Seedling Petri-dish inoculation method:A robust,easy-to-use and reliable assay for studying plant-Ralstonia solanacearum interactions

Ralstonia solanacearum causes a lethal bacterial wilt disease in many crops,leading to huge losses in crop production every year.Understanding of plant-R.solanacearum interactions will aid to develop efficient strategies to control the disease.As a soilborne pathogen,R.solanacearum naturally infects plants via roots.A huge limitation in studying plant-R.solanacearum interactions is the large variation of R.solanacearum infection assay due to the variable soil conditions and uneven inoculum exposure.Here,we developed a robust and reliable Petri-dish inoculation method which allows consistent and stable infection in young plant seedlings.This method is easy to use,takes about only 10 days from seed germination to the completion of inoculation assay,and requires less inoculum of bacteria as well as growth chamber space.We proved the efficacy of the seedling Petri-dish inoculation method by analyzing plant defense primed by molecular patterns,resistance of defense-related plant mutants,and virulence of R.solanacearum mutants.Furthermore,we demonstrated that the seedling Petri-dish inoculation method can be applied to other host plants such as tobacco and has great potential for high-throughput screening of resistant plant germplasms to bacterial wilt in the future.

9,10-KODA,an α-ketol produced by the tonoplastlocalized 9-lipoxygenase ZmLOX5,plays a signaling role in maize defense against insect herbivory

13-Lipoxygenases(LOXs)initiate the synthesis of jasmonic acid(JA),the best-understood oxylipin hormone in herbivory defense.However,the roles of 9-LOX-derived oxylipins in insect resistance remain unclear.Here,we report a novel anti-herbivory mechanism mediated by a tonoplast-localized 9-LOX,ZmLOX5,and its linolenic acid-derived product,9-hydroxy-10-oxo-12(Z),15(Z)-octadecadienoic acid(9,10-KODA).Transposon-insertional disruption of ZmLOX5 resulted in the loss of resistance to insect herbivory.lox5 knockout mutants displayed greatly reduced wound-induced accumulation of multiple oxylipins and defense metabolites,including benzoxazinoids,abscisic acid(ABA),and JA-isoleucine(JA-Ile).However,exogenous JA-Ile failed to rescue insect defense in lox5 mutants,while applications of 1 mM 9,10-KODA or the JA precursor,12-oxo-phytodienoic acid(12-OPDA),restored wild-type resistance levels.Metabolite profiling revealed that exogenous 9,10-KODA primed the plants for increased production of ABA and 12-OPDA,but not JA-Ile.While none of the 9-oxylipins were able to rescue JA-Ile induction,the lox5 mutant accumulated lower wound-induced levels of Ca^(2+),suggesting this as a potential explanation for lower wound-induced JA.Seedlings pretreated with 9,10-KODA exhibited rapid or more robust woundinduced defense gene expression.In addition,an artificial diet supplemented with 9,10-KODA arrested fall armyworm larvae growth.Finally,analysis of single and double lox5 and lox10 mutants showed that ZmLOX5 also contributed to insect defense by modulating ZmLOX10-mediated green leaf volatile signaling.Collectively,our study uncovered a previously unknown anti-herbivore defense and hormonelike signaling activity for a major 9-oxylipin α-ketol.

Priming of antiherbivore defensive responses in plants

Abstract Defense priming is defined as increased readiness of defense induction. A growing body of literature indicates that plants （or intact parts of a plant） are primed in anticipation of impending environmental stresses, both biotic and abiotic, and upon the following stimulus, induce defenses more quickly and strongly. For instance, some plants previously exposed to herbivore-inducible plant volatiles （HIPVs） from neighboring plants under herbivore attack show faster or stronger defense activation and enhanced insect resistance when challenged with secondary insect feeding. Research on priming of antiherbivore defense has been limited to the HIPV-mediated mechanism until recently, but significant advances were made in the past three years, including non-HIPV-mediated defense priming, epigenetic modifications as the molecular mechanism of priming, and others. It is timely to consider the advances in research on defense priming in the plant- insect interactions.

Trichoderma harzianum enhances tomato indirect defense against aphids

Many fungal root symbionts of the genus Trichoderma are well-known for their beneficial effects on agronomic performance and protection against plant pathogens; moreover, they may enhance protection from insect pests, by triggering plant resistance mechanisms. Defense barriers against insects are induced by the activation of metabolic pathways involved in the production of defense-related plant compounds, either directly active against herbivore insects, or exerting an indirect effect, by increasing the attrac- tion of herbivore natural enemies. In a model system composed of the tomato plant, the aphid Macrosiphum euphorbiae and the parasitoid Aphidius ervi, plant metabolic changes induced by Trichoderma harzianum and their effects on higher trophic levels have been assessed. T. harzianum T22 treatments induce a primed state that upon aphid attacks leads to an increased attraction of aphid parasitoids, mediated by the enhanced produc- tion of volatile organic compounds （VOCs） that are known to induce Aphidius ervi flight. Transcriptome sequencing of T22-treated plants infested by aphids showed a remarkable upregulation of genes involved in terpenoids biosynthesis and salicylic acid pathway, which are consistent with the observed flight response ofA. ervi and the VOC bouquet profile underlying this behavioral response.