Skip to main content

Enhanced resistance to soybean cyst nematode in transgenic soybean via host-induced silencing of vital Heterodera glycines genes

Abstract

Soybean cyst nematode (SCN, Heterodera glycines Ichinohe) is the most economically damaging pathogen affecting soybean production worldwide. Host-induced gene silencing provides a promising approach to confer resistance to plant parasitic nematodes. In the present study, we produced stable transgenic soybean plants individually harboring the inverted repeats of three essential H. glycines genes, Hg-rps23, Hg-snb1, and Hg-cpn1, and evaluated their resistance to SCN infection. Molecular characterization confirmed the stable integration of the hairpin double stranded (ds) RNA in host plants. Inoculation assays with SCN race 3 showed significant reduction of female index (FI, 11.84 ~ 17.47%) on the roots of T4 transgenic plants, with 73.29 ~ 81.90% reduction for the three RNA interference (RNAi) constructs, compared to non-transformed plants (NT, 65.43%). Enhanced resistance to SCN race 3 was further confirmed in subsequent generations (T5) of transgenic soybean. Moreover, when inoculated with SCN race 4 which was considered highly virulent to most of soybean germplasms and varieties, transgenic soybean plants also exhibited reduced FIs (9.96 ~ 23.67%) and increased resistance, relative to the NT plants (46.46%). Consistently, significant down-regulation in transcript levels of the Hg-rps23, Hg-snb1, Hg-cpn1 genes were observed in the nematodes feeding on the transgenic roots, suggesting a broad-spectrum resistance mediated by the host-mediated silencing of vital H. glycines genes. There were no significant differences in morphological traits between transgenic and NT soybean plants under conditions with negligible SCN infection. In summary, our results demonstrate the effectiveness of host-induced silencing of essential H. glycines genes to enhance broad-spectrum SCN resistance in stable transgenic soybean plants, without negative consequences on the agronomic performance.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  • Abad P, Gouzy J, Aury JM, Castagnone-Sereno P, Danchin EG, Deleury E, Perfus-Barbeoch L, Anthouard V, Artiguenave F, Blok VC, Caillaud MC, Coutinho PM, Dasilva C, De Luca F, Deau F, Esquibet M, Flutre T, Goldstone JV, Hamamouch N, Hewezi T, Jaillon O, Jubin C, Leonetti P, Magliano M, Maier TR, Markov GV, McVeigh P, Pesole G, Poulain J, Robinson-Rechavi M, Sallet E, Segurens B, Steinbach D, Tytgat T, Ugarte E, Van Ghelder E, Veronico P, Baum TJ, Blaxter M, Bleve-Zacheo T, Davis EL, Ewbank JJ, Favery B, Grenier E, Henrissat B, Jones JT, Laudet V, Maule AG, Quesneville H, Rosso MN, Schiex T, Smant G, Weissenbach J, Wincker P (2008) Genome sequence of the metazoan plant-parasitic nematode Mincognita. Nat Biotechnol 26:909–915

    CAS  Article  Google Scholar 

  • Alkharouf NW, Klink VP, Matthews BF (2007) Identification of Heterodera glycines (soybean cyst nematode [SCN]) cDNA sequences with high identity to those of Caenorhabditis elegans having lethal mutant or RNAi phenotypes. Exp Parasitol 115:247–258

    CAS  Article  Google Scholar 

  • Arelli PR, Sleper DA, Yue P, Wilcox JA (2000) Soybean reaction to races 1 and 2 of Heterodera glycines. Crop Sci 40:824–826

    Article  Google Scholar 

  • Concibido VC, Diers BW, Arelli PR (2004) A decade of QTL mapping for cyst nematode resistance in soybean. Crop Sci 44:1121–1131

    CAS  Article  Google Scholar 

  • Golden AM, Epps JM, Riggs RD, Duclos LA, Fox JA, Bernard RL (1970) Terminology and identity of infraspecific forms of the soybean cyst nematode (Heterodera glycines). Plant Disease Rep 54:544–546

    Google Scholar 

  • Gunadi A, Rushton PJ, Mchale LK, Gutek AH, Finer JJ (2016) Characterization of 40 soybean (Glycine max) promoters, isolated from across 5 thematic gene groups. Plant Cell Tissue Organ Cult 127:145–160

    CAS  Article  Google Scholar 

  • Guo X, Chronis D, De La Torre CM, Smeda J, Wang X, Mitchum MG (2015) Enhanced resistance to soybean cyst nematode Heterodera glycines in transgenic soybean by silencing putative CLE receptors. Plant Biotechnol J 13:801–810

    CAS  Article  Google Scholar 

  • Hershman DE, Heinz RD, Kennedy BS (2008) Soybean cyst nematode, Heterodera glycines, populations adapting to resistant soybean cultivars in Kentucky. Plant Dis 92:1475

    CAS  Article  Google Scholar 

  • Huang G, Allen R, Davis EL, Baum TJ, Hussey RS (2006) Engineering broad root-knot resistance in transgenic plants by RNAi silencing of a conserved and essential root-knot nematode parasitism gene. Proc Natl Acad Sci USA 103:14302–14306

    CAS  Article  Google Scholar 

  • Inagaki H, Tsutsumi M (1971) Survival of the soybean cyst nematode, Heterodera glycines Ichinohe (Tylenchida: Heteroderidae) under certain storage conditions. Appl Entomol Zool 6:156–162

    Article  Google Scholar 

  • Klink VP, Kim K-H, Martins VE, MacDonald MH, Beard HS, Alkharouf NW, Lee SK, Park S-C, Matthews BF (2009) A correlation between host-mediated expression of parasite genes as tandem inverted repeats and abrogation of the formation of female Heterodera glycines cysts during infection of Glycine max. Planta 230:53–71

    CAS  Article  Google Scholar 

  • Klink VP, Matsye PD, Lawrence GW (2011) Cell-specific studies of soybean resistance to its major pathogen, the soybean cyst nematode as revealed by laser capture microdissection, gene pathway analyses and functional studies. Soybean-Molecular Aspects of Breeding 397–428.

  • Koenning SR, Wrather JA (2010) Suppression of soybean yield potential in the continental United States by plant diseases from 2006 to 2009. Plant Health Progress. Vol. 11

  • Kumar A, Kakrana A, Sirohi A, Subramaniam K, Srinivasan R, Abdin MZ, Jain PK (2017) Host-delivered RNAi-mediated root-knot nematode resistance in Arabidopsis by targeting splicing factor and integrase genes. J Gen Plant Pathol 83:91–97

    CAS  Article  Google Scholar 

  • Li J, Todd TC, Oakley TR, Lee J, Trick HN (2010) Host derived suppression of nematode reproductive and fitness genes decreases fecundity of Heterodera glycines. Planta 232:775–785

    CAS  Article  Google Scholar 

  • Lin JY, Mazarei M, Zhao N, Zhu JJ, Zhuang XF, Liu WS, Pantalone VR, Arelli PR, Stewart CN, Chen F (2013) Overexpression of a soybean salicylic acid methyltransferase gene confers resistance to soybean cyst nematode. Plant Biotechnol J 11:1135–1145

    CAS  Article  Google Scholar 

  • Lin J, Mazarei M, Zhao N, Hatcher CN, Wuddineh WA, Rudis M, Tschaplinski TJ, Pantalone VR, Arelli PR, Hewezi T, Chen F, Stewart CN (2016) Transgenic soybean overexpressing GmSAMT1 exhibits resistance to multiple-HG types of soybean cyst nematode Heterodera glycines. Plant Biotechnol J 14:2100–2109

    CAS  Article  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25:402–408

    CAS  Article  Google Scholar 

  • Matsye PD, Lawrence GW, Youssef RM, Kim K-H, Lawrence KS, Matthews BF, Klink VP (2012) The expression of a naturally occurring, truncated allele of an α-SNAP gene suppresses plant parasitic nematode infection. Plant Mol Biol 80:131–155

    CAS  Article  Google Scholar 

  • McLean MD, Hoover GJ, Bancroft B, Makhmoudova A, Clark SM, Welacky T, Simmonds DH, Shelp BJ (2007) Identification of the full-length Hs1pro-1 coding sequence and preliminary evaluation of soybean cyst nematode resistance in soybean transformed with Hs1pro-1 cDNA. Can J Bot 85:437–441

    CAS  Article  Google Scholar 

  • Mcneece BT, Pant SR, Sharma K, Niruala P, Lawrence GW, Klink VP (2017) A Glycine max homolog of NON-RACE SPECIFIC DISEASE RESISTANCE 1 (NDR1) alters defense gene expression while functioning during a resistance response to different root pathogens in different genetic backgrounds. Plant Physiol Biochem 114:60–71

    CAS  Article  Google Scholar 

  • Niblack TL, Arelli PR, Noel GR, Opperman CH, Orf JH, Schmitt DP, Shannon JG, Tylka GL (2002) A revised classification scheme for genetically diverse populations of Heterodera glycines. J Nematol 34:279–288

    CAS  PubMed  PubMed Central  Google Scholar 

  • Noon JB, Hewezi T, Maier TR, Simmons C, Wei JZ, Wu G, Llaca V, Deschamps S, Davis EL, Mitchum MG, Hussey RS, Baum TJ (2015) Eighteen new candidate effectors of the phytonematode Heterodera glycines produced specifically in the secretory esophageal gland cells during parasitism. Phytopathology 105:1362–1372

    CAS  Article  Google Scholar 

  • Riggs RD, Schmitt DP (1991) Optimization of the Heterodera glycines race test procedure. J Nematol 23:149–154

    CAS  PubMed  PubMed Central  Google Scholar 

  • Sadia B, Domier LL, Biruk G, Gonfa B, Lakhssassi N, Meksem K, Lambert KN (2015) A SNARE-Like protein and biotin are complicated in soybean cyst nematode virulence. Plos One 10:e0145601

    Article  Google Scholar 

  • Sindhu AS, Maier TR, Mitchum MG, Hussey RS, Davis EL, Baum TJ (2009) Effective and specific in planta RNAi in cyst nematodes: expression interference of four parasitism genes reduces parasitic success. J Exp Bot 60:315–324

    CAS  Article  Google Scholar 

  • Srour A, Afzal AJ, Blahut-Beatty L, Hemmati N, Simmonds DH, Li W, Liu M, Town CD, Sharma H, Arelli P, Lightfoot DA (2012) The receptor like kinase at Rhg1-a /Rfs2 caused pleiotropic resistance to sudden death syndrome and soybean cyst nematode as a transgene by altering signaling responses. BMC Genomics 13:368–386

    CAS  Article  Google Scholar 

  • Steeves RM, Todd TC, Essig JS, Trick HN (2006) Transgenic soybeans expressing siRNAs specific to a major sperm protein gene suppress Heterodera glycines reproduction. Funct Plant Biol 33:991–999

    CAS  Article  Google Scholar 

  • Tel-Zur N, Abbo S, Myslabodski D, Mizrahi Y (1999) Modified CTAB procedure for DNA isolation from epiphytic cacti of the general hylocereus and selenicereus (cactaceae). Plant Mol Biol Report 17:249–254

    CAS  Article  Google Scholar 

  • Tian B, Li J, Vodkin LO, Todd TC, Finer JJ, Trick HN (2019) Host-derived gene silencing of parasite fitness genes improves resistance to soybean cyst nematodes in stable transgenic soybean. Theor Appl Genet 132:2651–2662

    Article  Google Scholar 

  • Urwin PE, Lilley CJ, Atkinson HJ (2002) Ingestion of double stranded RNA by pre-parasitic juvenile cyst nematodes leads to RNA interference. Mol Plant Microbe Interact 15:747–752

    CAS  Article  Google Scholar 

  • Wrather JA, Koenning SR (2006) Estimates of disease effects on soybean yields in the United States 2003–2005. J Nematol 38:173–180

    PubMed  PubMed Central  Google Scholar 

  • Yang X, Niu L, Zhang W, Yang J, Xing G, He H, Guo D, Du Q, Qian X, Yao Y, Li Q, Dong Y (2018) RNAi-mediated SMV P3 cistron silencing confers significantly enhanced resistance to multiple Potyvirus strains and isolates in transgenic soybean. Plant Cell Rep 37:103–114

    CAS  Article  Google Scholar 

  • Youssef RM, Kim KH, Haroon SA, Matthews BF (2013) Post-transcriptional gene silencing of the gene encoding aldolase from soybean cyst nematode by transformed soybean roots. Exp Parasitol 134:266–274

    CAS  Article  Google Scholar 

Download references

Acknowledgements

This research was supported by grants from Jilin Provincial Agricultural Science & Technology Innovation Project (KYJF2021ZR021, CXGC2021TD015), National Natural Science Foundation of China (31701448) and China National Novel Transgenic Organisms Breeding Project (2016ZX08004-004). We thank Editage (www.editage.cn) for English language editing.

Author information

Authors and Affiliations

Authors

Contributions

YX and ZX designed the experiments. ZY and ZQ conducted on molecular experiments and drafted the manuscript. NL, XG, and YJ conducted on Agrobacterium-mediated transformation. ZJ and LX conducted on SCN inoculation bioassay. All authors participated in the manuscript revision.

Corresponding authors

Correspondence to Xiaofang Zhong or Xiangdong Yang.

Ethics declarations

Conflict of interest

The authors claim no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Zhang, Y., Zhao, Q., Zhang, J. et al. Enhanced resistance to soybean cyst nematode in transgenic soybean via host-induced silencing of vital Heterodera glycines genes. Transgenic Res 31, 239–248 (2022). https://doi.org/10.1007/s11248-022-00298-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11248-022-00298-7

Keywords

  • Soybean cyst nematode
  • Heterodera glycines
  • Host-induced silencing