Plant Tissue Culture Strategies for Sustainable Conservation of Zanthoxylum Zanthoxyloides (Lam) Zepern & Timler in Nigeria: A Systematic Review

Zanthoxylum zanthoxyloides (Lam.) Zepern. & Timler (Rutaceae), commonly known as the African prickly ash or Senegal prickly ash, is one of the most ethnobotanically and pharmacologically important medicinal trees of sub-Saharan Africa. In Nigeria, it is used in various areas of therapy, including antimicrobial, analgesic, antipyretic, and anticancer applications, and forms part of the primary healthcare of many traditional communities. Although the species has significant socioeconomic and ecological importance, its populations are declining at an accelerating rate due to uncontrolled harvesting, substantial habitat fragmentation, forest clearance, and the recalcitrant nature of its seeds, which renders conventional ex situ seed banking largely ineffective. Plant tissue culture (PTC) provides a scientifically sound, scalable, and cost-effective platform to overcome these conservation deficits. This review systematically assesses the entire range of in vitro methods applicable to the conservation of Z. zanthoxyloides, including seed sterilisation and germination stimulation, callus induction and organogenesis, somatic embryogenesis, micropropagation via axillary shoot proliferation, synthetic seed technology, and cryopreservation—with specific reference to vitrification and encapsulation-dehydration protocols. The review critically evaluates published literature on the genus Zanthoxylum and the broader family Rutaceae, identifies important species-specific knowledge gaps, examines the infrastructural and regulatory environment for implementation in Nigeria, and proposes a prioritised ten-year research agenda. It is argued that long-term conservation of this species requires the integration of in vitro procedures with molecular characterisation, community engagement, and enabling policy reform—an approach that is both scientifically feasible and a national imperative.

cryopreservation; micropropagation; recalcitrant seeds; Rutaceae; somatic embryogenesis; traditional medicine; ex situ conservation; Nigeria

1. Abubakar, B. Y., Adebayo, M. A., & Oyelaran, O. A. (2019). Callus induction and plant regeneration from cotyledon explants of Zanthoxylum zanthoxyloides (Lam.) Zepern. & Timler. African Journal of Biotechnology, 18(12), 247–256. https://doi.org/10.5897/AJB2018.16631 [Google Scholar] [Crossref]

2. Agbo, M. O., Ukwueze, N. N., & Ezea, S. C. (2013). Phytochemistry and toxicological evaluation of aqueous and methanolic extracts of Zanthoxylum zanthoxyloides root bark on liver function indices of male Wistar rats. Journal of Chemical and Pharmaceutical Research, 5(1), 279–284. [Google Scholar] [Crossref]

3. Akinrinola, A. O., & Olawale, F. (2022). Ethnobotanical documentation of Zanthoxylum zanthoxyloides use in southwestern Nigeria: Implications for conservation planning. Journal of Ethnobiology and Ethnomedicine, 18(1), 34. https://doi.org/10.1186/s13002-022-00524-3 [Google Scholar] [Crossref]

4. Aker, S., & Mukherjee, P. K. (2020). Nanomaterials in plant tissue culture: Mechanisms of action and applications in plant biotechnology. Plant Cell, Tissue and Organ Culture, 141(2), 225–244. https://doi.org/10.1007/s11240-020-01762-0 [Google Scholar] [Crossref]

5. Al-Khayri, J. M., Jain, S. M., & Johnson, D. V. (Eds.). (2019). Advances in plant breeding strategies: Breeding, biotechnology and molecular tools. Springer. [Google Scholar] [Crossref]

6. Amoo, S. O., Aremu, A. O., & Van Staden, J. (2014). Unraveling the medicinal potential of South African Aloe species. Journal of Ethnopharmacology, 153(1), 19–41. https://doi.org/10.1016/j.jep.2014.01.039 [Google Scholar] [Crossref]

7. Andrade, J. C., Lima, E., Gomes-Carneiro, M. R., Bhatt, D. L., & Moura, M. D. (2021). Bioactive phytochemicals from Zanthoxylum genus: A review of traditional uses and recent advances in pharmacological investigations. Phytochemistry Reviews, 20(4), 877–918. https://doi.org/10.1007/s11101-021-09738-2 [Google Scholar] [Crossref]

8. Ashmore, S. E. (1997). Status report on the development and application of in vitro techniques for the conservation and use of plant genetic resources. International Plant Genetic Resources Institute. [Google Scholar] [Crossref]

9. Bajaj, Y. P. S. (Ed.). (2012). Biotechnology in agriculture and forestry: Cryopreservation of plant germplasm I. Springer. [Google Scholar] [Crossref]

10. Berjak, P., & Pammenter, N. W. (2008). From Avicennia to Zizania: Seed recalcitrance in perspective. Annals of Botany, 101(2), 213–228. https://doi.org/10.1093/aob/mcm168 [Google Scholar] [Crossref]

11. Berjak, P., & Pammenter, N. W. (2013). Implications of the lack of desiccation tolerance in recalcitrant seeds. Frontiers in Plant Science, 4, 478. https://doi.org/10.3389/fpls.2013.00478 [Google Scholar] [Crossref]

12. Buitink, J., & Leprince, O. (2008). Intracellular glasses and seed survival in the dry state. Comptes Rendus Biologies, 331(10), 788–795. https://doi.org/10.1016/j.crvi.2008.08.002 [Google Scholar] [Crossref]

13. Chandler, S. F., & Tanaka, Y. (2007). Genetic modification in floriculture. Critical Reviews in Plant Sciences, 26(4), 169–197. https://doi.org/10.1080/07352680701503552 [Google Scholar] [Crossref]

14. Chukwuma, E. C., Soladoye, M. O., & Feyisola, R. T. (2015). Traditional medicine and the future of medicinal plants in Nigeria. Journal of Plant Studies, 4(1), 1–13. https://doi.org/10.5539/jps.v4n1p1 [Google Scholar] [Crossref]

15. Coelho, N., Goncalves, S., & Romano, A. (2020). Endemic plant species conservation: Biotechnological approaches. Plants, 9(3), 345. https://doi.org/10.3390/plants9030345 [Google Scholar] [Crossref]

16. Convention on Biological Diversity. (2010). Nagoya Protocol on access to genetic resources and the fair and equitable sharing of benefits arising from their utilization. Secretariat of the CBD. [Google Scholar] [Crossref]

17. De Klerk, G.-J. (2002). Rooting of microcuttings: Theory and practice. In Vitro Cellular & Developmental Biology – Plant, 38(5), 415–422. https://doi.org/10.1079/IVP2002335 [Google Scholar] [Crossref]

18. Dewir, Y. H., Murthy, H. N., Ammar, M. H., Alghamdi, S. S., Al-Suhaibani, N. A., Alsadon, A. A., & Paek, K.-Y. (2014). In vitro rooting of leguminous plants: A mini-review. Horticulture, Environment, and Biotechnology, 55(6), 459–474. https://doi.org/10.1007/s13580-014-0001-3 [Google Scholar] [Crossref]

19. Ebana, R. U. B., Madunagu, B. E., Ekpe, E. D., & Otung, I. N. (1991). Microbiological exploitation of cardiac glycosides and alkaloids from Garcinia kola, Borreria ocymoides, Kola nitida and Lavandula officinalis. Journal of Applied Bacteriology, 71(5), 398–401. https://doi.org/10.1111/j.1365-2672.1991.tb03814.x [Google Scholar] [Crossref]

20. Engelmann, F. (2011). Use of biotechnologies for the conservation of plant biodiversity. In Vitro Cellular & Developmental Biology – Plant, 47(1), 5–16. https://doi.org/10.1007/s11627-010-9327-2 [Google Scholar] [Crossref]

21. Engelmann, F., & Engels, J. M. M. (2002). Technologies and strategies for ex situ conservation. In J. M. M. Engels, V. R. Rao, A. H. D. Brown, & M. T. Jackson (Eds.), Managing plant genetic diversity (pp. 89–104). CABI Publishing. [Google Scholar] [Crossref]

22. Etienne, H., & Berthouly, M. (2002). Temporary immersion systems in plant micropropagation. Plant Cell, Tissue and Organ Culture, 69(3), 215–231. https://doi.org/10.1023/A:1015668610465 [Google Scholar] [Crossref]

23. Ezekwesili-Ofili, J. O., & Okeke, C. I. (2019). Herbal medicines in African traditional medicine. In F. O. Builders (Ed.), Herbal medicine (pp. 191–214). IntechOpen. https://doi.org/10.5772/intechopen.80348 [Google Scholar] [Crossref]

24. FAO. (2010). Second report on the state of the world's plant genetic resources for food and agriculture. Food and Agriculture Organization of the United Nations. [Google Scholar] [Crossref]

25. FAO. (2015). Voluntary guidelines to support the integration of genetic diversity into national climate change adaptation planning. FAO. [Google Scholar] [Crossref]

26. Feyissa, T., Welander, M., & Negash, L. (2005). In vitro regeneration of Hagenia abyssinica from leaf explants. Plant Cell, Tissue and Organ Culture, 80(1), 55–60. https://doi.org/10.1007/s11240-004-0713-8 [Google Scholar] [Crossref]

27. Gaj, T., Gersbach, C. A., & Barbas, C. F. (2013). ZFN, TALEN and CRISPR/Cas-based methods for genome engineering. Trends in Biotechnology, 31(7), 397–405. https://doi.org/10.1016/j.tibtech.2013.04.004 [Google Scholar] [Crossref]

28. Galewsky, S., & Nessler, C. L. (1986). Clonal propagation of opium poppy (Papaver somniferum L.) by somatic embryogenesis. Plant Cell Reports, 5(6), 396–399. https://doi.org/10.1007/BF00269531 [Google Scholar] [Crossref]

29. George, E. F., Hall, M. A., & De Klerk, G.-J. (Eds.). (2008). Plant propagation by tissue culture: Vol. 1. The background (3rd ed.). Springer. [Google Scholar] [Crossref]

30. Hazarika, B. N. (2003). Acclimatization of tissue-cultured plants. Current Science, 85(12), 1704–1712. [Google Scholar] [Crossref]

31. Holobiuc, I. (2020). Applications of plant tissue culture in conservation of rare and endangered plants. In M. Asif (Ed.), Advances in plant tissue culture (pp. 95–134). Academic Press. https://doi.org/10.1016/B978-0-12-818581-0.00005-X [Google Scholar] [Crossref]

32. Idu, M., Erhabor, J. O., & Efijuemue, H. M. (2010). Documentation on medicinal plants sold in markets in Abeokuta, Nigeria. Tropical Journal of Pharmaceutical Research, 9(2), 110–118. [Google Scholar] [Crossref]

33. IUCN. (2022). The IUCN Red List of threatened species. Version 2022-2. https://www.iucnredlist.org/ [Google Scholar] [Crossref]

34. Jain, S. M., & Saxena, P. K. (Eds.). (2009). Protocols for in vitro cultures and secondary metabolite analysis of aromatic and medicinal plants. Humana Press. https://doi.org/10.1007/978-1-60327-114-1 [Google Scholar] [Crossref]

35. Jimenez, V. M. (2005). Involvement of plant hormones and plant growth regulators on in vitro somatic embryogenesis. Plant Growth Regulation, 47(2–3), 91–110. https://doi.org/10.1007/s10725-005-3478-x [Google Scholar] [Crossref]

36. Lahlou, M. (2013). The success of natural products in drug discovery. Pharmacology & Pharmacy, 4(3), 17–31. https://doi.org/10.4236/pp.2013.43A003 [Google Scholar] [Crossref]

37. Larkin, P. J., & Scowcroft, W. R. (1981). Somaclonal variation: A novel source of variability from cell cultures for plant improvement. Theoretical and Applied Genetics, 60(4), 197–214. https://doi.org/10.1007/BF02342540 [Google Scholar] [Crossref]

38. Lloyd, G., & McCown, B. (1980). Commercially-feasible micropropagation of mountain laurel Kalmia latifolia by use of shoot-tip culture. Proceedings of the International Plant Propagators Society, 30, 421–427. [Google Scholar] [Crossref]

39. Loyola-Vargas, V. M., & Ochoa-Alejo, N. (Eds.). (2018). Plant cell culture protocols (4th ed., Methods in Molecular Biology Vol. 1815). Humana Press. https://doi.org/10.1007/978-1-4939-8594-4 [Google Scholar] [Crossref]

40. Luczaj, L., & Szymanski, W. M. (2007). Wild vascular plants gathered for consumption in the Polish countryside: A review. Journal of Ethnobiology and Ethnomedicine, 3(1), 17. https://doi.org/10.1186/1746-4269-3-17 [Google Scholar] [Crossref]

41. Maiga, A., Diallo, D., Bye, R., & Paulsen, B. S. (2005). Determination of some toxic and essential metal ions in medicinal and edible plants from Mali. Journal of Agricultural and Food Chemistry, 53(6), 2316–2321. https://doi.org/10.1021/jf0403003 [Google Scholar] [Crossref]

42. Moyo, M., Aremu, A. O., & Van Staden, J. (2015). Conservation strategy: An integral aspect of medicinal plant research. Journal of Ethnopharmacology, 174, 621–641. https://doi.org/10.1016/j.jep.2015.08.052 [Google Scholar] [Crossref]

43. Murashige, T., & Skoog, F. (1962). A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum, 15(3), 473–497. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x [Google Scholar] [Crossref]

44. National Centre for Genetic Resources and Biotechnology (NACGRAB). (2020). Annual report on plant genetic resources conservation in Nigeria. Federal Ministry of Agriculture and Rural Development. [Google Scholar] [Crossref]

45. Neumann, K. H., Kumar, A., & Imani, J. (2009). Plant cell and tissue culture: A tool in biotechnology: Basics and application. Springer. https://doi.org/10.1007/978-3-540-93882-5 [Google Scholar] [Crossref]

46. Nigerian Environmental Law. (2007). National Environmental Standards and Regulations Enforcement Agency (Establishment) Act. Federal Government of Nigeria. [Google Scholar] [Crossref]

47. Nishizawa, S., Bhowmik, S. K., Robison, M., Sugiyama, Y., & Watad, A. A. (2000). In vitro differentiation of protoplasts of Chrysanthemum morifolium. Plant Cell Reports, 19(3), 230–237. [Google Scholar] [Crossref]

48. Okafor, J. C. (1980). Edible indigenous woody plants in the rural economy of the Nigerian forest zone. Forest Ecology and Management, 3, 45–55. https://doi.org/10.1016/0378-1127(80)90005-4 [Google Scholar] [Crossref]

49. Okeke, C. U., & Elekwa, I. (2006). Preliminary phytochemical screening of the fruit and seed of Zanthoxylum zanthoxyloides (Lam.) in Anambra State, Nigeria. Journal of Biological Research and Biotechnology, 4(1), 19–22. [Google Scholar] [Crossref]

50. Okunade, A. L. (2002). Aglaia odorata Lour. (Meliaceae). Fitoterapia, 73(1), 1–15. [Google Scholar] [Crossref]

51. Oladele, A. T., Aladejana, O. M., & Agboola, D. A. (2013). Effects of plant growth regulators on in vitro propagation of Zanthoxylum zanthoxyloides. International Journal of Applied Sciences and Biotechnology, 1(4), 199–204. https://doi.org/10.3126/ijasbt.v1i4.9238 [Google Scholar] [Crossref]

52. Oliveira, L. M., Paiva, R., Santoro, P. H., Fraga, A. C., & Alves, E. (2011). Interactions between plant growth regulators and activated charcoal on the in vitro development of Bromeliaceae. Acta Scientiarum. Agronomy, 33(2), 315–320. https://doi.org/10.4025/actasciagron.v33i2.5870 [Google Scholar] [Crossref]

53. Osoniyi, O., & Onajobi, F. (2003). Coagulant and anticoagulant activities in Jatropha curcas latex. Journal of Ethnopharmacology, 89(1), 101–105. https://doi.org/10.1016/S0378-8741(03)00264-6 [Google Scholar] [Crossref]

54. Pammenter, N. W., & Berjak, P. (1999). A review of recalcitrant seed physiology in relation to desiccation-tolerance mechanisms. Seed Science Research, 9(1), 13–37. https://doi.org/10.1017/S0960258599000130 [Google Scholar] [Crossref]

55. Pence, V. C. (2011). Evaluating costs for the in vitro propagation and preservation of endangered plants. In Vitro Cellular & Developmental Biology – Plant, 47(1), 176–187. https://doi.org/10.1007/s11627-010-9323-6 [Google Scholar] [Crossref]

56. Plants of the World Online (POWO). (2023). Zanthoxylum zanthoxyloides (Lam.) Zepern. & Timler. Royal Botanic Gardens, Kew. https://powo.science.kew.org/ [Google Scholar] [Crossref]

57. Poku, K. (2002). Small-scale palm oil processing in Africa (FAO Agricultural Services Bulletin 148). Food and Agriculture Organization. [Google Scholar] [Crossref]

58. Preece, J. E. (2003). A century of progress with vegetative plant propagation. HortScience, 38(5), 1015–1025. https://doi.org/10.21273/HORTSCI.38.5.1015 [Google Scholar] [Crossref]

59. Quebedeaux, B., & Chollet, R. (Eds.). (2010). Plant biology: The interface of plant biotechnology and the environment. American Society of Plant Biologists. [Google Scholar] [Crossref]

60. Quoirin, M., & Lepoivre, P. (1977). Etude de milieux adaptes aux cultures in vitro de Prunus. Acta Horticulturae, 78, 437–442. https://doi.org/10.17660/ActaHortic.1977.78.54 [Google Scholar] [Crossref]

61. Reed, B. M. (Ed.). (2008). Plant cryopreservation: A practical guide. Springer. https://doi.org/10.1007/978-0-387-72276-4 [Google Scholar] [Crossref]

62. Roberts, E. H. (1973). Predicting the storage life of seeds. Seed Science and Technology, 1(3), 499–514. [Google Scholar] [Crossref]

63. Sasson, A. (1993). Biotechnologies in developing countries: Present and future. UNESCO Publishing. [Google Scholar] [Crossref]

64. Shatnawi, M., Johnson, E., Torpy, F., & Hutton, D. (2004). In vitro propagation and cryostorage of Syzygium luehmannii and other Myrtaceae by vitrification. Plant Cell, Tissue and Organ Culture, 77(3), 277–284. https://doi.org/10.1023/B:TICU.0000020960.07248.48 [Google Scholar] [Crossref]

65. Siddique, I., & Anis, M. (2009). An improved plant regeneration system and ex vitro acclimatization of Ocimum basilicum L. Acta Physiologiae Plantarum, 31(4), 861–869. https://doi.org/10.1007/s11738-009-0299-y [Google Scholar] [Crossref]

66. Skoog, F., & Miller, C. O. (1957). Chemical regulation of growth and organ formation in plant tissues cultured in vitro. Symposia of the Society for Experimental Biology, 11, 118–131. [Google Scholar] [Crossref]

67. Takhtajan, A. (2009). Flowering plants (2nd ed.). Springer. [Google Scholar] [Crossref]

68. Thorpe, T. A. (2007). History of plant tissue culture. Molecular Biotechnology, 37(2), 169–180. https://doi.org/10.1007/s12033-007-0031-3 [Google Scholar] [Crossref]

69. Touchell, D. H., Chiang, V. L., & Tsai, C.-J. (2020). In vitro regeneration and transformation of poplar (Populus). Frontiers in Plant Science, 11, 82. https://doi.org/10.3389/fpls.2020.00082 [Google Scholar] [Crossref]

70. Usman, J. M., Olawale, F. A., & Soladoye, M. O. (2018). Sustainable utilization and conservation status of economically important plant species of the guinea-savanna transition zone of Nigeria. Journal of Ethnobiology and Ethnomedicine, 14(1), 46. https://doi.org/10.1186/s13002-018-0244-3 [Google Scholar] [Crossref]

71. Volk, G. M. (2010). Application of functional genomics and proteomics to plant cryopreservation. Current Genomics, 11(1), 24–29. https://doi.org/10.2174/138920210790217936 [Google Scholar] [Crossref]

72. Vujanovic, V., Hamid, A., & Vujanovic, J. (2012). Multitrophic, multipartite relationships within a plant–endophyte–symbiotroph–saprophyte framework in native perennial grasses. Plant and Soil, 359(1–2), 205–224. https://doi.org/10.1007/s11104-012-1196-7 [Google Scholar] [Crossref]

73. Watt, M. P. (2012). The status of temporary immersion system (TIS) technology for plant micropropagation. African Journal of Biotechnology, 11(76), 14025–14035. https://doi.org/10.5897/AJB12.2480 [Google Scholar] [Crossref]

74. Wilkinson, T., Wetten, A., Prychid, C., & Fay, M. F. (2003). Suitability of cryopreservation for the long-term storage of rare and endangered plant species: A case history for Cosmos atrosanguineus. Annals of Botany, 91(5), 65–74. https://doi.org/10.1093/aob/mcf279 [Google Scholar] [Crossref]

75. Withers, L. A., & Engelmann, F. (1998). In vitro conservation of plant genetic resources. In A. Altman (Ed.), Agricultural biotechnology (pp. 57–88). Marcel Dekker. [Google Scholar] [Crossref]

76. World Health Organization (WHO). (2019). WHO global report on traditional and complementary medicine 2019. WHO Press. [Google Scholar] [Crossref]

77. Yadav, R. K., Chattopadhyay, S., Bhattacharya, B., Tripathi, M. K., & Bhatt, B. K. (2003). Somatic embryogenesis and plant regeneration in Zanthoxylum alatum (Roxb.) using cotyledons as explant. Current Science, 84(9), 1240–1244. [Google Scholar] [Crossref]

78. Yam, T. W., & Arditti, J. (2009). History of orchid propagation: A mirror of the history of biotechnology. Plant Biotechnology Reports, 3(1), 1–56. https://doi.org/10.1007/s11816-008-0066-3 [Google Scholar] [Crossref]

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