Moloi, Makoena Joyce
English: A recent accumulating body of evidence points to the importance of nitric oxide (NO) as a signal molecule involved in the regulation of many physiological and biochemical processes in plants. This study was aimed at investigating the involvement of reactive nitrogen species (RNS), in particular NO and peroxynitrite (ONOO-), in the Russian wheat aphid (RWA), Diuraphis noxia (Mordvilko), resistance response of wheat (Triticum aestivum L.). Resistant (cv. Tugela DN) and near isogenic susceptible (cv. Tugela) wheat plants were grown under the greenhouse conditions in trays at 24 oC (± 2 oC) and were infested at an early three leaf stage with approximately 20 RWAs (biotype RWASA1) per plant. RWA infestation induced an early accumulation (3-9 hours post infestation) of NO to a higher level in the resistant than susceptible plants. A corresponding increase in the activity of nitrate reductase (NR, EC 1.6.6.1) and inhibition studies using sodium tungstate (NR inhibitor) confirmed its involvement in NO production during the RWA resistance response. In addition, results indicated that the reduction of nitrate (NO3-) to nitrite (NO2-) involves NR and subsequently, nitrite reductase (NiR, EC 1.7.7.1) is responsible for the direct production of NO from NO2-. Inhibition studies using a nitric oxide synthase (NOS, EC 1.14.13.39) inhibitor, N-nitrilo-L-arginine-methyl ester (L-NAME), further proved that NOS has no significant effect on NO production during the RWA resistance response. In an experimental approach using sodium nitroprusside (SNP, NO donor) and / or sodium tungstate (NR inhibitor), it was discovered that NO acts as a signal for the induction of activities of secondary defense related enzymes such as the intercellular β-1,3-glucanase and peroxidase. Higher β-1,3-glucanase transcript accumulation after SNP treatment in resistant plants further supported the involvement of NO as signal molecule during the RWA defense response. In addition, results revealed that NO acts upstream of salicylic acid (SA) during the signaling events. High production of ONOO- [a product of an interaction between superoxide anion (O2-) and NO] in resistant plants after RWA infestation, suggests its involvement in the RWA resistance response. Inhibition of β-1,3-glucanase and peroxidase activities by urate (a specific inhibitor of ONOO- production), further suggests the involvement of ONOO- in the induction of the secondary RWA defense response. Furthermore, it was found that SNP root application of wheat plants resulted in transient resistance only. However, in contrast to root treatments, application of SNP as a seed dressing was more effective and has indeed a great potential of alleviating RWA damage in the susceptible plants by reducing aphid attraction as well as the intrinsic rate of aphid increase.
Afrikaans: Die belangrikheid van stikstofoksied (NO) as ‘n seinmolekule by die regulering van fisiologiese en biochemiese prosesse in plante word huidiglik toenemend beklemtoon. Die doel van hierdie studie was om die rol van reaktiewe stikstofspesies (RNS), veral NO en peroksinitriet (ONOO-), in die weerstandsrespons van koring (Triticum aestivum L.) teen die Russiese koringluis (RKL), Diuraphis noxia (Mordvilko) te bepaal. Weerstandbiedende (cv. Tugela DN) en na-isogeniese vatbare (cv. Tugela) koringplanteplante is in saadlaaie in ‘n glashuis by 24 °C (± 2 °C) gekweek en op ‘n vroeë drieblaarstadium met nagenoeg 20 RKL (biotipe RWASA1) per plant geïnfekteer. RKL-infestering induseer ‘n vroeë akkumulasie (3-9 ure na infestering) van NO, met hoër vlakke in die weerstandbiedende plante as in die vatbare plante. ‘n Ooreenstemmende toename in die aktiwiteit van nitraatreduktase (NR, EC 1.6.6.1) en remmingsstudies met behulp van natriumtungstaat (NR-remstof) het die betrokkenheid van NR by NO-produksie tydens die RKL-weerstandsrespons bevestig. Hierbenewens het resultate aangetoon dat NR by die reduksie van nitraat (NO3-) na nitriet (NO2-) betrokke is en dat nitrietreduktase (NiR, EC 1.7.7.1) vir die direkte produksie van NO vanaf NO2- verantwoordelik is. Remmingstudies, waar ‘n stikstofoksiedsintase (NOS, EC 1.14.13.39) remstof, Nω-nitrilo-L-arginien-metielester (L-NAME), gebruik is, het verder bewys dat NOS geen betekenisvolle invloed op NO-produksie gedurende die RKL-weerstandsrespons gehad het nie. ‘n Eksperimentele benadering deur van natriumnitroprussied (SNP, NO skenker) en / of natriumtungstaat (NR-remstof) gebruik te maak, het getoon dat NO as ‘n sein vir die induksie van aktiwiteite van sekondêre verdedigingsverwante ensieme soos intersellulêre β-1,3-glukanase and peroksidase dien. Hoër β-1,3-glukanasetranskripakkumulasie na SNP-behandeling van weerstandbiedende plante ondersteun die betrokkenheid van NO as seinmolekule gedurende die RKL-verdedigingsrespons. Resultate het ook getoon dat NO stroomop van salisielsuur (SA) gedurende seintransduksie werksaam is. ‘n Hoë produksie van ONOO- [‘n produk van die interaksie tussen superoksiedanione (O2-) en NO] in weerstandbiedende plante na RKL-infestering was aanduidend van die betrokkenheid van ONOO- by die RKL-weerstandsrespons. Remming van β-1,3-glukanase- en peroksidase-aktiwiteite deur uraat (‘n spesifieke remstof van ONOO- produksie) toon verder dat ONOO- by die induksie van die sekondêre RKL-verdedigingsrespons betrokke is. Voorts is bevind dat SNP, as ‘n worteltoediening, net kortstondige weerstand by koringplante veroorsaak het. Daarenteen was SNP meer doeltreffend wanneer dit as ‘n saadbedekking aangewend was en het dit inderdaad groot potensiaal om RKL- skade in vatbare plante te verlig deur die luisaantrekking te verminder sowel as om die luistoenametempo te verlaag.