Welcome to Acta Agronomica Sinica,

Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (10): 2443-2450.doi: 10.3724/SP.J.1006.2022.14188

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Identification highly virulent population of soybean cyst nematode using China germplasms

LIAN Yun(), WEI He, WANG Jin-She, ZHANG Hui, LEI Chen-Fang, LI Jin-Ying, LU Wei-Guo()   

  1. Henan Academy of Crop Molecular Breeding / Zhengzhou Subcenter of National Soybean Improvement Center / Key Laboratory of Oil Crops in Huanghuaihai Plains, Ministry of Agriculture and Rural Affairs / Henan Provincial Key Laboratory for Oil Crops Improvement, Zhengzhou 450002, Henan, China
  • Received:2021-10-17 Accepted:2022-01-05 Online:2022-10-12 Published:2022-07-20
  • Contact: LU Wei-Guo E-mail:lianyun262@126.com;123bean@163.com
  • Supported by:
    National Key Research and Development Program of China(2017YFD0101400)

Abstract:

The disease caused by soybean cyst nematode (soybean cyst nematode, Heterodera glycines, SCN) is a worldwide soybean disease. The internationally used race identification models including Riggs model and Hg type for differentiate soybean cyst nematode population could not effectively distinguish the highly virulent race such as race 4, X12 population, and synthesized nematode population LY1. This study provides a simple identification method for distinguishing the three highly virulent population of the soybean cyst nematode, which would be greatly contribute to SCN research. This method included the following steps: First, we used the susceptible varieties and raised enough cysts in the soil infected highly virulent population of SCN for inoculation. Then, soybean varieties Xingxianhuipizhi (ZDD2315) and PI567516C were selected as differential hosts, and Lee was used as susceptible control for inoculation. Both ZDD2315 and PI567516C was sensitive in the inoculating reaction, indicating that the soil was infected by X12 population. If ZDD2315 was resistance while PI567516C were susceptible to the disease, indicating the soil was infected by race 4. If PI567516C was resistance to disease, it revealed that the soil was infected by LY1 population. These results indicated that the three highly virulent population including race 4, X12 population, and synthesized LY1 population could be effectively distinguished depending on the reaction of SCN on germplasms of ZDD2315 and PI567516C from China. In conclusion, these results of this study are of great significance for screening resistance germplasms, investigating the race distribution of SCN, and studying the pathogenic genes of SCN.

Key words: soybean cyst nematode, race, highly virulent, germplasms

Table 1

Identification Riggs differentials by inoculating X12 population and race 4, respectively"

寄主
Differential host
接种X12线虫群体 Inoculating X12 population 接种4号生理小种 Inoculating race 4 接种LY1线虫群体* Inoculating LY1 population*
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
每株上的平均胞囊数目
Average cysts number per plant
胞囊指数
FI
抗性反应
Reaction
Lee 185.4±26.9 (144-214) 330.6±62.8 (237-401) 100
Pickett 181.6±87.7 (52-264) 98 + 310.6±64.0 (251-413) 94 + /
Peking 95.2±91.5 (25-206) 51.3 + 292.0±21.3 (270-318) 88.3 + / 83 +
PI88788 177.6±94.1 (103-339) 95.8 + 290.0±93.8 (143-384) 87.7 + / 45 +
PI90763 93.0±47.9 (26-133) 50.2 + 99.6±21.0 (78-127) 30.1 + / 92 +

Table 2

Identification HG type differentials by inoculating X12 population and race 4, respectively"

寄主
Differential host
接种X12线虫群体Inoculating X12 population 接种4号生理小种 Inoculating race 4 接种LY1线虫群体* Inoculating LY1 population*
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
每株上的平均胞囊数目**
Average number of cysts per plant**
胞囊指数
FI
抗性反应 Reaction 每株上的平均胞囊数目
Average cysts number per plant
胞囊指数
FI
抗性反应
Reaction
Lee 185.4±26.9 (144-214) / 330.6±62.8 (237-401) / 100
Peking 95.2±91.5 (25-206) 51.3 + 292.0±21.3 (270-318) 88.3 + / 83 +
PI88788 177.6±94.1 (103-339) 95.8 + 290.0±93.8 (143-384) 87.7 + / 45 +
PI90763 93.0±47.9 (26-133) 50.2 + 99.6±21.0 (78-127) 30.1 + / 92 +
PI437654 51.2±23.1 (11-66) 27.6 + 23.6±6.4 (17-34) 7.1 - / 260 +
PI89772 179.5±51.5 (122-239) 96.8 + 54.0±5.7 (50-58) 16.3 + / 172 +
PI209332 235.0±46.7 (181-282) 126.8 + 421.2±86.8 (344-532) 127.4 + / 55 +
PI548316 224.8±49.2 (156-270) 121.2 + 251.8±98.6 (141-394) 76.2 + / 130 +

Table 3

Identification mode of distinguishing the population of soybean cyst nematode with strong pathogenicity using ZDD2315 and PI567516C"

寄主
Differential
接种X12线虫群体Inoculating X12 population 接种4号生理小种 Inoculating race 4 接种LY1线虫群体* Inoculating LY1 population*
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
每株上的平均胞囊数目**
Average cysts number per plant**
胞囊指数
FI
抗性反应
Reaction
Lee 185.4±26.9 (144-214) / 330.6±62.8 (237-401) / 100
ZDD2315 145.4±87.7 (52-264) 87.7 + 14.0±2.8 (12-16) 4.2 - /
PI567516C 225±51.5 (122-239) 121.4 + 80.2±32.8 (53-135) 24.2 + 7 -

Table 4

Verify race 4, race 2, and X12 population using multiple soil samples with ZDD2315 and PI567516C"

生理小种
Race
生理小种鉴定Race testing 抗性反应Reaction 土样编号
Soil code
土样来源 Origin of the soil 土样采集时间
Soil sample collecting time (month/day)
土样采集地点
Soil sample collecting location
Lee* PI567516C ZDD2315 经度
Longitude (°)
纬度
Latitude (°)
X12 185.4±26.9 (144-214) + + SX12 112.166 37.844 2012/7 山西省邢家社乡想儿岭村
Xiangerling village, Xingjiashe township, Shanxi province
X12 118.4±28.7 (62-158) + + GJ30 112.257 38.014 2019/8 山西省河口镇段家岭村
Duanjialing village, Hekou town, Shanxi province
X12 135±38.7 (85-176) + + GJ07 112.166 37.844 2019/8 山西省邢家社乡想儿岭村
Xiangerling village, Xingjiashe township, Shanxi province
X12 222.9±36.2 (159-265) + + SX12单胞囊繁殖
SX12 Raised from single cyst
112.166 37.844 2016/1 河南省农业科学院培育
Cultivated from Henan Academy of Agricultural Sciences
Race 4 122.4±23.2 (97-154) + - GJ05 112.190 37.758 2019/8 山西省邢家社乡郭家社村
Guojiashe village, Xingjiashe township, Shanxi province
Race 4 156.3±33.1 (118-192) + - GJ21 111.967 37.846 2019/8 山西省马兰镇南龙沟村
Nanlonggou village, Malan town, Shanxi province
Race 4 189±42.5 (129-229) + - GJ27 112.218 37.998 2019/8 山西省河口镇大坡村
Dapo village, Hekou town, Shanxi province
Race 4 110.4±21 (78-143) + - GJ14 112.070 37.876 2019/8 山西省马兰镇姬家庄村
Jijiazhuang village, Malan town, Shanxi province
Race 4 134±62.2 (45-260) + - GJ28 112.232 38.017 2019/8 山西省河口镇水头村
Shuitou village, Hekou town, Shanxi province
Race 4 124.6±26.1 (87-165) + - SX37 113.232 38.477 2012/7 山西省忻州市定襄县南庄村
Nanzhuang village, Dingxiang county, Xinzhou city, Shanxi province
Race 4 168±26.6 (135-212) + - SX49 112.802 39.484 2012/7 山西省朔州市山阴县安荣乡
An Rongxiang, Shanyin county, Shuozhou city, Shanxi province
Race 4 220±42.8 (138-270) + - SX51 112.468 39.511 2012/7 山西省朔州市平鲁区李西沟村
Lixigou village, Pinglu district, Shuozhou city, Shanxi province
Race 4 187.1±33.9 (130-222) + - SX59 112.613 36.935 2012/7 山西省长治市沁县西汤村
Xitang village, Qin county, Changzhi city, Shanxi province
Race 4 137.1±36 (90-196) + - SX37 113.232 38.477 2012/7 山西省忻州市定襄县南庄村
Nanzhuang village, Dingxiang county, Xinzhou city, Shanxi province
Race 2 170.4±50.6 (106-252) + - GJ 08 111.960 37.761 2019/8 山西省常安乡小娄峰村
Xiao Loufeng village, Chang'an township, Shanxi province
Race 2 95±32 (43-127) + - GJ 02 112.161 37.860 2019/8 山西省桃园办事处李家社村
Lijiashe village, Taoyuan office, Shanxi province
Race 2 659±283.5 (297-1138) + - GJ 04 112.188 37.785 2019/8 山西省邢家社乡龙子村
Longzi village, Xingjiashe township, Shanxi province
Race 2 286±80.5 (164-397) + - GJ 12 112.05 37.816 2019/8 山西省常安乡麻家口村
Majiakou village, Chang'an township, Shanxi province
Race 2 242.3±113 (74-454) + - GJ 16 111.924 37.861 2019/8 山西省岔口乡板四沟村
Bansigou village, Chakou township, Shanxi province
Race 2 112.3±44.7 (41-166) + - GJ 21 111.967 37.846 2019/8 山西省马兰镇南龙沟村
Nanlonggou village, Malan town, Shanxi province
[1] Bradley C A, Allen T, Sisson A J, Bergstrom G C, Wise K. Soybean yield loss estimates due to diseases in the United States and Ontario, Canada from 2015-2019. Plant Health Prog, 2021, 22: 483-495.
doi: 10.1094/PHP-01-21-0013-RS
[2] Woo M O, Beard H, MacDonald M H, Brewer E P, Youssef R M, Kim H, Matthews B F. Manipulation of two α-endo-β-1,4- glucanase genes, AtCel6 and GmCel7, reduces susceptibility to Heterodera glycines in soybean roots. Mol Plant Pathol, 2014, 15: 927-939.
doi: 10.1111/mpp.12157
[3] Koenning S R, Wrather J A. Suppression of soybean yield potential in the continental United States by plant diseases from 2006 to 2009. Plant Health Prog, 2010, 11. doi: 10.1094/PHP-2010- 1122-01-RS.
[4] Kim K S, Vuong T D, Qiu D, Robbins R T, Shannon J G, Li Z, Nguyen H T. Advancements in breeding, genetics, and genomics for resistance to three nematode species in soybean. Theor Appl Genet, 2016, 129: 2295-2311.
pmid: 27796432
[5] Tylka G L, Marett C C. Distribution of the soybean cyst nematode, Heterodera glycines, in the United States and Canada: 1954 to 2014. Plant Health Prog, 2014, 15: 85-87.
[6] 练云, 王金社, 李海朝, 魏荷, 李金英, 武永康, 雷晨芳, 张辉, 王树峰, 郭建秋, 李月霞, 李志辉, 靳巧玲, 徐淑霞, 张志民, 杨彩云, 于会勇, 耿臻, 舒文涛, 卢为国. 黄淮大豆主产区大豆胞囊线虫生理小种分布调查. 作物学报, 2016, 42: 1479-1486.
doi: 10.3724/SP.J.1006.2016.01479
Lian Y, Wang J S, Li H C, Wei H, Li J Y, Wu Y K, Lei C F, Zhang H, Wang S F, Guo J Q, Li Y X, Li Z H, Jin Q L, Xu S X, Zhang Z M, Yang C Y, Yu H Y, Geng Z, Shu W T, Lu W G. Race distribution of soybean cyst nematode in the main soybean producing area of Huang-Huai Rivers Valley. Acta Agron Sin, 2016, 42: 1479-1486. (in Chinese with English abstract)
doi: 10.3724/SP.J.1006.2016.01479
[7] Shaibu A S, Li B, Zhang S R, Sun J M. Soybean cyst nematode-resistance: gene identification and breeding strategies. Crop J, 2020, 8: 892-904.
doi: 10.1016/j.cj.2020.03.001
[8] Riggs R D, Schmitt D P. Complete characterization of the race scheme for Heterodera glycines. J Nematol, 1988, 20: 392-395.
pmid: 19290228
[9] Niblack T L, Arelli P R, Noel G R, Opperman C H, Orf J H. A revised classification scheme for genetically diverse populations of Heterodera glycines. J Nematol, 2002, 34: 279-288.
pmid: 19265945
[10] Lu W G, Gai J Y, Li W D. Sampling survey and identification of races of soybean cyst nematode (Heterodera glycines Ichinohe) in Huang-Huai valleys. Agric Sci China, 2006, 5: 615-621.
doi: 10.1016/S1671-2927(06)60101-6
[11] Howland A, Monnig N, Mathesius J, Nathan M, Mitchum M G. Survey of Heterodera glycines population densities and virulence phenotypes during 2015-2016 in Missouri. Plant Dis, 2018, 102: 2407-2410.
doi: 10.1094/PDIS-04-18-0650-SR
[12] Mitchum M G. Soybean resistance to the soybean cyst nematode Heterodera glycines: an update. Phytopathology, 2016, 106: 1444-1450.
doi: 10.1094/PHYTO-06-16-0227-RVW
[13] Mitchum M G, Wrather J. Variability in distribution and virulence phenotypes of Heterodera glycines in Missouri during 2005. Plant Dis, 2007, 91.
[14] Lian Y, Guo J Q, Li H C, Wu Y K, Wei H, Wang J S, Li J Y, Lu W G. A new race (X12) of soybean cyst nematode in China. J Nematol, 2017, 49: 321-326.
pmid: 29062156
[15] 大豆种质抗胞囊线虫鉴定协作组. 大豆种质资源对大豆胞囊线虫1、3和4号生理小种的抗性鉴定. 大豆科学, 1993, 12: 91-99.
Coordinative Group of Evaluation of SCN. Evaluation of soybean germplasm for resistance to race 1, 3 and 4 of the soybean cyst nematode. Soybean Sci, 1993, 12: 91-99. (in Chinese with English abstract)
[16] Lian Y, Wei H, Wang J S, Lei C F, Li H C, Li J Y, Wu Y K, Wang S F, Zhang H, Wang T F, Du P, Guo J Q, Lu W G. Chromosome-level reference genome of X12, a highly virulent race of the soybean cyst nematode Heterodera glycines. Mol Ecol Resour, 2019, 19: 1637-1646.
doi: 10.1111/1755-0998.13068 pmid: 31339217
[17] Arelli P R, Young L D, Concibido V C. Inheritance of resistance in soybean PI 567516C to LY1 nematode population infecting cv. Hartwig. Euphytica, 2009, 165: 1-4.
doi: 10.1007/s10681-008-9760-z
[18] Zhou L J, Song L, Lian Y, Ye H, Usovsky M, Wan J R, Vuong T D, Nguyen H T. Genetic characterization of qSCN10 from an exotic soybean accession PI 567516C reveals a novel source conferring broad-spectrum resistance to soybean cyst nematode. Theor Appl Genet, 2021, 134: 859-874.
doi: 10.1007/s00122-020-03736-4
[19] Usovsky M, Ye H, Vuong T D, Patil G B, Nguyen H T. Fine-mapping and characterization of qSCN18, a novel QTL controlling soybean cyst nematode resistance in PI 567516C. Theor Appl Genet, 2021, 134: 621-631.
doi: 10.1007/s00122-020-03718-6
[20] Vuong T D, Sleper D A, Shannon J G, Nguyen H T. Novel quantitative trait loci for broad-based resistance to soybean cyst nematode (Heterodera glycines Ichinohe) in soybean PI 567516C. Theor Appl Genet, 2010, 121: 1253-1266.
doi: 10.1007/s00122-010-1385-7 pmid: 20559815
[21] Arelli P R, Concibido V C, Young L D. QTLs associated with resistance in soybean PI567516C to synthetic nematode population infecting cv. Hartwig. J Crop Sci, 2010, 13: 163-167.
[22] Wang D, Duan Y X, Wang Y Y, Zhu X F, Chen L J, Liu X Y, Chen J S. First report of soybean cyst nematode, Heterodera glycines, on soybean from Guangxi, Guizhou, and Jiangxi provinces, China. Plant Dis, 2015, 99: 893.
[23] Peng D, Peng H, Wu D Q, Huang W, Cui J K. First report of soybean cyst nematode (Heterodera glycines) on soybean from Gansu and Ningxia China. Plant Dis, 2015, 100: 150727061835002. doi: 10.1094/PDIS-04-15-0451-PDN.
doi: 10.1094/PDIS-04-15-0451-PDN
[24] Shi H, Zheng H. First report of soybean cyst nematode (Heterodera glycines) on tobacco in Henan, central China. Plant Dis, 2013, 97: 852.
[25] Hua C, Li C J, Hu Y F, Mao Y Z, You J, Wang M Z, Tian Z Y, Chen J S, Wang C L. Identification of HG types of soybean cyst nematode Heterodera glycines and resistance screening on soybean genotypes in northeast China. J Nematol, 2018, 50: 41-50.
doi: 10.21307/jofnem-2018-007
[26] Chen J, Zhou Y, Wang Y, Fan H, Liu X, Zhao D, Wang D, Duan Y, Zhu X, Chen L. Characterization of virulence phenotypes of Heterodera glycines in Heilongjiang, northeast China. Plant Dis, 2021, 105: 2056-2060.
doi: 10.1094/PDIS-04-20-0820-SR
[27] Jiao Y, Vuong T D, Yang L, Li Z, Noe J, Robbins R T, Joshi T, Dong X, Shannon J G, Nguyen H T. Identification of quantitative trait loci underlying resistance to southern root-knot and reniform nematodes in soybean accession PI 567516C. Mol Breed, 2015, 35: 131.
[1] Zuo Tong-Hong, Zhang He-Cui, Zeng Jing, Zhu Li-Quan. Molecular cloning and expression analysis of BoPUB3L associated with self-incompatibility in Brasscia oleracea [J]. Acta Agronomica Sinica, 2026, 52(6): 1698-1710.
[2] Hou Jie, Fu Duo-Duo, Wu Hai-Feng, Hao Yu-Qiong, Zheng Xing-Wei, Wu Bang-Bang, Zhou Kai, Li Xiao-Hua, Zheng Jun, Zhao Jia-Jia. Chromosome diversity and its effects in wheat landraces from Shanxi province, China [J]. Acta Agronomica Sinica, 2026, 52(3): 746-763.
[3] Ma Ting-Ting, Guo Xiao-Jiang, Li Hao, Deng Mei, Pu Zhi-En, Li Wei, Zhang Ya-Zhou, Wang Feng-Tao, Cui Feng-Juan, Wei Yu-Ming, Wang Ji-Rui, Jiang Yun-Feng, Chen Guo-Yue. Breeding strategy for synergistic improvement of yield, disease resistance, and stress tolerance in Shumai 753 using the wheat landrace Xiaoganmai [J]. Acta Agronomica Sinica, 2026, 52(1): 56-71.
[4] WEN Xuan, ZHONG Xiu-Li, WANG Shang-Wen, JIN Tao, PENG Jun, LIU En-Ke. Screening of low nitrogen tolerant germplasm in seedling highland barley based on tolerance index and comprehensive evaluation of different nitrogen efficiency types [J]. Acta Agronomica Sinica, 2025, 51(7): 1949-1958.
[5] WANG Tian-Yi, YANG Xiu-Juan, ZHAO Jia-Jia, HAO Yu-Qiong, ZHENG Xing-Wei, WU Bang-Bang, LI Xiao-Hua, HAO Shui-Yuan, ZHENG Jun. Gliadin diversity and its effects on flour quality in wheat from Shanxi province, China [J]. Acta Agronomica Sinica, 2025, 51(7): 1784-1800.
[6] SHEN Wen-Jie, CHEN Qing-Qing, HU Yi-Qun, ZHANG Ai-Fang, ZHANG Man-Yu. Study on the sensitivity of various physiological races of Magnaporthe oryzae to isoprothiolane in Anhui province, China [J]. Acta Agronomica Sinica, 2025, 51(5): 1338-1346.
[7] ZHAN Zong-Bing, JIN Qi-Feng, LIU Di, LYU Ying-Chun, GUO Ying, ZHANG Xue-Ting, HU Meng-Xia, WANG Shang, YANG Fang-Ping. Molecular characterization and evaluation of important traits of landrace wheat Laomangmai in Gansu province, China [J]. Acta Agronomica Sinica, 2025, 51(3): 609-620.
[8] YANG Fang-Ping, GUO Ying, TIAN Yuan-Yuan, XU Yu-Feng, WANG Lan-Lan, BAI Bin, ZHAN Zong-Bing, ZHANG Xue-Ting, XU Yin-Ping, LIU Jin-Dong. Effect of vernalization and photoperiod genes and evaluation of cold tolerance for wheat landraces from Gansu province, China [J]. Acta Agronomica Sinica, 2025, 51(2): 370-382.
[9] XU Xiao-Wei, FENG Jing, WANG Feng-Tao, TONG Zhao-Yang, ZHANG Jian-Zhou, LI Chun-Ying, LIN Rui-Ming. QTL mapping of adult plant resistance to stripe rust in the Chinese wheat landrace Canlaomai [J]. Acta Agronomica Sinica, 2025, 51(11): 2933-2943.
[10] HUANG Lin-Yu, ZHANG Xiao-Yue, LI Hao, DENG Mei, KANG Hou-Yang, WEI Yu-Ming, WANG Ji-Rui, JIANG Yun-Feng, CHEN Guo-Yue. Mapping of QTL for adult plant stripe rust resistance genes in a Sichuan wheat landrace and the evaluation of their breeding effects [J]. Acta Agronomica Sinica, 2024, 50(9): 2167-2178.
[11] KE Hui-Feng, SU Hong-Mei, SUN Zheng-Wen, GU Qi-Shen, YANG Jun, WANG Guo-Ning, XU Dong-Yong, WANG Hong-Zhe, WU Li-Qiang, ZHANG Yan, ZHANG Gui-Yin, MA Zhi-Ying, WANG Xing-Fen. Identification for yield and fiber quality traits and evaluation of molecular markers in modern cotton varieties [J]. Acta Agronomica Sinica, 2024, 50(2): 280-293.
[12] YAO Zhu-Fang, ZHANG Xiong-Jian, YANG Yi-Ling, HUANG Li-Fei, CHEN Xin-Liang, YAO Xiao-Jian, LUO Zhong-Xia, CHEN Jing-Yi, WANG Zhang-Ying, FANG Bo-Ping. Genetic diversity of phenotypic traits in 177 sweetpotato landrace [J]. Acta Agronomica Sinica, 2022, 48(9): 2228-2241.
[13] ZHOU Qun, YUAN Rui, ZHU Kuan-Yu, WANG Zhi-Qin, YANG Jian-Chang. Characteristics of grain yield and nitrogen absorption and utilization of indica/japonica hybrid rice Yongyou 2640 under different nitrogen application rates [J]. Acta Agronomica Sinica, 2022, 48(9): 2285-2299.
[14] QIN Lu, HAN Pei-Pei, CHANG Hai-Bin, GU Chi-Ming, HUANG Wei, LI Yin-Shui, LIAO Xiang-Sheng, XIE Li-Hua, LIAO Xing. Screening of rapeseed germplasms with low nitrogen tolerance and the evaluation of its potential application as green manure [J]. Acta Agronomica Sinica, 2022, 48(6): 1488-1501.
[15] ZHANG Yi-Zhong, ZENG Wen-Yi, DENG Lin-Qiong, ZHANG He-Cui, LIU Qian-Ying, ZUO Tong-Hong, XIE Qin-Qin, HU Deng-Ke, YUAN Chong-Mo, LIAN Xiao-Ping, ZHU Li-Quan. Codon usage bias analysis of S-locus genes SRK, SLG, and SP11/SCR in Brassica oleracea [J]. Acta Agronomica Sinica, 2022, 48(5): 1152-1168.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
No Suggested Reading articles found!