水稻几丁质酶基因克隆RCH8的DNA结构分析

用DNA外切核酸酶Ⅲ和S1核酸酶生成连续缺失突变体,用Sanger双脱氧链终止法对该克隆进行双向DNA顺序测定,测序全长2049个碱基,初步确定了1057bp的5'端上游顺序,966bp不含内含子的完整编码区和可能的TATAbox等。所编码的322个氨基酸包括N-端20个氨基酸的信号肽,其后40个氨基酸长度的含8个半胱氨酸的hevein结构域和一个催化结构域。     

Transgenic tobacco plants which express thechiA gene fromSerratia marcescens have enhanced tolerance toRhizoctonia solani

We have prepared independent lines of transgenic tobacco plants which express high levels of theSerratia marcescens ChiA protein intracellulary or extracellularly (in glycosylated or unglycosylated forms). We have measured the susceptibility, of these plants toRhizoctonia solani infection in greenhouse trials and in the field. Transgenic tobacco plants which constitutively express theS. marcescens ChiA protein exhibit tolerance to the fungal pathogenR. solani. Disease tolerance is observed in transgenic tobacco plants which express the bacterial chitinase intra-or extracellulary. This is the first report to document disease reduction in the field in transgenic plants engineered for fungal disease tolerance.     

Mode of Parasitism of Meloidogyne and Other Nemaiode Eggs by Dactylella oviparasitica

Hyphae of Dactylella oviparasitica proliferated rapidly through MeIoidogyne egg masses, and appressoria formed when they contacted eggs. The fungus probably penetrated egg shells mechanically, although chitinase production detected in culture suggested that enzymatic penetration was also possible. In soil, D. oviparasitica invaded egg masses soon after they were deposited on the root surface and eventually parasitized most of the first eggs laid. Occasionally the fungus grew into Meloidogyne females, halting egg production prematurely. The fungus parasitized eggs in the gelatinous matrix or eggs freed from the matrix and placed on agar or in soil. Specificity in nematode egg parasitism was not displayed, for D. oviparasitica parasitized eggs of four Meloidogyne spp., Acrobeloides sp., Heterodera schachtii, and Tylenchulus semipenetrans. In tests in a growth chamber, parasitism by D. oviparasitica suppressed galling on M. incognita-infected tomato plants.     

保幼激素类似物对烟蚜茧蜂生长发育、寄生率和蜕皮相关酶活性的影响

【目的】烟蚜Myzus persicae是烟草上重要的害虫之一,烟蚜茧蜂Aphidius gifuensis是烟蚜的一种优势寄生蜂。本研究旨在筛选出可延迟烟蚜茧蜂羽化时间的最佳保幼激素,解决规模化繁殖过程中出现的烟蚜茧蜂羽化不一致、不整齐,生产上急需烟蚜茧蜂防控烟蚜时所需烟蚜茧蜂数量不足而影响防蚜效果等严重问题。【方法】利用液浸法测定了不同浓度(5 000, 1 000, 200, 40和8 ng/μL)的5种保幼激素类似物包括稀虫乙酯(ZR-512)、稀虫炔酯(ZR-777)、稀虫酯(ZR-515)、苯氧威［(对苯氧乙基)氨基甲酸乙酯)］和保幼激素Ⅲ(2,6-壬二烯酸)处理后对烟蚜茧蜂羽化率、羽化时间、成蜂寿命、雌蜂比例和寄生率的影响;通过生物化学方法测定1 000 ng/μL这5种保幼激素类似物处理后烟蚜茧蜂蛹内与蜕皮相关酶含量和活性,筛选能延迟烟蚜茧蜂羽化时间的最佳保幼激素类似物。【结果】测试的不同浓 度的5种保幼激素类似物中200 ng/μL ZR-777和1 000 ng/μL ZR-512处理后能显著延迟烟蚜茧蜂羽化时间,分别比对照(10%丙酮处理)延迟了44.00和56.00 h;不同浓度的ZR-515和苯氧威处理后烟蚜茧蜂羽化率较对照组显著降低。与对照组相比,测试的不同浓度的5种保幼激素类似物对成蜂寿命均没有显著影响;1 000和40 ng/μL ZR-777处理显著降低了雌蜂比例。200 ng/μL ZR-777, 1 000 ng/μL ZR-512和5 000 ng/μL ZR-512处理对烟蚜茧蜂的寄生率无显著影响。5种保幼激素类似物以1 000 ng/μL浓度处理时,ZR-512处理组中烟蚜茧蜂蛹内酚氧化酶含量和活性以及几丁质酶活性均最低。【结论】生产上可用1 000 ng/μL ZR-512和200 ng/μL ZR-777处理烟蚜茧蜂,以达到调控烟蚜茧蜂羽化时间,取得足量、一致的烟蚜茧蜂。结果为烟蚜茧蜂规模化繁殖提供了理论依据。     

Inducible Proteins in Citrus Rootstocks with Different Tolerance Towards the Root Rot Pathogen Phytophthora palmivora

Activities of defence‐related proteins (β‐1,3‐glucanases, chitinases and peroxidases) and concentrations of total soluble phenolics were measured in roots and leaves of non‐infected and infected plants to investigate the response of different citrus rootstock genotypes to the root rot pathogen Phytophthora palmivora Butler. Infection with the pathogen increased concentrations of total proteins, total phenolics and β‐1,3‐glucanase activity in roots of all genotypes, and increases were associated with the extent of root mass reductions and thus susceptibility of the plants. Root chitinase and root peroxidase levels were slightly reduced or unaltered upon infection. β‐1,3‐Glucanase activity was also elevated in leaves of infected plants, but increases did not differ between tolerant and susceptible rootstocks. Effects of root infection on leaves were typically the reverse of effects on roots for chitinase‐ and peroxidase levels and more pronounced in susceptible rootstock genotypes. Although differences in enzyme expression were observed between susceptible and tolerant citrus seedlings, effects were usually associated with disease progression, and not with resistance to P. palmivora. It is suggested that increased activities of the proteins and soluble phenolics studied are not implicated in the primary defence to Phytophthora root diseases, but may contribute to the inhibition of the pathogen during infection in tolerant citrus.     

Secondary cell wall formation in Cryptococcus neoformans as a rescue mechanism against acid-induced autolysis

Growth of the opportunistic yeast pathogen Cryptococcus neoformans in a synthetic medium containing yeast nitrogen base and 1.0–3.0% glucose is accompanied by spontaneous acidification of the medium, with its pH decreasing from the initial 5.5 to around 2.5 in the stationary phase. During the transition from the late exponential to the stationary phase of growth, many cells died as a consequence of autolytic erosion of their cell walls. Simultaneously, there was an increase in an ecto-glucanase active towards β-1,3-glucan and having a pH optimum between pH 3.0 and 3.5. As a response to cell wall degradation, some cells developed an unusual survival strategy by forming 'secondary' cell walls underneath the original ones. Electron microscopy revealed that the secondary cell walls were thicker than the primary ones, exposing bundles of polysaccharide microfibrils only partially masked by an amorphous cell wall matrix on their surfaces. The cells bearing secondary cell walls had a three to five times higher content of the alkali-insoluble cell wall polysaccharides glucan and chitin, and their chitin/glucan ratio was about twofold higher than in cells from the logarithmic phase of growth. The cell lysis and the formation of the secondary cell walls could be suppressed by buffering the growth medium between pH 4.5 and 6.5.     

Fate of bacteria, Aeromonas caviae, in the midgut of the housefly, Musca domestica

Abstract. The role of houseflies as agents in the spread of bacterial diseases has been thoroughly investigated, yet the fate of bacteria ingested by flies has not. We examined the physical location of the bacterial enteropathogen Aeromonas caviae in the midgut of laboratory‐reared adult houseflies. Food ingested by houseflies was separated from the midgut epithelium by a double‐layered peritrophic matrix (PM). The inner PM intimately enveloped the food as fecal pellets (food boluses), while the outer PM appeared as a long continuous tube. In flies fed a suspension of A. caviae, live bacteria were not observed within the inner PM, but were compartmentalized between folds of the PM in the inter‐PM space. Similar observations were made for flies fed a suspension of Serratia liquefaciens and for highly contaminated feral flies. Isolates of both A. caviae and S. liquefaciens were chitinolytic (as demonstrated by clearing zones on chitin agar), but the potential role of bacterial enzymes in the alteration of PM morphology or formation needs further investigation.     

Constitutive expression of two endochitinases from root nodules ofElaeagnus umbellata confers resistance on transgenicArabidopsis plants against the fungal pathogenBotrytis cinerea

Plant chitinases have been known as pathogenesis-related (PR) proteins, but recent studies suggest that they play functional roles during normal plant growth and development. We previously isolated two cDNA clones encoding endochitinases,EuNOD-CHT1 and -CHT2, from the root nodules ofElaeagnus umbellata. These genes show differential expression patterns, with theEuNOD-CHT1 gene being active in the root nodules and meristems, whileEuNOD-CHT2 is preferentially expressed in the infected cells of those nodules. To elucidate the functional roles of these two endochitinases, we have now constitutively expressed each gene in a heterologous plant system,Arabidopsis thaliana. Stable inheritance and expression of the transgenes were confirmed by genomic Southern hybridization and RT-PCR. Our transgenic plants did not differ morphologically from the wild types. However, constitutive expression ofEuNOD-CHT1 and -CHT2 inArabidopsis resulted in increased resistance against a fungal pathogen,Botrytis cinerea, but not against a bacterial agent,Pseudomonas syringae pv. Tomato DC3000. Expression levels were enhanced by both wounding and jasmonic acid treatments (forEuNOD-CHT1), or by jasmonic acid only (forEuNOD-CHT2). These data suggest thatEuNOD-CHT1 and -CHT2 primarily play defensive roles during root nodule development inE. umbellata.     

Differential inducible defense mechanisms against bacterial speck pathogen in Arabidopsis thaliana by plant-growth-promoting-fungus Penicillium sp. GP16-2 and its cell free filtrate

Although a wealth of information is available regarding resistance induced by plant growth-promoting rhizobacteria (PGPR), not much is known about plant growth-promoting fungi (PGPF). Hence, the goal of the present research was to provide more information on this matter. In Arabidopsis thaliana L., root colonizing PGPF Penicillium sp. GP16-2 or its cell free filtrate (CF) elicited an induced systemic resistance (ISR) against infection by Pseudomonas syringae pv. tomato DC3000 (Pst), leading to a restriction of pathogen growth and disease development. We demonstrate that signal transduction leading to GP16-2-mediated ISR requires responsiveness to JA and ET in a NPR1-dependent manner, while CF-mediated ISR shows dispensability of SA, JA, ET and NPR1-dependent signaling (at least individually). In addition, root colonization by GP16-2 is not associated with a direct effect on expression of known defense-related genes, but potentiates the activation of JA/ET-inducible ChitB, which only becomes apparent after infection by Pst. However, CF-mediated ISR was partly associated with the direct activation of marker genes responsive to both SA and JA/ET signaling pathways and partly associated with priming, leading to activation of JA-/ET-inducible ChitB and Hel genes. These suggest that CF may contain one or more elicitors that induce resistance by way where at least SA, JA and ET may play a role in defense signaling in Arabidopsis. Therefore, defense gene changes and underlying signaling pathways induced by Penicillium sp. GP16-2 root colonization and its CF application are not the same and only partially overlap.