30

3. Review of Literature

3.1. Major lepidopteran and mite pests of tea

The monographs by Green (1890) and Watt and Mann (1903) are the earliest

contributions on study of tea pests. Information on tea pests and their biology from

north-east India is subsequently given by Hainsworth (1952), Das (1965), Banerjee

(1983a, 1983b) and that of south India by Muraleedharan (1983). Present scenario of

insect and mite pests of tea throughout the world and their management has been

reviewed by Hazarika et al. (2009). Mukhopadhyay and Roy (2009) presented a recent

view on changing dimensions of climate, pest complex and pest management strategies

in sub Himalayan tea belt of North East India. Globally, 1031 arthropod species are

associated with tea (Chen and Chen, 1989). Due to the influence of climate, altitude,

nature of cultivation and age of plantation each geographic region may have its own

distinctive pest complex (Banerjee, 1983a; Muraleedharan, 1992; Anonymous, 1994;

Watt and Mann, 1903). In India only 300 species of insects belonging mainly to the

orders Lepidoptera, Hemiptera and Coleoptera, and mites (Acari) are recorded as tea

pests (Muraleedharan et al., 2001). Among these only 6 species are of major

importance from sub Himalayan West Bengal (Mukhopadhyay and Roy, 2009).

3.1.1. Major lepidopteran tea pests of Darjeeling Terai :

Among the tea attackers, the order Lepidoptera forms the largest group comprising

31.53% of the total pest species (Chen and Chen, 1989) and is enable to cause up to

40% crop loss (Banerjee, 1993). Recently even 100% crop loss by lepidopteran

defoliators is recorded in severely attacked sections of tea plantations of sub Himalayan

West Bengal (personal communication with tea estates). Major lepidopteran species

31

attacking tea in Terai and the Dooars regions of West Bengal are the species of looper

caterpillars, Buzura suppressaria, Hyposidra talaca and H. infixaria, belonging to the

family Geometridae (Anonymous, 1994; Mukhopadhyay and Roy, 2009). H. talaca

alone constitute 74% of the combined population of the lepidopterans attacking tea

(www.teaboard.gov.in). Some other members of the family Geometridae, such as,

Buzura bengaliaria, Boarmia sclenaria, B. acaciaria, Medasina strixaria, etc are

known to occur on tea, but none of them have attained the status of a pest (Anonymous,

1994). At the same time, some earlier known lepidopteran pests of tea, such as, jelly

grub, neetle grub etc. have lost their importance as pests. Some occasional and sporadic

lepidopteran pests of tea of Terai region include red slug caterpillars (Eterusia

magnifica), Ectropis bhurmitra, Cydia leucostoma, Caloptelia theivora, Ascotis sp.,

Euproctis latisfascia and others (Mukhopadhyay and Roy, 2009; Prasad and

Mukhopadhyay, 2013).

Buzura (Biston) suppressaria Guen.:

The common looper caterpillar, Buzura (Biston) suppressaria Guen., was first

recorded on tea in India by Cotes (1895) from Nowgong district of Assam. The

species had migrated to tea from jungle plants used as shade trees in the tea

plantations. Borthakur (1975) reported looper caterpillars as one of the important

pests of tea. Beeson (1941) in ‘The ecology and control of the forest insects of

India and neighboring countries’ recorded looper caterpillar on alternate hosts

such as Acacia modesta, A. catechu, Aleurites montana, Bauhinia variegata,

Cassia auriculata, Carissa diffusa, Dodonaea viscose, Lagerstroemia indica,

Dalbergia assamica, Deris robusta, Albizzia chinensis, A. odorotissima, A.

lebbek, Cajanus indicus and Priotropis cytisoides. By early 1990’s B.

suppressaria attained major pest status attacking tea in the plantations of Assam

and the Dooars region of West Bengal (Anonymous, 1994). However, in recent

32

past incidence of the species has become much lower than other loopers in Terai

region of West Bengal (Das, S. et al., 2010). Rare occurrence of looper

caterpillars has also been reported from tea growing areas of South India along

with their association with shade trees (Muraleedharan, 1991; 1993). Looper

caterpillars have been tainted as active defoliator of Indian and South-East Asian

tea (Hill, 1983).

Hyposidra talaca (Walker):

In recent past caterpillars of the geometrid species, Hyposidra talaca (Walker)

have emerged as major pest of tea from the Dooars region of West Bengal (Basu

Majumdar and Ghosh, 2004). Early report of occurrence of looper stages of H.

talaca on tea dates back to 1972, which also records the species on Cocoa,

Cinchona, Coffee and other fruit trees in tropical lowlands and highlads

(Entwistle, 1972). The species is polyphagous and is widely distributed. A brief

account on different species of Hyposidra is provided in the website,

www.mothsofborneo.com. Geographically, H. talaca is found in Indo-Australian

tropics from north-east Himalaya to Queensland and Solomons, and is recorded

from 96 plant species belonging to 28 plant families (Robinson et al., 2010) such

as Anacardiaceae (Anacardium sp.), Bombacaceae (Bombax sp.), Combretaceae

(Terminalia sp), Cupressaceae (Cupressus), Euphorbiaceae (Aleurites sp.,

Aporusa sp., Bischofia sp., Breynia sp., Glochidion sp., Manihot sp.), Moraceae

(Ficus sp.), Myrtaceae (Psidium sp.), Polygonaceae (Polygonum sp.), Rosaceae

(Rubus sp.), Rubiaceae (Cinchona sp., Coffea sp., Mussaenda sp.), Rutaceae

(Citrus sp., Euodia sp.), Sapindaceae (Schleichera sp.), Sterculiaceae

(Theobroma sp.), Theaceae (Camellia sp.), Verbenaceae (Tectona sp.) etc. to

name a few. The species is even found associated with narcotic plant coca

(Erythroxylum coca) (Rosen, 1991). The polyphagous species feeds on the foliage

33

of many forest trees in India (Mathew et al., 2005). Though about a hundred plant

species are recorded as alternate hosts of the species globally, it attained pest

status in a few cases. H. talaca is a quarantine pest of Litchi chinensis,

Dimocarpus longan and Garcinia mangostana from Thiland (Kuroko and

Lewvanich, 1993; Crop Protection Compendium (CPC), 2002; Thoda, 2004). It is

known as a pest of mango (Mangifera indica) (Butani, 1993; CPC, 2001) and

recently mentioned as a minor defoliator of forest trees, such as, Teak (Tectona

grandis) (Nair, 2007), Sal (Shoria robusta) (Sen Sharma and Thakur, 2008),

Quercus incana (Singh and Singh, 2004) etc. in India. H. talaca is also recorded

as a tea pest in Indonesia. Some work on the host association, biology and

oviposition behavior has been done by Das and Mukhopadhyay (2008), Das, S. et

al. (2010), Sinu et al. (2013).

Hyposidra infixaria Walker:

Different life stages (larva, pupa and adult) of H.infixaria apparently resemble

those of H.talaca, though distinguishing morphological features can be seen on

closer observation (Das, S. et al., 2010). H.infixaria is reported as a pest of tea in

West Bengal recently (Nair et al., 2008a; Das and Mukhopadhyay, 2009). The

occurrence of H. infixaria Walker ranges from north-east Himalaya to Taiwan and

Sundaland (www.mothsofborneo.com). The species had been reared on Pisum,

Desmos, Buchanania and Punica. Looper stage of the species was found

associated with castor bean, Ricinus communis and pomegranate, Punica

granatum and a weed, Rhodomyrtus tomentosa in Thiland (Winotai et al., 2005).

Nair et al. (2008a) reared the species on seven different host plants, wild jatropha,

jamun, tea, amra, jarul, citrus and water apple in Nadia district of West Bengal

and found lowest development period and highest growth index of the species on

34

tea. Difference in growth and survival of H. infixaria on different clonal varieties

of tea is also available (Basu Majumder et al., 2011).

3.1.2. Major mite pest of Darjeeling Terai:

Mites are persistent and the most serious pests of tea in almost all tea producing

countries (Cranham, 1966). Among twelve species of mites recorded on tea, the tea red

spider mite, Oligonychus coffeae Nietner (Acarina: Tetranychidae) is the major one and

is a serious pest of tea throughout the world (Banerjee, 1988; 1993) as well as in

Darjeeling Terai. O. coffeae was first discovered on tea in 1868 in Assam, India (Watt

and Mann, 1903) and besides India the mite is widely distributed in Bangladesh, Sri

Lanka, Taiwan, Burundi, Kenya, Malawai, Uganda, and Zimbabwe (Gotoh and Nagata,

2001).

Studies on life history and biology of O. coffeae on tea have been carried out in

different parts of the world including NE India (Das, 1959; Chakraborty et al., 2007;

Das et al., 2012; Mazid et al., 2013) and South India (Selvasundaram and

Muraleedharan, 2003; Muraleedharan et al., 2005). Recently, Roy et al. (2014) reviewed

the pest status, biology, ecology and management of O. coffeae in tea plantations.

The species inhabits upper leaf surface and are easily dislodged by heavy rainfall

(Hazarika et al., 2009). Life cycle of red spider mite, O. coffeae, consists of egg, larva,

nymph and adult stages. The egg is spherical with a curved filamentous process arising

from the upper pole and is red in colour, gradually changing to light orange before

hatching. Larva is six legged, almost round, yellowish at first which subsequently

change to pale orange. Nymphs are oval, eight legged and with deep reddish brown

abdomen. The adult female is somewhat elliptical; the posterior end of the abdomen is

broadly rounded and dark reddish brown in colour. The male is smaller with a narrower

abdomen tapering to the end (Fig. 3.1).

35

Fig. 3.1. Different stages of Oligonychus coffeae (red spider mite). 3.1a. Egg.

3.1b. Empty egg shells. 3.1c.Freshly hatched larva. 3.1d. Adult female

and male.

Leaf temperature and light penetration of tea bushes influence mite distribution

(Banerjee, 1979). Life cycle of red soider mite is dependent on temperature and relative

humidity (Das and Das, 1967). However, in their natural habitat other than abiotic

factors like temperature and relative humidity, the mite faces challenge from biotic

factors, such as, host plant and predators (Das et al., 2012). The optimal temperature for

growth and development is 30°C (Das and Das, 1967; Gotoh and Nagata, 2001). The

lower threshold for development is 10°C, and 232.6 degree days are required to

complete the life cycle from egg to egg (Gotoh and Nagata, 2001). The pest population

is found to be seasonally varying and dependent on the prevailing agroclimatic

conditions such as temperature and rainfall (Choudhury et al., 2006). In NE India, red

spider mite is found to be active and breed on tea throughout the year with highest

population density during late March and early April. Injury remains severe until the

monsoon rains wash off the active forms from the leaves (Das, 1959; Choudhury et al.,

♀

♂

3.1a

3.1b

3.1c 3.1d

36

2006; Mukhopadhyay and Roy, 2009). All stages of red spider mite persist on some of

the old leaves of un-pruned or skiffed tea bushes during cold weather and only a few

stay on clean pruned tea. It is the un-pruned tea fields that are primarily responsible for

the build up of red spider mite in spring (Das 1959). Though primarily a pest to tea, the

red spider mite is also known to attack jute, cotton, rubber, citrus, mango, oil palm and

many other tropical plants and weeds (Das, 1959). The mite has also been reported from

Deris robusta and Tephrosia candida in tea plantations of India (Andrews, 1928) and is

reported to attack Grevillea robusta and Albizzia falcata in Sri Lanka (Cranham, 1966).

Red spider mite of tea (O. coffeae) has gained economic importance because of crop

failure for a period of two or more months due to complete defoliation. It mainly attacks

the mature leaves, but under draught stress, tender leaves may also be attacked. Rao

(1974 a, 1974b) reported a loss of 340 – 511 kg of tea/ha due to red spider mite

infestation during draught. It causes considerable loss in tea production, the crop losses

ranging from 17 – 46% (Das, 1959, 1983; Lima et al., 1977; Mkwaila, 1983; Kilavuka,

1990).

3.2. Bio-ecology of polyphagous arthropod pests

The number of pestiferous species in agricultural crops is remarkably small in view of

the enormous pool of the potential invaders. Several biological characteristics of an

arthropod species, such as fecundity, diet breadth and voltinism, contribute to the

possibility of the species attaining pest status when a suitable habitat is made available

to it (Schoonhoven et al. 1998). Extensive bio-ecological studies enabled us to know

why among several dozen species or geographical subspecies of crucifer-feeding pierid

butterflies, only two species, Pieris brassicae and P. rapae have attained economic pest

status on crucifer crops worldwide. Among different aspects of bio-ecology, two

37

significant phenomena, host preference and post-embryonic development parameters,

which are important in the study of insect – plant interactions are discussed.

3.2.1. Host preference in polyphagous insects and mites:

Behavior is the link between physiology and ecology of animals (Bernays, 2001). Host

selection behavior of phytophagous insects is a catenary process, which is completed in

the acceptance of a plant suitable for oviposition and/or feeding (Schoonhoven, 1968).

An ability to use many plant species is an advantage. But at the same time, it is

suggested that generalist (polyphagous) insect herbivores find it relatively difficult to

choose among alternative host plants (for feeding or oviposition) due to neural

limitations giving rise to the problem of processing multiple sensory inputs (Levins and

Mac Arthur, 1969; Bernays, 2001), whereas a specialist (monophagous), who need not

to weigh among alternatives, is superior in detecting the appropriate host (Chapman et

al., 1981; Tingle et al., 1989). Studies on evolution of feeding pattern imply that

polyphagy may be ancestral to monophagy (Dethier, 1954; Bernays, 1998). In the order

Lepidoptera, the superfamilies Geometroidea and Noctuoidea have very high

proportions of generalists (Nielson and Common, 1991). In generalist lepidopterans and

other insect herbivores (except Orthopterans) different populations have relatively

restricted diets that are sometimes different in different regions such as in gypsy moth

(Fox and Morrow, 1981; Mauffette and Lechowicz, 1983). The term preference

involves a choice situation which relates a hierarchy of species of plants that can be

used as hosts (Thompson and Pellmyr, 1991). Exhibition of preference hierarchies for

host plants is common in polyphagous insects and it may be exercised by generalists if

they opt for the host species for supporting their development with varying success. In

addition to the substances of recognition (secondary plant metabolites), other conditions

provide a choice among viable hosts, such as the nitrogen content (White, 1984;

38

Bittencourt-Rodrigues and Zucoloto, 2005), the amount of attractive volatile substances

(Chew and Renwick, 1995), the plant physical characteristics (Bittencourt-Rodrigues

and Zucoloto, 2005) etc. However, changes in preference hierarchies through learning

can be favored for a variety of reasons such as variation of host quality or abundance

(Rausher, 1980; Papaj, 1986). Choice tests are simple and indispensable tool in any

insect-plant study including host preference (Schoonhoven et al., 2005). One of the

interesting aspects of insect feeding behavior is that feeding preference shows plasticity

(Ting and Hanson, 2002). In fact, it has often been reported in the entomological

literature that the food preferences of phytophagous insects can change following

feeding experience, such that the relative acceptability of plants already fed upon is

increased, i.e, induction of feeding preference occurs (Jermy et al., 1968; de Boer and

Hanson, 1984; Ting and Hanson, 2002). Most cases of such induced preference involve

minor changes; the food experienced becomes relatively more acceptable than

alternatives, although alternatives are also eaten (Szentesi and Jermy, 1990). In many

insect species, the induction of preference usually develops after one or more instars

remain on a particular host, with complex chemical stimuli participating in this process,

involving the central and peripheral nervous system (Hsiao, 1985). Host specific

phytochemical is found to modify chemoreceptors in Manduca sexta (del Campo et al.,

2001). Induction is quite common among lepidopteran larvae. Szentesi and Jermy

(1990) compiled examples of 22 lepidopterans and 12 insects belonging to other orders

where experience was found to change feeding preference. Other examples have since

been added to the list (Portillo et al., 1996; Ting and Hanson, 2002; Leal and Zucoloto,

2008). Induction of feeding preference due to experience has also been demonstrated in

mites such as two-spotted spider mite Tetranychus urticae (Gotoh et al., 1993; Egas and

Sabelis, 2001; Agrawal et al., 2002). Magowski et al. (2003) proposed associative

learning to be involved in the process of induction of feeding preference in T. urticae.

39

However, though induced preference is a fundamental type of behavioral change, it is

not universal among herbivores and could not be found in several insect species (Jermy,

1987; Sword and Chapman, 1994).

Schoonhoven et al. (2005) reviewed adaptive significance of induced feeding

preference. It has been assumed that it reflects an adaptation of insects in which

frequent changes of food type decreases the efficiency of food utilization. Induction

restricts the insect to the plant on which it is currently feeding (Ting and Hanson, 2002).

Restriction to a particular host plant following induction of feeding preference may lead

to evolution of host based biotypes in oligophagous/polyphagous insects and mites in

response to selection by the particular plant species (Mopper and Strauss, 1998; Ting

and Hanson, 2002). Formation of host-based biotypes are reported in many insects

belonging to different orders including lepidoptera and mite pests such as, peach-potato

aphid Myzus persicae (Hemiptera: Aphididae) (Saxena and Barrion, 1987), whitefly

Bemisia tabaci (Hemiptera: Aleyrodidae) (Baufeld and Unger, 1994; Cervera et al.,

2000), brown plant hopper Niliparvata lugens (Hemiptera: Delphacidae) (Saxena and

Barrion, 1987), European corn borer Ostrinia nubilalis (Lepidoptera: Pyralidae),

Walnut coddling moth Laspeyrsia pomonella (Lepidoptera: Tortricidae), red spider

mite Tetranychus urticae (Acarina: Tetranychidae) (Saxena and Barrion, 1987). The

entire panoply of evolutionary events form a spectrum, starting with ‘ecotypes’

(Thomas and Singer, 1998) and ‘biotypes’ (Eastop, 1973) at one extreme, passing

through races/strains, sub- and sibling species, and culminating in speciation itself

(Avise 1977, 1994, 2000).

40

3.2.2. Post- embryonic developmental parameters:

Environmental effects on phytophagous insects are largely exerted cumulatively

through their food (Safonkin, 2000). For successful feeding, the organism should be

capable of food assimilation and detoxification of some food components (Kondakova

and Strakhov, 1982; Rapport, 1988). So the study on the role of plants in adaptation of

phytophagous arthropod populations in a habitat is important.

Growth, development and reproduction of insects are strongly dependent on the quality

and quantity of food consumed (Scriber and Slansky, 1981). Variation in host-plant

quality may affect the body size of herbivorous insects which, in turn, can determine

life history traits, such as fecundity, longevity, and survival (Awmack and Leather,

2002; Saeed et al., 2010; Sequiera and Dixon, 1996). Post-embryonic development

period of an insect is an ideal parameter for interpreting the influence of its various

hosts. A good optimum diet results in faster postembryonic development. Previously

Naseri et al. (2009) examined life history and fecundity of Helicoverpa armigera on

different varieties of soybean. The data obtained in that study helped estimate the major

factors determining the susceptibility of soybean varieties. Farahani et al. (2011) studied

on the life table parameters of Spodoptera exigua and found that the development time

varied on different host plant. This observation is supported by the Azidah and Sofian-

Azirun (2006). Incubation period of eggs of Earias vitella varied considerably due to

host plant variation, which is shortest on Okra and longest in China rose (Syed et al.,

2011). Syed et al. (2011) also recorded considerable variation in the larval and pupal

period of E. vitella on different host plants, along with the adult longevity, fecundity

and life-cycle duration. Itoyama et al. (1999) found that the duration of the final larval

stadium of Spodoptera litura became significantly longer as diet quality decreased.

Delay in growth and development is an important symptom of feeding disturbances

41

(Levinson, 1976). Gypsy moth stands out as a study material in understanding the

complex mechanism of insect–host plant interactions in poly- and oligophagous

lepidopterans (Baranchikov, 1987). Host plant effect on gypsy moth performance and

its extremely polyphagous feeding habit have been well described. Host plant dependent

variation in larval growth and development is documented (Lazarević and Perić-

Mataruga, 2003). Larger body size is associated with higher fitness, i.e., higher

fecundity, flying and mating ability, stress tolerance, etc. Huge difference in body

weight was observed in Dectes taxanus when reared on different hosts. Higher adult

body weight could be correlated with female fecundity, male mating success and with

ability to survive adverse physical conditions (Michaud and Grant, 2005).

Polyphagous insects have the advantage that they can feed on different hosts that

provide different nutritional resources (Mozaffarian et al., 2007). The evolution of

polyphagy and its benefits have been studied in a number of insects (Sword and

Dopman, 1999; Bezerra et al., 2004). A number of studies on the biological parameters

of polyphagous lepidopteran pest of various crops, Spodoptera litura on different host

plants are done in India, Pakistan, China, Korea and other Asian countries. Larval

development of S. litura varied greatly depending on host plants, and the food

consumed by larva directly affected pupal size and weight (Xue et al., 2010a).

Parental nutrition may also affect population dynamics and trait evolution by

influencing quality of eggs. Researches on plastic responses to nutritive stress are

important for predicting insect outbreaks and understanding mechanisms of host plant

specialization. Presences of genotypic and phenotypic variations in natural populations

facilitate host-dependent specialization, host race formation and sympatric speciation in

herbivorous insects (Gorur, 2005). If host plant species constitute different selective

regimes to herbivorous insects, genetic differentiation and host plant-associated local

adaptation may occur (Ruiz-Monotoya et al., 2003). The existence of host-associated

42

populations has been examined in several insect pests (Downie et al., 2001; Abdullahi et

al., 2003; Sarafrazi et al., 2004).

3.3. Defense enzyme variability in populations of insects and

mites

Variation among populations may be caused by genetic factors, host plant and other

environmental influences (Agrawal et al., 2002). Toxic compounds ingested along with

the plant food can be hazardous, which the herbivores must overcome in order to

utilize the nutritional resources; another hazard results from exposure to pesticides used

for control of pest insects. Herbivorous species can tolerate these potentially toxic

compounds as they have evolved various physiological mechanisms to avoid their

harmful effects (Schoonhoven et al., 2005). They may either rapidly excrete the

unwanted compounds or degrade them through production of defense (=detoxification)

enzymes, or otherwise neutralize such chemicals before they can reach

pharmacologically active levels through development of target-site insensitivity

(Berenbaum et al., 1986; Brattsten, 1988a, 1988b). According to Yu (1986), enzyme

induction is a commonly occurring phenomenon representing an effective mechanism

of adaptation to external conditions. Some pesticides, especially insecticides and the

chemical constituents of host plants (plant secondary metabolites or allelochemicals), in

the case of phytophagous insects share a common metabolic detoxification process (Li

et al., 2007) and can have a great impact on inducing the enzymatic defense systems of

insects, thereby effecting insecticide resistance mechanisms (Yu, 1983, 1986; Zeng et

al., 2007). There are three major types of detoxification enzymes: 1) broad spectrum

oxidases such as mixed function oxidases or monoxygenases that include cytochrome P-

450 enzyme system, 2) hydrolases that break up esters, ethers and epoxides and 3)

conjugation systems such as glutathione S-transferase, which are mediated to cover up

43

the reactive part of the toxic chemical and further facilitate its removal. Major groups of

enzymes such as oxidases, hydrolases, transferases and reductases generally act in a

concerted way in the metabolism and conversion of foreign compounds to increasingly

polar metabolites in two arbitrary phases. Oxidation by cytochrome P450 dependent

monooxygenases and hydrolysis of ester bonds by carboxylesterases are conducted in

the phase I generating primary metabolites. Glutathione transferases are involved in

phase II conjugation reactions enabling conversion of primary metabolites into

secondary metabolites which are generally harmless.

Electrophoretic analysis of isozymes has contributed in the analysis of population

biology based on differences in geographic distribution, host plant association and

pesticide resistance status in many arthropod species (Loxdale and Hollander, 1989).

Esterase isozymes have been used in a number of insect population biology research,

for example, in Bemisia tabaci (Hemiptera: Aleyrodidae) (Guirao et al., 1997), Plutella

xylostella (Lepidoptera: Yponomeutidae) (Murai, 1993), Microtonus aethiopoides

(Hymenoptera: Braconidae) (Iline and Philips, 2003) etc. and are generally amongst the

most variable enzymes (Iline and Philips, 2003). Esterases also proved helpful in intra-

and inter-population variation studies in the mite, Tetranychus urticae (Acari:

Tetranychidae) (Goka and Takafuji, 1995a).

3.3.1. Host-based variability of defense enzymes:

Plants are suboptimal food due to inadequate nutrient ratios and the presence of

allelochemicals which the insect herbivores need to detoxify (Schoonhoven et al.,

1998). As herbivores are confronted by large amount of noxious chemicals in their plant

food, they literally are poisoned by every meal (Brattsten, 1979). Enzymatic

degradation of ingested plant toxic compounds by the herbivores is one of the

44

mechanisms to avoid their harmful effects. Herbivores, specially the polyphagous ones

have the opportunity to use several host plants as they can adapt to a heterogeneous

environment of diverse chemicals by efficient enzymatic detoxification mechanisms

(Ahmad et al., 1986). These enzymes are also known as ‘defense enzymes’ owing to the

protective role they play. Among these, variation of GST and GE in host-based

populations of arthropods, specially lepidopterans and mites is reviewed hereafter.

Variation of glutathione S- transferase (GST) and general esterase (GE)

enzymes in host-based populations of insects and mites:

Induction of GST by host plants has been reported by many authors. Yu (1982) reported

that in larval stages of fall armyworm, Spodoptera frugiperda reared on host plants,

such as, cowpeas, turnip and mustard, the activity of midgut GST was 7 – 10 fold higher

than the larvae of the same species reared on soybean, sorghum, millet, cucumber,

potato etc. In another polyphagous lepidopteran, Platynota idaeusalis, the tufted apple

bud moth, host plant affected activities of the detoxifying enzymes, glutathione

transferase and esterase (Dominguez-Gil and McPheron, 2000). Host plant mediated

variation in the activity of mid gut detoxification enzymes was also observed in larvae

of the eastern tiger swallowtail, Papilio glaucus glaucus when reared on leaves of black

cherry, tulip, paper birch, white ash or basswood with highest activity of GST and GE

activity on tulip and lowest on basswood leaves (Lindroth, 1989a). The same author

(1989b) suggested alteration in biochemical detoxification systems in evolutionary and

ecological adaptation of polyphagous luna moth, Actius luna to different food plants.

GST was found to be the key detoxification enzyme in metabolizing the chemical

components of sesame leaves in the larvae of S. litura as evident from a 6-fold increase

in GST level in the larvae fed with sesame leaves than those fed on an artificial diet

(Sintim et al. 2009). The success of polyphagous aphid pest, Myzus persicae to

different host plants has been related to the presence of enzymatic mechanisms of

45

detoxification responsible for the metabolisation of host-plant allelochemicals (Francis

et al. 2005; 2006). In a recent work by Cabrera-Brandt et al. (2010) a subspecies of

aphid, M. persicae nicotianae exhibited higher total esterase activities when reared on

tobacco than on pepper which suggested a participation of esterases on the ability of M.

persicae nicotianae to overcome the tobacco defenses. Mulin and Croft (1983) reported

host-related alterations of detoxification enzymes in the two spotted spider mite

Tetranychus urticae where these enzymes were stimulated by host plants such as carrot

and celery. Host based variation in the activity of esterase and GST in T. urticae is also

recorded by Yang et al. (2001) when they reared the species on lima bean, maize and

cucumber. Duration of host association is also found to influence the activities of these

enzymes in the mite species.

Pioneering research by Yu and co-workers could reveal specific role of an array of

allelochemicals in the induction of detoxification enzymes. The monoterpens (+)-α-

pinene, (-)-menthol and peppermint oil and the sesquiterpene lactone santonin were all

moderate inducers of the esterase, causing increases of 35 to 65 percent in activity in

fall armyworm larvae. The plant hormone analogs, (indole-3-acetonitrile and indole-3-

carbinol), as well as the flavonoids, (flavones and β-naphthoflavone), alkaloid (quinine),

along with furanocoumarin (xanthotoxin) stimulated the esterase, resulting in 35 to

114% increases of the enzyme (Yu and Hsu, 1985). Similar to this finding, plants such

as celery, potato and parsley were found active in inducing the esterase in the fall

armyworm larvae whereas corn, peanuts, cotton, soybeans, cowpeas, carrot, sweet

potato, peppermint, radish, turnip had no significant effect, thus reflecting the role of

host plant allelochemicals in inducing the hydrolase activity of the phytophag (Yu and

Hsu, 1985). Glutathione transferases also could metabolize toxic allelochemicals,

including α,β-unsaturated carbonyl compounds, isothiocyanates such as trans, trans-2,4-

decadienal, trans, trans-cinnamaldehyde, benzaldehyde, trans-2-hexenal, allyl

46

isothiocyanate, benzyl isothiocyanate and organothiocyanate such as benzyl

thiocyanate in lepidopterous insects (Wadleigh and Yu, 1987; 1988a; 1988b). The

glucosinolate, sinigrin, and the hydrolytic products of glucosinolates, β-

phenylethylisothiocyanate, indole 3-acetonitrile and indole 3-carbinol and flavones were

found to be potent inducers of the glutathione S-transferase in the armyworm (Yu,

1983). In addition, dietary coumarin and monoterpenes (α-pinene, β-pinene, limonene,

terpinene) induced GST in southern armyworm, Spodoptera eridania larvae (Brattsten

et al., 1984). However, monoterpenes were not inducers of transferase in fall armyworm

larvae (Yu 1982). Coumestrol, a coumerin analog found in a resistant soybean cultivar,

induced GST in soybean loopers (Rose et al., 1989). 2-Tridecanone found in wild

tomato leaves induced GST in tobacco budworm larvae (Riskallah et al., 1986).

Hemming and Lindroth (2000) studied the effects of phenolic glycosides on

detoxification activities of gypsy moth (Lymantria dispar) and forest tent caterpillar

(Malacosoma disstria) larvae. Esterase activities were induced by phenolic glycosides

only in gypsy moths, whereas GST activities were induced in both species. Ability to

detoxify phenolic glycosides enables these lepidopterans to adapt to such diet which is

otherwise toxic and unacceptable to folivores.

Apart from quantitative differences, electrophoretic variations in isoenzyme patterns

have also been documented in relation to host based populations of a particular species

of insects. Agarwala et al., (2002) could identify three host plant related clones of the

aphid, Lipaphis erysimi on the basis of variation in the banding pattern of esterase and

malic dehydrogenase. Electrophoretic pattern of esterase isozymes is used to study

variation in host- based populations of Bemisia tabaci (Wool et al., 1993; Helmi,

2010). Effect of host plants on the profile of detoxification enzymes of Helopeltis

theivora, a major sucking pest of tea could be documented through electrophoretic

analysis (Saha et al., 2012c).

47

3.3.2. Defense enzymes in pesticide-tolerant insect and mite

populations:

Applications of pesticides to control arthropod pests and vectors create selection

pressure on them leading to development of pesticide-tolerant and then pesticide-

resistant populations. Gene flow is an essential factor in spreading advantageous genes

such as pesticide-resistant genes in insect pests. Nearly 40 years of studies, all over the

world, suggest that, insecticide resistance could be correlated with quantitative and/or

qualitative changes in insecticide metabolizing enzymes. Efforts have been made to

estimate the activity of resistance associated enzyme with help of surrogate substrate,

whose products can be measured either in solution or cellulose filter paper or on

nitrocellulose membranes. Furthermore, these surrogate substrates can be used to probe

the isozymes and their mobility variance following electrophoresis and

electrofocussing. As stated earlier, there are three major types of detoxification

enzymes: 1) broad spectrum oxidases such as mixed function oxidases (MFOs) or

monoxygenases that include cytochrome P-450 (CYP450) enzyme system, 2)

hydrolases such as general esterases (GE) and 3) conjugation systems such as

glutathione S-transferases (GST). These three types of detoxification enzymes have

been documented to play a role selectively or in conjunction towards development of

resistance against different classes of insecticides in insects and mites, many of which

are of major pest status.

CYP450 dependent monoxygenases:

The CYP450 dependent monoxygenases are ubiquitous enzymes involved in

endogenous metabolism as well as metabolism of xenobiotics through oxidation

in phase I. CYP450 monoxygenase activities can be involved in the metabolism of

virtually all insecticides leading to an activation of the molecule or, more

48

generally to a detoxification and as a result may impart resistance to insecticides

(Feyerisen, 1999; Chen et al., 2005).

The role of GEs and GSTs as defense enzymes, the estimation of which has

mainly been done in the present thesis, are reviewed in details hereafter.

General esterases (GE) in insects:

GE are one the most significant enzymes in insects causing insecticide

detoxification in phase I metabolism. These defense enzyme groups have

repeatedly been implicated in metabolic resistance to the major organophosphate

(OP) and synthetic pyrethroids (SP) (Wheelock et al., 2005). OP and SP contain

carboxylester bonds that are subject to attack by esterase enzymes. Insect

carboxylesterases from the α-Esterase gene cluster is found to play an important

role in detoxification of OP insecticides (Jackson et al., 2013). Differences in

the amount of esterase activity between two strains of the same insect species is

considered as an indicator of the degree of sensitivity to certain insecticides,

subsequently, various biochemical assays have been used for insect populations

as possible indicators of insecticide resistance (Brown and Brogdon, 1987).

Several isozyme form of the esterases exist which can be detected by gel

electrophoresis. Besides higher production, change in electrophoretic banding

pattern such as occurrence of extra bands or bands with higher staining intensity

has been associated with resistance (Lalah et al., 1995; Neus et al., 2008). Role

of the GEs in imparting pesticide resistance in insects is summarized in table

3.1.

49

GST in insects:

GST is a family of multifunctional isozymes found in all eukaryotes. One of the

main functions of GST is to catalyze xenobiotics, including pesticides in the

marcapturic acid pathway leading to the elimination of toxic compounds (Hayes

and Pulford, 1995). In insects, this family of enzyme has been implicated as one

of the major factors in neutralizing the toxic effects of insecticides in phase II of

the metabolism process (Clark et al., 1986, Grant et al., 1991, Salinas and Wong,

1999). The majority of studies on insect GSTs have focused on their role in

detoxifying foreign compounds, in particular insecticides and plant

allelochemicals and more recently, their role in mediating oxidative stress

responses (Clark et al., 1986; Wang et al., 1991; Ranson et al., 2001; Vontas et

al., 2001; Sawicki et al., 2003). In insects, GST isozymes are present in three to

four forms in house flies (Clark and Dauterman 1982). Enhanced activities of

GSTs that confer insecticide resistance result from both quantitative and

qualitative alterations in gene expression (Chien et al., 1995; Wei et al., 2001).

There is evidence for over-expression of one or more GST isoforms in resistant

insects (Grant et al., 1991; Fournier et al., 1992; Marcombe et al., 2012). Role of

the GSTs in imparting pesticide resistance in insects is summarized in table 3.1.

50

Table 3.1. Insect pests showing enzyme-based (GE & GST) metabolic

resistance to insecticides (GE; general esterases and GST; glutathione S-

transferases)

Species of insect Resistant to Defense enzyme

imparting metabolic

resistance

Reference

Helicoverpa

armigera

SP GE

GST

Gunning et al., 1996;

Kranthi et al., 1997;

Achaleke et al., 2009; Wu

et al., 2011.

Omer et al., 2009

Plutella xylostella Malathione &

Phenthoate

SP

GE

GST

Chiang and Sun, 1993;

Maa and Liao, 2000

Dukare et al., 2009

Frankliniella

occidentalis

Methiocarb &

Acrinathin

GE Neus et al., 2008

Culex spp. OP GE Poirie et al., 1992;

Callaghan et al., 1994;

Jayawardena et al., 1994

Anopheles spp. Malathione

---

---

GE

GST

GST

Hemingway, 1982, 1983;

Herath and Davidson,

1981; Perera et al., 2008.

Ranson et al., 2001

Ganesh et al., 2003

Aedes aegypti OP, SP

---

GE

GST

Marcrombe et al., 2012

Grant et al., 1991;

Marcrombe et al., 2012

Musca domestica OP, SP

OP, SP, OC

GE

GST

Soderland and

Bloomquist, 1990; Funaki

et al., 1994

Oppenoorth et al.,1979;

Clark and Dauterman,

1982; Clark and Shaaman,

1984; Clark et al., 1986;

Fournier et al., 1992;

Chien et al., 1995; Wei et

al., 2001

51

Blattela germanica OP GE Valles, 1998

Myzus persicae OP, SP,

Carbamate

GE Davonshire and Field,

1991; 1995

Schizaphis

graminum

OP GE Ono et al., 1994; Zhu and

Gao, 1998; Zhu and He,

2000

Nilaparvata lugens SP GST Vontas et al., 2001

Oryzaephilus

surinamensis

Chlorpyriphos

---

GE

GST

Lee and Lees, 2001

Al-Dhaheri and Al-Deeb,

2012

Leptinotarsa

decemlineata

SP GE Argentine et al., 1995

Pediculus capitis Malathione GE Gao et al., 2005

Sitophilus oryzae OP, SP GE Iqbal et al., 2012 – 13

Bactocera

cucurbitae

OP, SP GE Rashid et al., 2012 – 13

Esterases and GSTs in mites:

Acaricides have been widely used for mite control in glasshouses, orchards and many

other cropping systems (Van Leeuwen et al., 2006). Frequent application of acaricides

to maintain mite populations below economic thresholds, as mites have a high

reproductive potential and extremely short life cycle. Such operations facilitate the

development of acaricide-resistance in them (Stumpf et al., 2001). Defense

(=detoxification) mechanisms to acaricides are also often attributed to enhanced activity

of defense enzymes, Esterases and GST. Higher Esterase and GST activities were

positively related with acaricide (OP and SP) resistance in two spotted spider mite, T.

urticae (Yang et al., 2001). Electrophoretic variations in Esterase-zymogram were

evident between abamectin resistant and susceptible T. urticae indicating possible

involvement of esterase-3 band in development of abamectin tolerance (Yorulmaz and

Ay, 2009). GSTs have been associated with macrocyclic lactone resistance in mites,

with elevated GST activity observed in abamectin resistant T. urticae (Konaz and

52

Nauen, 2004; Stumph and Nauen, 2002). Increased esterase and GST activity in

permethrin resistant Sarcoptes scabiei mite is reported (Pasay et al., 2009), of which,

GST appeared to be the most significant. Increased GST activity is associated with

permethrin tolerance in S. scabiei, GST inhibitors could restore susceptibility of the

mite species to permethrin and subsequently increased transcription of GST in

permethrin resistant S. scabiei was evident (Mounsey et al., 2010). Quantitative and

qualitative differences in esterase isozymes were recorded between pesticide-exposed

(tolerant) and unexposed (susceptible) populations of tea red spider mite, Oligonychus

coffeae from tea plantations of Terai region of West Bengal and South India (Sarker and

Mukhopadhyay, 2006b; Roobakkumar et al., 2012).

3.3.3. Host plants, defense enzymes and pesticide resistance: a

complex interrelation:

Host plants can modify the susceptibility of herbivorous arthropods to pesticides

(Brattsten, 1988a). A list some insect and mite crop pests that show host plant

dependent variation in the development of tolerance to pesticides is provided in Table

3.2. Physiological response of herbivores to host plants may lead to enhanced

metabolism of pesticides because underlying mechanisms that function in detoxification

of plant allelochemicals in their diets may also be effective in detoxifying pesticides

(Yang et al., 2001). Same set of defense enzymes, general esterases, GST and CYP450

dependent monooxygenase are involved in the metabolism of plant secondary chemicals

as well as pesticides in insect and mite systems. General esterases, which are capable of

degrading or sequestrating pesticides, can play a significant role in the detoxification of

OP and pyrethroid pesticides. Research works have demonstrated positive relation

between host-plant induced changes in defense enzyme level and susceptibility to

pesticides. Increased esterase level vis-á-vis tolerance in mite species T. urticae to

53

pesticide when using certain host plant species has been documented by Yang et al.

(2001). In S. litura higher esterase and GST activity on tobacco is correlated with higher

tolerance to several pesticides (Xue et al., 2010b). However, in tufted apple bud moth,

P. idaeusalis, though host plants could differentially induce detoxification enzymes,

patterns of enzyme activity and susceptibility to the pesticide azinphosmethyl could not

be clearly linked, inkling the complex relationship of the insect with the chemistry of its

host (Dominguez-Gil and McPheron, 2000).

Table 3.2. Changed pesticide susceptibility in arthropod-pests due to influence of

host plants

Species Reference

Aphis gossypii (Hemiptera) Furk et al., 1980; Godfray and Fuson,

2001

Myzus persicae (Hemiptera) Ambrose and Reghupathy, 1992

Nilaparvata lugens (Hemiptera) Heinrichs et al., 1984

Sogatella furcifera (Hemiptera) Heinrichs et al., 1984

Bemisia tabaci (Hemiptera) Castle et al., 2009; Liang et al., 2007; Xie

et al., 2010

Trialeurodes vaporariorum (Hemiptera) Liang et al., 2007

Leptinotarsa decemlineata (Coleoptera) Ghidiu et al., 1990; Mahdavi et al., 1991

Peridroma saucia (Lepidoptera) Berry et al., 1980

Spodoptera frugiperda (Lepidoptera) Wood et al., 1981

Heliothis armigera (Lepidoptera) Loganathan and Gopalan, 1985

Helicoverpa assulta (Lepidoptera) Wang et al., 2010

Epiphyas postvittana (Lepidoptera) Robertson et al., 1990

Spodoptera litura (Lepidoptera) Xue et al., 2010b

Tetranychus urticae (Acarina) Neiswander et al., 1950; Henneberry 1962;

Kady 1965; Gould et al., 1982; Yang et

al., 2001; Dermauw et al., 2012

54

3.4. Defense enzyme variability in insect and mite pests of tea in

North East India

In sub-Himalayan tea plantations of North-East India, most plantations are managed

conventionally by routine application of different organo-synthetic insecticides to

control pest populations. Organochlorines (OC), organophosphates (OP), synthetic

pyrethroids (SP) and neonicotinoids (NN)) are routinely applied round the year to keep

the insect pest populations under control (Sannigrahi and Talukdar, 2003;

Gurusubramanian et al., 2008). Repeated application of pesticides can result in the

resurgence of primary pests (Sivapalan, 1999), outbreak of secondary pest (Cranham,

1966) and development of resistance (Kawai, 1997; Roy et al., 2010a, 2010b). Many of

the tea pests being polyphagous feed on a number of plants besides tea, which indicate

that they can overcome challenge posed by a wide array of phytochemicals successfully.

There are evidences of physiological adaptation of some tea pests to both insecticides

and host plant chemicals involves induction of defense enzymes, such as, Esterases,

GST and Cytochrome P450 dependent monooxygenases (Saha et al. 2012a, 2012b,

2012c, Saha and Mukhopadhyay, 2013).

In recent days sucking insect pest, Helopeltis theivora has developed high tolerance to

some commonly used insecticides leading to control failures in sub Himalayan tea

plantations of West Bengal (Mukhopadhyay and Roy, 2009; Roy et al., 2011). Higher

levels of detoxifying enzyme (general esterases, GST and cytochrome P450

monooxygenase) activities in tea pests and their bearing on the level of susceptibility to

different synthetic insecticides have been documented (Sarker and Mukhopadhyay,

2006a; Saha et al., 2012a; Saha and Mukhopadhyay, 2013) from different sub-

Himalayan tea plantations of North-East India. There are reports of repeated control

failure of the sucking pests, E. flavescens and S.dorsalis in sub-Himalayan tea

55

plantations of North-East India. A positive relation to this effect has been established

between the population of the sucking pests showing high pesticide tolerance (LC50

values) and their titer of defense enzymes (Saha et al., 2012a; 2012b). Presence of

higher quantities of general esterases was reported in pesticide-exposed populations of

red slug caterpillar, Eterusia magnifica (Sarker and Mukhopadhyay, 2006a) and also in

tea red spider mite, Oligonychus coffeae (Sarker and Mukhopadhyay, 2006b).

Further, preliminary observations on difference in electrophoretic banding pattern of

general esterases in pesticide-exposed (field) population and pesticide-unexposed

(laboratory) population of the tea pests are available for red slug caterpillar, Eterusia

magnifica (Sarker and Mukhopadhyay, 2006a), tea mosquito bug, Helopeltis theivora

(Sarker and Mukhopadhyay, 2003) and tea red spider mite, Oligonychus coffeae (Sarker

and Mukhopadhyay, 2006b).

Host-based variation in H. theivora in terms of differential activity of three principal

xenobiotic detoxifying enzymes, the general esterases (GEs), glutathione S-transferases

(GSTs) and cytochrome P450 monooxygenases (CYPs) is documented (Saha et al.,

2012c). Further a host based differential activity of defense enzymes, esterases and GST

in tea looper pest H. talaca has also been observed (Das and Mukhopadhyay, 2008).

There are preliminary reports on the development of insecticide tolerance evident by

high LC values in H. talaca and H. infixaria (Nair et al., 2008b; Das, S. et al., 2010) and

in red spider mite, O. coffeae as well (Roy et al., 2008a; Roy et al., 2010a).

Literature on variability in defense enzymes in lepidopteran and mite pests of tea related

to their availability on different host plants or exposure to varying pesticides remains

scanty.