Strong evidence for a fire blight resistance gene of Malus robusta located on linkage group 3

Fire blight (FB), caused by the Gram-negative bacterium Erwinia amylovora is a dangerous disease on pome fruit, including apple. The FB-susceptible cultivar 'Idared' was crossed with the resistant wild species clone Malus  ×  robusta 5. A segregating population of 146 progeny has been tested by artificial shoot inoculation for susceptibility to FB. Progeny were infected from 0% to 100% of the shoot length. To identify chromosomal regions or loci responsible for resistance to FB of Malus  ×  robusta 5, a set of microsatellite markers (simple sequence repeat, SSRs) was chosen covering all linkage groups of apple. Up to eight different microsatellites were bulked to one mutliplex PCR using four different labels and a fifth label for a size standard. Fifty-nine microsatellite markers out of 72 SSRs were polymorphic. Fifty-four of 66 loci detected could be mapped and were useful for the detection of related resistant loci. Alleles of microsatellites Hi03d06, CH03g07 and CH03e03 originating from the resistant donor M. robusta were associated with resistance to Erwinia amylovora . Up to eighty percent of the phenotypic variation could be explained by the interval spanned by SSRs CH03g07 and CH03e03, indicating the presence of a major resistance gene. All three microsatellites are located on the distal part of linkage group 3, spanning 15 cM. The SSR marker CH03e03 can be regarded as diagnostic marker for FB resistance. Only seven progeny expressing allele b (184 bp) of CH03e03 showed blighted shoot lengths of more than 30% and only nine progeny lacking allele b showed blighted shoot lengths of <30%. By setting a threshold of 30% shoot necrosis for resistance to FB, the 146 individuals segregate into 71 susceptible and 75 resistant plants, and resistance to FB maps 9 cM away from marker CH03e03.     

蝇蛆几丁低聚糖抑菌作用的初步研究

 研究了蝇蛆几丁低聚糖对主要农作物种子病原菌和一些果蔬腐烂菌的抑制作用,并探索影响其抗菌活性的主要因子。结果表明,蝇蛆几丁低聚糖具有广谱的抗菌活性,对测试的32种农作物病原真菌和果蔬腐烂真菌中的花生白绢菌等31种有较好的抑制作用;蝇蛆几丁低聚糖的抗菌活性与脱乙酰度和分子量大小有关,随着脱乙酰度的增加,几丁低聚糖的抗菌活性逐渐增强,随着分子量的增大,几丁低聚糖的抗菌活性逐渐下降。盆栽试验结果表明,蝇蛆几丁低聚糖处理玉米、小麦和棉花种子,可以提高发芽率和出苗率。蝇蛆几丁低聚糖喷雾处理对玉米苗期小斑病具有一定的防病     

家蝇不同虫态附着蜡蚧轮枝菌分生孢子能力及与其体表结构的关系

【目的】研究家蝇成虫、幼虫和蛹附着蜡蚧轮枝菌分生孢子的能力及与其体表结构的关系,为解释虫生真菌对同一昆虫不同虫态间的致病性差异提供科学依据。【方法】采用2种不同接菌方法（浸泡虫体法、喷雾虫体法）,通过血球计测量家蝇3种虫态（成虫、幼虫和蛹）体表附着的蜡蚧轮枝菌分生孢子数,并利用扫描电子显微镜等显微技术观察3种虫态的体表超微结构及附着分生孢子情况。【结果】2种接菌方法测得家蝇3种虫态附着的蜡蚧轮枝菌分生孢子数量差异显著（P<0.05）,均表现为成虫＞蛹＞幼虫。显微观察发现,家蝇成虫周身密集着刚毛、鬃和微毛等体壁外长物,微毛上附着大量的蜡蚧轮枝菌分生孢子,孢子分泌粘液进一步加强附着作用;蛹体大量分布着呈环状的皱褶,在体节接合处有稀疏的棘刺带,蛹体上无分生孢子附着;幼虫体表几乎无皱褶,只在体节接合处附近有带状棘刺,其体表也无孢子附着。【结论】家蝇成虫、幼虫和蛹附着蜡蚧轮枝菌分生孢子的能力与其体表结构有密切关系。     

Cold pre-treatment is effective for 1-MCP efficacy in ‘Tsugaru’ apple fruit

1-Methylcyclopropene (1-MCP) treatment maintains apple fruit quality during storage, but its efficacy is dependent on a number of conditions. ‘Tsugaru’ apples are a major early season cultivar in Japan, but because ‘Tsugaru’ fruit produce abundant ethylene, they have a short shelf-life, and efficacy of 1-MCP is not as high with ‘Tsugaru’ as with other cultivars. To improve 1-MCP efficacy, ‘Tsugaru’ fruit were pre-cooled at −1 °C or −3 °C for 24 h before 1-MCP treatment. Ethylene production decreased with the cold treatment, resulting in better storage after 1-MCP treatment. Although ethylene production was low at the end of 24 h of the cold pre-treatment, expression of ACS1, the ethylene receptor genes ERS1, ETR1(a), ETR1b, ETR2 and ETR5, and the cell wall degradation-related gene PG1 all increased with a 24 h cold treatment. It is assumed that these elevated gene expression levels were not caused by ethylene, but more directly by cold stimulus. Thus, a short period of cold stimulus suppresses ethylene production, but induces expression of some genes. 1-MCP treatment was more effective with some initial fruit chilling.     

细菌诱导家蝇蛹抗菌物质的分离纯化

家蝇生境中充满病原菌,其有效的防防机制成为研究的热点。通过三氟乙酸缓冲液浸提、热处理、固相萃取以及两次RP-HPLC等方法,从混合细菌液诱导的家蝇蛹中分离纯化出4种具有抗菌活性的物质。结果表明,提取的家蝇蛹粗提液抗菌谱较广,对多种微生物具有不同程度的抗菌作用,且抗菌效果好于天然肽类防腐剂和一些化学防腐保鲜剂。4种抗菌物质中包括3个抗菌肽和1个抗菌化合物。Tricine-SDS-PAGE结果显示3个抗菌肽的分子量分别约为12kDa、8kDa和4kDa。本研究表明细菌诱导家蝇蛹中具有丰富的抗菌物质。     

几种植物生长物质对苹果果皮着色的影响

在7月下旬至8月下旬分别用不同浓度的ABA、BR与S3307对苹果果实进行喷施处理，以便筛选出促进果皮着色及增进果实品质的最佳药剂浓度。试验结果表明：ABA与S3307处理对果皮花青苷含量、果肉可溶性糖含量及单果重均有较好的促进作用，BR对果肉可溶性糖含量有明显促进作用。但各种浓度药剂处理对花青苷含量、可溶性糖含量及单果重影响效果不同，药剂对两个苹果品种的作用亦有差异。最终筛选出低浓度ABA、高浓度BR及高浓度S3307为最佳药剂处理浓度。     

Identification of an RAPD Marker Linked to the Vf Gene for Scab Resistance in Apples

Resistance to V. inaequalis, derived from the small-fruited species Malus floribunda 821, is determined by a major dominant gene, assigned as Vf. The material used in this paper is based on the introgression of the Vf gene from M. floribunda into commercial apple varieties. Comparing RAPD patterns of a genomic DNA sample of M. floribunda with a pooled DNA sample of resistant individuals and that of 10 susceptible commercial apple varieties, fragments were identified which are derived from M. floribunda. One of them, the fragment OPD20/600, proved to be linked to the Vf gene with a recombination value of about 0.20—0.25. It is the first DNA marker so far for scab resistance.     

一种新型家蝇幼虫抗菌肽基因(MDAP-2)的真核表达及抑菌活性检测

利用毕赤酵母表达系统表达新型家蝇抗菌肽MDAP-2基因(Musca domestica antimicrobial peptide,MDAP-2),为其作为新型抗菌物质应用于临床提供实验依据。根据已有的MDAP-2的氨基酸序列,采用PCR技术扩增获得MDAP-2基因,构建真核重组表达质粒pPIC9K-MDAP-2,将线性化的重组质粒电击转入毕赤酵母(P.pastoris)感受态GS115细胞内,将PCR鉴定为阳性的重组酵母菌进行甲醇诱导表达,并采用Tricine-SDS-PGAE检测表达产物的大小,经镍柱亲和层析纯化获得纯度较高的重组蛋白,采用管碟法检测重组蛋白的生物学活性。结果显示,重组质粒pPIC9K-MDAP-2获得高效表达,蛋白分子量为7.8 kD,管碟法检测重组蛋白对大肠杆菌与沙门氏菌均具有体外抑菌活性。MDAP-2基因的成功表达为进一步研究表达产物的生物学及免疫学活性奠定基础。     

Water use of young ‘Fuji’ apple trees at three soil moisture regimes in drainage lysimeters

Studies were conducted during 4 months of each growing season in 1994 and 1995 to measure water use of young apple trees (Malus domestica Borkh. cv ‘Fuji’) growing under different soil moisture regimes in temperate climate conditions and to evaluate monthly crop coefficients of such conditions. To do so, double pot lysimeters under a transparent rain shield were designed and installed. The three soil moisture regimes in three replicates each were: (A) drip-irrigation at −50 kPa of soil matric potential (IR50); (B) drip-irrigation at −80 kPa of soil matric potential (IR80); and (C) constant shallow water table at 0.45 m below the soil surface (WT45). In each treatment, soil surface was maintained with or without turf grasses. Monthly water use was not different in drip-irrigated treatments (IR50 and IR80), but greatest in the WT45 treatment. Monthly crop coefficients increased linearly in time for drip-irrigated apple trees (r² values of 0.76^*** for IR50 and of 0.77^*** for IR80), while those obtained in the WT45 treatment fluctuated. Leaf water potential (LWP) of drip-irrigated trees was similar until 63 days after treatment (DAT), but the values for IR80 trees began to decline thereafter. The LWP of WT45 trees decreased from 48 DAT. Temporal variations in leaf water content (LWC) was similar to that of LWP, except for two abrupt decreases in IR80 trees. The LWC of WT45 trees began to decrease from 59 DAT, and this occurred 2 weeks after the reduction in LWP. Average shoot length of IR50 trees was greater than that of IR80 and WT45 trees. The results of this study provided water use and crop coefficients for apple trees in relation to soil moisture regimes under temperate climate.     

Viability and release of Neonectria ditissima ascospores on apple fruit in Brazil

During European canker monitoring in an apple experimental orchard, 14 mummified fruit (two and three trees with 10 and four positive records in 2018 and 2019, respectively) showed perithecia. Perithecium production on apple fruit, confirmation of pathogenicity of Neonectria ditissima isolated from mummified fruit, and ascospore release from fruit tissues has rarely been reported, and their role in the epidemiology of European canker has been largely overlooked. Thus, the objectives of our study were to (a) prove the presence of both conidia and ascospores of N. ditissima in mummified fruit in an experimental field, confirming pathogenesis in different apple cultivars, and (b) monitor production of the two types of inoculum in infected apple fruit over time. Canker incidence in this orchard was 47% of trees with symptoms in 2018 and 48% in 2019. Molecular and morphological tests confirmed that the fungus detected in the mummified apple fruit was N. ditissima. Apple fruit with sporodochia and perithecia washed immediately after collection from the orchard showed conidia but no ascospores of N. ditissima. However, after 4 days’ incubation, perithecia on mummified fruit showed many ascospore cirri. Koch's postulates were fulfilled on apple plants and mature fruit. Fruit inoculated with N. ditissima released spores for over a year under Brazilian field conditions. The release of both spore types peaked in May (Brazilian leaf fall) and October (spring); release of conidia also peaked in February (early harvest). These results support our hypothesis that fruit can serve as primary inoculum for European canker in Brazilian apple orchards.