辣椒对疫霉抗性的快速鉴定

研究发现叶、茎、茎基部和果实接种4种方法均能用于鉴定辣椒对辣椒疫霉的抗性。叶与茎接种法操作简单,接种部位发病快,可同时检测大量品种,但要求空气湿度大;茎基部接种法模拟田间病害的发生过程,最能反映品种的真实抗性,但需要菌量大,工作量也大;果实接种法不仅操作简单、接种部位发病快、菌需求量小,而且可同时筛选大量品种,可以通过量化达到有效区分抗病和感病品种的目的,更具有完整保存原有植株的优势。     

Screening potential bacterial biocontrol agents towards Phytophthora capsici in pepper

A total of 1,487 bacterial isolates were obtained from the rhizosphere, phyllosphere, endorhiza and endosphere of field-grown pepper. In a dual assay, 232 isolates displayed the antagonistic activity towards Phytophthora capsici L.; 36.6?% and 39.2?% of them were obtained from the rhizosphere and phyllosphere, respectively. 40 of the 232 antagonistic isolates producing inhibition zones of at least 5?mm in diameter were assessed for production of siderophores and chitinase, cellulose, and protease activity. These 40 isolates fell into 15 groups according to 90?% similarity of the banding patterns obtained by amplified ribosomal DNA restriction analysis (ARDRA). Seventeen isolates spanning the 15 groups were evaluated in greenhouse tests for their ability to control Phytophthora blight of pepper. Biocontrol efficacy ranged from 0.7?% to 92.3?%, with three isolates (B1301, R98, and PX35) exhibiting maximum ability to reduce the disease severity (83.5?%, 92.3?% and 83.5?%, respectively). Based on 16S rDNA sequencing, these isolates were identified as Bacillus cereus (B1301), Chryseobacterium sp (R98) and Bacillus cereus (PX35). This is the first report that Chryseobacterium sp. (R98) can function as a biocontrol agent of Phytophthora blight.     

Molecular detection of Phytophthora capsici in infected plant tissues, soil and water

A species-specific PCR assay was developed for rapid and accurate detection of the pathogenic oomycete Phytophthora capsici in diseased plant tissues, soil and artificially infested irrigation water. Based on differences in internal transcribed spacer (ITS) sequences of Phytophthora spp. and other oomycetes, one pair of species-specific primers, PC-1/PC-2, was synthesized. After screening 15 isolates of P. capsici and 77 isolates from the Ascomycota, Basidiomycota, Deuteromycota and Oomycota, the PC-1/PC-2 primers amplified only a single PCR band of c . 560 bp from P. capsici . The detection sensitivity with primers PC-1/PC-2 was 1 pg genomic DNA (equivalent to half the genomic DNA of a single zoospore) per 25- µ L PCR reaction volume; traditional PCR could detect P. capsici in naturally infected plant tissues, diseased field soil and artificially inoculated irrigation water. Using ITS1/ITS4 as the first-round primers and PC-1/PC-2 in the second round, nested PCR procedures were developed, increasing detection sensitivity to 1 fg per 25- µ L reaction volume. The results suggested that the assay detected the pathogen more rapidly and accurately than standard isolation methods. The PCR-based methods developed here could simplify both plant disease diagnosis and pathogen monitoring, as well as guiding plant disease management.     

Effect of prior tillage and soil fertility amendments on dispersal of Phytophthora capsici and infection of pepper

Organic soil amendments including composted cotton gin trash, composted poultry manure, an incorporated rye-vetch green manure, or synthetic fertilizer were applied to subplots, and main plots were either tilled frequently or surface-mulched in experimental field plots between 1997 and 2004. Soil from each replication of the tillage and fertility treatments was sampled in August of 2001, 2002, and 2003, brought to the greenhouse, and infested with Phytophthora capsici to study the effect of previous soil treatments on disease incidence and dispersal of the pathogen. Both the previous tillage and fertility amendments affected the incidence of disease and dispersal of the pathogen. Final disease incidence, AUDPC and the distance of pathogen spread were significantly greater in soils with previous surface mulch applications than in frequently tilled soils. Final disease incidence, AUDPC and the distance of pathogen spread were also significantly higher in soils amended with cotton gin trash, than rye-vetch green manure, poultry manure, or synthetic fertilizer. Soils amended with cotton gin trash had higher soil water content, lower bulk density, higher humic matter content, higher porosity and higher levels of mineralizable N, than soils with other fertility amendments. Soil water content, soil porosity, humic matter content, and net mineralizable levels nitrogen were positively correlated and bulk density was negatively correlated with final incidence of disease.     

Evaluation of Trichoderma harzianum for controlling root rot caused by Phytophthora capsici in pepper plants

The ability of Trichoderma harzianum to control the rotting of pepper ( Capsicum annuum ) plant roots caused by Phytophthora capsici was studied. Interactions between the fungi were assessed in vitro on three culture media (V8c, Czapek and 2% water agar) and in vivo in plants grown in a substrate inoculated with P. capsici and T. harzianum . Studies on mutual antagonism in vitro showed that P. capsici was inhibited by T. harzianum ; however, the intensity of inhibition differed according to the medium used, being greatest on Czapek. Analysis of the fungal populations in the plant growth substrate showed that T. harzianum consistently reduced that of P. capsici over time. This reduction in the pathogen population was associated with a reduction in root rot of between 24 and 76%, although plant growth (dry weight) was still reduced by 21.2–24.7%, compared with the uninoculated control. In the absence of T. harzianum with the same pathogen inoculum levels, the reduction in dry weight was 59.8–68.6%, suggesting that T. harzianum reduced the damage.     

辣椒疫病抗性的浸根接种鉴定方法

为建立规模化的辣椒疫病苗期抗性鉴定技术, 将辣椒疫霉人工接种于抗疫病辣椒品种‘ICPN21-03’和感病辣椒品种‘B16144’, 研究了接种菌液浓度?接种时期?病情调查时期和接种方法等对人工接种鉴定效果的影响?结果表明, 接种菌液游动孢子浓度为10个/mL时, 感病品种病情指数为73.37, 可以满足寄主发病的需要; 1 ~ 10真叶期接种均能使植株发病, 但5 ~ 6真叶期接种能反映抗感病品种真实抗性; 接种5 d调查抗感病品种病情指数分别为9.32和73.37, 可明显区别抗感病品种?使用浸根法进行鉴定, 需菌量小, 对接种环境要求小?用37个辣椒品种进行验证, 鉴定结果表明该苗期抗性鉴定技术可很好地对辣椒品种进行抗性水平分级鉴定?     

辣椒中L型凝集素类受体激酶CaLecRKs鉴定及其对辣椒疫霉的响应

L型凝集素类受体激酶(LecRKs)广泛参与植物的先天免疫过程。目前未见在辣椒Capsicum annuum中全基因组鉴定LecRKs的报道。本研究对辣椒中的CaLecRK进行了全基因组鉴定, 并在接种辣椒疫霉Phytophthora capsici条件下通过基因表达分析探究其对辣椒疫霉的响应情况, 旨在挖掘参与辣椒抗疫病防御反应的CaLecRK基因。研究结果表明, 辣椒基因组中共鉴定出24个CaLecRK, 以其构建系统发育树发现, 可将24个CaLecRK分为7个分支。基因表达分析结果显示, 有4个CaLecRK基因(CaLecRK2.2、CaLecRK3.2、CaLecRK8.1和CaLecRK10.1)受辣椒疫霉诱导, 和接菌后0 h相比, 接菌处理后12 h 或36 h基因表达差异显著, 推测其在辣椒抗疫病防御反应中发挥了重要作用。     

Dispersal of Phytophthora capsici and P. parasitica in furrow-irrigated rows of bell pepper, tomato and squash

Dispersal of Phytophthora capsici and P. parasitica from point sources buried near the upper end of 74 m-long irrigation furrows was studied with three annual host crops. Furrows next to tomato were inoculated with either P. capsici or P. parasitica , while furrows next to pepper and squash were inoculated with P. capsici only. Irrigation was carried out on a 14-day cycle. Dispersal of each fungus in the water was monitored by transplants in the furrows and the incidence of infected tomato fruit along furrows. Disease gradients on roots and shoots of all hosts were also measured. Repeated irrigations dispersed P. capsici and P. parasitica up to 70 m from the source. Fruit infection increased with increasing distances downstream, suggesting an accumulation of secondary inoculum with the repeated flow of water. Conversely, gradients of disease severity on roots of tomato and pepper peaked at the source and rapidly decreased to low levels up to 32 m downstream. Fruit and root infection upstream from source was negligible. Root and crown rot in squash was highly variable, being confined to locations near the initial inoculum in some units while developing to severe levels several meters downstream in other units. Unlike the other hosts, squash petioles were in contact with the irrigation water and may have served as routes of invasion by P. capsici. Transport of inoculum from the furrow water to the roots was limited. P. capsici isolates were more virulent on tomato roots and caused a higher incidence of buckeye rot on tomato fruit than did isolates of P. parasitica.     

Factors affecting expression of resistance in pepper (Capsicum annuum) to blight caused by Phytophthora capsici in screening trials

The effects of plant age, fungus isolate, zoospore concentration and inoculation method on the resistance of pepper ( Capsicum annuum) to blight ( Phytophthora capsici) were studied under controlled conditions. Each of the factors studied produced differing responses in the pepper lines tested hut line CNPH 286 proved to be a reliable source of resistance and is now being used in a breeding programme. An inoculation method for screening pepper lines for blight resistance is proposed, using a standardized zoospore suspension applied to the base of plants 45 days from sowing.     

Defence responses of pepper (Capsicum annuum L.) infected with incompatible and compatible strains of Phytophthora capsici

In this study, we investigated the activities of β-1,3-glucanase and peroxidase enzymes in the leaves of pepper cultivar A3 infected with the incompatible strain PC and the compatible strain HX-9 of Phytophthora capsici. The activities of β-1,3-glucanase and peroxidase enzymes substantially increased in the incompatible interactions compared to the compatible interactions. We also analysed the expression patterns of four defence-related genes, including CABPR1, CABGLU, CAPO1 and CaRGA1, in the leaves and roots of pepper inoculated with different strains of P. capsici. All gene expression levels were higher in the leaves than in the roots. Markedly different expression patterns were observed between incompatible and compatible host-pathogen interactions. In the incompatible interactions, the expression levels of CABPR1, CABGLU and CAPO1 genes in leaves increased by a maximum of 17.2-, 13.2- and 20.5-fold at 24, 12 and 12 h, respectively, whereas the CaRGA1 gene expression level increased to a lesser degree, 6.0-fold at 24 h. However, in the compatible interactions, the expression levels of the four defence-related genes increased by a maximum of 11.2-, 8.6-, 7.9- and 2.0-fold at 48, 24, 48 and 72 h, respectively. Compared to the leaves, the expression levels of the four defence-related genes were much lower in the roots. The highest levels of mRNA were those of the CABPR1 gene, which increased 5.1-fold at 24 h in the incompatible and 3.2-fold at 48 h in the compatible interactions. The other three genes exhibited lower expression levels in the incompatible and compatible interactions. These results further confirmed that defence-related genes might be involved in the defence response of pepper to P. capsici attack.     

Survival and spread of Phytophthora capsici in Coastal Peru

Phytophthora capsici is a soilborne pathogen that causes significant losses to pepper production in Peru. Our objective was to investigate the mechanisms by which P. capsici is able to survive and spread. During 2005 to 2007, 227 isolates of P. capsici were collected from four species of pepper (Capsicum annum, C. baccatum, C. chinense, and C. pubescens) and tomato (Solanum lycopersicum) at 33 field sites in 13 provinces across coastal Peru. All 227 isolates were of the A2 mating type and amplified fragment length polymorphism (AFLP) analysis indicates that 221 of the isolates had the same genotype. Analyses of six polymorphic single nucleotide polymorphism (SNP) loci showed fixed heterozygosity suggesting a single clonal lineage is widely dispersed. Members of the same clonal lineage were recovered during 2005 to 2007 from geographically separate locations from each of the host types sampled. Our results indicate that clonal reproduction drives the population structure of P. capsici in Peru. The impact of continuous cropping and irrigation from common river sources on the population structure in Barranca Valley are discussed.     

Physiological races and soil population level of fusarium wilt of watermelon

Results of a comparative test oflocal isolates ofFusarium oxysporum f. sp.niveum with isolates from abroad indicate the existence of a highly virulent race of this fungus in Israel. All these isolates were found also pathogenic to resistant cultivars from the U.S.A. When inoculum density was tested, a one-hundredfold higher fungus population was required for extensive wilt of watermelon seedlings in freshly infested sterilized soil compared with the same soil kept dry for three months. In naturally infested soil, where almost 100% of the plants were infected toward the end of the season, the lowest count of the fungus population (400 propagules/g soil) was comparable to the inoculum density required for total wilt of watermelon seedlings in infested sterilized soil.     

Evidence for inbreeding and apomixis in close crosses of Phytophthora capsici

A series of inbreeding crosses, recurrent backcrosses and successive sibling crosses were completed up to the sixth generation in the plant pathogen Phytophthora capsici , generating a total of 692 oospore-derived isolates. All of the crosses stemmed from an initial mating between two wild-type P. capsici isolates. The heterozygosity level, as measured through the inheritance of 20 amplified fragment length polymorphism (AFLP) markers, decreased incrementally with continued inbreeding and was reduced by ~60–75% by the second consecutive sibling cross. Of the eight crosses analysed, all but one cross produced a proportion of oospore-derived progeny that were identical to one or other parent, indicating that apomixis can play a role in P. capsici intraspecific crosses. There was no evidence of isolates emerging through selfing or generation of homothallic isolates. Overall, these results indicate that back and sibling crosses are possible and that the large reservoirs of naturally occurring genetic variation in P. capsici may be useful for developing inbred lines to characterize complex genetic traits in Phytophthora .     

日光温室辣椒疫病发生规律及综合防治对策

辣椒疫病是本地区日光温室辣椒生产中的常见高发病害之一。笔者通过连续4年的多点温室田间调查研究与综合防治示范,初步明确和掌握了该病在本地区的田间发病症状、发病条件及发生规律,并从农业防治、物理防治、生态防治及化学防治等方面总结提出了一套较为系统完善的、科学实用的综合防治对策,以指导生产实践。     

Fusarium oxysporum Fo47 confers protection to pepper plants against Verticillium dahliae and Phytophthora capsici,and induces the expression of defence genes

The application of the nonpathogenic isolate Fusarium oxysporum 47 (Fo47) reduced the symptoms of verticillium wilt, phytophthora root rot and phytophthora blight in pepper plants. Botrytis cinerea was also tested on the leaves of plants treated with Fo47, but no protection was observed. Verticillium dahliae colonies cultured in the presence of Fo47 grew slower than control cultures, but Phytophthora capsici growth was unaffected by Fo47. At least part of the protection effect observed against V. dahliae could therefore be due to antagonism or competition. In order to search for induced resistance mechanisms, three defence genes previously related to pepper resistance were monitored over time. These genes encode a basic PR‐1 protein (CABPR1), a class II chitinase (CACHI2) and a sesquiterpene cyclase (CASC1) involved in the synthesis of capsidiol, a phytoalexin. These three genes were transiently up‐regulated in the roots by Fo47 in the absence of inoculation with the pathogen, but in the stem only CABPR1 was up‐regulated. In plants that were inoculated with V. dahliae after the Fo47 treatment, the three genes had a higher relative expression level than the control in both the roots and the stem.     

Phenylalanine ammonia lyase activity in chilli CM-334 infected by Phytophthora capsici and Nacobbus aberrans

We tested the hypothesis that PAL activity in chilli plants CM-334 inoculated with Nacobbus aberrans (Na) alone or in combination with Phytophthora capsici (Pc), is lower than in those inoculated only with Pc. At 21 days after nematode inoculation, inoculated plants showed a significant (P < 0.01) reduction of 48% in PAL activity compared to those non-inoculated in two separate experiments. In two other tests, where plants were inoculated with the oomycete 21 days after inoculation with the nematode, PAL activity at 2, 4, 6, 8 and 24 h after inoculation with Pc was significantly higher (Tukey, P < 0.01) in plants inoculated only with Pc than in plants inoculated only with Na or both pathogens (Na+Pc).     

Asexual Reproduction of Phytophthora capsici as Affected by Extracts from Agricultural and Nonagricultural Soils

ABSTRACT Formation of sporangia and zoospores in species of Phytophthora is known to be influenced by soil microbial and chemical composition. In Phytophthora capsici, the study of the relationship of soil chemical composition to production of sporangia and zoospores has been limited. P. capsici is a soilborne pathogen of a wide array of vegetable crops, including chile pepper (Capsicum annuum) in New Mexico. Production of sporangia and zoospores by P. capsici was evaluated in extracts of soils from three different environments in New Mexico: (i) agricultural environments with a long history of chile pepper cropping and occurrence of P. capsici (CP), (ii) agricultural environments with no history of chile pepper cropping and no occurrence of P. capsici (Non-CP), and (iii) nonagricultural environments consisting of forests and rangelands (Non-Ag). There was a significant difference in production of P. capsici asexual propagules, expressed as natural log (number of sporangia x number of zoospores), among the three environments (P = 0.0298). Production of propagules was 9 to 13% greater in Non-Ag than in CP or Non-CP environments. Stepwise multiple discriminant analysis and canonical discriminant analysis identified the edaphic variables Na, pH, P, organic matter content, and asexual propagule production as contributing the most to the separation of the three environments. Two significant (P < 0.0001) canonical discriminant functions were derived with the first function, accounting for approximately 75% of the explained variance. Based on the two discriminant functions, approximately 93, 86, and 89% of observations in CP, Non-CP, and Non-Ag environments, respectively, were classified correctly. Soils from agricultural and nonagricultural environments differentially influence production of sporangia and zoospores in P. capsici, and soil samples could be effectively classified into agricultural and nonagricultural environments based on soil chemical properties and the production of asexual propagules by P. capsici in soil extracts.     

重庆地区辣椒疫霉菌的分离培养及生理小种鉴定

本试验分离鉴定了来源于重庆不同辣椒产区的14个病原菌分离物,经形态特征鉴定及回接发病特征观察,确定这些菌株均为辣椒疫霉(Phytophthora capsici)。14个菌株在OMA培养基上诱导产生的孢子囊形态类似,形状多为卵圆形或长椭圆形,乳突明显。对PDA、OMA、CA及V8汁4种培养基上的培养性状观察表明,辣椒疫霉菌株在OMA培养基上生长速度最快,但在V8汁培养基上产孢最多。采用灌根法对14个菌株进行生理小种鉴别,其中10个为race 3,2个为race 2,2个为race 1,初步推定race 3为重庆地区辣椒疫病病原菌的优势小种。