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首页> 美国卫生研究院文献>Journal of Nematology >First Report of the Spiral Nematode Helicotylenchus microlobus Infecting Soybean in North Dakota
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First Report of the Spiral Nematode Helicotylenchus microlobus Infecting Soybean in North Dakota

机译：螺旋线虫螺旋线虫感染北达科他州大豆的首次报道

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摘要

Spiral nematodes (Helicotylenchus spp.) are common plant-parasitic nematodes in fields of many crops. In June 2015, two soil samples were collected from a soybean field in Richland County, ND. Nematodes were extracted from soil using the sugar centrifugal flotation method (). Plant-parasitic nematodes were identified to genus based on morphological features and counted. Both samples contained spiral nematodes from 1,500 to 3,300 per kilogram of soil. In June and August 2016, 10 soil samples were collected from the same field. Nematodes were extracted, and nine of the samples had spiral nematodes ranging from 125 to 3,065 per kilogram of soil. One soil sample with 1,500 spiral nematodes per kilogram was used to inoculate two soybean cultivars Sheyenne and Barnes each in four replicates. After 15 wk of growth at 22°C in a greenhouse room, the population of spiral nematodes was found to have increased greatly. The final density was 9,300 ± 1,701 spiral nematodes per kilogram of soil for Sheyenne and 9,451 ± 2,751 for Barnes. The reproductive factor in Sheyenne and Barnes was 6.2 and 6.3, respectively, indicating that this spiral nematode infects and reproduces well on these two soybean cultivars. Infected soybean roots had small brown lesions on the surface. Individual spiral nematodes were handpicked and examined morphologically and molecularly for species identification. Morphological measurements of adult females (n = 15) included body length (mean = 708.5 μm, range = 600.0–812.0 μm), stylet (27.6, 26.0–29.0), body width (28.3, 25.0–33.0), lip region end to posterior end of pharyngeal glands (142.5, 130.0–152.0), anal body width (15.8, 14.0–17.5), tail length (20.3, 15.0–25.0), tail annules (11.6, 10.0–14.0), a (25.0, 21.4–27.1), b (5.0, 4.4–5.7), c (35.4, 30.2–41.7), c′ (1.3, 1.0–1.6), and V (61.8%, 60.0–63.7). The spiral nematode was identified as Helicotylenchus microlobus according to morphological and morphometric characteristics (). DNA was extracted from single nematodes (n = 8) using the Proteinase K method (). The internal transcribed spacer (ITS) region of rDNA was amplified with the primers rDNA2/rDNA1.58S (). The PCR products were then purified and sequenced. The consensus ITS rDNA sequence (accession no. , 822 bp) that was deposited into the GenBank shared 99% identity with two isolates of H. microlobus from California ( and ) and one isolate of H. microlobus from Spain () (). It had only 91% sequence identity with seven isolates of H. pseudorobustus (, , , , , , and ) from California, Switzerland, and New Zealand, a spiral nematode species very closely related to H. microlobus in morphology. The molecular tests confirmed the identity of this spiral nematode as H. microlobus. The H. microlobus nematode was reported as one of the most commonly observed spiral nematodes in soil samples in the state of Minnesota, and all 13 soybean cultivars tested except Hawkeye were rated as hosts (href="#B5" rid="B5" class=" bibr popnode">Taylor, 1960). To our knowledge, this is the first report of H. microlobus in North Dakota.
机译：螺旋线虫（Helicotylenchus spp。）是许多农作物田间常见的植物寄生线虫。 2015年6月，从北达科他州里奇兰县的一个大豆田采集了两个土壤样品。使用糖离心浮选法从土壤中提取线虫（）。根据形态特征将植物寄生线虫鉴定为属并计数。两种样品都含有每公斤土壤1,500至3,300的螺旋线虫。 2016年6月和2016年8月，从同一田地中采集了10个土壤样品。提取了线虫，其中九个样品的螺旋线虫的范围为每公斤土壤125至3,065。一个土壤样品每公斤1500个螺旋线虫用于接种两个大豆品种Sheyenne和Barnes，一式四份。在温室中于22°C下生长15周后，发现螺旋线虫的数量大大增加。对于Sheyenne，每千克土壤的最终密度为9,300±1,701螺旋线虫，对于Barnes，最终密度为9,451±2,751。 Sheyenne和Barnes的生殖因子分别为6.2和6.3，表明该螺旋线虫感染并在这两个大豆品种上繁殖良好。感染的大豆根表面有小的棕色病变。手工挑选单个螺旋线虫，并进行形态和分子检查以鉴定种类。成年雌性（n = 15）的形态学测量包括体长（平均值= 708.5μm，范围= 600.0–812.0μm），管心针（27.6，26.0–29.0），体宽（28.3，25.0–33.0），嘴唇区域至咽后部（142.5，130.0–152.0），肛门体宽（15.8，14.0–17.5），尾巴长度（20.3，15.0–25.0），尾环（11.6，10.0–14.0），a（25.0，21.4– 27.1），b（5.0、4.4-5.7），c（35.4、30.2-41.7），c'（1.3、1.0-1.6）和V（61.8％，60.0-63.7）。根据形态和形态特征，螺旋线虫被鉴定为微螺旋螺旋线虫。使用蛋白酶K方法从单个线虫（n = 8）中提取DNA。用引物rDNA2 / rDNA1.58S（）扩增了rDNA的内部转录间隔区（ITS）。然后纯化PCR产物并测序。存入GenBank的共有ITS rDNA序列（登录号，822 bp）与来自加利福尼亚（和）的两株微球菌和来自西班牙（）的一株微球菌具有99％的同一性。它与来自加利福尼亚州，瑞士和新西兰的七个假丝酵母嗜血杆菌的分离株仅有91％的序列同一性，螺旋形线虫物种与形态学上的亲缘关系非常密切。分子测试证实了该螺旋线虫的身份为H. microlobus。据报道，H。microlobus线虫是明尼苏达州土壤样品中最常见的螺旋线虫之一，除Hawkeye外，所有测试的13个大豆品种均被定为宿主（href =“＃B5” rid =“ B5 “ class =” bibr popnode“>泰勒，1960年）。据我们所知，这是 H 的第一个报告。北达科他州的 microlobus 。

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