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Simultaneous detection of fungal, bacterial, and viral pathogens in insects by multiplex PCR and capillary electrophoresis 多重PCR和毛细管电泳同时检测昆虫真菌、细菌和病毒病原体
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.64
Kyu‐Won Kwak, S. Nam, Ji-Young Choi, Seokhyung Lee, H. Kim, Sung Hyun Kim, K. Park, Myung‐Sae Han
Beetles Protaetia brevitarsis seulensis Kolbe (Coleoptera: Cetoniidae) and Allomyrina dichotoma Linn. (Coleoptera: Scarabaeidae) are widely used in traditional medicine, and the number of insect-rearing farms is increasing in South Korea. The purpose of this study was to establish a multiplex PCR-based assay for rapid simultaneous detection of multiple pathogens causing insect diseases. Six insect parasites such as fungi Beauveria bassiana (Bals.-Criv.) Vuill. (Hypocreales: Cordycipitaceae) and Metarhizium anisopliae (Metschn.) Sorokin (Hypocreales: Clavicipitaceae), bacteria Bacillus thuringiensis Berliner (Bacillales: Bacillaceae), Pseudomonas aeruginosa Migula (Pseudomonadales: Pseudomonadaceae), and Serratia marcescens Bizio (Enterobacteriales: Enterobacteriaceae), and Oryctes rhinoceros nudivirus were chosen based on the severity and incidence rate of insect diseases in South Korea. Pathogen-specific primers were designed and successfully applied for simultaneous detection of multiple infectious agents in farm-bred insects P. b. seulensis and A. dichotoma using multiplex PCR and high resolution capillary electrophoresis. Our results indicate that multiplex PCR is an effective and time-saving method for simultaneous detection of multiple infections in insects, and the QIAxcel capillary electrophoresis system is useful for quantitative evaluation of the individual impact of each infectious agent on the severity of insect disease. The approach designed in this study can be utilized for rapid and accurate diagnostics of infection in insect farms.
白衣原甲(鞘翅目:拟蜂科)与异瓢虫。金龟子(鞘翅目:金龟子科)被广泛用于传统医学,韩国的昆虫养殖场正在增加。本研究的目的是建立一种基于多重聚合酶链反应(pcr)的多种病原菌快速同时检测方法。六种昆虫寄生虫,如真菌白僵菌(Bals.-Criv.)Vuill。(下creale:冬虫夏草科)和绿僵菌(Metschn.)根据韩国昆虫疾病的严重程度和发病率,选择了Sorokin (Hypocreales: Clavicipitaceae)、thuringiensis Berliner (Bacillales: Bacillaceae)、铜绿假单胞菌Migula (Pseudomonadaceae: Pseudomonadaceae)、粘质沙雷菌Bizio (Enterobacteriales: Enterobacteriaceae)和orytes rhinoceros nudivirus。设计了病原特异性引物,并应用多重PCR和高分辨毛细管电泳技术成功地同时检测了农种昆虫的多种病原。结果表明,多重PCR是同时检测昆虫多种感染的一种有效且省时的方法,QIAxcel毛细管电泳系统可用于定量评价每种感染因子对昆虫疾病严重程度的个体影响。本研究设计的方法可用于昆虫养殖场感染的快速准确诊断。
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引用次数: 4
Expression of porcine circovirus type 2 capsid protein fused with partial polyhedrin using baculovirus 猪圆环病毒2型衣壳蛋白与部分多面蛋白融合的杆状病毒表达
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.50
Jun Beom Lee, S. Bae, T. Shin, S. Woo
Porcine circovirus type 2 (PCV2) is an important infectious swine virus causing postweaning multisystemic wasting syndrome (PMWS). PCV2 capsid protein, encoded by ORF2 has type- specific epitopes, is very immunogenic, and is associated with the induction of neutralizing antibodies. For the efficient production of capsid protein, recombinant Autographa californica nucleopolyhedroviruses were generated to express ORF2 fused with two forms of a partial polyhedrin. Recombinant capsid protein was produced successfully with the partial polyhedrin fusion form and the yield was high, as was shown by SDS-PAGE. Production of recombinant capsid proteins in insect cells was confirmed by Western blot analysis using anti- His monoclonal antibody, anti-ORF2 monoclonal antibody, and anti-PCV2 porcine serum. Fusion expression with amino acids 19 to 110 of the polyhedrin increased the production of recombinant capsid protein, but fusion with amino acids 32 to 85 did not. Additionally, PCV2 capsid protein is a glycoprotein; however, the glycosylation of recombinant protein was not observed. The results of an Enzyme-linked immunosorbent assay (ELISA) showed that recombinant capsid proteins could be utilized as antigens for fast, large-scale diagnosis of PCV2-infected pigs. Our results suggest that the fusion expression of partial polyhedrin is able to increase the production of recombinant PCV2 capsid protein in insect cells.
猪圆环病毒2型(PCV2)是引起断奶后多系统消耗综合征(PMWS)的重要传染性猪病毒。由ORF2编码的PCV2衣壳蛋白具有类型特异性表位,具有很强的免疫原性,并与中和抗体的诱导有关。为了高效地生产衣壳蛋白,制备了重组加州签名虫核多角体病毒,表达ORF2与两种形式的部分多角体蛋白融合。SDS-PAGE结果表明,以部分多面蛋白融合形式成功制备了重组衣壳蛋白,产率高。利用抗His单克隆抗体、抗orf2单克隆抗体和抗pcv2猪血清进行Western blot分析,证实在昆虫细胞中产生重组衣壳蛋白。与氨基酸19 ~ 110的融合表达增加了重组衣壳蛋白的产量,而与氨基酸32 ~ 85的融合表达则没有增加重组衣壳蛋白的产量。此外,PCV2衣壳蛋白是一种糖蛋白;然而,重组蛋白的糖基化未被观察到。酶联免疫吸附试验(ELISA)结果表明,重组衣壳蛋白可作为快速、大规模诊断pcv2感染猪的抗原。我们的研究结果表明,融合表达部分多面蛋白能够提高重组PCV2衣壳蛋白在昆虫细胞中的产量。
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引用次数: 1
Silkworm-food plant-interaction: search for an alternate food plant for tasar silkworm (Antheraea mylitta Drury) rearing 蚕-食性植物-相互作用:寻找一种可替代的沙蚕(Antheraea mylitta Drury)饲养的食性植物
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.58
M. Deka, G. Gargi, Rajendra Kumar, H. Yadav, A. Sahay
An experimental rearing of tropical tasar silkworm, Antheraea mylitta Drury was conducted to study silkworm-food plant-interaction and thereby to search for an alternate silkworm food plant. The silkworm-food-plant-interaction was studied with six different food plant species viz. Terminalia tomentosa, Terminalia arjuna, Terminalia belerica, Terminalia chebula of Combretaceae family and Lagerstroemia speciosa, Lagerstroemia parviflora of Lythraceae family. The rearing performance of silkworm on Lagerstroemia speciosa in terms of cocoons per DFL and silk ratio was found comparable with Terminalia tomentosa and Termonalia arjuna, the primary tasar silkworm food plant species. These three plant species also possessed better results in terms of physiological (leaf moisture content and net photosynthesis rate) and biochemical (Chlorophyll, protein, carbohydrate and crude fibre contents) characteristics to support silkworm rearing than Terminalia belerica, Terminalia chebula and Lagerstroemia parviflora. The correlation study between silkworm rearing performance and food plant`s constituents indicates commercial perspective of Lagerstroemia speciosa as an alternate food plant for tasar silkworm rearing.
通过对热带柞蚕(Antheraea mylitta Drury)的实验饲养,研究了蚕与食性植物的相互作用,从而寻找替代的家蚕食性植物。研究了家蚕与6种不同食性植物的相互作用,分别是锦蚕科的毛毛终末草、阿朱终末草、belalia belerica终末草、chebula终末草和天蚕科的大紫薇(Lagerstroemia speciosa)、小紫薇。家蚕在紫花Lagerstroemia speciosa上的育茧率和产丝率与主要的沙蚕食用植物毛毛蕨和阿朱蕨相当。这三种植物在养蚕的生理(叶片含水量和净光合速率)和生化(叶绿素、蛋白质、碳水化合物和粗纤维含量)方面也比黄顶草、chebula和小花紫薇具有更好的效果。家蚕的饲养性能与食用植物成分的相关性研究表明,大紫薇作为一种可替代的沙蚕食用植物具有商业前景。
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引用次数: 1
Effect of different Bombyx mori silkworm varieties on the wet spinning of silk fibroin 不同家蚕品种对湿法纺丝丝素蛋白的影响
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.75
M. Jang, I. Um
The wet spinning of silk solution has attracted researchers’ attention because of 1) unique properties of silk as a biomedical material and 2) easy control of the structure and properties of the regenerated silk fiber. Recently, studies have reported that different silkworm varieties produce silk with differences in the molecular weight (MW) and other mechanical properties of the regenerated silk fibroin (SF) film. In this study, we look at the effect of different Bombyx mori varieties on the wet spinning of SF. Although five regenerated SFs from different silkworm varieties have different MWs and solution viscosity, the wet spinnability and post drawing performance of regenerated SFs were not different. This result is due to low variability in the MW of the regenerated SF samples from the different silkworm varieties. In addition, unlike regenerated SF films, the mechanical properties of wet spun regenerated SF filament were not affected by silkworm variety. This result suggests that the mechanical properties of wet spun SF filament are less affected by MW than those of SF film are.
蚕丝溶液的湿法纺丝技术由于蚕丝作为生物医学材料的独特性能和再生蚕丝纤维的结构和性能易于控制而受到研究人员的关注。近年来,有研究报道,不同品种的蚕所产的蚕丝在分子量(MW)和再生丝素(SF)膜的其他力学性能上存在差异。研究了不同家蚕品种对湿纺纱性能的影响。虽然不同蚕品种的5种再生丝的分子量和溶液粘度不同,但再生丝的湿纺性和拉伸后性能没有差异。这一结果是由于不同家蚕品种的再生SF样品的分子量变异性较低。此外,与再生丝绵膜不同,湿纺再生丝绵的力学性能不受品种的影响。结果表明,微波对湿纺顺丰长丝力学性能的影响小于顺丰膜。
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引用次数: 2
Developmental characteristics of Zophobas atratus (Coleoptera: Tenebrionidae) larvae in different instars 白蛉(鞘翅目:拟甲科)不同龄期幼虫的发育特征
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.45
Sunyoung Kim, H. Kim, S. Song, N. Kim
The giant mealworm beetle, Zophobas atratus (Coleoptera: Tenebrionidae), is reared for commercial purposes as a live feed for animals. In 2011, it was officially introduced in Korea, and since then it has been considered commercially important. This beetle is a good animal feed resource because of its high protein content with balanced nutrients. However, its life stage characteristics have not been clearly described, especially of the larval stage that can be used as commercial products. To this end, in our study, we determined the number of Z. atratus larval instars, and described their characteristics at each stage, providing basic information about this beetle. Z. atratus larvae required eight to nine d of incubation period before hatching. The first instar period comprised three to four d. There were relatively large variations in each instar period, except for the first instar. Before the adults emerged, most of the individuals passed through15 to 18 instars. The highest pupation rate, 25.71%, was observed in both 16th and 17th instars. Body length gradually increased with each successive instar and it reached its maximum at the 18th instar. The color of larvae was white at the first instar, and gradually turned brown after the second instar.
巨型粉虫甲虫(鞘翅目:拟粉虫科)被作为动物的活饲料用于商业目的而饲养。2011年,它被正式引入韩国,从那时起,它就被认为具有重要的商业价值。这种甲虫蛋白质含量高,营养均衡,是一种很好的动物饲料资源。然而,其生命阶段特征尚未得到明确的描述,特别是可作为商业产品的幼虫阶段。为此,在我们的研究中,我们确定了白斑天牛幼虫的龄数,并描述了它们在每个阶段的特征,为白斑天牛提供了基本的信息。白纹夜蛾幼虫需要8 ~ 9天的潜伏期才能孵化。第一生育期为3 ~ 4 d,除第一生育期外,其余各生育期变化较大。在成虫出现之前,大多数个体经历了15到18个阶段。16、17龄的化蛹率最高,为25.71%。体长随龄期的增加而逐渐增加,在第18龄达到最大值。幼虫一龄时呈白色,二龄后逐渐转为褐色。
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引用次数: 19
Structure and properties of silk sericin obtained from different silkworm varieties 不同家蚕品种丝胶蛋白的结构和性质
Pub Date : 2015-06-30 DOI: 10.7852/IJIE.2015.30.2.81
D. Chung, Ji Hye Lee, H. Kweon, Kwang-gill Lee, I. Um
【Recently, many researchers have studied silk sericin because of its high water retention, good wound healing, good cyto-compatibility, and blood-glucose- and cholesterol-lowering effects. Although sericin film can be used in wound dressing and cosmetic packs, its poor mechanical properties have prevented its use in industrial fields. In the present study, sericin was obtained from different silkworm varieties, and the effect of silkworm variety on the structure and properties of sericin was examined. Except for a small difference in serine content, no significant difference in sericin was noted among the silkworm varieties. In addition, silkworm variety almost had no effect on solution viscosity, implying that it does not influence the molecular weight of sericin. Mechanical properties of sericin film were strongly affected by silkworm variety. Wonwon 126 showed the best mechanical properties, while N74 and Geumgwangju displayed the worst properties.】
【近年来,许多研究人员对丝胶蛋白进行了研究,因为它具有高保水性,良好的伤口愈合,良好的细胞相容性,以及降低血糖和胆固醇的作用。】丝胶薄膜虽可用于伤口敷料和化妆品包装,但其机械性能差,阻碍了其在工业领域的应用。本研究从不同家蚕品种中提取丝胶蛋白,考察了不同家蚕品种对丝胶蛋白结构和性能的影响。蚕品种间除丝氨酸含量差异不大外,丝氨酸含量差异不显著。此外,蚕种对溶液粘度几乎没有影响,这意味着它不影响丝胶蛋白的分子量。蚕品种对丝胶膜的力学性能影响较大。元126的力学性能最好,而N74和锦光州的力学性能最差。
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引用次数: 8
Effects of silk fibroin hydrolysate on bone metabolism in ovariectomized rats 丝素蛋白水解物对去卵巢大鼠骨代谢的影响
Pub Date : 2015-03-31 DOI: 10.7852/IJIE.2015.30.1.17
H. Kweon, S. Shin, J. Chon, Kwang-gill Lee, You-Young Jo, J. Yoon, Y. Park, J. Jeon, Jong-Ho Kim, Bong-Seob Shin
This study aimed to investigate the effects of silk fibroin on bone metabolism in ovariectomized rats. A total of 30 Sprague-Dawley rats were randomized into sham-operated (SHAM), ovariectomized control (OVX), alendronate (OVX+ALEN, 10 mg/kg body weight/d), low silk fibroin (OVX+SF100, 100 mg/kg body weight/d), and high silk fibroin (OVX+SF300, 300 mg/kg body weight/d) groups. All the rats were fed by gavage for 12 wk. At the end of 12 wk, blood and urine were collected for analysis of bone turnover markers, and bone mineral density (BMD) was measured by micro-computed tomography. The results show that the OVX group (p < 0.05) displayed the highest mean body weight gain. Among the five groups, serum levels of bone alkaline phosphatase (ALP) and urine levels of deoxypyridinoline (DPD) were highest in the OVX group (p < 0.05). Bone ALP levels in the ALEN group were significantly lower than that of the silk-treated groups. On the other hand, DPD levels were not significantly different between the ALEN and silk-fibroin-treated groups ( p < 0.05). The trabecular BMD was significantly higher in the ALEN and silk-treated groups compared to the OVX group ( p < 0.05). In conclusion, this study showed that silk fibroin has similar effects as alendronate, which is used in osteoporosis medication. Therefore silk fibroin might be a new candidate for the prevention and treatment of osteoporosis in patients.
本研究旨在探讨丝素蛋白对去卵巢大鼠骨代谢的影响。将30只sd大鼠随机分为假手术组(SHAM)、去卵巢对照组(OVX)、阿隆膦酸钠组(OVX+ALEN, 10 mg/kg体重/d)、低丝素组(OVX+SF100, 100 mg/kg体重/d)和高丝素组(OVX+SF300, 300 mg/kg体重/d)。各组大鼠灌食12周。12周结束时,采集血液和尿液分析骨转换标志物,并通过显微计算机断层扫描测量骨矿物质密度(BMD)。结果表明,OVX组平均增重最高(p < 0.05)。血清骨碱性磷酸酶(ALP)和尿脱氧吡啶啉(DPD)水平均以OVX组最高(p < 0.05)。ALEN组骨ALP水平显著低于蚕丝处理组。另一方面,ALEN与丝素处理组DPD水平无显著差异(p < 0.05)。与OVX组相比,ALEN和蚕丝处理组的骨小梁骨密度显著升高(p < 0.05)。综上所述,本研究表明丝素蛋白与阿仑膦酸钠具有类似的骨质疏松药物作用。因此,丝素蛋白可能成为预防和治疗骨质疏松症的新候选药物。
{"title":"Effects of silk fibroin hydrolysate on bone metabolism in ovariectomized rats","authors":"H. Kweon, S. Shin, J. Chon, Kwang-gill Lee, You-Young Jo, J. Yoon, Y. Park, J. Jeon, Jong-Ho Kim, Bong-Seob Shin","doi":"10.7852/IJIE.2015.30.1.17","DOIUrl":"https://doi.org/10.7852/IJIE.2015.30.1.17","url":null,"abstract":"This study aimed to investigate the effects of silk fibroin on bone metabolism in ovariectomized rats. A total of 30 Sprague-Dawley rats were randomized into sham-operated (SHAM), ovariectomized control (OVX), alendronate (OVX+ALEN, 10 mg/kg body weight/d), low silk fibroin (OVX+SF100, 100 mg/kg body weight/d), and high silk fibroin (OVX+SF300, 300 mg/kg body weight/d) groups. All the rats were fed by gavage for 12 wk. At the end of 12 wk, blood and urine were collected for analysis of bone turnover markers, and bone mineral density (BMD) was measured by micro-computed tomography. The results show that the OVX group (p < 0.05) displayed the highest mean body weight gain. Among the five groups, serum levels of bone alkaline phosphatase (ALP) and urine levels of deoxypyridinoline (DPD) were highest in the OVX group (p < 0.05). Bone ALP levels in the ALEN group were significantly lower than that of the silk-treated groups. On the other hand, DPD levels were not significantly different between the ALEN and silk-fibroin-treated groups ( p < 0.05). The trabecular BMD was significantly higher in the ALEN and silk-treated groups compared to the OVX group ( p < 0.05). In conclusion, this study showed that silk fibroin has similar effects as alendronate, which is used in osteoporosis medication. Therefore silk fibroin might be a new candidate for the prevention and treatment of osteoporosis in patients.","PeriodicalId":14140,"journal":{"name":"International journal of industrial entomology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2015-03-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82492825","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 2
Monitoring of phase separation between silk fibroin and sericin using various dye system 不同染料体系下丝素与丝胶相分离的监测
Pub Date : 2015-03-31 DOI: 10.7852/IJIE.2015.30.1.1
H. Kwak, K. Lee
Keywords: Silk, fibroin, sericin, dye, interface Understanding the interactions between fibroin and sericin is crucial in solving the mechanism of silk spinning. In this study, various commercially available dyes were used to monitor the interface between fibroin and sericin during the gelation of fibroin. The phase separation between fibroin and sericin could be observed by the addition of azo dyes over a certain molecular weight. Furthermore, the addition of the dyes to the sericin layer showed vivid phase separation over addition to the fibroin layer.
关键词:丝,丝素蛋白,丝胶蛋白,染料,界面,了解丝素蛋白与丝胶蛋白之间的相互作用是解决丝绸纺丝机理的关键。在本研究中,使用各种市售染料来监测丝蛋白凝胶化过程中丝蛋白和丝胶蛋白之间的界面。在一定分子量的情况下,偶氮染料的加入可以观察到丝素蛋白和丝胶蛋白的相分离。此外,在丝胶层中添加染料比在丝素层中添加染料表现出明显的相分离。
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引用次数: 3
Biology of a predatory bug Eocanthecona furcellata Wolff (Hemiptera : Pentatomidae) on Vapourer tussock moth larvae: a major pest of tasar silkworm food plants 一种掠食性昆虫(半翅目:五蝽科)对塔沙蚕食性植物主要害虫——塔沙蚕蛾幼虫的生物学研究
Pub Date : 2015-03-31 DOI: 10.7852/IJIE.2015.30.1.26
Aruna Ambadahalli Siddaiah, A. Shanmuga Sundaram Devi
Antheraea is polyphagous in A large number of pests are reported to both the host plant as well as silkworm. As the rearing is conducted outdoor silkworms are exposed to attack of various pests during their life cycle. Eocanthecona furcellata (Hemiptera: Pentatomidae) is the major predator of tasar silkworm during young age rearing. Both nymphs and adults of the stink bug suck the heamolymph of silkworm leading to death. The present work was undertaken to study the life cycle of E. furcellata on larvae of vapourer tussock moth which is a major pest of tasar host plants. The incubation period, nymphal duration and total developmental period from egg to adult was 8.0 ± 0.45 d, 16.0 ± 3.24 and 22.0 ± 4.20 d, respectively. Pre-oviposition, oviposition and longevity of male and female stink bug was 3.20 ± 0.38, 17.60 ± 1.86, 42.40 ± 1.94, 37.00 ± 3.18 d, respectively. Analysis of the data revealed no significant difference in life cycle of the predator when provided silkworm and vapourer tussock moth larvae as hosts. Which clearly indicates that E. furcellata can multiply on vapourer tussock moth larvae during non rearing periods. Adoption of recommended pest control measures for defoliators of tasar host plants during non-rearing periods will help in reducing the incidence of host plant pests and also in reducing the incidence of stink bug during rearing season. During the study it was also observed that larvae semilooper, boll headed caterpillar and nymphs of leaf hopper serve as alternate/secondary hosts to E. furcellata during non rearing seasons. on vapourer tussock moth larvae.
据报道,在寄主植物和家蚕中都有大量的害虫。由于蚕是在室外饲养的,在其生命周期中会受到各种害虫的侵害。棘棘蚕(Eocanthecona furcellata,半翅目:蝽科)是沙蚕幼龄期的主要捕食者。臭虫的若虫和成虫都吸食蚕的血淋巴而死亡。本文研究了糠皮蛾对柞蚕寄主植物的主要害虫——斑蛾幼虫的生活史。卵到成虫的潜伏期为8.0±0.45 d,若虫期为16.0±3.24 d,总发育期为22.0±4.20 d。雌雄臭虫产卵前、产卵期、寿命分别为3.20±0.38、17.60±1.86、42.40±1.94、37.00±3.18 d。数据分析显示,以家蚕和飞蛾幼虫为寄主时,捕食者的生命周期无显著差异。这清楚地表明,在非饲养期,糠皮蛾可以在斑蛾幼虫上繁殖。采用建议的防治措施,在非育成期防治tasar寄主植物的脱叶虫,有助减少寄主植物害虫的发生,以及在育成季节减少臭虫的发生。研究还发现,在非饲养季节,半熟幼虫、棉头毛虫和叶跳虫若虫是糠毛虫的交替/次生寄主。对飞蛾幼虫。
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引用次数: 3
Genetic characterization of microsporidians infecting Indian non-mulberry silkworms (Antheraea assamensis and Samia cynthia ricini) by using PCR based ISSR and RAPD markers assay. 基于ISSR和RAPD标记的印度非桑蚕(Antheraea assamensis和Samia cynthia ricini)微孢子虫遗传分析
Pub Date : 2015-03-31 DOI: 10.7852/IJIE.2015.30.1.6
W. Hassan, B. S. Nath
This study established the genetic characterisation of 10 microsporidian isolates infecting non-mulberry silkworms (Antheraea assamensis and Samia cynthia ricini) collected from biogeographical forest locations in the State of Assam, India, using PCR-based markers assays: inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD). A Nosema type species (NIK-1s_mys) was used as control for comparison. The shape of mature microsporidian spores were observed oval to elongated, measuring 3.80 to 4.90 µm in length and 2.60 to 3.05 µm in width. Fourteen ISSR primers generated reproducible profiles and yielded 178 fragments, of which 175 were polymorphic (98%), while 16 RAPD primers generated reproducible profiles with 198 amplified fragments displaying 95% of polymorphism. Estimation of genetic distance coefficients based on dice coefficients method and clustering with un-weighted pair group method using arithmetic average (UPGMA) analysis was done to unravel the genetic diversity of microsporidians infecting Indian muga and eri silkworm. The similarity coefficients varied from 0.385 to 0.941 in ISSR and 0.083 to 0.938 in RAPD data. UPGMA analysis generated dendrograms with two microsporidian groups, which appear to be different from each other. Based on Euclidean distance matrix method, 2-dimensional distribution also revealed considerable variability among different identified microsporidians. Clustering of these microsporidian isolates was in accordance with their host and biogeographic origin. Both techniques represent a useful and efficient tool for taxonomical grouping as well as for phylogenetic classification of different microsporidians in general and genotyping of these pathogens in particular.
本研究利用基于pcr的标记技术:简单序列重复(ISSR)和随机扩增多态性DNA (RAPD),建立了从印度阿萨姆邦生物地理森林采集的10株感染非桑蚕(Antheraea assamensis和Samia cynthia ricini)的微孢子虫分离株的遗传特征。以Nosema型种NIK-1s_mys为对照。成熟小孢子虫孢子的形状为卵圆形至细长形,长3.80 ~ 4.90µm,宽2.60 ~ 3.05µm。14条ISSR引物共扩增出178个片段,其中多态性175个(98%);16条RAPD引物共扩增出198个片段,多态性为95%。采用骰子系数法估算遗传距离系数,并采用UPGMA聚类方法分析印度蚕种和黑蚕微孢子虫的遗传多样性。ISSR相似系数为0.385 ~ 0.941,RAPD相似系数为0.083 ~ 0.938。UPGMA分析生成了两个微孢子虫组的树突图,它们似乎彼此不同。基于欧几里得距离矩阵法,不同鉴定的微孢子虫的二维分布也显示出较大的差异性。这些微孢子虫分离物的聚类与其宿主和生物地理来源一致。这两种技术都是对不同微孢子虫进行分类和系统发育分类的有效工具,特别是对这些病原体进行基因分型。
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引用次数: 1
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International journal of industrial entomology
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