植物青枯病菌环介导等温扩增快速检测技术研究

为实现植物青枯病的早期诊断,需要建立一种适于田间快速便捷检测青枯病菌的方法。以细胞色素C基因为靶标设计一套特异性引物,建立了植物青枯病的LAMP检测方法。此方法最低检测极限为1 pg,可在1 h内完成,不依赖昂贵复杂的仪器,结果可经肉眼观察。利用此方法,在人工接种发病的茄子、番茄、花生、芝麻和凹头苋茎部浸出液和马铃薯病薯块茎组织液中均检测出青枯病菌的存在,尤其适用于田间疑似罹病的芝麻、花生、番茄、马铃薯和甘薯等植株的检测,且LAMP法的检出率远高于PCR法。应用LAMP技术检测青枯病菌快速高效、特异性强、灵敏度高,操作简单,适于在基层推广运用。     

PIRIN2 stabilizes cysteine protease XCP2 and increases susceptibility to the vascular pathogen Ralstonia solanacearum in Arabidopsis

PIRIN (PRN) is a member of the functionally diverse cupin protein superfamily. There are four members of the Arabidopsis thaliana PRN family, but the roles of these proteins are largely unknown. Here we describe a function of the Arabidopsis PIRIN2 (PRN2) that is related to susceptibility to the bacterial plant pathogen Ralstonia solanacearum. Two prn2 mutant alleles displayed decreased disease development and bacterial growth in response to R.  solanacearum infection. We elucidated the underlying molecular mechanism by analyzing PRN2 interactions with the papain‐like cysteine proteases (PLCPs) XCP2, RD21A, and RD21B, all of which bound to PRN2 in yeast two‐hybrid assays and in Arabidopsis protoplast co‐immunoprecipitation assays. We show that XCP2 is stabilized by PRN2 through inhibition of its autolysis on the basis of PLCP activity profiling assays and enzymatic assays with recombinant protein. The stabilization of XCP2 by PRN2 was also confirmed in planta. Like prn2 mutants, an xcp2 single knockout mutant and xcp2 prn2 double knockout mutant displayed decreased susceptibility to R. solanacearum, suggesting that stabilization of XCP2 by PRN2 underlies susceptibility to R. solanacearum in Arabidopsis.     

Tn-seq identifies Ralstonia solanacearum genes required for tolerance of plant immunity induced by exogenous salicylic acid

Ralstonia solanacearum, the causal agent of the devastating bacterial wilt disease, is of particular interest to the scientific community. The repertoire of type III effectors plays an important role in the evasion of plant immunity, but tolerance to plant immunity is also crucial for the survival and virulence of R. solanacearum. Nevertheless, a systematic study of R. solanacearum tolerance to plant immunity is lacking. In this study, we used exogenous salicylic acid (SA) to improve the immunity of tomato plants, followed by transposon insertion sequencing (Tn-seq) analysis and the identification of R. solanacearum genes associated with tolerance to plant immunity. Target gene deletion revealed that the lipopolysaccharide (LPS) production genes RS_RS02830, RS_RS03460, and RS_RS03465 are essential for R. solanacearum tolerance to plant immunity, and their expression is induced by plant immunity, thereby expanding our knowledge of the pathogenic function of R. solanacearum LPS. SA treatment increased the relative abundance of transposon insertion mutants of four genes, including two genes with unknown function, RS_RS11975 and RS_RS07760. Further verification revealed that deletion of RS_RS11975 or RS_RS07760 resulted in reduced in vivo competitive indexes but increased tolerance to plant immunity induced by SA treatment, suggesting that these two genes contribute to the trade-off between tolerance to plant immunity and fitness cost. In conclusion, this work identified and validated R. solanacearum genes required for tolerance to plant immunity and provided essential information for a more complete view of the interaction between R. solanacearum and the host plant.     

Diversity of Halogenated Secondary Metabolites in Okinawan Aplysia argus Including 12-Hydroxypinnaterpene C and Their Feeding Targets

A new irieane-type diterpene, 12-hydroxypinnaterpene C ( 1 ), and 21 known compounds, angasiol acetate ( 2 ), angasiol ( 3 ), 11-deacetylpinnaterpene C ( 4 ), palisadin A ( 5 ), 12-acetoxypalisadin B ( 6 ), 12-hydroxypalisadin B ( 7 ), aplysistatin ( 8 ), luzodiol ( 9 ), 5-acetoxy-2-bromo-3-chloro-chamigra-7(14),9-dien-8-one ( 10 ), neoirietriol ( 11 ), neoirietetraol ( 12 ), (3Z)-laurenyne ( 13 ), cupalaurenol ( 14 ), cupalaurenol acetate ( 15 ), (3Z)-venustinene ( 16 ), 10-hydroxykahukuene B ( 17 ), aplysiol B ( 18 ), (3Z)-13-epipinnatifidenyne ( 19 ), 3Z,6R,7R,12S,13S-obtusenyne ( 20 ), (3Z,9Z)-7-chloro-6-hydroxy-12-oxo-pentadeca-3,9-dien-1-yne ( 21 ), and cholest-7-en-3,5,7-triol ( 22 ) were isolated from the digestive diverticula of Aplysia argus from the Ikei Island in Okinawa, Japan. The structures of these compounds were determined using spectroscopic methods such as NMR and HR-ESI-MS. These compounds were tested for their antibacterial activity against the phytopathogen Ralstonia solanacearum. Compounds 11 and 21 exhibited antibacterial activity at 30 μg/disc. In this study, we also discuss the types of red algae that A. argus feeds on in the shallow waters of Okinawa Prefecture.