- Top of page
- 1 INTRODUCTION
- 2 MATERIALS AND METHODS
- 3 RESULTS
- 4 DISCUSSION
- 5 CONCLUSIONS
Plants have developed an elegant defence system against insect herbivory. The defence systems employed by plants against insects can be constitutive or induced. Constitutive resistance is present in plants all the time, whereas induced resistance occurs in response to various stimuli such as insect herbivory, pathogen infection and/or elicitor application.[1-3] Induced resistance is very important as it makes plants phenotypically plastic, thereby making it freakish for the insect pests to feed on it.[4, 5] Induced resistance can be direct or indirect. Direct induced resistance directly affects the insect pest through antixenosis and/or antibiosis mechanisms,[6, 7] whereas indirect induced resistance is mediated through volatiles emitted by the plants in response to insect damage, which attract the natural enemies (parasitoids and predators) of the insect pests.[4, 8, 9]
Although many plant hormones act as elicitors of induced resistance, the most important and widely used phytohormones are jasmonic acid (JA) and salicylic acid (SA).[3, 10] The use of these phytohormones in inducing plant resistance against insect pests has raised the possibility of their implications for insect pest management. Exogenous application of JA results in the induction of plant responses that are almost similar to herbivore feeding. The JA-mediated octadecanoid pathway leads to the production of many defensive components, such as plant defensive proteins, oxidative enzymes, glandular trichomes, flavonoids, terpenoids, alkaloids, volatile compounds, etc.[1, 4, 9] SA, a benzoic acid derivative, is an endogenous plant growth regulator that generates in plants a wide range of metabolic and physiological responses involved in plant growth and development, and defence against various stresses, including insect herbivory.[3, 10, 12]
Groundnut (Arachis hypogaea L.) is an annual herbaceous plant belonging to the family Fabaceae. It is cultivated mostly in semi-arid tropical and subtropical regions. It is damaged by several insect pests, of which the legume pod borer, Helicoverpa armigera (Hübner), is an important defoliator during the vegetative stage. H. armigera is widely distributed in Asia, Africa, southern Europe and Australia. In semi-arid tropics, H. armigera causes an estimated loss of over $US 2 billion annually, in spite of the $US 500 million worth of pesticides applied for controlling this pest. It has developed high levels of resistance to several commonly used insecticides. Therefore, there is a need for alternative methods of pest control to reduce overdependence on insecticides and to conserve biodiversity. It is in this context that host plant resistance, which is economic and environmental friendly, assumes a central role in integrated pest management.
Host plant resistance plays an important role in groundnut defence against a variety of insect pests. Many biochemical parameters have been associated with resistance in groundnut against insect pests. Higher levels of antioxidative enzymes, phenols and tannins contribute to groundnut resistance against Spodoptera litura (Fab.) and H. armigera.[15-18] Stevenson et al. observed that quercetin, caffeoylquinic acids and diglycosides contribute to resistance in groundnut against S. litura. Procyanidin in groundnut plants provides resistance against Aphis craccivora (Koch).[16, 19] Nitrogen, soluble sugars and polyphenols are involved in groundnut resistance against leaf miner Aproraema modicella Dev. Understanding the mechanisms of induced resistance can help in building up the natural defences in plants by the application of elicitors and/or mild damage by the herbivores. Although it has been well documented that phytohormones induce plant resistance in plants through the expression of a number of proteins and non-protein-based compounds, such studies are limited in groundnut. To test this hypothesis, JA and SA were exogenously applied to groundnut plants with differential levels of resistance to H. armigera to study the induced resistance. The plants were pre- and/or simultaneously treated with JA and SA and infested with H. armigera. Various plant defensive enzymes and plant secondary metabolites were investigated.
- Top of page
- 1 INTRODUCTION
- 2 MATERIALS AND METHODS
- 3 RESULTS
- 4 DISCUSSION
- 5 CONCLUSIONS
Although several phytohormones are involved in host plant defence against biotic and abiotic stresses, JA and SA play an important role in modulating plant defence against insect herbivory.[1, 3-5, 12] The JA- and SA-mediated induced resistance operates through octadecanoid and phenylpropanoid pathways, respectively, resulting in increased production of secondary metabolites and plant volatiles.[4, 37] JA also regulates the activity of calcium-dependent protein kinases involved in plant defence against a variety of biotic and abiotic stresses through signal transduction. JA accumulates in plants in response to insect damage and also by exogenous application. During this process, several secondary metabolites and volatiles are produced. Further, JA also activates antioxidative enzymes, such as POD, PPO and LOX, and the production of PIs. SA regulates reactive oxygen species (ROS) metabolism in plants and the oxidation of certain substrates of POD, CAT, SOD and other antioxidative enzymes, thus altering the hormonal balance and cell-wall lignifications.[3, 10-12] Increase in host plant resistance to herbivores has been observed through exogenous application of JA or MeJA[4, 37] and SA.[10, 12] Elucidation of various defensive responses in plants by exogenous application of JA and SA is essential for gaining an understanding of the induced plant resistance against insect pests that is mediated by these hormones and the implications for insect pest management.
The present results showed that plants pretreated with JA had greater activity of defensive enzymes such as POD and PPO than the plants pretreated with SA. Increase in POD activity is regarded as the initial response of plants to insect attack.[5, 8] Increased activities of these enzymes in response to JA might be due to the greater accumulation of JA after insect infestation, and the subsequent activation of plant defensive pathways, resulting in increased activity of defensive enzymes such as POD and PPO. Higher levels of POD activity enhance cell lignification, wound healing and production of secondary metabolites, besides detoxifying the peroxides, thus defending the plants against insects, pathogens and other stresses.[8, 39, 40] The reduced nutritional quality of plant tissues on account of PPO has also been reported to play an important role in plant defence against insect herbivory.[10, 41, 42] Moreover, toxic but highly reactive quinines produced from phenol oxidation interact with the nucleophilic side chain of amino acids and crosslink the proteins in plant tissues, thus reducing their digestibility.
PAL activity is induced by various stresses, including insect herbivory. PAL activity was greater in groundnut plants pretreated with JA and SA and in plants simultaneously treated with JA compared with insect-infested and uninfested control plants. The increase in PAL activity by JA and SA can be attributed to their identical effect on the activation of defensive pathways in response to damage by H. armigera. These pathways produce various plant secondary metabolites, which on oxidation form several defensive compounds. In addition, the phenylpropanoid pathway, of which PAL is a central enzyme, also leads to lignin synthesis. Lipoxygenase gene expression is regulated by JA and different biotic/abiotic stresses, including insect herbivory. LOX catalyses the production of JA from linolenic acid in the octadecanoid pathway. It also elicits the production of various plant defensive secondary metabolites and plant volatiles. The present study revealed that PJA + HIN-treated and JA + HIN-treated plants had significantly greater levels of LOX activity than the rest of the treatments. This increased LOX activity in plants pre- and/or simultaneously treated with JA might be due to signalling of the octadecanoid pathway by exogenous application of JA. Oxylipins produced from fatty-acid oxidation by LOX play a wide array of functions in plant growth and development, senescence and defence against biotic and abiotic stresses, including insect herbivory. Compounds formed from LOX-mediated reactions are either directly deterrent to insect pests and/or produce post-ingestive toxicity in Insects.
The antioxidative enzymes involved in plant oxidative stress due to biotic and abiotic factors are SOD, APX and CAT. The present study revealed greater increase in APX activity in plants pretreated with JA and SA, and JA + HIN. Insect-resistant genotypes exhibited significantly greater APX activity than the susceptible check JL 24. Pretreatment with JA followed by insect infestation and simultaneous application of JA and insect infestation resulted in greater increase in CAT and SOD activities across the genotypes. Pre- and/or simultaneous treatment with SA also increased the activities of these enzymes; however, induction was lower compared with that of JA. Insect-resistant genotypes showed greater increase in the activities of antioxidative enzymes compared with the susceptible check JL 24, but the levels of induction varied. The differential responses across the genotypes might be due to the differential ability of groundnut genotypes to perceive insect damage and/or the ability to mount a defensive response. Greater increase in SOD, APX and CAT following JA or SA treatment could be due to signalling of transduction pathways modulated by these phytohormones, which leads to the production of antioxidative enzymes to scavenge the toxic-free radicals produced by herbivory. The higher constitutive levels of these enzymes in insect-resistant genotypes might protect them from initial oxidative damage before the induced defence system is activated. APX decreases the ascorbate content in plant tissues by utilising ascorbic acid as the electron donor in ascorbate-glutathione recycling while catalysing the reduction of H2O2 to water, which in turn reduces insect growth and development. Greater APX activity in soybean leaves removes ascorbate from the H. zea larval midgut, thereby reducing insect growth and development. Scirpophaga incertulas (Walk.) and Cnaphalocrosis medinalis (Guenee) damage induces higher levels of CAT in rice. CAT resists the oxidative stress in soybean caused by H. zea infestation. SOD converts the toxic-free radicals, especially of oxygen, into less toxic and relatively stable H2O2. Induction of SOD activity by SA has been found to reduce plant oxidative damage in maize. H. zea infestation increases SOD activity in tomato and soybean.
Plants produce many non-enzymatic defensive proteins against insect pests. However, PIs are the most exploited plant defensive proteins that confer resistance to insect pests. The in vitro PI activity of groundnut plants pre- and/or simultaneously treated with JA and infested with H. armigera was significantly greater than that of uninfested control plants. Overall, insect-resistant genotypes showed greater PI activity than JL 24 in almost all the treatments. The reduction in protein digestibility by PIs and deprivation of insects of essential amino acids lead to retarded growth and development of Insects. PIs are strongly upregulated in plants in response to wounding or herbivore damage and/or elicitor application. For example, exogenous application of MeJA in Nicotina attenuata Torr. ex S. Watson results in quick accumulation of JA and the induction of trypsin proteinase inhibitors against M. sexta.
Phenols constitute one of the most important and extensively studied groups of secondary metabolites against insect pests.[7, 17, 48] An abrupt increase in phenolic content occurs in plants damaged by insects and/or treated with elicitors, including JA and SA.[21, 22] PJA + HIN-treated, PSA + HIN-treated and HIN-treated plants exhibited greater phenolic content than the SA + HIN-treated and untreated plants; however, some genotypes, such as ICG 2271, ICG 1697 and JL 24, responded similarly to pre- and/or simultaneous treatments of JA and SA. Further, insect-resistant genotypes showed a greater increase compared with the susceptible check JL 24. This might be due to the strong induction of the octadecanoid and phenylpropanoid signalling pathways by JA and SA, respectively. Flavonoids have been reported to confer resistance against Spodoptera frugiperda (J.E. Smith) in Arabidopsis thaliana (L.). Higher levels of flavonoids, such as daidezin and genistin, have been observed in soybean plants infested with Nezara viridula (L.). Tannins have been reported to be systemically induced in insect-damaged plants. In N. attenuata, application of MeJA induced greater accumulation of JA, which in turn activated the production of phenols, flavonoids, nicotine and trypsin proteinase inhibitors against M. sexta.
The oxidative state of the host plants is associated with plant resistance to insects,[5, 10] which results in the production of ROS, which are toxic to herbivores. The present results showed that both JA and SA induced higher levels of H2O2 in all the genotypes infested with H. armigera. However, the induction was greater in plants pretreated with JA and SA and in plants simultaneously treated with JA and infested with H. armigera. Insect-resistant genotypes showed a strong response in terms of accumulation of H2O2. The higher induction of H2O2 by pretreatment with JA and SA could be attributed to the increased activity of antioxidative enzymes in the treated plants, and conversion of toxic-free radicals into H2O2. JA and SA induce oxidative burst in plants,[10-12] which happens to be the first and foremost defence against insect herbivory.[5, 8, 17, 48] Transduction pathways signalled by H2O2 produce many defensive compounds, which results in the oxidation of phenols and other compounds producing many defensive compounds. Oxidative damage in the midgut of insects feeding on pre-wounded plants is due to the accumulation of H2O2 through JA- and SA-mediated pathways.[12, 57]
Malondialdehyde is an important lipid peroxidation product that indicates the extent of plant defensive response to stress. The plants infested with H. armigera and pre- and/or simultaneously treated with SA had a higher MDA content. Overall, JL 24 showed higher amounts of MDA among all the genotypes. This could be due to greater stress experienced by this genotype and the higher levels of lipid peroxidation. Lipid peroxidation and hydroxyl ion formation (OH·−) have been proposed to play an important role in plant defence by increasing the activity of oxidative enzymes. MDA is also involved in volatile emission, and thus has a role in indirect plant defence as well. Hao et al. reported higher amounts of MDA in rice plants in response to rice stripe virus and small brown planthopper, Nilaparvata lugens (Stål.). Induction of proteins and their role in induced resistance against insect pests have been well established.[5, 41, 48] The present studies indicated that there was a significant increase in proteins in plants treated with PJA + HIN, followed by JA + HIN-treated plants. Increase in protein concentration may be due to the increase in antioxidative enzymes and other non-enzymatic defensive proteins. Defence-related enzymes and other protein-based defensive compounds accumulate in plants in response to oxidative stress[39, 41] and on the application of elicitors,[4, 21, 22, 37] which defend them from various biotic and abiotic stresses.
Expression of resistance to insects and insect growth and development are closely related. The PJA + HIN-treated plants suffered lower damage due to H. armigera across genotypes. The insect-resistant genotypes showed greater reduction in plant damage than the susceptible check JL 24. Similar results were observed in terms of larval survival and larval weights of H. armigera. Reduced damage and lower larval survival and larval weights might be due to the greater production of toxic secondary metabolites in the insect-resistant genotypes by insect damage and JA application.[41, 42, 44, 45] Reduced damage and lower larval growth and development were correlated with increased activity of POD, PPO and other defensive enzymes induced following insect attack and/or elicitor application. Larvae of Manduca sexta (L.) and Spodoptera exigua (Hüb.) fed on JA-deficient mutant (def1) tomato plants exhibited higher survival and weight gain compared with those fed on wild-type tomato.[60, 61] Increased levels of POD, PPO and LOX in plants have been correlated with reduction in insect growth and development.[39, 42, 52] Plant defensive compounds induced in insect-resistant genotypes reduced the survival and development of S. frugiperda larvae. Reduced larval weights due to antibiosis and antixenosis against H. armigera have also been observed in chickpea.