Hostname: page-component-8448b6f56d-wq2xx Total loading time: 0 Render date: 2024-04-19T21:48:59.575Z Has data issue: false hasContentIssue false

Seed germination of Cistanche armena (Orobanchaceae), a rare endangered holoparasitic species endemic to Armenia

Published online by Cambridge University Press:  23 March 2023

Yuliya Krasylenko*
Department of Biotechnology, Faculty of Science, Palacký University Olomouc, Šlechtitelů 241/27, 783-71 Olomouc, Czech Republic
Adelá Hýlová
Department of Chemical Biology and Genetics, Faculty of Science, Palacký University, Center of the Region Haná for Biotechnological and Agricultural Research, Šlechtitelů 241/27, 783-71 Olomouc, Czech Republic
Yevhen Sosnovsky
Botanical Garden, Ivan Franko National University of Lviv, Cheremshyny St. 44, 79014 Lviv, Ukraine
Markéta Ulbrichová
Centre of the Region Haná for Biotechnological and Agricultural Research, Czech Advanced Technology and Research Institute (CATRIN), Palacký University Olomouc, Šlechtitelů 241/27, 783-71 Olomouc, Czech Republic
Lukáš Spíchal
Centre of the Region Haná for Biotechnological and Agricultural Research, Czech Advanced Technology and Research Institute (CATRIN), Palacký University Olomouc, Šlechtitelů 241/27, 783-71 Olomouc, Czech Republic
Renata Piwowarczyk
Center for Research and Conservation of Biodiversity, Institute of Biology, Jan Kochanowski University, 7 Uniwersytecka St., 25-406 Kielce, Poland
*Author for Correspondence: Yuliya Krasylenko, E-mail:


The obligate root parasite Cistanche armena is a recently rediscovered, extremely rare endangered species endemic to Armenia, specifically parasitizing camelthorn (Alhagi maurorum, Fabaceae) and saltwort (Salsola dendroides, Chenopodiaceae). Its populations are reputedly declining due to habitat destruction and biotic impacts. Since the only known means of its reproduction is via the seeds, understanding the mechanisms of breaking C. armena seed dormancy and germination along with the related aspects of the species’ biology is highly important both from fundamental (functional ecology and evolution) and applied (conservation and management) perspectives. Here, we present the first in vitro seed germination protocol for C. armena involving fluridone, a systemic herbicide targeting the carotenoid biosynthetic pathway. In addition, the seed micromorphology of C. armena is described using both light microscopy and lignin autofluorescence visualized by confocal laser scanning microscopy. The actin cytoskeleton in radicle cells of germinated C. armena seedlings is described for the first time, being the proof of seed viability. Further elaboration and application of the proposed germination protocol with the cultivation of C. armena on susceptible hosts are altogether seen as a valuable tool for the conservation of this species.

Research Paper
Copyright © The Author(s), 2023. Published by Cambridge University Press

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)


These authors contributed equally.


Ali-Rachedi, S, Bouinot, D, Wagner, MH, Bonnet, M, Sotta, B, Grappin, P and Jullien, M (2004) Changes in endogenous abscisic acid levels during dormancy release and maintenance of mature seeds: studies with the Cape Verde Islands ecotype, the dormant model of Arabidopsis thaliana. Planta 219, 479488.CrossRefGoogle ScholarPubMed
Bao, YZ, Yao, ZQ, Cao, XL, Peng, JF, Xu, Y, Chen, MX and Zhao, SF (2017) Transcriptome analysis of Phelipanche aegyptiaca seed germination mechanisms stimulated by fluridone, TIS108, and GR24. PLoS ONE 12, e0187539.10.1371/journal.pone.0187539CrossRefGoogle ScholarPubMed
Barua, D, Butler, C, Tisdale, TE and Donohue, K (2012) Natural variation in germination responses of Arabidopsis to seasonal cues and their associated physiological mechanisms. Annals of Botany 109, 209226.10.1093/aob/mcr264CrossRefGoogle ScholarPubMed
Baskin, JM and Baskin, CC (2022) Germination and seed/embryo size in holoparasitic flowering plants with ‘dust seeds’ and an undifferentiated embryo. The Botanical Review 88, 149.CrossRefGoogle Scholar
Bouwmeester, H, Li, C, Thiombiano, B, Rahimi, M and Dong, L (2021) Adaptation of the parasitic plant lifecycle: germination is controlled by essential host signaling molecules. Plant Physiology 185,12921308.CrossRefGoogle ScholarPubMed
Brun, G, Braem, L, Thoiron, S, Gevaert, K, Goormachtig, S and Delavault, P (2018). Seed germination in parasitic plants: what insights can we expect from strigolactone research? Journal of Experimental Botany 69, 22652280.CrossRefGoogle ScholarPubMed
Chae, SH, Yoneyama, K, Takeuchi, Y and Joel, DM (2004) Fluridone and norflurazon, carotenoid-biosynthesis inhibitors, promote seed conditioning and germination of the holoparasite Orobanche minor. Physiologia Plantarum 120, 328337.CrossRefGoogle ScholarPubMed
Chen, JS and Hsiao, SC (2011) Study on seed morphogenesis of Orobanchaceae in Taiwan. Taiwania 56, 267278.Google Scholar
Chen, QL, Wang, HL, Wang, ZF, Shan, CG, Zhai, ZX and Guo, YH (2009) Effects of cold stratification and exogenous gibberellic acid (GA3) on seed germination and contents of endogenous gibberellins (GAs) and abscisic acid (ABA) in Cistanche deserticola. Plant Physiology Communications 45, 270272.Google Scholar
Chen, QL, Jia, YM, Wang, ZF, Shan, CG, Zhu, JB and Guo, YH (2011) Postembryonic development of Cistanche tubulosa (Schrenk) Whigt. Pakistan Journal of Botany 43, 18231830.Google Scholar
Chen, YC, Li, M, Chen, XJ, Zhang, L and Song, YX (2012) Effects of exogenous signal substances on seed germination and haustorium formation of Cistanche deserticola. Plant Physiology Communications 48, 260264.Google Scholar
Chen, QL, Guo, Y, Jiang, Y and Tu, P (2016) Mechanism of fluridone-induced seed germination of Cistanche tubulosa. Pakistan Journal of Botany 48, 971976.Google Scholar
Das, M, Fernández-Aparicio, M, Yang, Z, Huang, K, Wickett, NJ, Alford, SR, Wafula, EC, dePamphilis, CW, Bouwmeester, H, Timko, M, Yoder, J and Westwood, JH (2015) Parasitic plants Striga and Phelipanche dependent upon exogenous strigolactones for germination have retained genes for strigolactone biosynthesis. American Journal of Plant Sciences 6, 11511166.CrossRefGoogle Scholar
Durlik, K, Żarnowiec, P, Piwowarczyk, R and Kaca, W (2021) Culturable endophytic bacteria from Phelipanche ramosa (Orobanchaceae) seeds. Seed Science Research 31, 6975. doi:10.1017/S0960258520000343CrossRefGoogle Scholar
Eriksson, O and Kainulainen, K (2011) The evolutionary ecology of dust seeds. Perspectives in Plant Ecology, Evolution and Systematics 13, 7387.CrossRefGoogle Scholar
Fitzpatrick, CR and Schneider, AC (2020) Unique bacterial assembly, composition, and interactions in a parasitic plant and its host. Journal of Experimental Botany 71, 21982209.CrossRefGoogle Scholar
Halouzka, R, Zeljkovic, SC, Klejdus, B and Tarkowski, P (2020) Analytical methods in strigolactone research. Plant Methods 16, 76.CrossRefGoogle ScholarPubMed
Hristeva, T, Dekalska, T and Denev, I (2013) Structural and functional biodiversity of microbial communities in the rhizosphere of plants infected with broomrapes (Orobanchaceae). Biotechnology & Biotechnological Equipment 27, 40824086.CrossRefGoogle Scholar
Huet, S, Pouvreau, J-B, Delage, E, Delgrange, S, Marais, C, Bahut, M, Delavault, P, Simier, P and Poulin, L (2020) Populations of the parasitic plant Phelipanche ramosa influence their seed microbiota. Frontiers in Plant Science 11, 1075.CrossRefGoogle ScholarPubMed
Iasur Kruh, L, Lahav, T, Abu-Nassar, J, Achdari, G, Salami, R, Freilich, S and Aly, R (2017) Host-parasite-bacteria triangle: the microbiome of the parasitic weed Phelipanche aegyptiaca and tomato-Solanum lycopersicum (Mill.) as a host. Frontiers in Plant Science 8, 269.10.3389/fpls.2017.00269CrossRefGoogle ScholarPubMed
Ito, S, Yamagami, D, Umehara, M, Hanada, A, Yoshida, S, Sasaki, Y, Yajima, S, Kyozukam, J, Ueguchi-Tanaka, M, Matsuoka, M, Shirasu, K, Yamaguchi, S and Asami, T (2017) Regulation of strigolactone biosynthesis by gibberellin signaling. Plant Physiology 174, 12501259.CrossRefGoogle ScholarPubMed
Jamil, M, Wang, JY, Yonli, D, Patil, RH, Riyazaddin, M, Gangashetty, P, Berqdar, L, Chen, G-TE, Traore, H, Margueritte, O, Zwanenburg, B, Bhoge, SE and Al-Babili, S (2022) A new formulation for strigolactone suicidal germination agents, towards successful Striga management. Plants 11, 808.CrossRefGoogle ScholarPubMed
Kaštier, P, Krasylenko, YA, Martinčová, M, Panteris, E, Šamaj, J and Blehová, A (2018) Cytoskeleton in the parasitic plant Cuscuta during germination and prehaustorium formation. Frontiers in Plant Science 9, 794.CrossRefGoogle ScholarPubMed
Kato, Y and Hisano, K (1983) In vitro culture of a root parasite, Aeginetia indica L. The Botanical Magazine – Shokubutsu-gaku-zasshi 96, 203209.CrossRefGoogle Scholar
Kato, Y, Inoue, T and Onishi, Y (1984) In vitro culture of a root parasite, Aeginetia indica L. II. The plane of cell division in the tendril. Plant and Cell Physiology 25, 981987.Google Scholar
Kusumoto, D, Chae, SH, Mukaida, K, Yoneyama, K, Yoneyama, K, Joel, DM and Takeuchi, Y (2006) Effects of fluridone and norflurazon on conditioning and germination of Striga asiatica seeds. Plant Growth Regulation 48, 7378.CrossRefGoogle Scholar
Li, TR, Xu, YY, Ge, JX and Xu, MY (1989) The seed germination of Cistanche deserticola Ma and it's relationship with host plant Haloxylon ammodendron Bunge. Acta Scientiarum Naturalium Universitatis Intramongolicae 20, 395400.Google Scholar
Li, Z, Lin, H, Gu, L, Gao, J and Tzeng, CM (2016) Herba Cistanche (Rou Cong-Rong): one of the best pharmaceutical gifts of traditional Chinese medicine. Frontiers in Pharmacology 7, 41.CrossRefGoogle ScholarPubMed
Liebst, B and Schneller, J (2008) Seed dormancy and germination behaviour in two Euphrasia species (Orobanchaceae) occurring in the Swiss Alps. Botanical Journal of the Linnean Society 156, 649656.CrossRefGoogle Scholar
Matusova, R, Mourik, T, and Bouwmeester, H (2004) Changes in the sensitivity of parasitic weed seeds to germination stimulants. Seed Science Research 14, 335344.CrossRefGoogle Scholar
Matusova, R, Kullacova, D and Toth, P (2014) Response of wild and weedy broomrapes to synthetic strigolactone analogue GR24. Journal of Central European Agriculture 14, 7282.CrossRefGoogle Scholar
Merckx, VSFT, Mennes, CB, Peay, KG and Geml, J (2013) Evolution and diversification, pp. 215244 in Merckx, VSFT (Ed.), Mycoheterotrophy: the biology of plants living on fungi. New York, Springer.CrossRefGoogle Scholar
Müller-Stöver, D, Nybroe, O, Baraibar, B, Loddo, D, Eizenberg, H, French, K and Christensen, S (2016) Contribution of the seed microbiome to weed management. Weed Research 56, 335339.CrossRefGoogle Scholar
Murashige, T and Skoog, F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum 15, 473497.CrossRefGoogle Scholar
Niu, DL, Song, YX, Guo, SH, Ma, HA, Li, M, Zheng, GQ, Gao, XY (2006) Original studies on dormancy and germination properties of Cistanche deserticola seeds. Seed 25, 1721.Google Scholar
Panteris, E, Apostolakos, P and Galatis, B (2006) Cytoskeletal asymmetry in Zea mays subsidiary cell mother cells: a monopolar prophase microtubule half-spindle anchors the nucleus to its polar position. Cell Motility and the Cytoskeleton 63, 696709.CrossRefGoogle Scholar
Petrosyan, K, Thijs, S, Piwowarczyk, R, Ruraż, K, Vangronsveld, J and Kaca, W (2022) Characterization and diversity of seed endophytic bacteria of the endemic holoparasitic plant Cistanche armena (Orobanchaceae) from a semi-desert area in Armenia. Seed Science Research 32, 264273.CrossRefGoogle Scholar
Piwowarczyk, R and Mielczarek, Ł (2018) First report of Eumerus mucidus (Diptera: Syrphidae) on Cistanche armena (Orobanchaceae) and from Armenia. Florida Entomologist 101, 519521.CrossRefGoogle Scholar
Piwowarczyk, R, Kwolek, D, Góralski, G, Denysenko, M, Joachimiak, AJ and Aleksanyan, A (2017) First report of the holoparasitic flowering plant Cistanche armena on Caspian manna (Alhagi maurorum) in Armenia. Plant Disease 101, 512.CrossRefGoogle Scholar
Piwowarczyk, R, Góralski, G, Denysenko-Bennett, M, Kwolek, D, Joachimiak, AJ and Fayvush, G (2018) The first report of eastern dodder (Cuscuta monogyna) parasitizing licorice (Glycyrrhiza glabra) in Armenia. Plant Disease 102, 2664.CrossRefGoogle Scholar
Piwowarczyk, R, Sanchez Pedraja, Ó, Moreno Moral, G, Fayvush, G, Zakaryan, N, Kartashyan, N, and Aleksanyan, A (2019) Holoparasitic Orobanchaceae (Cistanche, Diphelypaea, Orobanche, Phelipanche) in Armenia: distribution, habitats, host range and taxonomic problems. Phytotaxa 386, 101106.CrossRefGoogle Scholar
Piwowarczyk, R, Ochmian, I, Lachowicz, S, Kapusta, I, Sotek, Z and Błaszak, M (2020) Phytochemical parasite-host relations and interactions: a Cistanche armena case study. Science of the Total Environment 716, 137071.10.1016/j.scitotenv.2020.137071CrossRefGoogle ScholarPubMed
Pouvreau, JB, Gaudin, Z, Auger, B, Lechat, MM, Gauthier, M, Delavault, P and Simier, P (2013) A high-throughput seed germination assay for root parasitic plants. Plant Methods 9, 112.CrossRefGoogle ScholarPubMed
Qiao, XY, Wang, HL and Guo, YH (2007) Study on conditions of seed germination of Cistanche. Zhongguo Zhong Yao Za Zhi 32, 18481850.Google Scholar
Šamaj, J, Müller, J, Beck, M, Böhm, N and Menzel, D (2006) Vesicular trafficking, cytoskeleton and signaling in root hairs and pollen tubes. Trends in Plant Science 11, 594600.CrossRefGoogle ScholarPubMed
Schneider, CA, Rasband, WS and Eliceiri, KW (2012) NIH Image to ImageJ: 25 years of image analysis. Nature Methods 9, 671675.CrossRefGoogle ScholarPubMed
Smertenko, AP, Deeks, MJ and Hussey, PJ (2010) Strategies of actin reorganisation in plant cells. Journal of Cell Science 123(17), 30193028.CrossRefGoogle ScholarPubMed
Song, WJ, Cao, DD, Jin, ZL and Zhou, WJ (2005) Studies on factors influencing the seed germination of root parasitic plant Orobanche. Seed 24, 4447.Google Scholar
Suwa, H, Suzuki, Y, Zhang, YH, Murofushi, N and Takeuchi, Y (1995) Endogenous gibberellins in Aeginetia indica, a parasitic plant, and its host. Bioscience, Biotechnology, and Biochemistry 59, 17121715.CrossRefGoogle Scholar
Szymanski, D and Staiger, CJ (2018) The actin cytoskeleton: functional arrays for cytoplasmic organization and cell shape control. Plant Physiology 176, 106118.CrossRefGoogle ScholarPubMed
Wang, J, Baskin, JM, Baskin, CC, Liu, G, Yang, X and Huang, Z (2017) Seed dormancy and germination of the medicinal holoparasitic plant Cistanche deserticola from the cold desert of northwest China. Plant Physiology and Biochemistry 115, 279285.10.1016/j.plaphy.2017.04.010CrossRefGoogle ScholarPubMed
Xu, R, Chen, J, Chen, SL, Liu, TN, Zhu, WC and Xu, J (2009) Cistanche deserticola Ma cultivated as a new crop in China. Genetic Resources and Crop Evolution 56, 137142.CrossRefGoogle Scholar
Yang, CH, Zhang, XS and Zhuang, ZK (2007) Effect of acetylcholine on seed germination and endogenous IAA and ABA contents of Cistanche tubulosa. Journal of Plant Physiology 43, 295297.Google Scholar
Yoneyama, K, Xie, X, Sekimoto, H, Takeuchi, Y, Ogasawara, S, Akiyama, K, Hayashi, H and Yoneyama, K (2008) Strigolactones, host recognition signals for root parasitic plants and arbuscular mycorrhizal fungi, from Fabaceae plants. New Phytologist 179, 484494.CrossRefGoogle ScholarPubMed
Zhang, RM, Bai, J, , CL, Chen, HW and Gao, Y (2008) Fluctuating temperature stratification induced seed germination of Cistanche deserticola. Scientia Silvae Sinica 44, 170173.Google Scholar
Zhang, RM, Chen, HW, Zhang, D, Bai, J and Gao, Y (2009) Chemical induction on the seed germination and haustorium formation of Cistanche deserticola. Scientia Silvae Sinica 9, 170173.Google Scholar