西格斯贝基亚东方L.,属于菊科科,也被称为“Xi-XianCao”或HerbaSiegesbeckiae,自唐代以来一直是重要的中药(Wang等人。,2021)。由于干燥的地上部分具有药用价值,美国东方白杨在中国广泛种植,Japan,韩国,和越南。在洛南县发现了一个近600平方米的具有发育迟缓和叶片枯萎症状的东方链球菌植物(110.26E,34.06N),陕西省,2022年8月。在这些植物的根上观察到许多gall虫,第二阶段幼鱼(J2s)的密度为每100cm3土壤260〜370。从感染的根部解剖雌性和卵,并从土壤中提取J2s和雄性进行物种鉴定。女性的会阴模式(n=20)是椭圆形的,有小背弓,不同的横向场,和肛门周围的微小穿孔。雄性的头帽很高,明显比头部区域窄,头部区域从第一个身体环扩大。女性(n=20)的形态学测量为:体长(L)=897.66±50.89(860.96-949.74)μm,阀体宽度(BW)=577.69±51.01(489.91-638.65)μm,探针长度(ST)=14.03±0.63(13.25-14.97)μm,咽背腺孔至管心针基部(DGO)=4.96±0.47(4.08-5.37)μm,外阴狭缝长度=18.82±1.97(17.24-22.02)μm,外阴狭缝至肛门距离=13.62±1.22(12.34-16.18)μm。男性(n=10)的测量结果为:L=1298.73±95.96(1202.77-1394.69)μm,BW=28.24±2.38(25.93-30.55)μm,ST=20.23±0.78(19.42-21.04)μm,DGO=4.89±0.44(4.56-5.22)μm,针长=28.98±1.68(26.94-31.02)μm。J2s的测量值:L=375.35±14.02(341.01-400.46)μm,BW=15.09±1.47(12.02-16.82)μm,ST=12.74±0.61(11.46-13.84)μm,DGO=2.58±0.59(1.61-3.7)μm,尾长=74.15±13.73(50.92-95.09)μm,透明尾端=11.36±2.27(9.53-17.85)μm。这些形态特征与MeloidogynehaplaChitwood的形态特征一致,1949年,如Whitehead(1968)所述。使用蛋白酶K方法(Kumari和Subbotin,2012).扩增rDNA-ITS区的序列,并用引物rDNA-F/R(TTGATTACGCTGCCCCCTTT/TTTCACTCGCCGTACTACTAGG)进行测序(Vrain等人。,1992).768bp序列(GenBankOP542552)与hapla分枝杆菌的rDNA-ITS序列(JX024147和OQ269692)具有99.74%的同一性。然后扩增28SrRNA的D2/D3片段,并用引物D2A/D3B(ACAAGTACCGTGAGGAGGAAAGTTG/TCGGAAGGAACCAGCTACTA)(McClure等人。,2012).762bp的片段(OP554218)显示出与单峰分枝杆菌(MN752204和OM744204)的序列100%相同。为了确认人群的致病性,在无菌沙中培养的六个2周龄健康的东方链球菌幼苗分别接种2,000个从卵中孵化出来的J2s。四个未接种的幼苗用作阴性对照。在25°C下保持60天后,galls出现在接种植物的根部,与野外观察到的症状一致,而阴性对照没有症状。用物种特异性引物JWV1/JWV将从接种的植物中收集的雌性鉴定为M.hapla(Adam等人。,2007),扩增出440bp的片段。每株接种植物的平均回收率为3,814个J2s,繁殖因子为1.91,也证实了寄生现象。这是东方沙门氏菌是Hapla的宿主的第一份报告。该疾病降低了东方紫草的质量和产量,并将为其生产控制做出更多努力。
Siegesbeckia orientalis L., belonging to the family of Asteraceae and also known as \'Xi-Xian Cao\' or Herba Siegesbeckiae, has been an important traditional Chinese medicine since the Tang Dynasty (Wang et al., 2021). As the dried aerial parts have medicinal values, S. orientalis is widely grown in China, Japan, Korea, and Vietnam. One almost 600 m2 block of S. orientalis plants with stunting and leaf withering symptoms was found in Luonan County (110.26 E, 34.06 N), Shaanxi Province, in August 2022. Many galls were observed on the roots of these plants, and densities of second-stage juveniles (J2s) were 260~370 per 100 cm3 of soil. Females and eggs were dissected from infected roots, and J2s and males were extracted from the soil for species identification. The perineal patterns of females (n=20) were oval-shaped, with minor dorsal arches, distinct lateral fields, and tiny punctations around anus. The head caps of males were high and obviously narrower than head region which broadened out of the first body annuli. Morphological measurements of females (n=20) were: body length (L) = 897.66 ± 50.89 (860.96-949.74) μm, body width (BW) = 577.69 ± 51.01 (489.91-638.65) μm, stylet length (ST) = 14.03 ± 0.63 (13.25-14.97) μm, dorsal pharyngeal gland orifice to stylet base (DGO) = 4.96 ± 0.47 (4.08-5.37) μm, vulval slit length = 18.82 ± 1.97 (17.24-22.02) μm, vulval slit to anus distance = 13.62 ± 1.22 (12.34-16.18) μm. Measurements of males (n=10) were: L = 1298.73 ± 95.96 (1202.77-1394.69) μm, BW = 28.24 ± 2.38 (25.93-30.55) μm, ST = 20.23 ± 0.78 (19.42-21.04) μm, DGO = 4.89 ± 0.44 (4.56-5.22) μm, spicule length = 28.98 ± 1.68 (26.94-31.02) μm. Measurements of J2s: L = 375.35 ± 14.02 (341.01-400.46) μm, BW = 15.09 ± 1.47 (12.02-16.82) μm, ST = 12.74 ± 0.61(11.46-13.84) μm, DGO = 2.58 ± 0.59 (1.61-3.7) μm, tail length= 74.15 ± 13.73 (50.92-95.09) μm, hyaline tail terminus= 11.36 ± 2.27 (9.53-17.85) μm. These morphological characteristics were consistent with those of Meloidogyne hapla Chitwood, 1949 as described by Whitehead (1968). The DNA of single females (n=10) was isolated using the Proteinase K method for molecular identification (Kumari and Subbotin, 2012). The sequence of rDNA-ITS region was amplified and sequenced with the primers rDNA-F/R (TTGATTACGTCCCTGCCCTTT/TTTCACTCGCCGTTACTAAGG) (Vrain et al., 1992). The 768 bp sequence (GenBank OP542552) was 99.74% identical to the rDNA-ITS sequences of M. hapla (JX024147 and OQ269692). Then the D2/D3 fragments of the 28S rRNA were amplified and sequenced with the primers D2A/D3B (ACAAGTACCGTGAGGGAAAGTTG/TCGGAAGGAACCAGCTACTA) (McClure et al., 2012). The 762 bp fragment (OP554218) showed 100% identical to sequences of M. hapla (MN752204 and OM744204). To confirm the pathogenicity of the population, six 2-week-old healthy S. orientalis seedlings cultured in sterilized sand were each inoculated with 2,000 J2s hatched from egg masses. Four non-inoculated seedlings served as negative controls. After maintenance at 25°C for 60 days, galls appeared on the roots of inoculated plants, being consistent with the symptoms observed in field, while the negative controls showed no symptoms. Females collected from inoculated plants were identified as M. hapla with species-specific primer JWV1/ JWV (Adam et al., 2007), which amplified a fragment of 440 bp. Parasitism was also confirmed by the average recovery of 3,814 J2s per inoculated plant with the reproductive factor of 1.91. This is the first report of S. orientalis being a host of M. hapla. The disease reduces the quality and yield of S. orientalis, and much more efforts would be made for its control in production.