Compendium on Integrated Disease Management

Compendium on Two Days Training Programme on

Integrated Disease Management (18-19 February, 2021) Course coordinator Dr. Ranbir Singh

Organized by SAMETI-J in collaboration with Division of Plant Pathology

Sher-e-Kashmir University of Agricultural Sciences and Technology of Jammu

Compendium on Two Days Training Programme on

Integrated Disease Management (18-19 February, 2021)

Course coordinator Dr. Ranbir Singh Compiled by Ranbir Singh Uma Shankar S.K. Singh

Organized by SAMETI-J in collaboration with Division of Plant Pathology

Sher-e-Kashmir University of Agricultural Sciences and Technology of Jammu

Contents Chapter(s) Title(s)

Page Nos.

1

Integrated Disease Management

1-5

Dr. Ranbir Singh 2

IDM of Major Horticultural Crops

6-11

Dr. A. K. Singh 3

Integrated Pest Management in Major Rabi Crops

12-26

Dr. Uma Shankar 4

Integrated Disease Management in wheat

27-32

Dr. S. K. Singh 5

Role of Biological Control in Plant Disease Management

33-47

Dr. Vishal Gupta 6

Diseases of Mushroom and their Management

48-54

Dr. Sachin Gupta 7

IDM of Vegetables and Floricultural crops

55-62

Dr. Amrish Vaid 8

Isolation, Identification and Culturing, of Plant Pathogens (Practical) Dr. Ranbir Singh

63-68

Chapter-1

Integrated Disease Management Ranbir Singh, Sachin Gupta, S. K. Singh, A. K. Singh and Uma Shankar Division of Plant Pathology, Faculty of Agriculture SKUAST-Jammu _____________________________________________________________________________ Introduction Integrated disease management can be defined as a decision-based process involving

co-

ordinated use of multiple tactics for optimizing the control of pathogen in an ecologically and economically manner. In most cases IDM consists of scouting with timely application of a combination of strategies and tactics. These may include site selection and preparation, utilizing resistant cultivars, altering planting practices, modifying the environment by drainage, irrigation, pruning, thinning, shading and applying pesticides, if necessary. But in addition to these traditional measures, monitoring environmental factors (temperature, moisture, soil pH, nutrients, etc.), disease forecasting, and establishing economic thresholds are important to the management scheme (Khoury and Makkouk, 2010). These measures should be applied in a coordinated integrated and harmonized manner to maximize the benefits of each component. The basic objectives of any IDM program should be to achieve at least the following: 1.

Reduce the possibility of introducing diseases into the crop

2.

Avoid creating conditions suitable for disease establishment and spread

3.

Simultaneous management of multiple pathogens

4.

Regular monitoring of pathogen effects, and their natural enemies and antagonists as well

5.

Use of economic or treatment thresholds when applying chemicals

6.

Integrated use of multiple, suppressive tactics.

Principles of Plant Disease Control 1.

Avoidance—prevents disease by selecting a time of the year or a site where there is no inoculum or where the environment is not favorable for infection.

2.

Exclusion—prevents the introduction of inoculum.

3.

Eradication—eliminates, destroy, or inactivate the inoculum.

4.

Protection—prevents infection by means of a toxicant or some other barrier to infection.

5.

Resistance—utilizes cultivars that are resistant to or tolerant of infection.

1

6.

Therapy—cure plants that are already infected

Advantages of Integrated Disease Management Integrated approach integrates preventive and corrective measures to keep pathogen from causing significant problems, with minimum risk or hazard to human and desirable components of their environment. Some of the benefits of an integrated approach are as follows: 

Promotes sound structures and healthy plants



Promotes the sustainable bio based disease management alternatives.



Reduces the environmental risk associated with management by encouraging the adoption of more ecologically benign control tactics



Reduces the potential for air and ground water contamination



Protects the non-target species through reduced impact of plant disease management activities.



Reduces the need for pesticides and fungicides by using several management methods



Reduces or eliminates issues related to pesticide residue



Reduces or eliminates re-entry interval restrictions



Decreases workers, tenants and public exposure to chemicals



Alleviates concern of the public about pest & pesticide related practices.



Maintains or increases the cost-effectiveness of disease management programs

Components of Integrated Disease Management The major components of disease management are host-plant resistance, cultural practices, biological control and chemical control. Even though these components will be dealt with individually, it should be mentioned that often the different components are complementary to each other with strong interaction among and between them and the environment. The major components of Integrated Disease Management are Cultural practices Cultural practices such as cultivation techniques, mulching, intercropping, plant density, planting date, crop rotation, strip farming, timing of harvest, barrier crops, crop mixtures, roguing, healthy planting material, soil solarization, soil amendments, fertilizer management, and water management have been used singly and in combination as tools for disease management.

Cultural control methods not only serve in promoting the healthy growth of the crop, but are also 2

effective in directly reducing inoculum potential and in enhancing the biological activities of antagonists in the soil. Crop rotation is a very important practice, especially for soil borne disease control. For many soil borne diseases, at least a 3-year-rotation using a non-host crop reduces pathogen populations. This practice is beneficial for Phytophthora blight of pepper and Fusarium wilt of watermelon. Vegetable fields should be located as far as away as possible from inoculum and insect vector sources. Weed control is important for the management of viral diseases. Weeds may be alternate hosts for several important vegetable viruses and their vectors.

Elimination of weeds might reduce primary inoculum. Cover crops help to reduce weed populations that may harbor pathogens between seasons. For this purpose use cover crops that grow fast and provide maximum biomass. Excessive handling of plants such as thinning, pruning and tying may be involved in spread of pathogens, particularly bacteria. Because some pathogens can only enter the host through wounds, situations which promote plant injury should be avoided. During pruning process and harvest, workers should periodically clean their hands and tools with a disinfectant, such as isopropyl alcohol. If applicable, plants can be staked and tied for improved air movement in the foliar canopy. A more open canopy results in less wetness, discouraging growth of most pathogens. Soil aeration and drying can be enhanced through incorporation of composted organic amendments in the soil. Build up of inoculum can be reduced by removing all plant materials (infected and apparently healthy) after harvest. Betweenrow cover crops reduce plant injury from blowing sand. Polyethylene mulch can be used as a physical barrier between soil and above-ground parts of plants. This is an important practice for fruit rot control in the field. Highly UV-reflective (metalized) mulches repel some insects. It is beneficial to use metalized mulch during certain times of the year when insect vectors of some viral diseases are prevalent. Tomato spotted wilt virus (TSWV) incidence and associated vector thrips populations have been demonstrated to be effectively reduced by using metalized mulches on tomatoes. Plant nutrition and soil pH can also impact some diseases. Fertilizers with a higher proportion of nitrate nitrogen (NO3) than ammoniacal nitrogen (NH4) will help to reduce the incidence of Fusarium wilt on tomato. Increasing soil pH by liming is a good management strategy to reduce Fusarium wilt incidence as well as Botryis gray mold severity. Optimum calcium nutrition and higher soil pH may reduce the incidence of bacterial wilt in the field.

Adequate calcium is necessary to minimize blossom end rot and to provide for overall healthy growth. Avoiding excessive nitrogen leads to less dense canopies, thus improving air movement 3

in the canopy. High soil moisture enhances the development of soil borne pathogens including Phytophthora, Pythium and the bacterial wilt pathogen. Excess water damages roots by depriving them of oxygen and creates conditions that favor infection by certain soil borne pathogens.

Irrigation management, based on plant needs, will help to create an environment unfavorable for pathogen survival and disease development.

Chemical control For many decades fungicides played an important role in disease control. In the1960s, systemic fungicides started gradually to replace the older non-systemic chemicals with more effectiveness and specificity in disease control. Very quickly, triazole fungicides gained 24% of the total fungicides market (Hewitt, 1998). However, the non-systemic fungicides such as mancozeb and chlorothalonil plus copper and sulpher-based products continued to have a good share of the market, especially in developing countries because of their lower cost.

Biological control Success in using microorganisms against plant pathogens started with the control of crown gall with Agrobacterium radiobacter K84 (Kerr, 1980), and that of seedling blights caused by Pythium and Rhizoctonia with Trichoderma harizanum (Harman and Bjorkman, 1998), Gliocaladium virens (Lumsden and Walter, 1995) and Streptomyces griseus (Cook et al., 1996).

However, the use of naturally occurring bio-control agents (antagonists) of plant pathogens can be traced back to many centuries through the traditional practice of crop rotations that primarily permit the reduction of pathogens’ inoculum potential in the soil below injury level. This approach is still the most important single component, in both developed and developing countries used to manage root pathogens. This process is often accelerated by adding composts or manures, which enrich the soil with antagonistic microflora (Baker and Cook, 1974).

Host-plant resistance Host plant resistance is an important tool to control diseases of major food crops in developing countries, especially wheat, rice, potato, cassava, chickpea, peanuts and cowpea. The use of resistant varieties is very much welcomed by resource poor farmers because it does not require additional cost and it is environment-friendly. Rice varieties resistant to rice blast, bacterial blight, rice tungro and brown spot are widely used. Rusts have been known to cause serious

4

disease on wheat since its domestication. The use of genetic resistance is still the most economic and feasible mode of disease control.

Conclusion Integrated disease management (IDM) is a disease control approach that uses all available management strategies to maintain disease pressures below an economic injury threshold. It does not advocate a routine chemical application program to prevent disease, but promotes the integration of cultural, physical, biological and chemical control strategies.

5

Chapter- 2

Integrated Diseases Management of Major Horticultural Crops Ashok K. Singh, S. K. Singh, Ranbir Singh and Sachin Gupta Division of Plant Pathology, SKUAST of Jammu ______________________________________________________________________________ Introduction The Horticulture (fruits including nuts, and plantation crops medicinal) has become a key drivers for economic development in many of the states in the country and it contributes 30.4 per cent to GDP of agriculture, which calls for knowledge and technical backstopping. Intensive cultivation of the high valued horticultural crops, resulted in the outbreak of several diseases of National importance. In recent days, stakeholders import planting materials from North American Countries. Introduction of planting materials also impose threat in the introduction of new diseases not known to be present earlier. However, the diseases, if not managed on a war foot, it will result in drrastic yield reduction and quality of the produces. Hence adoption of suitable management measures with low residue levels in the final produces becomes as a need of the hour and gives emphasis on the diagnosis of plant diseases and their management.

Diseases of Mango and their Management 1. Anthracnose Anthracnose symptoms occur on leaves, twigs, petioles, flower clusters (panicles), and fruits. On leaves, lesions start as small, angular, brown to black spots and later enlarge to form extensive dead areas. Panicles develop small black or dark-brown spots, which can enlarge, coalesce, and kill the flowers .Petioles, twigs, and stems are also susceptible and develop the typical black, expanding lesions. Fruits may also drop from trees prematurely due to rotting. Green fruit infections that take place at mature stage remain latent and invisible until ripening and carry the fungus into storage. This disease is caused by Colletotrichum gloeosporioides.

Integrated Management 

Proper sanitation of orchard by periodical removal of fallen plant debris and pruning of trees eradicates the fungus and checks further spread of the disease.

6



Maintaining tree vigour with proper irrigation and fertilization.



Fungicide sprays should begin when panicles first appear and continue at the recommended intervals until fruits are picked.



Spraying the trees twice with Carbendazim (0.1%) or Mancozeb (0.2 %) or combination of Carbendazim 12 % + Mancozeb 63 % @ 0.1 % at 15 days interval during flowering to control blossom infection and twice during pea nut stage to prevent fruit infection.



Alternate sprayings of Carbendazim and Mancozeb to avoid development of resistance in pathogen to fungicides.



For post harvest anthrcanose, fruits are dipped in hot water at 50 C for 30 min. in combination with 0.05 % carbendazim.

2. Die back / Fruit Stem end rot The disease is characterized by drying of twigs and branches followed by complete defoliation, which gives the tree an appearance of scorching by fire. Tip die back disease occurs on the branches, trunk of infested trees that start drying slowly at first and suddenly branches become completely dried /killed resulting gummy substance oozes out or remains hanging on the tree .The dark area advances and young green twigs start withering first at the base and then the twig or branch dies, shrivels and falls called die back. This may be accompanied by exudation of gum.

In old branches, brown streaking of vascular tissue is seen on splitting it longitudinally. Disease caused by Lasiodiplodia theobromae.

Stem end rot appears as rotting from pedicel end of fruit during ripening until pathogen remains latent forming appressoria that remains quiescent in the subcuticular layer in green fruits.

Integrated Management 

Pruning of infected plant parts from 7- 10 cm below the infection site and pasting the cut ends with Bordeaux paste.



Spraying the trees twice at 15 days interval with Carbendazim (0.1%) or combination of Carbendazim 12 % + Mancozeb 63 % @ 0.1 % during pea nut stage to prevent fruit infection.



Fruits should be harvested with stalk (5 cm), otherwise, the opening must be sealed with wax.

3. Powdery mildew 7

The disease affects inflorescence, leaves and young fruits. Appearance of a whitish, powdery growth of the fungus on leaves, panicles and young fruit which later turns brown and fall. The white growth can also be seen on the undersurface of young infected leaves which becomes distorted. Severe infection of young leaves results in premature leaf drop. On mature leaves, the spots turn purplish brown, as the white fungal mass eventually disappears. On developing inflorescence powdery growth leads to drying of flowers . Young fruits at peanut stage are covered with mildew that leads to corky tissue and drops. The disease is caused by Oidium mangiferae.

Integrated Management 

Pruning of diseased leaves and panicles.



Three sprays of fungicides at different stages starting with Wettable Sulphur (0.2%) at the time of panicle initiation followed by Dinocap (0.1%) subsequently followed Tridemorph (0.1%) at 15-20 days interval.



Sparying with mycobutanil @0.1% or Triademefon @0.1% or carbendazim @0.1% or Thiopahante methyl 0.1% found effective against disease

4.Grey leaf Blight Brown spots develop on the margin and at the tip of the leaf lamina which coalesce covering the leaf margin and becomes dark brown. Black dots appear on the spots which are acervuli of the fungus. If infection starts from tip, it advances on either side of mid rib and within 3-4 months severe defoliation results. This disease is caused by Pestalotia mangiferae.

Integrated Management 

Removal of infected plant parts. Spraying one time with Copper oxychloride @ 0.25 % or Mancozeb @ 0.25% or Bordeaux mixture @ 1.0% at the visual appearance of disease.

5 . Sooty Mould The disease is common in the orchards where mealy bug, scale insects and hoppers are not controlled efficiently. Honey dew secretion by insects make the fungi produce mycelium which is superficial and dark and forms black encrustation on leaves. In severe cases, the trees turn completely black due to the presence of mould over the entire surface of twigs and leaves. The severity of infection depends on the honey dew secretion of the above insects. Presence of a black sooty mould on the leaf surface adversely affects the photosynthetic activity of the leaf and

8

thereby fruit set is reduced. Causal organism of the disease is Capnodium mangiferae, C. ramosum.

Integrated Management 

Pruning of affected branches and their prompt destruction .Spraying systemic insecticides like to control insects.



Spraying of 5 per cent starch (1kg Starch/Maida in 5 litres of water. Boiled and dilute to 20 liters) helps to control the disease as dried starch flakes removes the fungus.

6. Mango malformation Three types of symptoms: bunchy top phase, floral malformation and vegetative malformation.

In bunchy top phase in nursery at 40-5 months old. Bunching of thickened small shoots, bearing small rudimentally leaves. Shoots remain short and stunted giving a bunchy top appearance. In vegetative malformation induces excessive vegetative branches of limited growth in seedlings.

They are swollen with short internodes forming bunches of various size and the top of the seedlings shows bunchy top appearance. In malformation of inflorescens, shows variation in the panicle. Reduction in length of primary axis and secondary branches of panicle makes the flowers to appear in clusters. Secondary branches are transformed into number of small leaves giving a witches broome appearance. Malformed head dries up in black mass and persist for long time. Such panicles dot not bear. The infection is localized. The disease is caused by Fusarium moniliforme var. subglutinans.

Integrated Management 

Diseased plants should be destroyed.



Use of disease free planting material



Incidence reduced by spraying 100-200ppm NAA during October.



Pruning of diseased parts along the basal 15-20 cm apparently healthy portions followed by the spraying of Carbendazim (0.1%).

Diseases of Citrus 1. Gummosis 9

The symptoms appear as yellowing of leaves, followed by cracking of bark and profuse gumming on the surface. The main source of infection is infected planting material. As a result of severe gumming, the bark becomes completely rotten and the tree dries owing to girdling effect. Prior to death, the plant usually blossoms heavily and dies before the fruits mature. In such cases, the disease is called foot rot or collar-rot. The disease caused by Phytophthora parasitica, P. palmivora,P. citrophthora.

Integrated Management 

Preventive measures like selection of proper site with adequate drainage, use of resistant rootstocks and avoiding contact of water with the tree trunk by adopting ring method of irrigation are effective.



Alternatively the disease portions are scraped-out with a sharp knife and the cut surface is disinfected with Mercuric chloride (0.1%) or Potassium permanganate solution (1%) using a swab of cotton.



Painting 1 m of the stem above the ground level with Bordeaux helps in controlling the disease.



Also spraying and drenching with Ridomil MZ 72@ 2.75 g/l or Aliette (2.5 g/l) is effective in controlling the disease.

2. Canker Acid lime, lemon and grapefruit are affected. Rare on sweet orange and mandarins. Affects leaf, twig and fruits. In canker, leaves are not distorted. Lesions are typically circular with yellow halo; appear on both sides of leaf, severe in acid lime (difference from scab). When lesions are produced on twigs, they are girdled and die. On fruits, canker lesions reduce market value.

Disease is caused by bacterium Xanthomonas campestris pv citri.

Integrated Management 

Spray Streptomycin sulphate 500-1000 ppm or Phytomycin 2500 ppm or Copper oxychloride 0.2% at fortnight intervals.



Control leaf miner when young flush is produced.



Prune badly infected twigs before the onset of monsoon.

Diseases of Guava 1. Anthracnose

10

Symptoms of this disease are observed on mature fruits on the tree. The characteristic symptoms consist of sunken, dark colored, necrotic lesions. Under humid conditions, the necrotic lesions become covered with pinkish spore masses. As the disease progresses, the small sunken lesions coalesce to form large necrotic patches affecting the flesh of the fruit. This disease is caused by Colletotrichum gloeosporioides.

Integrated Management 

Spray with Mancozeb @ 0.25% at appearance of symptoms.

2. Algal leaf spot Disease symptoms are exhibited on both abaxial and adaxial leaf surfaces asorange, rust-colored, dense silky tufts ranging from 5 to 8 mm in diameter. Upon scraping away these spots, a thin, grayish white to dark-colored, necrotic crust remains on the leaf. These spots usually come together to form large irregular patches on a leaf. As the spots mature they take on a dull, grayish green color. Twigs and branches are also affected causing the bark to crack due to the growth and expansion of the pathogens filaments into the cortical tissues of the host. Algal leaf spot is caused by Cephaleuros virescens.

Integrated Management 

Algal leaf spot can be reduced by maintaining tree vigor with cultural techniques such as proper fertilization and irrigation, proper pruning to enhance air circulation within the canopy and sunlight penetration, managing weeds and wider tree spacing.



Managing insect, mite and other foliar diseases increases tree vigor and lessens susceptibility to this disease. Spray Copper oxychloride 0.25%.

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Chapter-3

Integrated Pest Management in Major Rabi Crops Dr. Uma Shankar Division of Entomology, Faculty of Agriculture, Sher-e-Kashmir University of Agricultural sciences & Technology of Jammu, Chatha, Jammu180 009, UT of J&K, India, Email:umashankar.ento@gmail.com, Mobile: 9419202151 ____________________________________________________________________________ Introduction Vegetables and field crops play an important role in food and nutritional security and now a day it has been instrumental for generating employment opportunities in the form of entrepreneurship development. The alarming rate of growing population has resulted into so many consequences wherein, with the limited land and depleting water resources, India has to feed the burgeoning population without destroying the ecological balances. In this context, we must have to look after diversification in cropping system, mixed and intercropping of vegetable crops, increase in intensity which also aggravates the insect pest problems. Insect pest and disease infestation is one of the major limiting factors in increasing the yield potential in many vegetable crops.

Although, the extent of crop losses in vegetables varies with the crop type, crop location, damage potential of the insect pests involved and cropping season. With the introduction of high yielding varieties and hybrids vegetable seeds resulted in dramatic changes in insect pest scenario leading to minor pests assuming the status of major pests. Overall, the insect pests inflict crop losses to the tune of 30- 40 per cent in vegetable production and in many cases, there is 100 per cent yield loss due to viral diseases vectored by insects. Vegetable growers by and large depend on chemical pesticides to counter the problem of insect pests. It accounts for 13-14 per cent of total pesticides consumption, as against 2.6 per cent of cropped area (Sardana, 2001).

The

continuous, excessive and injudicious use of pesticides has triggered several ecological consequences like resistance (currently, more than 500 species of insects) and resurgence of insect pests, pesticide poisoning, environmental pollution, destruction and elimination of natural enemies, effect on non target organisms, disruption in food cycle and food web and residues in food commodities.

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Keeping the above facts in view, Integrated Pest Management is the only way to avoid such problems. It is a systems approach to pest control that combines biological, cultural, and other alternatives to chemical control with the judicious use of pesticides and maintain pest levels below economically.

Insect Pest Management in Wheat 1.

Wheat Aphid : Sitobion avenae Fab.(Hemiptera : Aphididae)

In India, wheat Aphid is reported from Delhi, Haryana, Himachal Pradesh, Jammu and Kashmir, Madhya Pradesh, Punjab, Rajasthan, Uttar Pradesh and West Bengal. It is the major pest of cereal crops in the winter/spring. Aphids are soft bodied insects that can be found in wheat anytime during the growing season. The most common aphids found on wheat are the bird cherry oat aphid, root aphid, green bug, corn leaf aphid and English grain aphid. The first four occur mostly in the fall and winter. Only the green bug aphid causes direct feeding damage that appears speckled brown and discoloured with some leaf curling. Aphids also act as vector for viral disease named barley yellow dwarf (BYD). Wheat and barley can be severely damaged, but oats are mostly susceptible to this disease. Infection can occur from seedling emergence through heading, but yield loss is greatest when plants are infected in the fall. Yield losses of 5-15% are common but losses can exceed 30% during epidemics.

Management: 1. For the management of aphids, foliar spray of imidacloprid 200SL @20g a.i./ha on border rows at the start of the aphid colonization be given.

2. A well timed application of a persistent pyrethroid insecticide such as Lamda cyhalothrin can reduce the incidence of BYD and increase yields.

3. Spraying formothion or oxydemeton methyl @ 250ml a.i./ha or 2 ml per litre of water.

4. Growing aphid resistant varieties, if available.

2. Fall army worm: Spodoptera frugiperda The scientific name of Fall armyworm derives from frugiperda, which is a Latin word meaning lost fruit. It is named for its capability to damage and destroy a large variety of crops. The genus “Spodoptera” has 25 species but S. littura, S. muritia, S. exempta and S. frugiperda are economically significant species for agricultural crops. S. frugiperda (Fall Armyworm) is an

13

invasive pest of many crops but most serious pest for wheat and maize. It is migratory lepidopteran insect pest which infest more than 100 plant species in India.

IPM of Fall Armyworm (FAW), Spodoptera frugiperda Monitoring: Installation of pheromone traps @5/acre in the current and potential area of spread in crop season and off-season.

Cultural Measures: 1. Deep ploughing is recommended before sowing.

This will expose FAW pupae to

predators.

2. Timely sowing is advised. Avoid staggered sowings.

3. Erection of bird perches @ 10/acre during early stage of the crop (up to 30 days).

4. Clean cultivation and balanced use of fertilizers.

Mechanical control: 1.

Hand picking and destruction of egg masses and neonate larvae in mass by crushing or immersing in kerosene water.

2.

Mass trapping of male moths using pheromone traps @ 15/acre.

Bio-Control: 1.

Augmentative release of Trichogramma pretiosum or Telenomus remus @ 50,000 per acre at weekly intervals or based on trap catch of 3 moths trap.

2.

Biopesticides: Suitable at 5% damage in seedling to early stage and 10% panicle damage with entomopathogenic fungi and bacteria.

a.

Entomopathogenic fungal formulations:

Application of Metarhizium anisopliae talc

formulation (1x108cfu/g) @ 5g/litre at 15-25 days after sowing. Another 1-2 sprays may also be given at an interval of 10 days depending on pest damage. b.

Bacillus thuringiensis var. kurstaki formulations @ 2g/l (or) 400 g/acre.

Chemical Control: 1. To manage 2nd and 3rd instars larvae at 10-20% damage, spray Emamectin benzoate @ 0.4 g/l of water or Spinosad @ 0.3 ml/l of water or Thiamethoxam 12.6% + lambdacyhalothrin 9.5 %@ 0.5 ml/l of water or Chlorantraniliprole 18.5% SC @ 0.4 ml/l of water.

14

2. Insecticide management is not cost effective at this stage. Hand picking of the larvae is advisable. All the sprays should be directed towards early hours of the day or in the evening time.

3. Termites: Microtermes obesi Hol., Odontotermes Obesus Ramb. (Isoptera : Termitidae) These are social insects like honey bees and ants live in colonies. They build up nests consisting of small chambers connected with narrow passages make superficial tunnels on the ground to attack growing plants, timber wood, dry leaves and grasses. In the plants, generally the roots and stems are damaged with the result that they don’t grow properly, wilt soon and dry up. An attacked plants when cut or dug up shows irregular galleries on the bark which are made by termites eaten up the woody tissues. A new termite colony is usually started at the beginning of rains when sexually mature winged males and females which emerge in vast numbers come out of their colonies. The male and female termites shed their wings and copulate; the female burrows into the ground to build a new nest. The queen may lay 30,000 to 80,000 eggs/day and 15 million eggs throughout her life span of five years.

Management: 1. Do not apply raw or undecomposed farmyard manure; if possible irrigate the field when infestation is serious.

2. Treat the seed with chlorpyriphos @ 5 ml/kg of seed before sowing.

3. Mounds of termites if any in the area should be treated with 0.5% chlorpyriphos after breaking open the earthen structure and making holes with an iron bar. The insecticidal emulsion should be used @4 litres /m3 of the mound.

4. In the termite prone areas, (especially the northern states) seed treatment with chlorpyriphos @ 0.9g a.i /kg seed.

5. Fipronil (Regent 5FS @ 0.3 g a.i./kg seed) is also very effective.

6. In the standing crop, the broadcasting of the insecticide treated soil 15 DAS be practiced.

For this, chloropyriphos @ 3 ml mixed in 50 Kg soil be used for one hectare field.

4. Rodent pest management in wheat Rodents (order: Rodentia) possess a great feeding potential as they generally consume 5–10% of their body weight on a daily basis. Agricultural fields serve as a highly productive rodent habitat and crops such as sugarcane, rice, wheat, groundnut and fodder serve as an ideal habitat for 15

rodent pests. They are one of the most successful animals on earth due to their vast breeding potential and adaptability to a variety of living conditions. Of over 2000 species reported globally, only a limited number of species (~10%) are considered as serious impediments causing significant losses to food production and storage. The pest rodents cause 5–10% losses to various production systems such as agriculture, horticulture, forestry and stored food grains.

Indian Field Mouse, Mus booduga occupies second position after Bandicota bengalensis in rice, wheat and sugarcane fields in many regions causing serious damage.

Management: 1. Proper sanitation should be maintained.

2. Mechanical removal of the rodent population from any habitat is mainly done using different types of trap. Trapping of B. indica and B. bengalensis in flooded deep water rice in Bangladesh and of B. bengalensis, G. ellioti, M. meltada, etc. in vegetable crops in Himachal Pradesh, India was found to be an effective method of rodent control. An ecofriendly mechanical ‘burrow fumigator’ device has been developed by the All India Network Project on Rodent Control. The device utilizes farm wastes for generating smoke, which is pushed into rodent burrows through an inbuilt blower. The rodents die of asphyxiation.

3. Habitat Manipulation methods are low cost treatments and involve little modification in crop husbandry practices, such as ploughing, puddling, removal of wild vegetation and refuge of previous crops and reduction in bund size, etc. It helps in the migration of pest rodents from crop fields.

4. Several species of vertebrates, mainly birds and mammals, such as snakes, owls and mongoose are the predominant vertebrate predators of rodents.

5. Some plant products have been found to possess anti-rodent properties. Neem leaf powder (5%) showed an antifeedant action on rodents, whereas neem oil repelled the rats to the tune of 18–48%. Neem ormulation BBR (3% concentration) recorded a repellency index up to 87% in baits against rodents.

6. Chemosterilants: Some of these are antiestrogen U–11, diphenylindane derivatives, metepa, tepa, tetradifon, furadentin, colchicine, glyzophrol, etc.

16

7. Use of rodenticides is the most common, expedient and humane method to control pest rodents and therefore forms the basis of present day rodent management strategy in most parts of the world. Among acute rodent poisons, only zinc phosphide, barium carbonate and aluminium phosphide are registered by the Government of India for common use.

8. Non-anticoagulant rodenticides include

Calciferol, cholecalciferol, bromethalin and

flupropadine.

9. Anticoagulant rodenticides: As the name indicates, this group of rodenticides interfere with the blood coagulation process, leading to internal bleeding and haemorrhage in the target

animals.

They

include

chlorophacinone,

diphacinone,

pindone

and

hydroxycoumarins. The hydroxycoumarin rodenticides are further divided into (i) first generation (coumachlor, coumatetralyl and warfarin) and (ii) second generation (bromadiolone, brodifacoum, difencoum, flocoumafen, difethialone) rodenticides.

Insect pests of pulses A variety of insect pests infest pulses and the annual yield loss is estimated to be 20 per cent in pigeonpea, 15 per cent in chickpea and 30 per cent in urdbean and mungbean. On an average 2.5 to 3.0 million tonnes of pulses are lost annually due to pests (Ali, 1998). The important insect pests which causes economic losses on pulse crops are listed below       

Gram pod borer : Helicoverpa armigera Spotted pod borer : Maruca virtata Plume moth : Exalastis atomosa Blue butterfly : Lamides boeticus Whitefly : Bemisia tabaci Podfly : Melanagromyza obtusa Pod bug : Clavigralla gibbosa; Riptortus spp.

Temite : Odentotermes obesus

Helicoverpa is the key pest causing an average yield loss of 10 per cent for the entire country on chickpea. There was even 90 per cent damage under severe cases. The annual loss due to this pest was estimated to be Rs.20.30 crores (Lal et al. 1985) Integrated Pest management Monitoring of Helicoverpa armigera

17

The seasonal cycle of this pest varied in different parts of the country and also with cropping pattern. Studies so far conducted have indicated that these traps can be used as a monitoring device to design the management strategies against H. armigera.

Inflorescence and pod borers Spraying can be taken up based on Economic Threshold (ETL). For example , Maruca 3/plant, Exalastis 5/plant. Whenever H. armigera is predominant apply NPV 500LE/ha to control the pest in the early stage.

Spraying of NSKE 5% twice followed by triazophos 0.05% is also effective. Recent studies have indicated that following IPM methods have controlled the pest and increased the yield.

1. Installation of bird perches @ 50/ha 2. H. armigera pheromone trap @ 10/ha 3. Collection and destruction of fully grown larvae 4. Spraying NSKE 5% at 50% flowering 5. Spraying HaNPV at 500 LE/ha at 15 days after first spray 6. Application of the following insecticides at 15 days interval depending on the intensity of pest. Chlorpyriphos 0.05%.

Insect pests of Tomato is a very popular constituent of Indian diet. It is devastated by an array of insect pests, however the major damage is caused by fruit borer, Helicoverpa armigera, serpentine leaf miner, Lariomyza trifoli and white flies, Bemesia tabaci (Shankar et al., 2009b).

1.

Tomato fruit borer, Helicoverpa armigera Hubner

Helicoverpa has been recorded on more than 200 hosts in India (Pawar, 1998) and is most damaging to cotton, pigeonpea, chickpea, groundnut, sorghum, pearl millet, tomato and many other crops of economic importance. Caterpillars are the immature stage of the insect which causes the considerable damage to the crops in various stages. In the pre-fruiting stage, the caterpillar feeds on the tender foliage including leaves, flowers, buds and even sometimes they make holes in the tender twigs which reflects the perforated look to the crops. After fruiting, the larvae bore large, clear, circular holes into fruits and feed on the pulp. Its name has been changed according to its host. Its larvae are stoutly built and robust. In tomato, it is responsible to destroy many fruits and make them unfit for the human consumption. Its larvae have peculiar feeding 18

habit on host crop. The larvae thrusts its half of the body into the fruits and feeds the inner contents and rendering them unfit for the human consumption. It causes damage up to the tune of 50 to 80 per cent if not intervened at right time.

Management 

Soil solarization in nursery beds as well as in the main field.



Plant one row of marigold as trap crop for every 16 rows of tomato.



Monitor and clip top three tender leaves for Helicoverpa eggs destruction.



Early maturing varieties should be planted to escape the damage by borers.



Installation of pheromone traps @ 5-7 per ha for early detection and 12-15 per ha for trapping and mass destruction for mating disruption.



Release Trichogramma chilonis @ 50,000 adults /ha per release (6 times) at weekly interval when eggs of Helicoverpa are noticed.



Conservation of Campoletus chloridae, a potential parasitoid of H.armigera.



Spraying of EPN Steinernema carpocapsae against Helicoverpa gave an excellent result in killing the larvae within 24 to 48 hrs.



Spray Ha-NPV 250 to 500 LE/ha (freshly prepared) 2-3 times at 10 days interval at evening hours.



Spraying of bacterial formulation Bt @ 500 g /ha also may proved beneficial in regulating the Helicoverpa larvae in the field condition.

2.

Serpentine leaf miner, Liriomyza trifoli

Leaf miners lay eggs below the epidermal layer of leaves and cause heavy leaf mining on upper or lower side of the leaves. Mines starts from the margins of leaves and progress towards the base/centre of leaf. Mines are initially narrow and gradually enlarge often twisting through leaf which causes stippling of young seedlings. Severe leaf mining slows down the plant growth and accelerates the leaf drop.

Management 

Dimethoate @ 2 ml per lit of water are found to be effective against leaf miner.



Hymenoptera (Eulophidae) an endo-larval parasiotid of leafminer, Hemiptarsenus varicornis is found to be very effective.

19



Neem seed kernel extract (NSKE) @ 5 per cent concentration found promising in controlling leaf miner followed by Karanj, Pongamia pinnata oil next in the order of efficacy.

Insect pests of chillies 1.

Chilli mites, Polyphagotarsonemus latus

Chilli crops are attacked by yellow mite, Polyphagotarsonemus latus which has become a serious problem in the entire country. Mite affected leaves of the chilli plant turn towards underside and look like an inverted boat. As young and adult mites attack the tender leaves, almost all new leaves curled downward and lower surface appears silvery whereas, the upper surface of leaves look more dark green as compared to healthy ones. Further the plant growth is checked and the internodes become shorter.

Management 

Since last few decades, farmers have normally been using different kinds of insecticides for the control of mites. Being insecticides these were not able to give satisfactory control of mites, due to which mites also developed resistance.



Some insecticides have been reported to be effective for mite control (chilli pest complex) - triazophos 40 EC @ 2ml per lit of water or Dicofol 18.5 EC @ 2ml per lit of water or propargite 57 EC @ 3-4 ml per lit of water at the beginning of mite attack followed by another spray at an interval of 12 days.



Seedling root dip treatment with imidacloprid 200 SL @ 1ml per lit of water before planting and 3 sprays with the same at 15 days interval.



The following lines/varieties have been reported resistant/ tolerant to mites viz., Pusa Jwala.

2.

Thrips, Scirtothrips dorsalis

Thrips cause severe damage to the plants in the nursery and in transplanted crop and is the main constraint in the chilli and Capcicum cultivation. Both the nymphs and adults of chilli thrips lacerate the leaf and imbibe the oozing sap. Sometimes even buds and flowers are also attacked.

Tender leaves and growing shoots are preferred for feeding, while the older leaves are rarely attacked. The infested leaves start curling and crumbling and ultimately shed while, buds become brittle and drop down. Main visible symptom are curling of leaves upward (normal boat shaped), 20

small leaf size and shorter internodal space with stunting growth of plants, no fruiting or small deformed fruiting in infected plants takes place.

Management 

Rouge out the infected plants and burn them.



Grow resistant varieties.



Seed treatment of imidacloprid 70 WS at 10, 20 and 30g per kg protected the seedlings in the nursery up to 45 days and foliar treatment of imidacloprid 200 SL at 250, 375 and 500 ml /ha reduced the thrips population significantly and also recorded higher yield of chilli fruits (Santharam et al., 2003).



Periodic spray of metasystox @ 2 ml per lit of water or imidacloprid @ 0.5 ml per lit water at 10 days interval for vector (Bemesia tabaci) Management.

Cruciferous vegetable crops The cruciferous crops (cole), which includes economically important crops like Cabbage, Brassica oleracea var. capitata; Cauliflower, B. oleracea var. botrytis; Knol khol, B. caulorapa (L.) throughout the world. However, poor productivity is mainly attributed to the incidence of diamondback moth (DBM), Plutella xylostella; cabbage caterpillar, Pieris brassicae; leaf webber, Crocidolomia binotalis; cabbage head borer, Hellula undalis and aphid Brevicoryne brassicae infesting the crop in various seasons (Shankar et al., 2009a).

1.

Cabbage butterfly, Pieris brassicae and P. rapae)

It is also known as imported cabbage worm. Two species of cabbage butterflies i.e. large butterfly, Pieris brassicae and small butterfly, P. rapae have been recorded and found damaging the cole crops in Jammu province. They are serious threats to the crops grown for seed purpose especially in hills as well as in plain areas of Jammu. Adults are small to large sized butterflies, pale white with black markings. Female butterflies have two black circular dots on the dorsal side of each forewing. After mating, female butterfly lay about 100-150 conical, yellowish eggs in clusters on lower or upper side of the leaf. The eggs hatch in 3-15 days depending upon the environmental temperature. Just after hatching, the caterpillars feed gregariously on leaves in early stages. They passes from five larval instars and become full grown in 2-3 weeks. Their incidence has been recorded in fields from October to April- May except in cool winter months 21

during December-January. They completely skeletonize the leaves leaving only mid ribs. Due to complete loss of photosynthetic area crops failed to produce the economic yields.

Management 

Hand picking and mechanical destruction of caterpillars during early stage insect pest attack is beneficial.



Cotesia glomeratus are recorded as a potential parasitoids against cabbage caterpillar larva.



Phryxe vulgaris (dipteran fly) are also observed abundantly in cole crops fields to parasitize the caterpillars.



Foliar spray of Bt formulations @ 500 g/ ha along with sticker (0.5 ml/lit of water) is found promising to control all lepidopteran insect pests.



Need based application of cypermethrin @ 1 ml per lit or malathion @ 2ml per lit or endosulfan @ 3ml per lit of water are beneficial in case of severe infestation.

2.

Cabbage head borer (Hellula undalis and Helicoverpa armigera)

Cabbage head borer is seen sporadically but its attack coincides with the head/curd initiation stage and a single larva is responsible to damage the entire head/curd. The early instar may feed on the growing point, and bore inside the head and damaging the head and excrements which prevents head formation. When fully grown, larvae pupate inside the head, or on stems, or in soil. Helicoverpa armigera larvae have also been recorded at several places in Jammu region as head borer of cabbage. It may be due to absence of preferred host in winter months or some environmental and behavioural change.

Management 

Monitoring and mechanical destruction of larvae at the time head /curd initiation may proved to be good.



Bracon spp. are important larval parasitoids against head borer.



Fungal formulations like Beauvaria bassiana and entomopathogenic nematodes (EPN) may be beneficial to suppress the attack of head borer.



Spraying of synthetic pyrethroids like cypermethrin, fenvalerate, deltamethrin @ 1 ml per lit of water is effective to control head borer.

22

3.

Diamondback moth (Plutella xylostella)

Diamondback moth is the universal pest of crucifers’ crops and remains one of the most serious problems for the intensive cole crops growing areas. Plants at all stages of growth may be attacked by and completely devastated by this pest. In Jammu province, diamondback moth is recorded only on early and on late season cole crops and on crucifers i.e., during SeptemberNovember and from February onwards.

Management 

Use of pheromone traps are useful in monitoring and detection of the DBM pest popualtion in early stages.



Practice of trap or intercropping of mustard, coriander and marigold with cole crops may reduce the DBM pest infestation.



Release of Cotesia plutellae, natural bio-agent of DBM larvae is effective to suppress its population in field condition (1000 adults per release every 2 week interval up to harvest).



Use of resistant varieties is also effective against DBM.



Some safer insecticides and neem products are recorded to be effective against DBM.



Spray cypermethrin@ 1ml, cartap hydrochloride@ 2ml, spinosad@ 1ml and Bt formulation@ 500 g/ha may be promising in controlling the DBM pest population.

4.

Tobacco catepillar : Spodoptera Litura

Spodoptera litura has also wide host range and economic insect pest of cole crops and tomato but it is usually kept under damaging levels by management practices targeted to Helicoverpa.

They are voracious feeders and scrape the leaves and later on it may completely defoliate the leaves. The insect is active during the entire year except severe cold months and females can lay up to 300-600 eggs each, usually in groups of about 100. Eggs are laid on the underside of lower leaves and are covered with fuzzy, white creamy scales. Under warm conditions, eggs hatch within three to four days. The neonate larvae feed from one to three weeks, in groups at younger stage and scattered on the plants when larger.

Management 

Summer ploughing is beneficial to expose the hibernating pupae to natural enemies and for bird predation.

23



Hand picking and mechanical destruction of egg masses, caterpillars and spraying of NSKE 5% during early stage may proved beneficial.



Installation of pheromone traps @ 25 to 30 /ha may be the useful tool for early detection and also for mass trapping and destruction of tobacco caterpillar.



Spraying of Splt NPV @ 250 LE with gur or jaggary (10 g/lit) and sticker during evening hours may be the promising to control tobacco caterpillar (Monobrullah and Shankar, 2008).



Foliar spray of Bt formulations @ 500 g/ ha is the good for controlling the caterpillars.



EPN may be quite promising to suppress the Spodoptera larval population in the field condition.



Chemical sprays recommended for the DBM control will also work against this pest but in this case, spot application may be enough if the attack is not wide spread in the field.

Onion and Garlic Onion thrips, Thrips tabaci Besides onion, several other species of thrips infest tomato, brinjal, capsicum, and on cucurbits.

They are tiny, slender insects that may vary in colour from yellow to dark brown or black. Both nymphs and adults rasp the epidermis of tender leaves and imbibe the oozing sap. As a result of which leaves develop numerous white specks. Heavily infested leaves curl up and get distorted and ultimately the top of attacked plants wither away. The symptoms develop by feeding on foliage may cause a silvering of foliage.

Management 

In the case of onion, the only reliable method of controlling onion thrips is through the use of tolerant varieties. Advantages of pest-resistant crop varieties include ease of use; compatibility with other integrated pest management tactics; low cost; cumulative impact on the pest (each subsequent generation of the pest is further reduced); and reduced negative impact on the environment.



The following lines/varieties have been reported resistant/ tolerant to onion thrips: White Persian, Guano, Sweet Spanish, Crystal Wax, Spanish White, Bombay White, Pusa Red Udaipur 103, N 780-1, Pbr-3, PBR-4, VL-1, No. 18, No. 19, Pusa Ratnar.



Some insecticides and neem products have been reported to be effective against thrips.

24



Biological control agents like minute pirate bugs, lacewing larvae and ladybirds prey on thrips.



Application of systemic insecticides like imidacloprid or thimethoxam or dimethoate after 10 days of transplanting may help to suppress the thrip population.

Insect Pests of Cucurbits Cutworms in cucurbits: Agrotis ipsilon

1.

It is a polyphagous insect pest which feeds mainly in seedling stages of many vegetables, potato, maize and other crops. Caterpillar is the only damaging stage of this pest. The short stout larvae are highly destructive. At night, larvae cut the plant at ground level at an early stage of their growth. A few caterpillars in the field kill many small plants during one night. They fell/cut more plants rather consume.

Management 

The population of cut worm larvae can be considerably suppressed by applying cartap hydrochloride @ 20 kg /ha or drench the soil with chlorpyriphos @ 2.5 ml per lit of water

2.

Red Pumpkin Beetle, Aulacophora foveicollis

The beetles eat leaf lamina, cause perforation, irregular holes or complete defoliation and damage is severe, particularly at cotyledonary stage of crop. The grubs feed on the underground stem and root portion of the host plant forming holes/ galleries leading to dry up of branches or whole plant. They may also attack the fruits touching the ground surface by making holes.

Management 

Plant the crop in November to avoid the damage from this insect.



After harvesting the cucurbits, the field should be ploughed to exposed and kill the grubs in soil.



Spray Of the following insecticides are recommended against red pumpkin beetlescarbaryl @ 2g per lit of water or endosulfan @ 2ml per lit of water

3.

Fruit flies, Bactrocera cucurbitae

Cucurbit fruit fly is a major pest of all cucurbitaceous vegetables. It is serious and destructive pest of cucurbits like melon, pumpkin, cucumber and gourds. Adult flies are reddish brown with lemon yellow markings on thorax. Female fly lacerates or puncture the fruits by its pointed ovipositor and laid eggs on soft fruits. Female secrets the gummy secretion which cements the 25

punctured tissue and make the entrance water proof. The most damaging stage is the maggots.

Maggots feed on the pulp of fruit and cause premature dropping of fruits and also make them unfit for the human consumption. Sometimes secondary bacterial infection also takes place.

Oozing of brown resinous fluid from the fruits and fruits become distorted and malformed. In severe condition or in rainy season 50-100 % damages are reported.

Management 

In endemic areas, change the sowing dates, as the fly population is low in hot dry conditions and its peak during rainy season.



Collect infested fruits and dried leaves and burn it in deep pits. Expose the pupae by deep ploughing and turning over soil after harvest.



Use ribbed gourd as trap crop and apply acetamiprid (0.2 g/lit) on the congregating adult flies on the undersurface of leaves.



Use poison baiting in severe infestation: Saturated sugar solution 5 ml+ Malathion 50EC 5ml+ gur or jaggary 100 ml are kept in the container and put it to the four corners of the field as a poison baits.



Use fruit fly traps@ 50 traps /ha : Use 5 g wet fish meal in a polythin bag (20×15 cm in size and having 6 holes of 3 mm in diameter) and add 0.1 ml diclorovos (1 drop) in the cooton plug and put it in the bag. For better performance, dichlorovos is added every week and fishmeal should be renewed after 20 days.

26

Chapter-4

Integrated Disease Management in Wheat S. K. Singh, Vishal Gupta, Ranbir Singh and A. K. Singh Division of Plant Pathology, Sher-e-Kashmir University of Agricultural Sciences and Technology of Jammu, FoA, Chatha-180 009 ________________________________________________________________________ Introduction Wheat is attached by a number of fungi, bacteria, viruses etc. But the fungal diseases are the most predominant biotic stress as compared to other microorganisms. The most important fungal diseases that affect wheat are stem, leaf and stripe rusts, loose smut, karnal bunts, leaf blight, powdery mildew, flag smut and hill bunt. In wheat, brown rust, yellow rust, black rust, loose smut, kamal bunt and powdery mildew were observed to be important diseases in different states. Kamal bunt and foliar blights were more damaging whereas stem rust became relatively less important after the green revolution.

Wheat Rusts The three important rust diseases occurs on wheat viz., stem or black rust (Puccinia graminis f. sp. tritici), leaf or brown rust (Puccinia recondita) and stripe or yellow rust (Puccinia striiformis). Rust diseases are responsible for huge economics losses.

In India leaf rust are most important diseases and prevalent all over the country. The disease appears in north-western region in late January and assumes epidemic growth rate in the month of February- march when the crop is in anthesis or grain filling stage but in case of stem rust it is mainly prevalent in central and peninsular region of country but it appear very late with low intensity in the northern belt.

Stripe rust is restricted only in northern parts of the country particularly in foot hills of Himalayas and adjoining plains of north-western region. Spores of both Puccinia recondita and P. striiformis (brown rust and yellow rust of wheat) are thought to be primarily dispersed by wind and rainfall has the potential to spread both brown rust and yellow rust of wheat.

27

Symptoms The pustules of leaf rust are small, oval shaped, orange to orange brown in colour, scattered on leaf and leaf sheath. Uredial pustules are may also developed on awns, glumes, peduncles, and internodes. Mostly uredial pustules are generally found on upper surface of leaf and the epidermis are not ruptures.

In case of stripe rust, uredial pustules

are developed as narrow yellow, linear stripe of leaves, leaf sheath, outer and inner sides of glumes and kernels. Rusty pustules transform into black teleutospores as plant gets maturity.

Stem rust has been a serious disease of wheat, barley, oat and rye, as well as various important grasses. Infections in cereals occur mainly on stems and leaf sheaths, but occasionally they may be found on leaf blades and glumes as well. The first macroscopic symptom usually appears as a small chlorotic fleck, after a few days of infection. About 8-10 days after infection, a long pustule is formed by rupturing the host epidermis from pressure of a mass urediospores of brick-red colour produced in the infection. The powdery masses of urediospores appear similar to rust spots on a weathered iron surface.

With age, the infection ceases production of brick-red urediospores and produces a layer of black teliospores in their place, causing the stems of heavily infected plants to appear black in the late season.

Integrated management 

Grow resistant / tolerant varieties of wheat recommended for particular area like DBW 222, DBW 187, WB-2, DBW-88, and DBW-90.



Grow different varieties at the farm level (Gene deployment) and application of balanced dose of nitrogen and potash fertilizer that also helpful to minimize the rust severity on plant.



Spray the crop in case of severe infection with fungicides viz., propiconazole or tebuconazole @ 0.1 per cent.

Smuts and bunts Five bunt and smut diseases namely Loose smut (Ustilago segetum (Pers.) Roussel tritici, Flag smut (Urocystis agropyri (preuss) schroet), Common bunt (Tilletia caries (DC) Tul. and Tilletia floetida Wallr. Liro.), Dwarf bunt (T. controversa Kuhn) and Karnal bunt (Neovossia indica (Mitra) Mundkur) attack wheat crops.

28

Loose smut present all over the country, is more severe in north than south India and can causes yield losses up to 2-4 percent in north western India. Karnal bunt which is minor disease has assumed importance in recent years due to its quarantine importance.

Reduction in yield due to karnal bunt in India is not beyond 0.2 to 0.5 percent but 3-5 percent infected grains can cause blackening of whole meal. Hill bunt and flag smut are of minor importance because of their restricted distribution in isolated pockets of Rajasthan, Haryana and Punjab.

Symptoms In case of loose smut entire inflorescence, except the rachis, is replaced by masses of smut spores. These black teliospores often are blown away by the wind, leaving only the bare rachis and remnants of other floral structure.

Karnal bunt is not easily detected prior to harvest, since it is usual for only a few kernels per spike to be affected by the disease. Following harvest, disease kernels can be easily detected by visual infections. A mass of black teliospores replaces a portion of endosperm, and the pericarp may be intact or ruptured. Disease kernels give off a fetid or fishy smell when crushed. Karnal bunt of wheat is caused by the fungus Tilletia indica, which partially converts kernels into sori filled with teliospores. Despite minor overall yield and quality losses, the disease is of considerable international quarantine concern.

The main symptom of caused by hill bunt or common bunt are fungal structure are called “bunt balls” which resemble as kernels but are completely filled with black teliospores.

The bunt balls of common bunt caused by T. caries and T. floetida are about the same size as shape as the kernel they replaced. When bunt balls are crushed they give off a fetid or fishy odour. Infected spike tend to be bluish green in colour, and the glumes tend to spread apart slighty; the bunt ball often visible after the soft douf stage and a slight reduction in plant height is typical of common or hill bunt.

Integrated management Flag smut Flag smut of wheat incited by Urocystis agropyri could be managed by growing resistant cultivars. Seed treatment with tebuconazole 2DS @ 1.25 g / kg, or carboxin and carbendazim @ 2.5 g / kg seed reduced cent per cent disease incidence. Application of

29

poultry manure @ 4–6 t per ha managed flag smut. Nl60P50 kg per ha and early or late sowing was found effective in management of the disease.

Loose smut Sowing of resistant varieties was mainly adopted for the management of the disease. Seed treatment with Vitavax @ 2-2.5 g / kg or tebuconazole 2DS @1.25 g / kg seed before sowing can manage the disease by reducing the seed borne primary inoculum. Roguing out suspected disease ears at the time of ear emergence and destroyed them properly by burning or burring under the ground. Soaking of seeds and solar heat treatment of seeds is practicable in hot summer areas of plains. During bright sunny days in the last weeks of May or first week of June soak the seed in water for about four hours in the morning and dry it under hot sun in the afternoon. Storage of properly dried seed for use in the next season is also reducing the disease incidence.

Karnal bunt Sowing of resistant varieties and use of disease free seeds are the important control measure for the management of the disease. Treatment of the seeds before sowing with Vitavax @ 2-2.5 g / kg or tebuconazole 2DS @ 1.25 g / kg seeds minimizes the disease intensity through elimination of seed borne infection. Avoidance of excessive irrigation at the time of flowering also reduces the disease intensity.

Common bunt 

Treatment of the seeds before sowing with Vitavax @ 2-2.5 g / kg or tebuconazole 2DS @ 1.25 g / kg seeds reduces the disease intensity by elimination of seed borne primary inoculums.

Foliar blight complex A number of pathogens causing blight, blotch and spots are associated with wheat in India. Bipolaris sorokiniana is the major pathogen followed by Alternaria triticina prasada and prabhu. Drechslera tritici repentis and Aternaria alternata are also associated with wheat but are not very prominent. Both the pathogen is widely prevelant in Bihar, West Bengal, Uttar Pradesh, Rajasthan and Orissa. Of late B. sorokiniana has made appearance in North western region of the Indo-Gangatic plains also.

The importance of foliar blights must be expressed in terms of yield losses. Yield losses due to B. sorokiniana revealed that yield of wheat cultivars UP 2338, UP 262, HP 1633 30

may be reduced upto 36.24, 20.93 and 22.26 per cent respectively due to spot blotch disease.

Symptoms Alternaria blight symptoms appear as small, oval, discoloured lesion, irregularly scattered on leaves. As lesion enlarged, they become irregular and dark brown to grey surrounded by yellow margin. Later several lesions coalesce and cover large areas exhibiting bunt appearance.

The lesion on leaves caused by B. sorokiniana are light brown to almost black, which are circular linear or elliptical. A well developed lesion is typically elliptical with abundant sporulation.

Integrated management 

Cultivation of resistant varieties is an important component of disease management



Hot water treatment of seeds at 520 C for 10 minutes before sowing provides good control of the disease by reducing primary inoculum



Seed treatment with systemic fungicide viz., carboxin @ 2.5 g / kg or tebuconazole 2DS @ 1.25 g / kg of seeds also effective for the management of the disease in the field



Use of balanced NPK fertilizers and irrigation reduce the intensity



Spraying of plants with propiconazole or tebuconazole @ 0.1% at 25-35 days interval starting from the first symptoms appearance in the field

Powdery mildew Powdery mildew of wheat is caused by Erysiphe graminis and is widely distributed in the humid and sub humid wheat growing regions of the world. It is basically important disease of northern hilly region of India, Mount Abu, Palney and Nilgiri hills. This disease is also occurs in the north-western region of the Indo Gangatic Plains where stripe and wheat rust are the potential threat to wheat production. It is believed that heavy attack of powdery mildew in hilly and north-west plain regions result in yield losses through reduced grain size.

Symptom All the green part of the plant, the upper surface or undersize of the leaves, sometimes both surface together, the stem or even the ears are affected by the pathogens, exhibiting 31

white to grey coloured powdery masses on the plant surface. The cottony powdery mass contains the conidia and conidiophores of the fungus. As the season progresses, small black cleistothecia developed on the powdery mass. The infected leaves are twisted, crinkled and deformed, infected plant become yellow and photosynthetic rates are severely affected.

Integrated management 

Sowing of resistant varieties is the major control measure.



Burning of the previous crop refuses in the field after harvesting the crop could reduce the disease intensity.



Spray the crop in case of severe infection with fungicides viz., propiconazole or tebuconazole @ 0.1 per cent.



Two to three sprays should be given at an interval of 15-20 days if losses are expected to very high.

32

Chapter-5

Role of Biological Control in Plant Disease Management Vishal Gupta, Ranbir Singh and Satish Kumar Sharma Sher-e-Kashmir University of Agricultural Sciences and Technology of Jammu, Chatha 180 009, vishal94gupta@rediffmail.com __________________________________________________________________________________

Introduction Global population has been projected to reach 9 billion by 2050 with an increase of 30 per cent. To fulfil the demand of food for this increasing population, the food production shall be increase by 70 per cent on the limited resources. Urbanization, Infrastructure development, fluctuating fuel prices, shrinkages natural resources, food crisis and climate change are the present challenges which aggravates the situations more. Out-break of biotic and abiotic stresses driven by climate change at the same time reduced the agriculture production by 20 per cent. An 10–30 per cent

yield losses from pests have been estimated (Thind, 2015),

which can be significant by additional of post-harvest losses. In recent, losses up to 6, 10, 18 and 13 per cent per cent have been estimated in cereals and pulses, oilseeds, fruits and vegetables during harvesting, handling and storage (Jha et al., 2015). To avert the losses caused by plant pathogens, fungicides of approx. 16 billion US dollars has been used per annum. Although chemical pesticides have been successfully use for the control of pests as these have been effective, but their indiscriminate and non-judicious uses has resulted in several problems such as development of resistance in pests against pesticides, resurgence of minor pests, pollution in water, air and soil, elimination of natural enemies, disruption of ecosystem and residual effects in the produce also. Further, Phytosanitary standards, environmental and socially unacceptable cultivation practices emerged as barriers to international trade. Besides, the consumers of the importing countries are rejecting questionable products (residue in produce) resulting in heavy losses to the exporters. In the meantime, many traditional chemical pesticides have been withdrawn from use as a result of environmental and health concerns (Damalas and Eleftherohorinos, 2011). In India, thirtytwo pesticide (active ingredients) are banned for manufacture, importation and use, with a further eight withdrawn from the market and thirteen more restricted for use (CIBRC, 2017).

33

All these created an urgent need to adopt suitable non-toxic and eco-friendly method as alternative to chemical pesticides.

Biopesticides, the naturally occurring formulations made from the substances that control pests by non toxic mechanisms and in ecofriendly manner, are not newer technologies. They have been used in various forms since human civilization. Biopesticides being a living organisms (natural enemies) or products there of pose less threat to the environment and to human health, hence can be used for the management of pests.

Benefits of Fungal & Bacteria Antagonists  It offers efficient control of plant diseases.  It has a wide range of action.  High level of propagation in the soil, increasing its populations and exerting long-term control over phytopathogenic fungi.  It helps to decompose organic matter, converting nutrients into forms that are available to the plant, so having an indirect effect on crop nutrition.  It stimulates crop growth because it has metabolites that promote the development processes in plants.  It can be applied to decomposing compost or organic matter to accelerate the process of maturation.  It favors the proliferation of beneficial soil organisms, such as other antagonistic fungi.  It does not need a safety period before harvesting.  It preserves the environment and reduces the use of fungicides.  It economizes the costs of crop production.  It prevents disease by protecting the roots and leaves.  It promotes root and root hair growth.  It improves nutrition and water absorption.  It reduces or eliminates the dependence on chemical fungicides.  No phyto-toxic effect has been recorded.  It mobilizes soil nutrients for plants.  It acts as a bio- degradant of agrochemicals.  It can be used as weedicide.  It protects farm and botanical seeds from phytopathogens.  It is compatible with Mycorrhizae, Azotobacter and other biofertilizers.

34

 It is also compatible with bioagents that control pests and diseases Application of fungal antagonists Seed treatment: The talc-based formulation is used as a dry seed treatment @ 4 g/kg and the treated seed can be sown immediately.

Seedling root dipping: Apply 10 g of the formulation to the 10 lt. water. The seedlings, after pulling out from the nursery can be dipped in the water containing the bacteria/ fungus. A minimum period of 10 minutes is necessary for soaking the roots and prolonged soaking will enhance the efficacy.

Nursery treatment: Mix 500 g of the formulation in the 100 lt. of water and drench nursery beds (10 cm depth), immediately after sowing seeds. Use1.25 kg of prepared FYM containing formulation in the nursery beds at the time of mulching of nursery beds.

Soil application: Mix 1 kg of formulation with 50 kg of FYM spread in 3 X 6 beds and moisten it. Keep covered for 3-4 days. Turn FYM and moisten again 3-4 times. After 15 days apply/spread the prepared FYM mixture over one acre land Foliar application: Spray the product @ 2.5 kg / ha in 800 lt. of water at 10 days interval for 2 times depending on disease / pest infestation / intensity. If there is no disease incidence, a single spray is sufficient.

Mass production of Trichoderma spp.: Pure culture of Trichoderma spp.

Mass production on liquid media (PD-Broth) Incubate in B.O.D at 26±2OC for ten days Harvest the growth in mixer grinder Mix with inert material (1:3) along with CMC @1 per cent Quality control 1x 108 cfu/g

35

Table 1: Plant diseases management by bio-control agents: A. Cereal Crops

Pathogen

Bio-Control Agents

Mechanism of action

Reference

Wheat Take-All

Gaeumannomyces

Disease

var. tritici

graminis Pseudomonas fluorescens (Pf32- Antibiotic

Fusarium Head Fusarium graminearum

(2,4- Hiddink

gfp).

diacetylphloroglucinol)

(2005b)

Microsphaeropsis sp. (P130A)

Reduced ascospore production

Bujold

Blight Seedling Blight

et

al.

et

al.

(2001) Fusarium culmorum

and

Increase wheat yield

Pantoea sp. MF626

Microdochium nivale

et

al.(2003)

leaf blotch

Mycosphaerella graminicola

tan spot

Pyrenophora

tritici

Johansson

Induce systemic resistance

Trichoderma harzianum

Perelló

et

al.

et

al.

(2006)

– T. koningii

repentis Corn Ear Rot

Fusarium verticillioides

B.

amyloliquefaciens

and By reducing toxin levels

Pereira (2010)

Enterobacter hormaechei Sorghum and Millets Charcoal Rot

Macrophomina phaseolina

P. chlororaphis SRB127

Downy Mildew

Sclerospora graminicola

Trichoderma lignorum,

harzianum, Glocladium

Trichoderma )

virens

Enhance vigour index T

.

Das et al. (2008) Raj et al. (2005)

(= and 36

Bacillus subtilis Cotton Verticilium

Verticillium dahlia

Penicillium chrysogenum

induction of resistance

Dong et al. (2006)

Wilt Root Rot

Fusarium oxysporum ( Fo ) Trichoderma hamatum + and Pythium debaryanum ( harzianum Pd)

+

T. Enhanced control efficacy in Abo-Elyousr et al.

Paecilomyces combination

lilacinus

with

disease (2009)

resistance inducers

Seedling

Rhizoctonia solani , Pythium T . virens Q

antibiosis and/or induction of

Lumsden

Diseases

aphanidermatum ,

systemic resistance

(1992); Howell et

ultimum

and

P .

et

al.

al. (2000)

Rhozopus

oryzae.

Black Root Rot

Thielaviopsis basicola

Paenibacillus alvei K-165

Inhibits root colonization by T. Schoina

et

al.

systemic (Raghavendra

et

(2011)

Basicola Bacterial Blight

Xanthomonas

axonopodis

Sargassum wightii

Induction

of

resistance

pv. Malvacearum

al. (2007)

Pulse Crops Damping-Off of soybean

Rhizoctonia solani

Binucleate Rhizoctonia (BNR)

Suppression of R. Solani along Khan and Nelson with

increased

seedling (2005)

emergence and survival of soybean plants 37

Sclerotinia Stem

Rot

Sclerotinia sclerotiorum

Antibiotic cyclosporine A

Fusarium oxysporumS6

of

Rodriguez et al.

(2006).

soybean Fusarium Root F .

Rot of soybean

oxysporum

and

graminearum

Fusarium Wilt Fusarium oxysporum of Chickpea

F . f.sp.

ciceris

fluorescens Suppressed

Pseudomonas

development

RGAF101, P . fluorescens RG26

disease Landa et al. (2001, by

delayed 2004b)

expression of symptoms Root

Rot

of Macrophomina phaseolina

Chickpea

Glomus

intraradices

Rhizoctonia

sp.

Pseudomonas

putida

, Increased nutrients uptake and Akhtar and root colonization by BCA

Siddiqui

and

2008)

and (2007,

Paenibacillus polymyxa Damping-Off

Pythium ultimum

of Chickpea

Bacillus pumilus GB34 B . Production of enzymes subtilis

GB03, B . subtilis

MBI600,

Streptomyces lydicus

WYEC108 , K61),

Leisso et al. (2009)

S . grieseoviridis

Trichoderma harzianum

Rifai strain KRL-AG2 Ascochyta Blight

Ascochyta rabiei

Aureobasidium pullulans

Antagonism

Dugan et al. (2009)

of 38

Chickpea Pigeonpea Wilt

Fusarium udum

Aspergillus niger , citrinum

,

Penicillium Inhibitary effect on F. udum

Singh et al. (2002)

Trichoderma

harzianum and T .virens Root

Rot

of Macrophomina phaseolina

Mungbean

T. viride and T . harzianum or Antagonism

Choudhary et al.

(2010)

T. harzianum and Aspergillus versicolor

Oilseed Crops Root

Rot

of

Macrophomina phaseolina

peanut

Shanmugam et al.

Pseudomonas fluorescens Pf1

(2002)

+ Rhizobium TNAU-14 Fusarium solani

T. harzianum ITEM 3636 and

Antagonism

Rojo et al. (2006)

T. longibrachiatum ITEM 3635 Sclerotinia

Sclerotinia minor

Coniothyrium minitans

Blight of peanut

Mycoparasitism (infection and Budge et al. (1995) colonization of the sclerotia of Gerlagh pathogen)

Collar Rot of Sclerotium rolfsii

et

al.

(1999) Abd-Allah and El-

Bacillus subtilis

peanut

Didamony (2007) Aspergillus niger

Bacillus megaterium

Reduces the biosynthesis of Kong et al. (2010) aflatoxin by inhibiting

aflS 39

and aflR gene expresiion Sclerotinia Stem

Rot

Sclerotinia sclerotiorum of

Pseudomonas chlororaphis

PA- induction

of

systemic Fernando

resistance

23

et

al.

(2007)

oilseed rape Blackleg

of Leptosphaeria

Canola

biglobosa weakly

virulent

PG-1

isolate Systemic Acquired Resistance Chen and Fernando (SAR)

(PG-1) and L. maculans inoculated at 6 leaf stage

(2006)

(PG-2,PG-3 and PG-4) Head

Rot

of Sclerotinia sclerotiorum

Sun flower

Trichoderma

spp. With honey Suppression of development of Escande the sclerotia

bees as its vector

et

al.

et

al.

(1994 )

Tomato Damping-Off

Pythium spp., Rhizoctonia , Pseudomonas fluorescens , P. solani and Sclerotium rolfsii putida ,

P. marginalis ,

P.

corrugata and P. viridiflava Southern Blight

Sclerotium rolfsii

Bacillus

amyloliquefaciens

IN937a and B. pumilus IN937b,

Jetiyanon (2003)

SE34, SE49, T4 and INR Anthracnose

Stem Canker

Colletotrichum coccodes

Botrytis cinerea

Compost amendments stimulating Systemic induced resistance

Tränkner (1992)

the proliferation of antagonistic

Abbasi

microbes

(2002)

Rhodosporidium

Utkhede

et

al. and 40

Mathur (2006)

diobovatum Bacterial Wilt

Ralstonia solanacearum

Antagonism

Pseudomonas fluorescens

Ambadar and Sood (2010)

Paenibacillus polymyxa strains

Strong antibacterial activity

Algam

et

al.

(2010)

Bacterial Spot

Xanthomonas

filamentous bacteriophages

Infects and kills the pathogenic Addy et al. (2012)

ØRSM3

bacteria

axonopodis Bacillus pumilus SE34

Enhance the level of resistance

Ji et al. (2006)

Cross-protection

Wilson

pv. vesicatoria Bacterial Speck

Pseudomonas syringae

pv. P. syringae Cit7

et

al.

(2002)

Tomato Bacterial

Clavibacter

michiganensis fluorescent pseudomonads

Canker

subsp . michiganensis

Induced resistance

Antoniou

et

al.

(1995) Trichoderma

harzianum

and Suppresses

Rhodosporidium diobovatum

bacteria

the

pathogenic Utkhede and Koch (2004)

Potato Verticillium

Verticillium dahliae

Uppal et al. (2008)

F isolate DF37

Wilt Stem Rot

Pseudomonas fluorescens biotype

Sclerotinia sclerotiorum

Trichoderma koningii , T. virens ,

Ojaghian (2011)

T. ceramicum and T. viridescens 41

Late Blight

Phytophthora infestans

Pseudomonas putida

Induced systemic resistance

Daayf et al. (2003)

Serratia plymuthica

Antibiosis

Daayf et al. (2003)

Trichoderma spp. ,

Antagonism

Basu (2009)

Pseudomonas spp and Bacillus spp.

Dry Rot

Fusarium sambucinum

Bacillus

cereus

X16

+

B. Antagonism

Sadfi et al.( 2002)

thuringiensis var. galleriae and Trichoderma viride 55T Pink Rot

Phytophthora erythroseptica

Trichoderma virens DAR74290 Antagonistic

Etebarian

and T. harzianum T-39

(2000)

Enterobacter

cloacae

Enterobacter

spp.

Pseudomonas spp.

, Prevents

entery

of

the Schisler

et et

al. al.

and zoospores via potato tuber (2009) eyes or lenticels, cracks or cuts on the tuber

Potato Scab

Streptomyces scabies

green manures (buck wheat and Enriches the abundance or Wiggins canola)

and

activity of antibiotic producers Kinkel (2005) within

the

soil

microbial

community

42

Pepper Phytophthora

Phytophthora capsici

Production

Serratia plymuthica

Blight Verticillium

of

cell

wall Kim et al. (2008)

degrading enzymes Verticillium dahliae

Pythium oligandrum

Colletotrichum acutatum

Myxococcus spp. KYC1126, 1136 Produces

Wilt Anthracnose

and 2001

a

large

bioactive

no.

of Kim

and

Yun

secondary (2011)

metabolites Fusarium

Fusarium solani

Wilt

Bacillus subtilis, Pseudomonas Induced systemic resistance

Sundaramoorthy

fluorescens

et al. (2012)

Bacillus subtilis, Pseudomonas Antagonists

Georgakopoulos et

fluorescens, P. corrugate,

al. (2002)

Cucurbitacous Crops Pythium Root

Pythium aphanidermatum

Trichoderma viride and T. ( Gliocladium ) virens Fusarium Wilt

Fusarium oxysporum f.sp . Composts harbouring BCAs like Antagonistic activity against Bradley and Punja radicis

–

cucumerinum Pseudomonas

aeruginosa

and FORC and FOM

(2010)

(FORC) in cucumber while Aspergillus spp.

Suárez-Estrella

F. oxysporum f.sp. melonis

al. (2007)

et

(FOM) in melon.

43

Anthracnose

Colletotrichum orbiculare

Powdery

Sphaerotheca fuliginea

Seratia marcescens Ampelomyces quisqualis

Induced systemic resistance

Neher et al. 2009

Mycoparasites

Romero

Mildew

et

al.

et

al.

2007

Allium Crops White Rot

Sclerotium cepivorum

Degrade sclerotia

Trichoderma viride

Clarkson (2002)

Onion

leaf Botrytis squamosa

Colonizing onion leaves and Carisse

Microsphaeropsis ochracea

blight Black mold

reducing production of conidia Aspergillus niger

Production

Pantoea agglomerans

of

Fusarium oxysporum

al.

(2006)

antifungal Özer and Köycii

compounds Basal Rot

et

(2006)

Antibiosis

f.sp. T. harzianum

cepae (FOC) Xanthomonas Leaf Blight

Xanthomonas

axonopodis

pv . allii ( Xaa) c

Pantoea agglomerans C9-1 and

strain

Pseudomonas fl u

orescens A506 Apple Root Rot

R. solani AG-5

Mazzola,1998,

Pseudomonas putida

1999 Scab

Venturia inaequalis

Microsphaeropsis sp

Restricts ascospore production

Carisse et al. 2000

Powdery

Podosphaera leucotricha

Ampelomyces sp.

mycoparasites

Szentiványi

and 44

Mildew Fire Blight

Kiss, 2003 Erwinia amylovora

Pantoea agglomerans Eh252

Antibiotic

Stockwell

et

al.

2002) Grapevine Root and Stem

Armillaria mellea

Trichoderma harzianum

Powdery

Uncinula necator

Pseudozyma fl occulosa

(John et al. (2005) Antagonist

Bélagner

Mildew Downy Mildew

and

Labbe (2002) Plasmopara viticola

Penicillium chrysogenum

Inducing resistance

Thuerig

et

al.

(2006) Gray Mold

Botrytis cinerea

Ulocladium oudemansii

Colonization

Elmer

et

al.

(2003); Shorten et al. (2003) Crown gall

Nonpathogenic

Agrobacterium vitis,

Agrobacterium

strains vitis

of Competitors ARK- strain

of

pathogenic

Kawaguchi

and

Inoue (2012)

1,ARK-2 and ARK-3 Citrus Phytophthora Root Rot

P. parasitica nicotianae)

( = P. Pseudomonas putida 06909 and

P.

Parasitism (bacteria adhered to Turney the hyphae)

et

al.

(1994)

citrophthora.

45

Banana Panama Wilt

F. oxysporum f.sp . cubense

Nel et al. (2006)

P. fl uorescens strain WCS417; Trichoderma harzianum

Root rot

Cylindrocladium spathiphylli

Glomus proliferatum

Colonization of banana roots

Declerck

et

al.

(2002) Leaf Spot

Pseudocercospora

musae Bacillus subtilis

(teleomorph

Antagonism

Fu et al. (2010)

:

Mycosphaerella musicola ) Strawberry Verticillium

Verticillium dahliae

Wilt

Serratia plymuthica strain HRO- Chitinolytic activity

Kurze et al. (2001)

C48

Phytophthora

Phytophthora cactorum

Trichoderma spp.

Gray Mold

Botrytis cinerea

Ulocladium

atrum,

Porras et al. (2007) Bacillus Colonozier, Antagonist

Cook, 1993; Hang et al. (2005)

subtilis isolate S1-0210 Plantation Crops Blister Blight of Exobasidium vexans

Pseudomonas fl uorescens strain Induction

tea

Pf1

Phomopsis

Phomopsis theae,

Trichoderma harzianum

canker of tea Tea bird’s eye Cercospora ocellata

resistance

of

systemic Saravanakumar et al. (2007) Ponmurugan

and

Baby (2007) Streptomyces sannanensis

Gnanambigai

and 46

spot

Ponmurugan (2012)

Anthracnose of Colletotrichum tea

sinensis

Coffee Rust

Hemileia vastatrix

theae

– Bacillus subtilis BD0310 Bacillus

sp. isolate B157 and

Pseudomonas sp. isolate 286

Antagonism

Kim et al. (2009) (Haddad

et

al.(2009)

47

Chapter-6

Mushroom Diseases and Their Management Sachin Gupta and Ranbir Singh Division of Plant Pathology Sher-e-Kashmir University of Agricultural Sciences and Technology of Jammu, Main Campus, Chatha-180009 (J&K) ____________________________________________________________________________ Introduction Diseases and disorders of button mushroom (Agaricus bisporus) can be categorized into two factors namely, biotic and abiotic. Biotic causes include parasitic and antagonistic fungi, bacteria and insect pests. The abiotic causes of various diseases include temperature, relative humidity, carbon dioxide concentration in the air, moisture level in compost and casing and presence of toxic chemicals in atmosphere, compost or casing.

Biotic Causes (A)

Fungal Diseases

The general categorization of mushroom moulds and pathogenic fungi are as under: 1.

Those occurring mainly in compost

a)

Olive green mould

b)

Ink caps

c)

Brown plaster mould

d)

Green mould

2.

Those occurring in compost and in casing soil

a)

Brown plaster mould

b)

False truffle

c)

White plaster mould

d)

Green mould

3.

Those occurring on and in casing soil and on growing mushrooms

a)

Dry bubble

b)

Wet bubble, and

c)

Cobweb 48

(B)

Bacterial Diseases a) Bacterial blotch of button mushroom

Abiotic Causes a)

Rose comb

b)

Stoma

c)

Hollow core/stem

d)

Long stem/stipe

e)

Brown discoloration

f)

Scales or crocodiles

g)

Purple stem

Disease

and Symptoms

Management

Pathogen FUNGAL DISEASES/ DISORDERS Olive Green Mould

The early symptoms of the fungus  The fermentation period of the

(Chaetomium spp.)

consist

of

appearance

of

compost should not be too short.

inconspicuous greyish white fine  Avoid too wet compost mixture mycelium in the compost ten days

and should not be pressed too

after spawning that delays spawn

hard during composting.

growth. Fruiting structures of the  Sufficient oxygen should be fungus

look

like

grey-green,

applied during pasteurization.

cockle-burns that develop on the  Temperature straw in isolated spots of the

in

the

tunnel

should not exceed 620C.

affected compost. The infected  In severe cases, respawning of compost gives a musty odour. The

the compost should be done to

fungus grows in compost that

get good yield.

remains too wet and is not properly prepared. This can be attributed to ammonia left in the compost after peak heating or due to undercomposting.

49

Ink caps appear in the  Rogue out young fruit bodies of

Ink Caps (Coprinus spp.)

compost during spawn run or

the weed fungus to avoid further

newly cased beds and in the

spread.

compost pile during composting.  Properly pasteurize the compost.

These are slender, bell-shaped,  Compost mixture should not be cream coloured mushrooms later

pressed

turning

composting.

bluish

ultimately

black,

decay

and

which

too

hard

during

produce  Do not add insufficient gypsum

blackish slimy mass resembling

in the compost.

inky fluid. It depletes the food material from the compost and hampers spawn run and yield.

Brown

The fungus is characterized  Localized treatment of infected

Plaster

Mould

by producing large and dense

spots with 2 per cent formalin or

(Papulaspora

white patches initially on the

Bavistin 0.05% in initial stages

byssina)

compost and then upon casing

after the removal of affected

surface, later turning brown and

portions.

powdery with age. No mushroom  Prepare compost from fresh grows on places where plaster mould occurs.

materials.  Regulate

compost

moisture

between 62-65%.

The

Green Mould

fungus

appears

in  Maintain

proper

hygienic

(Trichoderma

spawn bottles, compost, casing soil

conditions in and around the

species)

and on grains after spawning. A

growing house.

dense, pure white

growth of  Proper

mycelium

on

appears

casing

pasteurization

and

conditioning of the compost

surface or in compost, which  Lower down the temperature in resemble

mushroom

mycelium.

the growing room to 15-180C.

Later the mycelium turns to green  Treat the affected patches with colour due to heavy sporulation, which

is

the

formalin 2% or Bavistin 0.05%

characteristic  Destroy the affected bags.

50

symptom of the disease. Sometime the fungus develops on the newly formed primordia and also grows around

dead

butts

of

the

mushroom. In severe cases, there is complete crop failure. The fungus also causes dry sunken brown spots on stem and cap.

The disease is also known  Prepare compost on a concrete

False Truffle (Diehliomyces

as calve’s brains grows with a

microsporus)

creamy

white

floor.

mycelium  Avoid temperature above 250C

resembling mushroom mycelium

during spawn run and after

in the initial stage. This may

casing.

appear mostly at the juncture of  Lower down the temperature to compost layer and the casing

15-180C in the growing room

mixture.

during cropping.

Initially

the

fungal

fruiting body appear like small pin  Formalin 2% or Bavistin 0.05% heads of mushroom and later

should be used to check the

turning to thick, bold, yellowish,

initial infection by treating the

grayish wrinkled mass resembling

infected patches.

brain like structures. At maturity, fruit bodies emit chlorine like odour and spreads fast on the compost and on casing soil. It usually

appears

when

the

temperature of the growing room exceeds 25 0C White

Plaster

The disease appears as  Maintain compost pH less than

Mould

dense white patches of mycelium

8.0.

(Scopulariopsis

on compost or casing soil giving  Spot touch of formalin 2% or

fimicola)

floor like appearance. The fungus

Bavistin 0.05% after scrapping 51

does not change its colour with age

the fungal mat.

as in case of brown plaster mould.  Proper composting and adequate Spawn run is hampered and yield

gypsum in compost preparation

is reduced. It occurs when compost

reduces growth of mould.

pH is more than 8.0.

Whitish mycelial growth is  Use properly sterilized casing

Dry Bubble (Verticillium

initially noticed on the casing soil

fungicola)

which turns grayish yellow. If  Proper infection takes place in early stage,

soil. disposal

of

spent

compost.

typical onion shape mushrooms are  Dithane M-45 0.2% or bavistin produced. Infection at later stage

0.05%

leads to malformed mushrooms

interval

with distorted shaggy swollen

controlling dry bubble.

treatment is

at

10-day

effective

in

stipes and tilted caps. In severe form, it produces light brown depressed necrotic spots on fully developed sporophores. In the later stages

these

become

irregular

swollen masses of leathery tissues.

When infection takes place  Cover the infected spots with

Wet Bubble (Mycogone

early before differentiation, of

common salt and plastic caps to

perniciosa)

stipe and pileus, the sclerodermoid

prevent the spread of infection.

form results. If infection takes  Maintain place

after

differentiation

proper

hygienic

conditions

mushrooms with thickened stipe  Spray Dithane M-45 at 0.2% at and deformed gills develop. In

7 day interval in severe cases.

severe cases, there is putrefaction of

fruiting

bodies

giving

unpleasant smell. The infected pinheads decay with brown to 52

umber colour liquid ooze.

It

Cobweb

produces grayish

to

conspicuous  Avoid

(Dactylium

cottony

rose

red

dendroides)

mycelial patches on casing soil.  Treat

excessive

humidity

during cropping. infected

portion

with

Later it engulfs the developing pin

0.15%

heads and buttons which results in

solution or spray with 0.2%

their rottage.

Dithane M-45.

calcium

hypochlorite

BACTERIAL DISEASE OF WHITE BUTTON MUSHROOM Bacteria

The diseased mushroom  Avoid

Blotch

spraying

water

on

(Pseudomonas

shows yellowish brown patches on

developing pinheads.

tolassii)

cap and stipe. In severe cases  Keep fans on or doors and whole mushroom turns brown in

windows open after water spray

colour. Sometimes brown sunken

to

pits are also seen on mushrooms.

mushrooms.

allow

quick

drying

of

The young pinheads turns brown  Remove diseased mushrooms in colour and die prematurely. The

and spray bleaching powder at

disease

0.05% at seven day interval.

spreads

rapidly

mushrooms stay wet for long.

when

 Avoid temperature above 180C and humidity more than 85 % during cropping.  Streptomycin

200

ppm

or

oxytetracycline 300 ppm is also effective

in

controlling

the

disease

53

ABIOTIC DISORDERS OF WHITE BUTTON MUSHROOM Disease Rose Comb

Cause

Symptoms

Management

 The deformity occurs  Pink gill tissues with  The

of

diesel/

due to contamination

porous

by hydrocarbons (diesel

develop

on

the

growing rooms should

or kerosene)

surface

of

the

be avoided.

 Contaminated soil

with

appearance

use

kerosene for heating the

casing

mushroom cap giving  Heat the growing room

heavy

comb like appearance

overdose of pesticide

with steam and the

 Mushrooms

are

vapours

distorted

and

maintain the humidity.

unsaleable.

will

also

 Use of pesticides in the casing soil should be limited

Stroma

 It is due to high carbon  Mycelium

after  Ruffle

the

affected

dioxide concentration,

penetrating the casing

high relative humidity

soil

and high temperature in

pinheads but contiue

dioxide

the growing room.

to grow to form thick

below 300 ppm.

do

not

casing layer and recase.

form  Maintain

mycelial mat above  Bring the casing layer.

the

carbon

concentration down

temperature

the below

150C in the growing room.

Hollow Stem

 This abnormality arises  Stems are not solid,  Avoid low humidity in due to low humidity,

instead

excess of dry air and

hollow.

they

are

less moisture in the  Stipe begins to split casing.

the growing room  Keep the casing soil moist.

and forms scale on stipe and pileus.

54

Chapter-7

Integrated Disease Management in Vegetables and Flowers Amrish Vaid and Ranbir Singh Division of Plant Pathology, Faculty of Agriculture, Sher - e - Kashmir University of Agriculture Sciences and Technology of Jammu, Chatha, Jammu-180009 __________________________________________________________________________ Introduction The term Integrated Disease Management was first given by Smith and van den Bosch (1967), based on the ‘integrated control’ concept given by Stern, et al. (1959), entomologists from University of California, who defined it as “applied pest control which combines and integrates biological and chemical control supplemented with the plantation of the resistant varieties. Once these preventive measures are taken, the fields should be regularly scouting to determine the levels of pests and the environmental factors. When the disease is above the threshold level then only the suppressive measures should be taken with most suitable technique or combinations of techniques, such as biological control, genetically resistant hosts, environmental modifications and when necessary an appropriate, selective use of pesticides, while causing minimum disruption of natural enemies. This approach is markedly different from the traditional application of pesticides on a fixed schedule. The various diseases of flowers and their management is discussed as under Rose Black spot- Diplocarbon rosae Symptoms 

Black lesions with feathery margins surrounded by yellow tissue are found on the leaves.

Infected leaves drop prematurely.



Purple/red bumpy areas on first year canes may be evident.



Plants may be weakened due to defoliation and reduced flower production may be observed

55

Management 

Roses should be planted where the sun can quickly dry the night's dew.



Space roses far enough apart for good air circulation Avoid overhead watering and keep foliage as dry as possible.



Remove infected canes and burn diseased leaves.



Spraying with Mancozeb (or) Chlorothalonil 0.2% (or) Benomyl 0.1% or a copper dust .

Powdery mildew – Sphaerotheca pannosa Symptoms: 

The symptom appears as grayish-white powdery substance on the surfaces of young leaves, shoots and buds.



Infected leaves may be distorted, and some leaf drop may occur.



Flower buds may fail to open, and those that do may produce poor-quality flowers.



It can occur almost anytime during the growing season when temperatures are mild (70 to 80 °F), and the relative humidity is high at night and low during the day.



It is most severe in shady areas and during cooler periods

Management: 

Collection and burning of fallen leaves.



Spray with Wettable sulphur 0.3% (or) Dinocap 0.07% (or) Carbendazim 0.1% 2-3 sprays at 15 days interval is effective.

Die back – Diplodia rosarum3 Symptoms: 

Drying of twigs from tip down wards.



Blackening of the twigs.



The disease spreads to root and causes complete killing of the plants

Management: 

Pruning should be done so that lesions on the young shoots will be eliminated. Apply chaubatia paste in the pruned area.



Spray with Difolatan 0.2% (or) Chlorothalonil 0.2% (or) Mancozeb 0.2%

56

Rust – Phragmidum mucronatum Symptoms: 

Damage to lemon yellow pustules appear on lower surface of the leaves and stems. Then the colour changes to blackish red.



The affected leaves turn yellow deformed and fall prematurely.

Management: 

Collection and burning of fallen leaves



Spray with Carboxin 0.1% or Wettable sulphur 0.25% or Captan 0.2%

CHRYSANTHEMUM Wilt -Fusarium oxysporum f.sp. chrysanthemi Symptoms 

Initial symptoms are in the form of yellowing and browning of leaves.



Affected leaves die from the base of the plant upward. Infected plants are stunted and often fail to produce flower.



Wilting may cause rotting of root or the base of the stem.

Management 

Drenching the soil with or Carbendazim 0.1% is effective.



Before planting dipping the rooted cuttings in a solution of Thiram @1.5g/litre of water.

Since the disease spreads mostly through cuttings, it is important to use disease free planting material.



Disease can further be minimized by following strict sanitation; periodical monitoring; crop rotation and roguing of infected plants.

Rust -Puccinia chrysanthemi Symptoms 

The disease symptoms are in the form of brown blister-like swellings, which appear on the undersides of leaves.



These burst open releasing masses of brown, powdery spores. Severely infected plants become very weak and fail to bloom properly

.Management 

Early removal of infected leaves/plants helps to prevent the further spread of the disease.

57



Spraying the plants with Karathane @ 0.025% or Wettable Sulphur @ 0.3 % is effective in controlling the disease.

Septoria Leaf Spot -Sepotria chrysanthemella Symptoms 

Leaf spots occur during cool-wet periods of the rainy season.



Since the pathogens are spread through rain splashes the lowermost leaves get infected first.



Serious infection may result in premature withering of the leaves; the dead leaves hang to the stem for some time.



When flowering starts the infection occurs on flower buds, which rot completely.

Management 

This disease can be controlled by spraying Carbendazim 0.1% six times at 15 days intervals from the end of July or spraying Benomyl (0.1%) followed by Captafol (0.2%)



Destruction of disease debris and avoiding excessive irrigation is recommended.

Powdery Mildew -Oidium chrysanthemi Symptoms 

Infection is more severe in older plants under humid conditions.



The growth of the fungus on the leaves appears as powdery coating. Infected leaves turn yellow and dry out.



Infected plants remains stunted and fail to flower.

Management 

Disease can be effectively controlled with Sulphur fungicides or Captan (0.2%).



Good ventilation and proper spacing for free circulation of air is recommended.

Jasmine Cercospora leaf spot – Cercospora jasminicola Symptoms: 

Circular to irregular reddish brown spots of 2-8 mm dia appear on the appear surface of the leaves.



Later the spots become irregular covering larger areas of the leaves.

58

Management: 

Spraying with Mancozeb 0.25% (or) Carbendazim 0.1%

Alternaria leaf blight – Alternaria jasmine, A. alternate Symptoms: 

In the leaves dark brown spots appear.



On fumed condition the spots enclanges covering larges area causing blighting of leaves.

Concentric rings can be seen the lesions. The disease also affects stem, petiole and flowers.

Management: 

Collection and removal of fallen leaves



Spray with Copper oxychloride 0.25% or Mancozeb 0.25%

Phyllody – Phytoplasma Symptoms: 

Leaves become small malformed and bushy. In the place of flowers green leaf like malformed flowers are formed.

Management: 

Selection of cuttings from healthy plants.



Spraying insecticide to control the vector.

TUBEROSE Stem Rot: Sclerotium rolfsii Symptoms 

The disease symptoms are preceded by the appearance of prominent spots of loose green colour due to rotting which extend and cover the entire leaf. The infected leaves get detached from the plant.

Management 

The disease can be controlled by soil application of Brassicol (20%) @ 30kg/hectare.

Botrytis Spot and Blight: Botrytis elliptic Symptoms 

The disease appears during the rainy season.

Infected flowers show dark brown spots and ultimately the entire inflorescence dries up. The infection also occurs on the leaves and stalks.

59

Management 

Spraying the plants with Carbendazim @2g/litre of water effectively controls the disease.



The treatment should be repeated at 15 days interval.

In general the varius strategies to manage the diseases in vegetables and flowers are Prevention and Management Practices Site Selection and Preparation: Soil borne diseases remain a major limiting factor for the production of vegetables. It is important to start with clean soil and proper sites for crops. Plowing and disking will reduce pathogen carryover in old crop refuse. The longer the fallow period, the more pathogen populations are reduced. It is also essential to follow the latest recommendations for soil fumigation, cultural practices and biological control options to eliminate or reduce initial inoculum of soil borne pathogens. It is important that soil compaction is avoided, since this interferes with root growth, encourages soil moisture retention and promotes root diseases.

Preparation of raised beds generally allows for better drainage. Prior to planting, soil should be tested for nutrient levels and nematode populations (and other pathogens if tests are available).

Planting times can be altered to avoid or reduce development of certain diseases.

Host Resistance: Development of resistant varieties through hybridization is a cheap method of disease management for the farmers. With the development in biotechnology it is possible to introduce distant resistance genes in the plant by manipulation, genetic modification and multiplication of plants through techniques such as tissue culture and genetic engineering.

Soil and Fertilizer Management: Plant nutrition and soil pH can also impact some diseases. Fertilizers with a higher proportion of nitrate nitrogen (NO3) than ammoniacal nitrogen (NH4) will help to reduce the incidence of Fusarium wilt on tomato. Increasing soil pH by liming is a good management strategy to reduce Fusarium wilt incidence as well as Botryis gray mold severity. Optimum calcium nutrition and higher soil pH may reduce the incidence of bacterial wilt in the field. Adequate calcium is necessary to minimize blossom end rot and to provide for overall healthy growth. Avoiding excessive nitrogen leads to less dense canopies, thus improving air movement in the canopy.

60

Cultural Practices: Cultural practices serve an important role in prevention and management of plant diseases. The benefits of cultural control begin with the establishment of a growing environment that favors the crop over the pathogen. Reducing plant stress through environmental modification promotes good plant health and aids in reducing damage from some plant diseases. Sanitation practices aimed at excluding, reducing or eliminating pathogen populations are critical for management of infectious plant diseases. It is important to use only pathogen-free transplants, especially for late blight, bacterial spot, viral diseases and early blight. In order to reduce dispersal of soil borne pathogens between fields, stakes and farm equipment should be decontaminated before moving from one field to the next. Reduction of pathogen survival from one season to another may be achieved by destruction of volunteer plants and crop rotation. Removal of cull piles and prompt destruction of crops should be applied as a general practice. Avoid movement of soil from one site to another to reduce the risk of moving pathogens. For example, sclerotia of Sclerotinia sclerotiorum and Sclerotium rolfsii, are transported primarily in contaminated soil. Minimizing wounds during harvest and packing reduces post-harvest disease problems. Depending on crops and other factors, sanitation of soil can be achieved to some degree by solarization.

Crop Rotation : Crop rotation is a very important practice, especially for soil borne disease control. For many soil borne diseases, at least a 3-year-rotation using a non-host crop reduces pathogen populations.

This practice is beneficial for Phytophthora blight of pepper and Fusarium wilt of watermelon.

Land previously cropped to alternate and reservoir hosts should be avoided whenever possible.

Vegetable fields should be located as far as away as possible from inoculum and insect vector sources.

Biological Control: Biocontrol agents for use in vegetable disease management are increasing in use especially among organic growers. These products are considered safer for the environment and the applicator, than conventional chemicals and are mainly used against soil borne diseases.

Examples of commercially available biocontrol agents include the fungi Trichoderma harzianum and T. virens, an actinomycete Streptomyces griseoviridis and a bacterium Bacillus subtilis.

61

Chemical Control: Fungicides and bactericides are an important component of many disease management programmes. It is important to remember that chemical use should be integrated with all other appropriate tactics mentioned above.

Information regarding physical mode of action of a

fungicide is helpful in timing of fungicide applications. Physical modes of action of fungicides can be classified into four categories: protective, after infection, pre-symptom, and anti-sporulant (post-symptom). Protectant fungicides include the bulk of the foliar spray materials available. In order to be effective, protectant fungicides, such as copper compounds, mancozeb, etc., need to be on the leaf (or plant) surface prior to arrival of the pathogen. Systemic (therapeutic) fungicides, based on their level of systemically are active inside of the leaf (can penetrate at different rates through the cuticle). Systemic fungicides may stop an infection after it starts and prevent further disease development. If necessary, fungicides must be used based on recommended fungicide resistance management strategies. A new strategy to chemically manage plant diseases without direct interference with the pathogen is the triggering of plant defense reaction. Chemicals must be used at recommended rates and application frequencies. Besides selection of the most efficacious material, equipment must be properly calibrated and attention paid to the appropriate application technique. As always, the key to effective disease management is correct diagnosis of the problem. Always read the pesticide labels and follow the instructions carefully. Effective management of whiteflies, thrips, and aphids should be practiced to reduce the incidence and secondary infections of viral diseases vectored by these insects.

62

Chapter-8

Isolation, Identification and Culturing of Plant Pathogens Ranbir Singh, Sneha Choudhary, Dechan Choskit and Irrha Sheikh Division of Plant Pathology, SKUAST-Jammu ______________________________________________________________________ Introduction Isolation of the fungal pathogens from diseased material is made by surface sterilizing the diseased area with surface sterilizing agents, removing a small portion of the infected tissue (leaves, stems, fruits etc.) with a sterile scalpel, and plating it in a plate containing a nutrient medium. The most common method, for isolating fungal pathogens from infected leaves as well as other plant parts involves cutting several small sections 5-10 mm-square from the margin of the infected lesion to contain both diseased and healthy looking tissue. These are placed in surface sterilizing agents solutions for about 15-30 seconds the sections are taken out aseptically and blotted dry on clean, sterile paper towels or washed in three changes of sterile water and are finally placed on the nutrient medium, usually three to five per dish. The pathogen will grow from the sections and the colonies of the pathogen are sub cultured aseptically for further study.

Materials required Infected young leaves,sterile Petri -dishes, PDA slants, sodium hypochlorite solution ( 1 % ), sterile water, razor blade, forceps, inoculation needle, burner/spirit lamp, spirit, incubator, PDA medium.

Procedure 1. Select infected host tissue from the advancing margin of the lesions.

2. Cut into small pieces (2-5 mm ) containing both the diseased and healthy tissue and keep in sterile Petri dishes 3. Dip the pieces into 1 % sodium hypochlorite solution for about one minute.

4. Transfer the pieces to Petri - dishes containing sterile distilled water and wash thoroughly in two changes of sterile water to free them from the chemicals if any.

5. Wash hands with rectified spirit and wipe the table top of inoculation chamber-'with rectified spirit.

63

6. Lit the burner 7. Hold the flask containing sterile Luke warm PDA in the right hand and remove plug near the flame. Lift the lid of Petri dish gently with left hand and pour about 20 ml of medium. Close the mouth of the flask with plug near the flame 8. After solidification of the medium, place four sterilized pieces at different distance in a single PDA plate.

9. Incubate the Petri dishes in an inverted position at 25° C and examine for 3-5 days.

Observations and results Observe the incubated plates from the second day onwards for the growth of the fungus.

Aseptically transfer the bits of mycelia from the margin of the colonies on fresh PDA slants for further study. Mycelia growth on the medium from the infected tissues, indicates that the disease may be due to a fungus.

Isolation of phyto-pathogenic bacteria from diseased plants Isolation and identification of bacteria associated with diseased plant is important to determine whether bacteria are involved in plant disease. The method normally used to isolate phytopathogenic bacteria differs from that used for fungi. A suspension of bacterium is prepared from the infected material and loopfuls of this are streaked onto nutrient agar plates. The aim is to produce single colony that can be sub-cultured pure. Pure cultures are absolutely essential for pathogenicity assays and characterizing the pathogen for identification. The serial dilution method is used for isolating bacteria from diseased tissues contaminated with other bacteria.

After surface sterilization of sections of diseased tissues, the sections are ground in small volumes of sterile water and then part of this homogenate is diluted serially. Finally, plates containing nutrient agar are streaked with a loop dipped in each of the different serial dilutions and single colonies of the pathogenic bacterium are obtained from the higher dilutions that still contain bacteria.

Choice of material: Selection of the diseased tissue is important because pathogenic bacteria may occupy different locations in the plant. In isolation of bacteria, it is generally better to use newly collected material. The earliest stages of symptom development should be used. Old lesions and dead areas usually contain few pathogens and many saprophytes. Necrotic diseases usually start with tiny, dark greenish, spots, which are excellent for isolations. Cankers and soft rots should either be at an early stage, or if, older lesions only are available, the advancing edge 64

must be used, where the disease is spreading into healthy tissue. When crown gall is suspected in a woody plant a search must be made for young galls on young green stems. With wilts and other vascular infections small pieces of infected stem are usually good for isolation.

Preparation of material: Clean leaves and stems, carefully chosen and "handled aseptically, can often be used without surface sterilization. Roots and parts contaminated with soil should be gently washed with clean water as soon as possible after collection.

Medium: Nutrient agar is suitable for the isolation of most plant pathogens. The medium used for isolations must have a dry surface. If water is present the bacteria move around and a carpet of mixed growth results instead of the required single colony. This exercise deals with the isolation of bacterium, Xanthomonas axonopodis pv. citri causal agent of citrus canker.

Materials Fresh citrus leaves infected by Xanthomonas axonopodis pv. citri, nutrient agar medium, surface sterilizing agents (1 % sodium hypochlorite), sterile razor blade, glass rod, sterile water, sterile test tubes and Petri-dishes, sterile pipettes (I ml), inoculation loop.

Procedure Put on the U.V lamp of inoculation chamber for 5 mts. Wipe the table top with rectified spirit Wash hands with rectified spirit and air dry. Lit the burner or spirit lamp, arrange sterile Petri dishes near the burner.

1. Select a diseased citrus leaf infected by canker and cut out a small portion of the diseased tissue from the advancing lesion using sterile razor blade in a drop of sterile water and after several minutes, examine under microscope . If bacterial ooze is seen, proceed for isolation.

2. Surface disinfests the cut portions by dipping in sodium hypochlorite solution for 60 sec. and then immediately rinse three times with sterile water.

3. Immerse the disinfested cut portions in I ml of sterile water taken in a clean sterilised test tube.

4. Crush the cut portions of the leaf with a sterile glass rod. Allow it to stand for 5 minutes to allow the bacteria to diffuse out of the cut tissue and into the water.

5. Gently lift the lid of a Petri dish with left hand and using inoculation loop transfer several loopfuls of the bacterial suspension to sterile Petri-dishes (three) containing 1 ml of sterile water and mix thoroughly.

65

6. Hold flask filled with sterile Luke warm nutrient agar medium in the right hand and remove cotton plug near the flame and pour about 20 ml of medium into each dish and mix thoroughly by gentle rotation. Allow time for solidification of medium.

7. Incubate the dishes in an inverted position at 25°C for 36 to 72 hours.

8. Observation: Observe the dishes for appearance of desired bacterial colonies. If colonies appear, select consistently found and well isolated colonies of the pathogen, for sub-culturing and further studies.

9. Select the isolated colonies and streak on the surface of a solidified medium in a zigzag manner and incubate the dishes at 25oC. Bacteria isolated from nature may be contaminated with saprophytic species; hence, re-streaking for isolation ensures a pure culture. Transfer some of the purified colonies to NA slants and grow them for further use.

Preservation of living cultures Living cultures are stored for use as reference cultures, or for later use in pathogenicity tests or other experiments. Cultures are stored in national culture collections as part of reference materials that support a national database of plant pathogens.

Storage in sterile water—Pythium and Phytophthora This is a low-cost, simple method that is particularly suitable for Pythium and Phytophthora. A sterile work chamber should be used for this procedure. Agar blocks 1 cm square are cut from the margin of a young, actively growing fungal colony. These are placed in sterile water in a McCartney bottle and the cap is screwed down. The bottles are stored under cool conditions. Do not store in a refrigerator as some species are killed at low temperatures. Cultures can be stored between 6 months to 2 years, depending on the species. Cultures are revived by removing a block of agar from the bottle and placing mycelium side down on fresh medium. It is essential to ensure that the water and agar blocks are not contaminated by bacteria—the presence of bacteria will lead to rapid death of the fungus.

Storage of sclerotia Sclerotia can be stored for long periods under cool dry conditions in a small screw cap glass bottle or ampoule. This is a suitable technique for storage of species such as Sclerotium rolfsii, Sclerotinia sclerotiorum, Rhizoctonia spp. (sclerotial forming isolates).

In tropical regions it is best to store sclerotia on sterile paper tissue over blue silica gel in a McCartney bottle (or similar screw cap bottle) to ensure very low humidity for storage.

66

Storage as colonized pieces of plant stem or leaves Cultures are grown on sterile WA containing pieces of sterile plant tissue or seeds. The colonised pieces are air dried and then stored in a small glass tube. Alternatively they can be stored on a sealed container on sterile paper above blue silica gel to ensure very dry storage conditions.

Lyophilisation by freeze drying Lyophilisation, or freeze-drying, is the method of choice for long-term preservation of many fungi and is used routinely in most major culture collections. Its major drawback is the requirement and expense of specialised equipment. It is best suited to fungi which grow and sporulate well in culture on sterile plant tissue such as green rice stem-pieces or carnation leafpieces. There are also many fungi which cannot be freeze-dried successfully, such as oomycetes, rusts and mildews.

Cultures are lyophilised by drying colonised stem or leaf pieces in small glass ampoules under high vacuum (10–1 to 10–2 Torr). The ampoules are prepared by inserting a small cotton wool plug and then autoclaving in a loosely covered beaker. Five stem or leaf pieces are taken from a culture (which is two weeks old and initiated from a single conidium), and aseptically transferred to the ampoule. The ampoule is replugged, labelled (with an internal label) then heated and drawn out to an hourglass shape using a gas torch. The ampoules are attached to the freeze dryer for 12–24 hours, then sealed under high vacuum and stored at room temperature or at 5 °C. Many species of Fusarium and other fungal genera have been successfully lyophilised using this technique and have retained viability for many years. Cultures can be revived by aseptically plating the dried stem or leaf pieces onto a suitable medium. The ampoule is first surface sterilised before it is shattered to release the leaf pieces.

Other preservation techniques for living cultures Cultures can also be stored as spore suspensions in glycerol in a –80 °C freezer for long-term storage. Many species have also been stored successfully in liquid nitrogen. However, these are very expensive techniques.

Preservation of fungal cultures for herbarium records Cultures are initiated from single germinated conidia and grown under standard conditions of temperature and light for 2 to 3 weeks. Cultures are then killed by exposing the plates to formalin in a closed container for 3 days. Preservation of the culture is achieved using agar and glycerine.

Three grams of agar are dissolved in 147 mL water, which is then dispensed as 6 mL aliquots 67

into test tubes before autoclaving. The lid of the culture dish is inverted, 1.5–1.75 mL glycerine is added and then the 6 mL aliquot of hot agar is poured over the glycerine. The culture is aseptically lifted from the Petri dish and floated on the mixture in the lid. Cultures are then allowed to dry in a drawer for 3–5 days covered with a sheet of paper. When dried, the culture is rubbery and can be removed from the Petri dish for storage. This procedure was originally developed for use with Fusarium species at the Fusarium Research Centre, Pennsylvania State University. It is suitable for many fungi.

68

Compendium on Two Days Training Programme on

Integrated Disease Management (18-19 February, 2021)

Blight of Potato

Yellow Rust of Wheat

Loose smut of wheat

Leaf curl of Peach