IBR   13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
capítulos de libros
Título:
Section II. Plant-microorganisms interactions. Hypersensitive response.
Autor/es:
DAURELIO, L.D.; TONDO, M.L.; DUNGER, G.; GOTTIG, N.; OTTADO, J.; ORELLANO, E.G.
Libro:
Plant Bioassays
Editorial:
Series Editor: S. S. Narwal. Co – Editors: C. A. N. Catalán, D. A. Sampietro, M. A. Vattuone and B. Polyticka.
Referencias:
Año: 2007;
Resumen:
Plants have evolved to defense themselves against a variety of pathogens like bacteria, viruses and fungi. Hypersensitive response (HR) is a rapid and localized cell death associated with disease resistance including: race-specific (classical R-Avr interaction), race-non-specific and non-host resistance (Flor, 1971). The HR was first described by Stakman (1915). It is characterized by the rapid death of a limited number of cells in the vicinity of the invading pathogen and its function is thought to restrict or delay further pathogen spread and eventually kill the pathogen (Heath, 2000). Then, subsequent signalling events induce local and systemic activation of a set of defence responses that play a role in resistance. One of the most rapid plant responses is associated with the oxidative burst, which constitutes the production and accumulation of active oxygen species (AOS), primarily superoxide (O2-) and hydrogen peroxide (H2O2), at the site of invasion (Lamb and Dixon, 1997) which can produce toxic effects via DNA damage, protein degradation and modification and lipid peroxidation (Imlay and Linn, 1988). Other physiological and molecular modifications correlated with the HR are ion leakage from dead cells into the apoplast (Mysore and Ryu, 2004), deposition of lignin and callose into the plant cell wall (Verma and Hong, 2001) and production of phytoalexins, hydrolytic enzymes and pathogenesis-related proteins (PRs) (van Loon et al., 2006). Plant cell morphological modifications observed during HR are schematized in Figure 1.Xanthomonas axonopodis pv. citri (Xac), the bacterium responsible for citrus canker, induces a non-host resistance with HR in non-host plants such as pepper, tobacco, tomato, bean and cotton (Dunger et al., 2005). This chapter describes a variety of techniques to evaluate the HR in the interaction between bacteria and non-host plants using Xac and tobacco, pepper or cotton. A detailed description of plant growth and inoculation techniques is shown (Experiment 1). Pathogen growth restriction in inoculated plant leaves is evaluated using growth curves (Experiment 1). Also the AOS produced during the HR are biochemically detected. Hydrogen peroxide is detected using DAB (3,3´-Diaminobenzidine) staining that is oxidized by endogenous peroxidases in the presence of H2O2 (Experiment 2), while superoxide production is characterized by NBT (Nitro-blue tetrazolium) reduction and the formation of insoluble blue staining (Experiment 3). Ion leakage that reveals cellular membrane damage is determined by conductivity measurements (Experiment 4) and cell death can be monitored by trypan blue staining since the dye only passes through membranes of dead cells (Experiment 5). Finally, cell wall alterations revealed by callose deposition as a form of a non-host resistance is determined by staining with aniline blue, that selectively stains b-1-3 glycosidic linkages of the glucose units in callose (Experiment 6).