Impact of Russet Mite on the Induced Defences of Tomato Against Spider Mites
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Università degli Studi di Napoli “Federico II” MABROUK BOUNEB European PhD “Insect Science and Biotechnology” Course XXVI Tri-Trophic Interactions: Impact of Russet Mite on the Induced Defences of Tomato against Spider Mites Supervisors Candidate Dr. Sauro SIMONI Mabrouk BOUNEB Prof. Francesco PENNACCHIO Academic year 2013-2014 _______________________________________________________________________ ِ ِ ِ ِ َِّ ِ ِ ﴿ُق ْل إ َّن َصََلتي َوُن ُسكي َوَم ْحَيا َي َوَم َماتي لله َر ِّب اْل َعالَمي َن﴾ [ سورة اﻷنعام اﻵية: 261] _______________________________________________________________________ Index Summary _________________________________________________ 1 Riassunto _________________________________________________ 3 I. Introduction ____________________________________________ 5 1.1. Plant defences against herbivores ........................................................................................ 5 1.1.1. Direct defence ............................................................................................................................. 5 1.1.1.1. Anatomical defence ............................................................................................................. 5 1.1.1.2. Production of secondary metabolites .................................................................................. 6 1.1.1.3. Digestibility reducers ......................................................................................................... 11 1.1.1.4. Antinutritive enzymes ........................................................................................................ 11 1.1.2. Indirect defence ......................................................................................................................... 12 1.1.3. Plant responses mediating signal transduction pathways ........................................................ 14 1.1.3.1. The octadecanoid pathway ................................................................................................ 14 1.1.3.2. Cross-talk between signal transduction pathways ............................................................ 16 1.1.4. Plant responses to multiple herbivory ...................................................................................... 18 1.2. Herbivore offences ............................................................................................................. 18 1.2.1. Feeding and oviposition choices ................................................................................................ 19 1.2.2. Suppression of defence signaling .............................................................................................. 20 1.2.3. Detoxification ............................................................................................................................ 20 II. Study system _____________________________________________ 22 2.1. The host plant ..................................................................................................................... 22 2.1.1. Overview .................................................................................................................................... 22 2.1.2. Tomato defences against herbivores ........................................................................................ 23 2.2. Herbivores ........................................................................................................................... 27 2.2.1. Spider mite ................................................................................................................................ 27 2.2.2. Tomato russet mite ................................................................................................................... 29 2.3. Predatory mites .................................................................................................................. 31 2.3.1. Phytoseiulus persimilis Athias-Henriot ...................................................................................... 31 2.3.2. Neoseiulus californicus (McGregor) .......................................................................................... 32 III. Research objectives ______________________________________ 34 IV. Material and methods ______________________________________ 35 4.1.Material ............................................................................................................................... 35 4.1.1. Plant material ............................................................................................................................ 35 4.1.2. Herbivores ................................................................................................................................. 35 4.1.3. Predatory mites ......................................................................................................................... 35 4.2. Methods .............................................................................................................................. 36 4.2.1. Evaluation of microscopic damages caused by spider mite and russet mite ............................ 36 4.2.2. Effects of russet mites and SA treatment on the feeding choice of spider mites ..................... 36 4.2.3. Effects of russet mites and SA treatment on the performance of spider mites ........................ 37 4.2.4. Olfactory choice of predatory mites .......................................................................................... 37 4.2.4.1. Y-tube olfactometer set-up ............................................................................................... 37 _______________________________________________________________________ 4.2.4.2. Mites infestations and olfactometer experiments ............................................................ 38 4.2.5. Gene expression analysis ........................................................................................................... 39 4.2.5.1. Infestation of plants ........................................................................................................... 39 4.2.5.2. RNA extraction and DNase treatment ............................................................................... 39 4.2.5.3. RNA quantification ............................................................................................................. 40 4.2.5.4. Complementary DNA (cDNA) synthesis ............................................................................. 40 4.2.5.5. Target genes and primers design ....................................................................................... 40 4.2.6. Volatile organic compounds (VOCs) emission ........................................................................... 43 4.2.6.1. Volatile sampling ............................................................................................................... 43 4.2.6.2. VOCs chemical analysis ...................................................................................................... 45 4.3. Statistical analysis ............................................................................................................... 45 V. Results ________________________________________________ 47 5.1. Spider mites and russet mites effects on tomato leaf tissue ............................................. 47 5.2. Spider mites feeding preference and performance ........................................................... 48 5.3. Transcriptional analysis of defence genes .......................................................................... 49 5.3.1. Upstream of JA synthesis ........................................................................................................... 50 5.3.2. JA-responsive proteinase inhibitor: WIPI-II ............................................................................... 52 5.3.3. SA-dependent gene: PR-1 ........................................................................................................ 53 5.3.4. Tomato geranylgeranyl pyrophosphate synthase 1 .................................................................. 54 5.3.5. Systemic responses.................................................................................................................... 55 5.4. Predatory mites olfactory preference behavior ................................................................. 56 5.6. Headspace volatiles emitted from tomato plants .............................................................. 58 VI. Discussion and conclusions _________________________________ 61 6.1. Differential gene expression in response to mite herbivory .............................................. 61 6.2. Terpene synthase: GGPS1 ................................................................................................... 62 6.3. Tri-trophic interaction ......................................................................................................... 63 6.4. How russet mites overcome the tomato defence? ............................................................ 65 Literature cited _____________________________________________ 67 _______________________________________________________________________ Summary Plant and herbivores coexist for millions of years and have developed an arsenal of complex interactions. They can be mutually beneficial or antagonistic. In antagonistic interaction, plants have evolved a wide array of constitutive morphological, biochemical and molecular defences to defend themselves from herbivore attacks (Karban