKS.

Communities seldom appear as discrete units. In many cases

interspecefic and intraspecfic communities integrade one another both in time and

space and often exhibit no distinct boundaries between them. This is specially

true of aquatic systems and in instances of algal bloom formation and symbiotic

relations between algae and bacteria.

The present remarks concern largely with those aspects of planktonic blue-

green algae where generalizations are possible in the light of present findings.

Caution is always necessary in drawing conclusions based on one few lake

studies. In the following lines however, an attempt is made to draw general

remarks, summary and conclusion of the work under investigation.

In the present study on Khateshwar lake an investigation was carried out on

at four week interval and data was collected from Jun- 2005 to May 2006.

The investigation was carried out to record physicochemical and biological

changes occurring in lake Khateshwar. It concludes following remarks that, the

small bodies of water can have intense intermix can provide more or less similar

biota. Pollutional algae may exist even at, temperature at 190 c to 300 C, higher

temperature recorded high crowding of microorganism in euphotic zone. This

lake accordingly is define as eutrophic.

Algal crowding can cut light to a depth of 6 cm photic zone.

Pollutional algal forms can thrive in the water well at higher buffering action

and biological activity may bring about pH increase to as alkaline as 9 pH.

The present water study recorded well buffered situation because of higher

run off or higher biological activities and supported pollutional forms. Carbonates

and bicarbonates never remained limiting to control overgrowth of biota in natural

water may be soil dependent on local peculiarities.

The lake flora under study showed the dominance of eutrophic algae.

Higher chloride values of the lake in summer are in conformity with other

and with other Indian lakes literates.

This study DO reflects the photosynthetic activity carried out by

phytoplankton.

B.O.D. value of the lakes explains constant decomposition and supply of

nutrients throughout the season during study period.

Higher nitrates of lake during winter and lower nitrite during same period of

time reflect on the nitrite to nitrate during winter and nitrate degradation during

summer as recorded in the lake.

In the present instance the lake was represented by several groups of algae

Microsystic recorded higher during July and August; when Microsystis was absent

Chroococcus, Merismopedia, Oscillatria, were recorded. The lake was also

represented by desmids and diatoms.

The lake understudy was represented by the presence of pollutinal grade

species like, Anabaena, Microcystic, Merismopedia and Scenedesmus.

Strain isolated from Khateshwar lake at different pH. is capable of

producing antifungal compounds against pathogenic fungei and showed maximum

antifungal activity against Penicillium sp., Rhizopus sp., Candiada sp., and

Fusarium sp. at pH 10.

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List of graphs

List of photographs

List of tables