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LITERATURE REVIEW

Fungal diseases of rhizome of ginger

Fungi are significant destroyers of foodstuffs and grains during storage, rendering them

unfit for human consumption by retarding their nutritive value and often by producing

mycotoxins (Janardhana et al., 1998 and Marin et al., 1999). Eckert and Ratnayake

(1983) estimated that out of 100,000 species of fungi, less than 10% are plant pathogens

and more than 100 species of fungi are responsible for the majority of postharvest

diseases.

A huge amount of ginger is affected mostly by deuteromycetous group of fungi leading

to variable symptoms in storage condition despite its own antifungal property

(Sharififar et al., 2009; Dohroo, 2000).

In India the disease reduces potential yield to a greater extent in field, storage and

market and may cause losses of even more than 50 per cent (Joshi and Sharma, 1980).

Ginger rhizomes rot during storage was first reported in India by Merhotra in 1952.

Storage rots along with other major diseases cause loss in ginger economy (Nada et al.,

1996). Under storage condition, the white mycelium of fungi grew and covered ginger

rhizomes. The causal fungus was identified as Fusarium roseum and able to infect

through the wounds but not in healthy rhizomes. Therefore, the causal fungi that

concluded are secondary invader or would invader (Merhotra, 1952).

Rhizome rot of ginger caused by species of Pythium, was first reported by Butler in

1907, which has been a persistent threat to the cultivation of ginger in Surat (Gujrat) in

India. Although the disease is prevalent in India, Japan, China, Nigeria, Fiji, Taiwan,

Australia, Hawaii, Sri Lanka and Korea, but a very few information about the incidence

of ginger disease in abroad was reported in literature. In Nepal, the losses due to

rhizome rot in storage condition were 24% (Nepali et al., 2000). In Thailand, Tanboon-

Ek et al. (1978) reported that ginger rhizomes those stored under the experimental

storage conditions (13°C, 80% RH) for export were infected by the pathogen that

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identified as F. oxysporum. A new disease of ginger, characterized by an extensive

magenta-pink discolouration of the periderm and a dry rot of the cortical tissue which

become purple red in underground rhizome, was noticed in the yearly 1980’s and the

causal organism was later identified as Phoma hawaiiensis (Trujillo et al., 1996). Belay

et al. (2012) isolated spp. of Aspergillus, Fusarium, Penicillium, Rhizopus, Eurotium

and Mucor associated with post-harvest deterioration of Ginger rhizome samples,

collected from Hadaro-Tunto and Boloso-Bombae, Southern Ethiopia. According to

NARI (2004), infection by Fusarium sp. is typically associated with wounds or insect

and nematode damaged tissue. Small, brownish, irregular, water-soaked patches

characterize the Fusarium rot over the rhizomes and white mycelium is observed over

the infected areas (Cherian, 2002). Watery rot, caused by the fungus Rhizopus, is one of

the most rapidly developing storage rots of ginger. Symptoms include a soft watery rot

that progress rapidly and may rot the entire rhizome within a week. Infected tissue is

mottled brown and soft. In a humid atmosphere, the infected area is soon covered with

large amounts of white moulds and the mould will eventually turn black. Rhizopus is a

wound pathogen and is not effective in colonizing healthy tissue (NARI, 2004). In

different parts of Southern Ethiopia, dry rot in rhizome of ginger due to Rhizopus was

also reported by Cherian, (2002).

Several species of Pythium have been reported to cause the rot disease in different parts

of the world. P. aphanidermatum (Edson) Fitz. was responsible for soft rot of ginger in

Pusa (Bihar) (Mitra and Subramanian,1928), in Kerala (Sarma et al.,1979), in Nagpur

(Maharashtra) (Shahare and Asthana,1962) and in Madhya Pradesh (Haware and Joshi,

1974). Rhizome rot caused by P. butleri Subram. was known to exist from 1918 in the

Malabar and South Kanara districts of South India (Thomas, 1938) and it was reported

later on in Ceylone (Park,1934). The soft rot causing fungi, P. complectans Braun was

isolated from infected ginger in Ceylon (Park, 1934). P. delense Meurs was reported

from Madhya Pradesh (Haware and Joshi, 1974). P. gracile (de Bary) was found in

Bengal, Gujarat (Butler, 1907), Kerala (Sen, 1930), Assam and Fiji (Parham, 1935). P.

graminicolum Subram was reported from Ceylon (Park, 1935). In Kerala, Poona,

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Bombay, Nagpur, Taiwan, Ceylon and Hong Kong the ginger rhizome was mainly

affected by P. myriotylum Drechsler (Dake and Edison, 1989; Uppal, 1940; Patel et al.,

1949; Shahare and Asthana, 1962; Lin et al., 1971; Park, 1937 and Bertus, 1942). P.

zingiberum was reported from Osaka (Japan) and Korea (Takahashi, 1954 and Yang et

al., 1988). Simmonds (1955) described ginger yellows, a serious rot disease, for the first

time in Queensland and later in Hawaii (Trujillo, 1963) and India (Haware and Joshi,

1973). In South Africa, F. oxysporum f. sp. zingiberi was found to cause yellows

(Manicom, 1998). P. vexans de Bary was found to be responsible for fungal infection in

rhizome of ginger in Kerala (Ramakrishnan, 1949). P. pleroticum T Ito causes disease

in Solan of Himachal Pradesh (Sharma and Dohroo, 1985). P. ultimum affected the

rhizomes of ginger in Himachal Pradesh (Dohroo, 1987). In Kerala, heavy infection of

Pythium may cause losses up to 90 percent (Rajan and Agnihotri, 1989). Sinha and

Mukhopadhyay (1988) reported losses up to 50 to 90 percent due to fungal infection in

rhizomes of ginger under storage condition. In Rajasthan P. myriotylum was found in

association with Fusarium solani causing soft rot of ginger (Mathur et al., 1985 and

Doorjee, 1986). Butler and Bisby (1931) considered P. butleri and P. gracile to be

identical with P. aphanidermatum. In Madras (now Chennai), Pythium spp. occur in

association with Sclerotium rolfsii and causes rhizome rot (Anonymous, 1953). In

Himachal Pradesh, P. pleroticum was found in association with Fusarium equiseti. P.

pleroticum causes wet rot, whereas F. equiseti was responsible for dry rot under field

and storage conditions (Sharma and Dohroo, 1980). Other species of Fusarium such as

F. solani (Mart.) Sacc., F. equiseti (Corda) Sacc., and other unidentified Fusarium spp.

were also reported to be associated with the diseases of ginger rhizomes (Rosenberg,

1962). Bhardwaj et al. (1988b) reported five pathogens of rhizome rot of ginger in

Himachal Pradesh which include P. pleroticum, P. aphanidermatum, P. ultimum

(Dohroo, 1987), F. equiseti and F. solani. Sharma and Dohroo (1990) also isolated five

species of Fusarium associated with the ginger diseases in Himachal Pradesh, which

include F. solani, F. oxysporum, F. moniliforme, F. graminearum and F. equiseti.

Besides P. aphanidermatum (Edson) Fitz. (Subramanian 1919), P. myriotylum Drech.

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and P. vexans de Bary (Ramakrishnan, 1949), other pathogens reported to cause

rhizome rot are F. oxysporum Schl. f. zingiberi Trujillo from Madhya Pradesh; F. solani

(Mart.) Sacc. from Karnataka (Kumar, 1977); and Pseudomonas solanacearum E.F.

Smith from Kerala (Sarma et al., 1978; Mathew et al., 1979). Rath and Mishra (1993)

found that ginger rhizome samples collected from local markets and ginger fields of

Orissa were infected by F. oxysporum, F. equiseti, Nectria inventa, Cylindrocladium

scoparium and Cylindrocarpon sp. Sarma and Nambiar (1974) reported that

Macrophomina phaseolona was the causal organism of dry rot of ginger rhizome,

which infected in both field and storage conditions.

Sharma and Joshi (1976) reported that post-harvest diseases and pathogens of ginger

rhizomes were occurred with different symptoms such as, red rot (N. inventa), gray rot

(Trichorus spiralis) and black rot (Memnoniella echinata). In Korea, post-harvest

diseases of ginger rhizomes were found in variable symptoms and pathogens, yellow

soft rot (Erwinia carotovora and Pseudomonas aeraginosa), brown rot (F. solani and P.

aeraginosa), localized ring rot (F. solani) and water soaked rot (P. ultimum) etc. (Kim

et al., 1998). Rhizomes when infected by spp. of Pythium, turn brown and gradually

decompose, forming a watery mass of putrefying tissue enclosed by the tough skin of

rhizome. The fibrovascular strands are not affected and remain isolated within the

decaying mass (Dohroo, 1982). Penicillium developed dry rot covered with blue mould,

Aspergillus produced dry rot which is brownish yellow and dark brown, Eurotium sp.

similarly caused dry rot which is brown in colour and Mucor induced similar rots like

dry rots with black colour (Pandey, et al., 1997; Cherian.2002; Dohroo, 2005; Stirling,

2004).

Factors responsible for fungal infection

Climatic factors play a vital role in disease development and severity of pathogenesis.

Amongst the various climatic factors, temperature, relative humidity, rainfall etc. have

profound influence in initialization and spreading of pathogenic fungi. The invasion of

different storage mould at different level of relative humidity was studied by

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(Christensen, 1972) and found that it is different with different fungi. Moubeshwar et al.

(1979) found that groundnut seeds at 28oC showed maximum incidence of Aspergillus

flavus, A. niger, A.terreus, A. fumigatus, and Penicillum sp., but at 40oC temperature

only A. fumigatus and A. terreus could grow.

Malik and Singh (2004) studied the effect of temperature and relative humidity on fungi

and reported that these factors play important role in germination of spores. Wardlaw

(1972) pointed out that temperature and humidity play important role in determining the

type of decay caused by fungi in storage. Very high humidity usually favours the

growth of many fungi and causes subsequent reduction in cost of fruits and vegetables

in market (Harvey, 1978; Wadia et al., 1986). The rotting of onion is due to microbial

decay and bruising was found maximum when high temperature is coupled with high

humidity and result the losses (NHRDF 2008).

The infection of P.vexans is maximum when the temperature is low; the maximum

tolerance limit of that pathogen is being 34oC but in case of P. aphanidermatum and P.

myriotylum optimum temperature is 34oC and for P. pleroticum, it is 25 to 30

oC,

however, the optimum temperature required for proper growth and multiplication of

F.equiseti is 30oC (Dohroo,1979). A temperature range of 15

oC to 30

oC (optimum 23 to

29oC), accompanied by very high humidity is responsible for the development of yellow

disease caused by F. oxysporum f. sp. zingiberi (Sharma and Jain, 1978b).

Three traditional methods of storage of ginger i.e., storage in soil pits, dry shady place

and field by delayed harvesting were practised in Sikkim and Darjeeling against

rhizome rot (Rai and Hossain, 1998). Lana et al. (1993) found less infection in the

rhizomes that stored at room temperature (17-25°C) and 40-80 % RH than the rhizomes

stored at lower temperature (13 ±1°C) and 80 % RH. Mishra and Rath (1989) found that

the ginger rhizomes samples collected from local markets in Orissa, India was infected

by Gleocladium candidum, the pathogens were infected and caused ginger rhizomes rot

15 days after inoculation under controlled conditions (temperature, 25°C and 100 %

RH).

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Post-harvest spoilage in ginger is normally due to rough harvesting and handling

practices, which result in injury to the skin and flesh of the rhizome (FAO, 2004).

Apart from temperature, rainfall, humidity, the fungal contaminated air may be more or

less responsible for fungal infection in storage condition. A study of bio-aerosol in a

market environment showed that most of the fungi found on vegetables and fruits

originated in the field or developed during transportation (Panduranjan and

Suryanarayanan, 1985). The spoiled materials, dumping plant material and debris

present in the market places are likely to act as reservoirs of plant pathogens. The fungi

takes part in biodegradation and deterioration of the food stuff because of their

requirement for prime sources of carbon, nitrogen and other nutrients (Pitt and Hocking,

1985). Ageing and ripening induce the susceptibility to the vegetables for infection in

post harvest condition. No systematic studies on the incidence of post- harvest disease

and airborne fungal spores in the vegetable market have been published.

Change in physic-chemical constituents of rhizomes due to fungal

infection

Nutritional value of crops chiefly depends on their bio-chemical constituents. Bio-

chemical changes in post infection stages reduce the nutritional value and ultimately

decrease the market value of the crops. Most of the workers have established the fact

that both qualitative and quantitative changes occur in infected crops.

The reduction of carbohydrate, protein, oil, moisture, fibre etc. in crops during

pathogenesis has received remarkable attention from the mycologists and pathologists

during the last few decades.

Reduction in the moisture and carbohydrate contents in infected coconut may be

attributed to the fact that the fungi utilized the water and carbohydrate for metabolic

activities (Burnett, 1976).

Fungal association brings certain biochemical changes in seeds during storage by

decreasing reducing sugars and oil content (Mathur and Sinha, 1978).

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According to Hwang (1983), Patil et al., (1985) and Jeun and Hwang (1991) the

carbohydrates increase the severity of the infection and that they may serve as easily

metabolized carbon substrates for the pathogen.

Jayabalan and Rao (1983) attributed to changes in amino acids, soluble sugar and

phenolic content of different fruits.

Total phenolic contet was found to be changed in Aspergillus flavus, A. niger, Fusarium

moniliforme and Pencillium oxalicum infected chick pea (Dwivedi, 1990). It was also

reported that, although the amount of phenolic compound increased at the initial stage

of infection, later the amount was found to be decreased.

Gradual loss of protein and carbohydrate content were reported by Saxena and Karan

(1991) in sesame and sunflower seeds due to A. flavus and A. niger infection during

storage.

Azad (1991) observed that susceptible Chilli variety contains maximum sugar and

nitrogen while resistant variety shows minimum sugar and nitrogen and maximum

ascorbic acid, capsaicin, sulphur and phenols.

The carbohydrate content (sucrose, reducing sugars and polysaccharides) was

significantly decreased in fungal infected leaves of Senecio when compared with

healthy ones (Aldesuquy and Baka, 1992).

Healthy and infected fruits of brinjal were analysed by Sharma et al. (1993) and

reported progressive decrease in the quantity of total non-reducing and reducing sugar,

phenols with an increase in the infection grade.

Ndoumou et al. (1996) studied cocoa pods infected by Phytophthora megakaraya, the

causal agent of black pod, and observed variations in the content of carbohydrates,

amino acids, and phenolics. In general, higher levels of soluble sugars (SS), starch,

chlorophylls, phenolics, and tannins were observed in healthy leaves compared with

infected leaves.

Proximate analysis of healthy and fungal infected coconut fruit samples showed the

reduction in carbohydrate (17.75% healthy, 10.62% infected) and crude fibre (13.13%

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healthy, 10.01% infected) and also found the moisture content of healthy fruit (36.44%)

was higher than that of the infected fruit (10.39%) (Onifade and Jeff- Agboola, 2003).

The Alternaria blight disease of mustard adversely affects seed quality by reducing seed

size, seed discolouration and oil content (Prasad et al., 2006). The disease also reduces

germination ability and protein content of seeds.

A relative decrease in the carbohydrates, protein, fiber, fat and vitamin A and C

contents of the fruits of Colletotrichium gloeosporioids was found after infection with

Rhizopus stolonifer and Aspergillus niger causing soft-rot diseases (Nweke and Ibiam,

2012).

Parihar (2012) observed the flavonols contents decreased with increase in infection and

plant age in Brassica juncea.

Control of ginger diseases

Fungicides are the primary means of controlling post-harvest diseases. Further, the use

of synthetic chemicals to control postharvest deterioration has been restricted due to

their carcinogenicity, teratogenicity, high and acute residual toxicity, long degradation

period, environmental pollution and their direct effects on food and other side effects on

humans (Lingk, 1991; Unnikrishnan and Nath, 2002).

Another problem with these synthetic chemicals is that as their potency has been

enhanced, so have been their side effects, and their cost (Tyler, 1992; Castro et al.,

1999; Falandysz, 2000; Kast-Hutcheson et al., 2001; Sorour and Larink, 2001). In

addition, synthetic fungicides can leave significant residues in treated commodities

(Parmar and Devkumar, 1993; Fernandez et al., 2001; Dogheim et al., 2002).

Development of resistance to commonly used fungicides within populations of

postharvest pathogens has also become a significant problem (Reimann and Deising,

2000; Dianz et al., 2002). For example, many synthetic fungicides are currently used to

control blue mould rot of citrus fruit. However, acquired resistance by Penicillium

italicum and P. digitatum to fungicides used on citrus fruit has become a matter of much

concern in recent years (Fogliata et al., 2001). The side-effects of synthetic fungicides

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means that alternative strategies need to be developed for reducing losses due to

postharvest decay that are perceived as safe by the public and pose negligible risk to

human health and environment (Wilson et al., 1999). Recently, several promising

biological approaches that include microbial antagonists (Schena et al., 1999; Xi and

Tian, 2005) and plant extracts have been advanced as potential alternatives to synthetic

fungicides to control postharvest decay of fruits and vegetables.

Trichoderma as antifungal agent

Antagonistic Trichoderma species are considered as promising biological control agents

against numerous phytopathogenic fungi (Sarhan et al., 1999; Mohamed and Haggag,

2006).

The antagonistic nature of Trichoderma was demonstrated more than seven decades ago

(Weindling, 1934). There are high levels of diversity among the species of

Trichoderma. Total 104 species of Trichoderma have been recorded internationally

(www.isth.info.in) and 13 species from India which were isolated from various

substrates and locations. It was suggested that the production of antifungal metabolites,

extracellular enzymes, and antibiotics are responsible for the ability of Trichoderma to

control the growth of pathogens (El-Katatny et al., 2001; El-Katatny et al., 2006;

Shoulkamy et al., 2006; Montealegre et al., 2010). Antagonistic Trichoderma species

are successful biocontrol agent of storage fungal pathogens and it shows its efficiency at

relatively low concentration (Wisniewski and Wilson, 1992). T. harzianum strains were

generally found to be effective at low concentration of 106-10

8 conidia/ml. These

concentrations are even lower than the recommended concentrations of other biocontrol

agents (Janisiewicz, 1988; Wang et al., 2008), thus considered suitable for commercial

use. Although, Trichoderma spp. are under intensive research because of their abundant

natural occurrence, biocontrol potential against fungal and nematode diseases as well as

host defense inducing ability (Haraman and Kubicek 1998). Two strains of T.

harzianum (T3 and T24) have potential biocontrol activity against postharvest rot

caused by different fungal pathogens in cherry tomato fruit. Therefore, the use of these

isolates offer a promising, safe and effective alternative to fungicides in treatment of

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postharvest fungal diseases of tomato fruits (El-Katatny and Emam 2012). Thomas

(1939) reported the antagonistic affect of T. lignorum on Pythium sp., causal organism

of rhizome rot of ginger. Dohroo and Sharma (1984) controlled the rhizome rot caused

by P. pleroticum and F. equiseti up to 80 per cent by using T.viride under storage

condition. In vitro antifungal property of Trichoderma sp against the growth of P.

aphanidermatum, F. equiseti, F. solani, Cladosporium cladosporioides and Mucor

hiemalis were established (Bharadwaj and Gupta, 1987). The activity of P.

aphanidermatum and F. equiseti were found to be inhibited by steeping in spore

suspension or smearing with T. viride and T. hamatum (Bharadwaj et al., 1988a). The

growth of F. oxysporum f. sp. zingiberi can be inhibited by the application of

T.harzianum and Gliocladium virens (Sharma and Dohroo, 1991). According to Rathore

et al.,(1992) the soil borne antagonistic fungi, T. viride and T. harzianum have been

identified as naturally existing potential biological agent against F. oxysporum. Some

nonvolatile substance produced by T. viride might be responsible for inhibiting the

activity of P. myriotylum and F. solani (Dubey et al., 2007; Shanmugam et al.,

2008; Garcia et al., 1997). Antagonistic effect of T. harzianum, T. aureoviride and

Gliocladium virens can reduce the ginger rot caused by F. solani and P. myriotylum

(Ram et al., 2000). T. harzianum and T.viride can be considered as potent antagonists

against P. aphanidermatum (Shanmugam et al., 2000).

Phytoextract as antifungal agent

There is evidence that Neanderthals living 60,000 years ago in present-day Iraq used

plants such as hollyback (Stockwell, 1988 and Thomson, 1978); these plants are still

widely used in ethnomedicine around the world. Medicinal plants are part of human

medicine since the dawn of civilization. These plants are making backbone of

traditional medicinal systems in India (Nayak et al., 2011). The Use of plant extracts to

treat microbial infections is also reported in our ancient Ayurvedi compendium ‘Charak

Samhita’ and ‘Sushrut Samhita” (Chatterjee and Pakrashi, 1994; Atkinson and

Ramsturd, 1946; Dhar et al., 1968). Indian literature is wealthy concerning the scientific

information and knowledge about plants and their uses. Many ancient texts provide vital

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information about plants. Brihatsamhita of Varahmihira (6th century A.D.) described

the remedies for plant diseases, in the chapter No.55. ‘Vrikshayurveda’ of Surapala,

describes about various plant diseases, classifies them and suggests the possible

measure to cure them. ‘Sarngadharapaddhati’ of Sarngadhara also provides the

remedies for plant disease in 20 versus (Sadhale, 1996).

Works on the medicinal properties of the plants did not remain a monopoly of one

region. The civilization in Egypt, Greece and China acknowledge the utility of plants as

medicine (Patwardhan and Hooper, 1992). Many British people explored the plant for

their medicinal values. Around 15th and 16th century A.D. many books were published,

which gave information about ‘Herbs’ and that period is recognized as ‘Age of

Herbals’. As time passed, the scientific community tried to gather the knowledge about

medicinal properties of plants, which led to screen the available compounds present in

the plants, responsible for antipathogenic activities. Osborn (1943) screened 2300 plant

species to know the antibacterial activity against some bacteria. In 1946, Atkinson and

Ramsturd tried to evaluate antibacterial properties of 1100 species of higher plants.

Relatively 1–10% of plants is used by humans out of the estimated 250,000–500,000

species of plants on earth (Borris, 1996) and has been screened for the phytoactivity

(Oluwalana and Adekunle, 1998; Oluwalana et al., 1999; Khafagi and Dewedar, 2000).

Medicinal plants represent a rich source of antimicrobial agents (Mahesh and Satish,

2008), which may be alternatives to currently used disease control agents (Swain 1977

and Wink 1993). Some recent researches on antifungal activity of extracts of several

higher plants have indicated the possibility of their exploitation as natural antifungal

agents for control of plant diseases (Naidu and John, 1981; Gundidza, 1986; Shetty and

Shetty, 1987; Miah et al., 1990; Anwar et al., 1994; Qureshi et al., 1997). The plant-

based pesticides are locally available, non-toxic, and easily biodegradable and cheaper

than modern chemical pesticides (Mann et al., 2008; Kelmanson and Staden, 2000 and

Srinivasan et al., 2001). Due to increased prevalence of drug resistant microorganisms,

there is great need to search for new effective drugs having natural or synthetic origin

(Pai et al., 2004). Over the last two decades, intensive effort has been made to discover

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chemically useful antibacterial or antifungal drugs of plant origin, (Sofowara, 1993;

Valsaraj et al., 1997; Perumalsamy et al., 1999). There are evidences from earlier works

that several plant species possess antifungal and antibacterial properties

(Manoharachary and Gourinath, 1988; Bandara et al., 1989; Srivastava and Lal, 1997;

Maji et al., 2005; Harlapur et al., 2007; Nduagu et al., 2008; Yasmin et al., 2008 and

Akinbode and Ikotun, 2008). Plants are also the sources of natural pesticides that make

excellent leads for new pesticide development (Arokiyaraj et al., 2008; Gangadevi et

al., 2008; Satish et al., 2008; Brinda et al., 2009; Jagadish et al., 2009; Pande et al.,

2009; Shanmugavalli et al., 2009; Swarna Latha and Neelakanta Reddy, 2009; Vetrivel

Rajan et al., 2009).

A number of compounds isolated from plants such as dimethyl pyrrole,

hydroxydihydrocornin-aglycones, indole derivatives, etc., are reported to have

antifungal activities, however, development of useful antifungal drugs from these

compounds has not yet been possible (Schultes, 1978). Phytoconstituents present in

plants viz. flavonoids, alkaloids, tannins and triterpenoids are producing exciting

opportunity for the expansion of modern chemotherapies against wide range of

microorganisms (Lutterodt et al., 1999 and Marjorie, 1999). Tannins and salicylic acid

are polyphenol compounds extracted from Gaullher procumbens, Rhammus purshiand,

and Anacardium pulsatilla with high antifungal activity (Schultes, 1978).

Shivpuri et al., (1997) carried out screening of ethanol extracts of 10 plant species

(Allium cepa, A. sativum, Azadirachta indica, Calotropis procera, Datura

stramonium, Ocimum sanctum, Polyalthia longifolia, Tagetes erecta, Vinca rosea and

Withania somnifera) against five pathogenic fungi viz. Alternaria brassicola,

Colletotrichum capsici, Fusarium oxysporum, Rhizoctonia solani and Sclerotinia

sclerotiorum. Under laboratory conditions at two different concentrations (500 and

1000 µg/ml), leaf extracts of A. indica, D. stramonium, O. sanctum, P. longifolia and C.

roseus were found to be the most fungitoxic against all the test fungi. Their efficacy was

more pronounced at 1000 µg/ml.

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Oxyspora paniculata extracts showed highest inhibition agains P. aphanidermatum

while Pythium sp. was completely inhibited by Macaranga dentic-ulata extracts (Bhat,

2000).

The methanol extract, obtained from Tagetes patula plant was evaluated by Mares et

al., (2004), against three phytopathogenic fungi viz. Botrytis cinerea, Fusarium

moniliforme and Pythium ultimum and the extract proved to have a dose-dependent

activity on all the test fungi.

Begum et al.,(2007) investigated the ethanolic extract of 40 higher plant materials

(whole plant, aerial parts, leaf, root, bark and rhizome) were collected fresh from

various localities in Chittagong, Cox’s Bazar, Rangamati and Moheshkhali,

representing 23 families were tested for antifungal activity against six phytopathogenic

fungi. The two most active plants showing potent antifungal activity were Acorus

calamus and Piper betel. The rhizome extract of A. calamus exhibited highest

antifungal activity inhibiting complete mycelial growth (100%) against all the six test

pathogens. P. betel exhibited more than 50% inhibition against most of the test fungi.

The ethanolic extract of several higher plants could be used as alternative source of

antifungal agents for protection of plants or crops against fungal infection.

In 2007, Dababneh and Khalil studied the inhibitory effect of five Jordanian medicinal

plants against five plant pathogenic fungi which include Crupina crupinastrum,

Teucrium polium, Achillea santolina, Micromeria nervosa and Ballota philistaea. All

plants showed antifungal activity against the test fungi. The activity of the extracts

increased with the increase of concentration (from 100 to 1000ppm.). The highest

activity on all fungi found with Achillea santolina at 1000ppm concentration which

gave 42.2 and 42.0 percent inhibition on Fusarium oxysporum and Rhizoctonia solani

respectively. Lowest inhibition of 3.6 and 3.5 percent was found against Penicillum sp

with the application of M. nervosa and B. philistaea. The results clearly indicate that the

medicinal plants used in the study may be some promising sources of antifungal

compounds.

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The antipathogenic efficacy of the methanol extracts from 27 medicinal plant species at

concentrations of 0.5, 1 and 2 mg/mL for their in vivo activities were evaluated against

six phytopathogenic fungi which caused serious damage to crops in Korea. Their

efficacy varied with the variation of plant pathogen, tissue sampled and plant species.

The extracts of Boswellia carterii, Saussurea lappa, Glycyrrhiza uralensis, Piper

nigrum, Rheum coreanum, Lysimachia foenum-graecum, Evodia officinalis, Santalum

album and Curcuma longa at 2 mg/mL showed very strong fungicidal activity. At 1

mg/mL, S. album, P. nigrum and L. foenum-graecum showed potent fungicidal activity

against Blumeria graminis f. sp. hordei, Puccinia recondita and Magnaporthe grisea,

respectively whereas, L. foenum-graecum exhibited strong fungicidal activity against M.

grisea at 0.5 mg/mL (Park et al., 2008).

By using agar well diffusion method, Bobbarala et al.,(2009) evaluated the antifungal

activity of the methanolic extracts prepared from forty nine different plants collected

from different places in Visakhapatnam district of Andhrapradesh, used in traditional

Indian medicine, against Aspergillus niger. Among the forty-nine plants, 86% of the

plants had antifungal activity while the remaining 14% had no activity. Four plants

Grewia arborea, Melia azadiracta, Peltophorum pterophorus and Terminalia chebula,

showed exceptionally remarkable antifungal properties, and among them the extract of

G. arborea showed maximum activity even at lower concentration.

Schuster et al., (2010), used ethanolic plant extract from Glycyrrhiza glabra (2.5% w/v)

against different important plant pathogenic fungi. In a previous investigation he could

able to report 100% efficacy against late blight (Phytophthora infestans) on detached

tomato leaves. Against another Oomycetes, Pseudoperonospora cubensis, on cucumber,

application of a 2.5% extract led to an efficacy of above 90%. In a trial on beans against

bean rust (Uromyces appendiculatus), G. glabra extract (5% concentration) showed

92% efficacy. Overall, the results indicate a high potentiality of G. glabra extract to

control a number of important plant pathogens.

10 plant species used in traditional Uruguayan medicine, were screened by Dellavalle et

al. (2011), to evaluate the antifungal activity against the phytopathogenic fungus

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Alternaria spp. The plants were selected based on their reported ethnobotanical uses. In

vitro antifungal activity of aqueous, saline buffer and acid extracts of different plant

species were examined against Alternaria spp. 31% of the extracts inhibited growth,

which is almost similar to the effects of a chemical fungicide. Acid extracts of the plants

were more effective than the aqueous or buffer extracts against Alternaria spp. MICs

and MFCs values obtained from leaves (Salvia officinalis L. and Rosmarinus

officinalis L.) and seeds extracts (S. sclarea L.) were quite comparable to values

obtained with the conventional fungicide Captan (2.5 µg mL-1). The extracts of S.

sclarea, S. officinalis and R. officinalis could be considered as potential sources of

antifungal compounds for treating fungal diseases in plants. These extracts showed

maximum activity, even at very low concentrations and their strong fungicidal

properties support the traditional use as antiseptics.

Tapwal et al. (2011) investigated the antifungal activity of aqueous extract of Cannabis

sativa, Parthenium hysterophorus, Urtica dioeca, Polystichum squarrosum and

Adiantum venustum was investigated against Alternaria solani, A. zinniae, Curvularia

lunata, Rhizoctonia solani and Fusarium oxysporum at different concentrations (5, 10,

15 and 20%). At 20% concentration, maximum antifungal potential was observed with

the extracts of C. sativa, which recorded excellent inhibitory activity against C. lunata

(100%), A. zinniae (59.68%), followed by leaf extract of P. hysterophorus (50%)

against A. solani.

Bhardwaj et al. (2012) evaluate the antifungal activity of aqueous extracts from 20

plants against Coriolus versicolor, wood rotting fungi. Results showed varying activity

of the plant extracts against the mycelium growth. The combined seed extracts

of Azadirachta indica and rhizome extracts of Curcuma domestica in general showed a

strong enhancement in activities over the individual seed extracts of A. indica and

rhizome extracts of C. domestica against the mycelium growth. The seed extracts

of Albizia stipulata and seed extracts of Acacia arabicae showed strong inhibitory

effect against the test fungi. The leaf extracts of Adhatoda vasika, leaf extracts of Aegle

marmelos, whole plant extracts of Cuscuta reflexa, leaf extracts of Clerodendron

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inerme and root extracts of Acacia catechu showed appreciable good inhibitory effect

against the test fungi.

In vitro studies were carried out by Hadian (2012) to test the antifungal activity of 5

plant extracts of Azadirachta indica, Melia azadiracta, Allium sativum, Curcuma longa

and Caryophilium aromaticus and found concentration dependent activities of the

extracts. All plant extracts except C. aromaticus showed significant reduction in the

growth of species of Fusarium and Rhizoctonia. A. indica and A. sativum, extract

(100% concentration) were the most effective to inhibit the growth of tested fungi.

Findings from this study confirmed that plant extract can be used as natural fungicides

to control pathogenic fungi, thus reducing dependence on the synthetic fungicides. A.

indica, which was found to be the most efficient extract, 98% inhibition on Fusarium

and 96% inhibition on Rhizoctonia could be a promising material for controlling these

fungi.

Al-Askar and Aziz in 2012, screened the in vitro antifungal activity of Alhagi

maurorum Medic, Capparis spinosa L. and Punica granatum L, against Alternaria

alternate, F. oxysporum, P. destructive, R. solani and Sclerotium rolfsii at concentration

of 0, 3, 6 and 9% (v/v). All the plant extracts prepared from seeds, roots and rinds had

different degree of antifungal activity against the tested fungi. When compared with the

control, the highest antifungal activity was found in case of A. maurorum seed extract at

a concentration of 9% while at 9% P. granatum rinds extract showed less activity. The

ethanolic extract of A. maurorum seed may be recommended as potent bio-fungicide.

A significant antifungal activity was reported by Singh et al. in 2013 when screening

was done by using acetone, methanol, benzene, ethyl acetate and chloroform extracts of

five plants viz., Foeniculum vulgare, Trachyspermum ammi, Cuminum cyminum,

Syzygium aromaticum and Cinnamomum tamala against 3 Candida species.

Plants used in the present study for inhibitory effect

Andrographis paniculata Burm.f.

Andrographis paniculata (Burm.f.) Wall ex. Nees (Family-Acanthaceae) is an erect

branched annual herb, 0.3-0.9m in height with quadrangular branches. Leaves are

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simple, lanceolate, acute at both ends, glabrous, with 4-6 pairs of main nerves. Flowers

are small, pale but blotched and spotted with brown and purple distant in lax spreading

axillary and terminal racemes or panicles. Calyx-lobes are glandular pubescent with

anthers bearded at the base. Fruits are linear capsules and acute at both ends. Seeds are

numerous, yellowish brown and sub-quadrate (Warrier et al, 1993). It contains

flavones- echiodinin and its glucoside-echioidin (Husain et al, 1992).

Leaves contain two bitter substances lactone “andrographolid” and “kalmeghin”.

Kalmeghin is the active principle that contains 0.6% alkaloid of the crude plant. The

plant is antifungal, antityphoid, hepatoprotective, antidiabetic and cholinergic. Shoot is

antibacterial and leaf is hypotensive (Garcia et al., 1980). This is used for the

inflammation of the respiratory tract. The plant demonstrated high antimalarial effect in

vitro and in vivo (Nik Abdul Rahman et al., 1999) and as a remedy for snakebite

(Selvanayagam et al., 1994).

Callistemon linearis DC.

Callistemon linearis DC. (Family- Myrtaceae) is a small, evergreen tree with rough

fissured bark and dropping branches. It is native to the states of New South

Wales and Queensland in Australia. Although it is grown in India as an ornamental

plant, it has also great medicinal value. It is also known as the narrow-leaved

Bottlebrush because of their cylindrical brush like flowers resembling a traditional

bottlebrush. It grows up to around 3 metres in height and has a stiff appearance. It has

very narrow leaves up to 100-120 mm long, with a rigid point. Spikes having red flower

are about 10-11 cm long and 6-7 cm wide. Blooms appear in late spring and summer.

Phytochemical studies of different Callistemon species revealed that the presence of

different monoterpenes, sesquiterpenes flavonoids. There are several reports of the oil

exhibiting fungi toxicity, inhibit the growth of cowpea mosaic virus, mung bean mosaic

virus (Aburjai and Hudaib, 2006). The ethnic tribal communities have been using this

plant for many generations and information regarding the efficacy remains primarily

anecdotal. The methanolic extract of leaf showed good antimicrobial activity against

gram‘-’ and gram‘+’ bacteria and moderate activity against fungal strains (Seyyednejad

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and Motamedi, 2010). In preminary antifungal assay of ethanol extract showed

moderate zone of inhibition against Candida albicans and weak activity against

Aspergillus aegyptiacus. The seeds of a sample from Japan yielded an oil containing ß-

sitosterol (Kanjilal and Das, 1992 and Kokate, 2001) and 1, 8-cineole has been reported

as their major constituent in the oil. It has been proved that the volatile oil obtained

from C. linearis leaf contained neither antifungal nor antibacterial properties against

some strain of microorganisms. (Das et al., 2009).

Cinnamomum porrectum Roxb.

Cinnamomum porrectum (Roxb.) (Family- Lauraceae), a native of India and Malesia, is

a lofty tree, 9-30 meters tall with whitish rough bark. Young leaves are red, the adult

leaves are dark green, glaucous beneath, elliptic-ovate, acute or acuminate, base acute

or round; 5-10 cm long, 2.5-4.5 cm wide; petioles slender, 2.5-3.2 cm. long (Kirtikar

and Basu,1980). The inflorescence is an axillary or pseudo-terminal panicle measuring

2.5-15 cm long. The flowers are smooth or sparingly hairy. The fruit is spherical to

slightly depressed spherical, measuring 0.8-1 cm across and seated on a funnel-shaped

perianth cup and with an entire margin. Its geographical distribution is limited to India,

Burma (Myanmar), Thailand and southern China, Peninsular Malaysia, Singapore,

Sumatra, Java and Borneo.

The leaves of this particular species were used as carminative, tonic, and stomachic in

Thai local markets (Pongboonrod, 1976). Many parts of this plant, such as roots, bark,

wood, and leafy branch lets contain camphor and volatile oil. The wood is used for

furniture and cabinets due to its fine grain. Silkworm consumes the leaves. The fruit

kernel has fat and oil up to 60% and major component oil is safrole which is utilized in

soap production (Xiwen et al., 2008) and also in perfumes, foods and drinks. The oil

from root obviously has antibacterial and antifungal activities (Phongpaichit et al.,

2007). The oil from C. porrectum has been used as a topical antiseptic and pediculicide,

but it may also be carcinogenic (Rocha and Ming, 1999). The methanolic extracts

heartwood of this plant showed moderate antifungal activity against a brown rot fungus,

Gloeophyllum trabeum (Kawamura et al., 2011).

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Kaempheria galanga Linn.

Kaempheria galanga L. (Family-Zingiberaceae) is a small perennial herb that grows

abundantly in the lowlands or mountains particularly in southern China, Indochina,

Malaysia, India and Bangladesh (Kanjanpothi et al., 2004). The plant is characterized

by the absence of stem and presence of dark brown, rounded rhizomes having a specific

aroma. The number of strands of K. galanga leaves no more than 2-3 pages dealing

with the composition. The flowers are arranged and a half sitting with a crown of

flowers amounted to between 4 to 12 pieces, violet flowers with white colour is more

dominant. It grows and develops in the rainy season.

Being a source of valuable bioactive compounds, the plant is famous for its medicinal

as well as edible use (Techaprasan et al., 2010). It is one of those medicinal herbs which

are still comparatively less known and are underutilized (Peter, 2004). It is also used in

Chinese cooking and Chinese medicine, and is sold in Chinese groceries under the name

Sha jiang (Van Wyk and Ben-Erik, 2005). The rhizome of this plant has been used for

the treatment of allergy and gastro intestinal disorders as well as an aphrodisiac and for

fungal infections (Pengcharoen, 2002). The rhizomes of the plant, which contains

essential oils, have been used in a powdered form for indigestion, cold, pectoral and

abdominal pains and headache. Its alcoholic maceration has also been applied as

ointment for rheumatism (Keys, 1976 and Lieu, 1990). Medhi et al. (2012) reported the

antifungal activity of the acetone, chloroform, methanol, petroleum ether and water

extracts of the rhizome of K. galanga, against six fungal pathogens of Ginger. The

extract of the plant was also found to be effective against Curvularia sp. The methanol

and petroleum ether crude extracts exhibited an antibiotic potential against Escherichia

coli. (Parvez et al., 2005).

Naravelia zeylanica Linn.

Naravelia zeylanica (L) DC. (Family-Ranunculaceae) is a woody climber with tuberous

roots; wiry stem strong tendrils, leaves 3-foliate, opposite, terminal leaflet modified

into a 3 branched tendril, leaflets ovate-lanceolate, serrate or crenate, prominently

nerved; opposite, ovate, cordate leaflets; small flowers arranged in panicles and flowers

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yellow and fragrant. The plant occurring wild in the warm regions of Eastern

Himalayas, Assam, Bengal, Bihar and greater parts of Deccan Peninsula in India

particularly and in Indo-Malaysian region (The Wealth of India, 1998).

In Ayurveda it is mentioned that the plant has been extensively used by native people as

an astringent, bitter, antipyritic and anti-inflammatory (Harsha et al., 2003). It is also

useful in pitta, helminthiasis, dermatopathy, leprosy, rheumatalgia, odontalgia,

cephalalgia, colic inflammation, wound healing and ulcer protection (Sivarajan and

Balachandran, 1958). The root and stem have a strong penetrating smell and is used to

relieve malarial fever and headache. Root and stem paste is applied externally for

psoriasis, itches and skin allergies (Anis et al., 2003). The traditional medicine

practioners using the leaf and stem juices for treating intestinal worms, psoriasis and

dermatitis (Saldanha and Nicolson, 1976). Plant contains alkaloids, berberine and

sterols (Gagliardo et al., 2003). The ethanolic and benzene extracts of leaves of N.

zeylanica showed significant antimicrobial activity on some pathogenic bacteria

(Lalitha and V.Alex, 2011). The leaf extracts of leaves found to have antifungal

property against five pathogenic fungi of ginger (Kotoky et al., 2012). Uvarani et al., in

2012 reported that the ethanolic extract was exhibited moderate activity against

Salmonella typhi and Staphylococcus aureus. The potent antibacterial activity may be

due the presence of phytoconstituents such as flavanoids, triterpenoids and phenolic

compounds in this plant.

Pongamia pinnata Linn.

Pongamia pinnata Linn. (Family-Leguminaceae), a medium sized glabrous, perennial

tree grows in the littoral region of South Eastern Asia and Australia (Satyavati et al.,

1987; Allen and Allen, 1981). Leaves imparipinnate, shiny, flowers fragrant, white to

pinkish, paired along rachis in axillary, pod short stalked, oblique-oblong, flat, smooth,

thickly leathery to subwoody, indehiscent, 1-seeded; seed thick, reniform (Allen and

Allen, 1981). It is a preferred species for controlling soil erosion and binding sand

dunes because of its dense network of lateral roots. Plants growing in arid zones are

good source, not only for fiber, food and feed but also for the production of various

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types of secondary metabolites like flavonoids, steroids, antifertility compounds which

make them resistant against drought, salinity and pathogen (Sharma et al., 2011). All

parts of the plant have medicinal properties and traditionally used as crude drug for the

treatment of tumor, piles, skin diseases, wound and ulcers (Tanaka et al., 1992). In the

traditional system of medicine, such as Ayurveda and Unani, P. pinnata is used as anti-

inflammatory, anti-plasmodial, anti-nonciceptive, anti-hyperglycamic, anti-

lipidperoxidative, anti-diarrhoeal, anti-ulcer, anti-hyperammonic due to its antioxidant

activity (Chopade et al., 2008). Chandrashekar and Prasanna in 2010 reported the

antimicrobial activity of P. pinnata leaf extract against Escherichia coli, Staphylococcus

aureus, Aspergillus niger, Candida albicans, Pseudomonas aeruginosa and Salmonella

typhi. Free flavonoid fractions of leaves of P.pinnata also showed significant activity

against eight bacterial and two fungal strains viz. Bacillus cereus, E. coli,

Mycobacterium smegmatis, Proteus vulgaris, P. aeruginosa, S. typhimurium, S. aureus,

S. epidermidis, C. albicans and Trichoderma viride (Sharma et al., 2011).

Solanum indicum Linn.

Solanum indicum Linn. (Family-Solanaceae) is found throughout the tropics. It is a

spiny shrub, found wild along roadsides, forest margins and forest floors throughout

North East India (Handique, 2009). It is much branched, very prickly undershrub,

leaves are simple, large, and ovate, and flowers are blue in extra-axillary cymes having

stellately hairy and prickly peduncles. Fruits are globose berries, reddish or dark yellow

with smooth or minutely pitted seeds.

Leaves of the plant are eaten as vegetables, the juice from the leaves is applied

externally on skin problem. In the Ayurveda and Yunani medicinal system, the fruit is

used in pruitus, leucoderma, bronchitis, asthama, fever, vomiting, and loss of appetite

and diseases of the eye. Half - ripe fruits are employed in the preparation of curries and

chutneys. Its roots are useful in vitiated conditions of vata and kapha, odontalgia,

dyspepsia, flatulence, colic, verminosis, diarrhoea, pruritus, leprosy, skin diseases,

strangury, cough, asthma, bronchitis, amenorrhoea, dysmenorrhoea, fever, cardiac

disorders, vomiting etc. (The useful plants of India, 1992; Kumar, 2009). Roots bitter,

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acrid, astringent, thermogenic, anodyne, digestive, carminative, anthelmintic,

stomachic, constipating, resolvent, demulcent, depurative, diuretic, expectorant,

aphrodisiac, emmenagogue, febrifuge and cardiotonic. The chloroform, methanol and

aqueous leaf extracts of S. indicum were found to be effective against five

dermatophytic fungi (Kotoky et al., 2012).