Plant Growth Promoting Rhizobacteria Having 1-Aminocyclopropane-1-Carboxylic Acid Deaminase to Induce Salt Tolerance in Sunflower (<i>Helianthus annus L.</i>)

Soil salinity badly affects agriculture productivity through accumulation of salts in upper layers of soils. The harmful effects of salts in arable lands have influenced modern as well as ancient civilizations. A pot study was carried out to test the performance of two PGPR isolates (KS 8, KS 28) on sunflower (SMH-0917) under different salinity levels (8, 10 and 12 dS·m-1). These salinity levels were developed by adding calculated amount of salts (NaCl, Na2SO4, CaCl2 and MgSO4) with ratio of 3:4:2:1. The bacterial strains KS 8 and KS 28 were applied separately in two treatments while third treatment was co-inoculation (KS mix). Completely randomized experimental design (CRD) was used and data were collected at flowering stage about pre-decided plant growth parameters (plant height, shoot dry weight and root dry weight). The bacterial isolate KS 8 showed an increase of 26, 102% and 83% in plant height, shoot dry weight and root dry weight at EC 8 dS·m-1, while this improvement was 67%, 163% and 296% at EC 10 dS·m-1, however an increase of 100%, 74% and 382% was recorded over control respectively at EC 12 dS·m-1. Similarly isolate KS 28 exhibited an increase of 14%, 69% and 54% in plant height;shoot dry weight and root dry weight at EC 8 dS·m-1, whereas this improvement was 56%, 163% and 188% at EC 10 dS·m-1, while an increase of 60%, 41% and 282% was registered respectively over control at EC 12 dS·m-1. The increase due to mixture treatments was 4%, 41% and 16% in plant height, shoot dry weight and root dry weight at EC 8 dS·m-1, while an increase of 33%, 57% and 100% at EC 10 dS·m-1, whereas an improvement of 53%, 33% and 164% respectively was noted at EC 12 dS·m-1 over un-inoculated. The isolate KS 8 performed better than KS 28 and mixture treatment. These two PGPR strains could be used to mitigate the adverse impact caused by salinity stress on sunflower.

假单胞菌株M18分泌羧基吩嗪抑制黄瓜枯萎病害

设计并应用新的高效筛选程序 ,从植物根际土壤分离出同时具有促进植物生长和抑制黄瓜枯萎病害的假单胞菌株 M1 8.它对黄瓜幼苗的根系具有明显的促进作用 ,在培养皿中 ,M1 8能有效地抑制黄瓜枯萎病菌以及多种其他植物病原菌菌丝体的生长 .经 M1 8活菌浸种处理春黄瓜种子并在大棚内浇灌定植后的根际土壤 ,与对照相比 ,黄瓜枯萎病的株发病率降低 70 %～ 80 % ,霜霉病的发病率和病情指数都同时平均下降 70 %左右 ,黄瓜产量提高 2 0 %以上 .M1 8的生物防治效果达到极显著程度 .对培养液中纯化的抗真菌物质进行光谱分析和 LC- MC测定的结果表明 ,M1

Isolation of a Tomato Protease that May Be Involved in Proteolysis of 1-Aminocyclopropane-l-Carboxylate Synthase

1-aminocyclopropane- 1-carboxylate （ACC） synthase is a principal enzyme that catalyses the committed step in phytohormone ethylene biosynthesis. Previous evidence indicates that the hypervariable C-terminus of ACC synthase is most likely to be processed proteolytically in vivo. However, the protease responsible has not been identified thus far. In the present study, we detected proteolytic activity against ACC synthase （LeACS2） in tomato （Lycopersicon esculentum Mill.） fruit extract based on a newly established in vitro assay system. Purification of the protease through DEAE, gel filtration and MonoQ chromatography resulted in considerable enrichment of a 64-kDa protein species. Subsequent biochemical analysis of the purified tomato protease revealed that the optimal conditions for its proteolytic activity were at pH 8.0 and at 37 ~C. In addition, the protease activity was blocked completely by the metalloprotease inhibitor 1,10-phenanthroline. The present study represents the first report on the isolation of an ACC synthase- processing protease from plant tissues.

Mutation in the gene encoding 1-aminocyclopropane-1-carboxylate synthase 4 (CitACS4) led to andromonoecy in watermelon

Summary Although it has been reported previously that ethylene plays a critical role in sex determination in cucurbit species, how the andromonoecy that carries both the male and hermaphroditic flowers is determined in watermelon is still unknown. Here we showed that the watermelon gene 1-aminocyclopropane-1-carboxylate syn- thase 4 CCitACS4）, expressed specifically in carpel primor- dia, determines the andromonoecy in watermelon. Among four single nucleotide polymorphism （SNPs） and one lnDel identified in the coding region of CitACS4, the C364W mutation located in the conserved box 6 was co- segregated with andromonoecy. Enzymatic analyses showed that the C364W mutation caused a reduced activity in CitACS4. We believe that the reduced CitACS4 activity may hamper the programmed cell death in stamen primordia, leading to the formation of hermaphroditic flowers.

辣椒根际促生菌的分离筛选及生物育苗基质研制

[目的]将根际促生菌(plant growth-promoting rhizobacteria,PGPR)与普通育苗基质联合,研制成生物育苗基质,进而促进功能菌株苗期的根际定殖和移苗后促生功能的发挥。[方法]利用从辣椒根际分离筛选的产IAA和ACC脱氨酶菌株,保活添加至普通育苗基质研制成生物育苗基质,通过比较育苗效果筛选出生物育苗基质的最佳配伍菌株。[结果]从辣椒根际分离获得6株IAA产生量大于5 mg·L-1的菌株,其中菌株NJAU-G10、NJAU-N5和NJAU-N1同时能产ACC脱氨酶,且能力高于其他菌株;两季苗盘育苗试验结果表明,添加菌株NJAU-G10的生物基质,表现出较其他菌株更为突出的根际定殖和促进幼苗生长的能力,并确定其最佳接种量为5%;盆栽试验结果表明,菌株NJAU-G10在移苗后对辣椒的生长仍具有显著的促进作用,且能够有效地在根际定殖;结合形态、生理生化特征和16S r DNA基因序列分析,初步鉴定菌株NJAU-G10为枯草芽孢杆菌。[结论]分离获得一株枯草芽孢杆菌NJAU-G10,添加其研制的生物育苗基质能够有效促进辣椒苗盘期种苗及其移栽后的生长,研究结果能够为含PGPR蔬菜育苗基质的开发提供理论支撑,同时为PGPR新产品的开发提供新的思路。

Quinclorac Resistance in Echinochloa crus-galli from China

Echinochloa crus-galli is a major weed in rice fields in China,and quinclorac has been long used for its control.Over-reliance of quinclorac has resulted in quinclorac resistance in E.crus-galli.Two resistant(R)E.crus-galli populations from Hunan,China were confirmed to be at least 78-fold more resistant to quinclorac than the susceptible(S)population.No difference in foliar uptake of 14C-labelled quinclorac was detected between the R and S plants.However,a higher level of 14C translocation and a lower level of quinclorac metabolism were found in the R plants.Basal and induced expression levels ofβ-cyanoalanine synthase(β-CAS)gene andβ-CAS activity were not significantly different between the R and S plants.However,the induction expression of 1-aminocyclopropane-1-carboxylic acid oxidase(ACO1)gene by quinclorac treatment was evident in the S plants but not in the R plants.Quinclorac resistance in the two resistant E.crus-galli populations was not likely to be related to foliar uptake,translocation or metabolism of quinclorac,nor to cyanide detoxification viaβ-CAS.Thus,target-site based quinclorac signal reception and transduction and regulation of the ethylene synthesis pathway should be the focus for further research.

The Dependence of <i>N</i>-Malonyltryptophan Formation in Plants on Water Deficit (Review)

Drought stress in plants is accompanied by several metabolic changes. One of them is the appearance of N-malonyltryptophan (MT) during leaf wilting of many species, but there is a significant number of plant species in which the appearance of MT did not occur. Plants of some species were able to synthesize also N-acetyltryptophan (AT). Excised tomato leaves incubated with D-amino acids (including D-Trp) transform them into malonyl- and acetyl-derivatives even without water deficit. However, MT which appeared during water deficit has been shown to contain L-Trp. Amino acid—1-amino-cyclopropane-1-carboxylic acid (ACC) is also malonylated during water deficit, but other L-amino acids were not malonylated. N-malonyl transferases specific for Trp and ACC have been found in several plants. The existence of N-malonyltransferase specific to L-Trp and appeared during water deficit in plants forming MT is supposed, but clear experimental proof has not been obtained yet. Plants can transform MT applied exogenously into Trp and further to indole-3-acetic acid (IAA). But no evidence has been appeared up to now that endogenous MT may be a source of IAA. It is unknown till now why it is necessary for plants of many species to malonylate only Trp during water deficit. How MT metabolized in animals and if it affects them is also unknown. The necessity to use molecular-genetic approaches for the elucidation of the physiological significance of MT formation during water deficit is underlined.

耐盐菌Pseudomonas brassicacearum YZX4的筛选、鉴定及其植物促生特性

植物根际促生菌(PGPR)具有促进植物生长的作用.从盐碱地植物根际土壤中分离筛选耐盐菌,测定其在盐胁迫下的1-氨基环丙烷-1-羧酸(ACC)脱氨酶活性、吲哚乙酸(IAA)合成能力、嗜铁素合成能力、无机磷溶解能力,以及在Ashby无氮培养基上的生长情况;并对同时具有以上促生功能的耐盐菌进行不同盐浓度下的促生功能测定、小黄白(白菜Brassica pekinensis的一个品种)种子萌发促生实验和菌株鉴定.结果显示,在筛选得到的15株耐盐菌中,菌株YZX4在10 g/L NaCl浓度下同时具有多种促生特性.在不同盐浓度下促生功能测定实验中,当盐浓度为10 g/L时,菌株的ACC脱氨酶活性(以α-KA/Pr计)、IAA合成量和嗜铁素相对含量最高,分别为11.07(±1.89)μmol mg^-1h^-1、36.42(±1.81)mg/L和0.61(±0.15),且随着盐浓度的增加而降低;在20 g/L盐浓度下,该菌株的固氮量、有机磷溶解量和无机磷溶解量最高,分别为4.79(±1.61)mg/L、1.68±(0.04)mg/L和23.77(±1.30)mg/L.在小黄白种子萌发促生实验中,当盐浓度为5.84 g/L时,YZX4的菌液(105 CFU/mL)对小黄白的种子萌发率、幼苗根、茎长和平均鲜重分别提高了7.19%、17.33%、23.85%和22.69%.根据形态特征、生理生化鉴定结果和16S rDNA序列分析,初步确定菌株YZX4为油菜假单胞菌(Pseudowonas brassicacearum).上述研究结果表明在盐胁迫下同时具备多种促生特性的菌株YZX4可作为盐碱地改良微生物菌剂的优良菌源.

Exogenous ethylene influences flower opening of cut roses (Rosa hybrida) by regulating the genes encoding ethylene biosynthesis enzymes

The purpose of this paper is to investigate the differential responses of flower opening to ethylene in two cut rose cultivars, ‘Samantha’, whose opening process is promoted, and ‘Kardinal’, whose opening process is inhibited by ethylene. Ethylene production and 1- aminocyclopropane-1-carboxylate (ACC) synthase and oxidase activities were determined first. After ethylene treatment, ethylene production, ACC synthase (ACS) and ACC oxidase (ACO) activities in petals increased and peaked at the earlier stage (stage 3) in ‘Samantha’, and they were much more dramatically enhanced and peaked at the later stage (stage 4) in ‘Kardinal’ than control during vasing. cDNA fragments of three Rh-ACSs and one Rh- ACO genes were cloned and designated as Rh-ACS1, Rh-ACS2, Rh-ACS3 and Rh-ACO1 respectively. Northern blotting analysis revealed that, among three genes of ACS, ethylene-in- duced expression patterns of Rh-ACS3 gene corresponded to ACS activity and ethylene production in both cultivars. A more dramatic accumulation of Rh-ACS3 mRNA was induced by ethylene in ‘Kardinal’ than that of ‘Samantha’. As an ethylene action inhibitor, STS at concentration of 0.2 mmol/L generally inhib-ited the expression of Rh-ACSs and Rh-ACO in both cultivars, although it induced the expression of Rh-ACS3 transiently in ‘Kardinal’. Our results suggests that ‘Kardinal’ is more sensitive to ethylene than ‘Samantha’; and the changes of Rh-ACS3 expression caused by ethylene might be related to the acceleration of flower opening in ‘Samantha’ and the inhibition in ‘Kardinal’. Additional results indicated that three Rh-ACSs genes were differentially associated with flower opening and senescence as well as wounding.

Seed priming with Pseudomonas putida isolated from rhizosphere triggers innate resistance against Fusarium wilt in tomato through pathogenesis-related protein activation and phenylpropanoid pathway

Pathogenesis-related(PR)proteins are one of the major and preliminary proteins accumulated as a defense against biotic stress.This defense response can be induced by using beneficial rhizobacteria,which has been studied in various host-pathogen interactions.In the present study,eleven Pseudomonas isolates were assessed for their potential to ferment sorbitol,reduce nitrate,and produce mycolytic enzymes,1-aminocyclopropane-l-carboxylic acid(ACC)deaminase,phenazine antibiotics,and N-acyl homoserine lactones(AHLs).All isolates were tested against the host-specific pathogen Fusarium oxysporum MTCC1755 in tomato under greenhouse conditions,and shortlisted isolates were tested for their rhizosphere competence.In-vitro test results showed that the isolates were able to produce mycolytic enzymes,including protease,lipase,chitinase,cellulase,and amylase,and the antibiotic phenazine and were negative for pyoluteorin.All the isolates except two were positive for ACC deaminase production.Greenhouse results showed that the isolates M80,M96,and T109 significantly reduced symptoms of Fusarium wilt.Extended greenhouse tests under autoclaved and unautoclaved soil conditions showed that M80,M96,and T109 were excellent rhizosphere competitors and were identified as Pseudomonas putida.In brief,the defense-specific biochemical variations in the host could describe the improved defense against Fusarium wilt occurring in the primed plants.These three Pseudomonas strains could be used as potential biocontrol agents,along with their rhizosphere competence.