• Baradwaj Rg Department of Plant Science, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India.
  • Rao Mv Department of Plant Science, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India.
  • Senthil Kumar T Department of Plant Science, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India.


Objective: Elimination of endophytic actinomycetes before micropropagation using antibiotic pre-treatment in rhizome bud explants of Alpinia galanga. Then, the formulation of an operative protocol for Micropropagation of the same void of endophytic actinomycetes.

Methods: A treatment of mercury chloride and carbendazim, alone and in combination was used as surface sterilants. A pre-treatment of rifampicin and fusidic acid was used against actinomycete endophyte disinfection of rhizome bud explants. Then, Murashige and Skoog (MS) medium supplemented with various concentrations of cytokinins were used for micropropagation of disinfected explants.

Results: A treatment of 0.1% (w/v) mercury chloride and 0.1% (w/v) carbendazim, one after the other for 5 minutes gave the best sterility of 83.3%. A pre-treatment of Rifampicin 100 mg/l and fusidic acid 100 mg/l for 2 hrs gave the best disinfection of 70% against actinomycete endophytes. A combination of thidiazuron (TDZ) 0.45 μM and 6-benzyladenine 13.32 μM in MS medium resulted in 9.4 shoots per explant. MS medium fortified with 10.74 μMof 1-naphthaleneacetic acid gave the best rooting of 20 roots/shoot. inter simple sequence repeat marker genetic similarity of regenerants with the mother plant was confirmed.

Conclusion: This study shows the potency of Rifampicin and Fusidic acid to disinfect explants from actinomycete endophytes and is significant as the first report on curbing actinomycetes endophytes in plant tissue culture of A. galanga. This is also the first report conferring the dissimilar regeneration capabilities of TDZ in comparison to other cytokinins in Zingiberaceae.

Keywords: Alpinia galanga, Rifampicin, Fusidic acid, Actinomycetes, Thidiazuron, Inter simple sequence repeat.


1. Litz RE, Gray DJ. Organogenesis and somatic embryogenesis. In: Hammerschlag FA, Litz RE, editors. Biotechnology of Perennial Fruit Crops. Wallingford, UK: CABI International; 1992. p. 3-34.
2. Borthakur M, Hazarika J, Singh RS. A protocol for micropropagation of Alpinia galanga. Plant Cell Tissue Organ Cult 1998;55(3):231-3.
3. Parida R, Mohanty S, Nayak S. Evaluation of genetic fidelity of in vitro propagated greater galangal (Alpinia galanga L.) using DNA based markers. Int J Plant Anim Environ Sci 2011;1(3):123-33.
4. Rao K, Chodisetti B, Gandi S, Mangamoori LN, Giri A. Direct and indirect organogenesis of Alpinia galanga and the phytochemical analysis. Appl Biochem Biotechnol 2011;165(5-6):1366-78.
5. Singh NM, Chanu LA, Devi YP. Micropropagation-an in vitro technique for the conservation of Alpinia galanga. Adv Appl Sci Res 2014;5(3):259-63.
6. Taechowisan T, Chuaychot N, Chanaphat S, Wanbanjob A, Shen Y. Biological activity of chemical constituents isolated from Streptomyces sp. Tc052, an endophyte in Alpinia galanga. Int J Pharmacol 2008;4(2):95-101.
7. Taechowisan T, Lumyong S. Activity of endophytic actinomycetes from roots of Zingiber officinale and Alpinia galanga against phytopathogenic fungi. Ann Microbiol 2003;53(3):291-8.
8. Taechowisan T, Wanbanjob A, Tuntiwachwuttikul P, Taylor WC. Identification of Streptomyces sp. Tc022, an endophyte in Alpinia galanga, and the isolation of actinomycin D. Ann Microbiol 2006;56(2):113-7.
9. El-Tarabily KA, Nassar AH, Hardy GE, Sivasithamparam K. Plant growth promotion and biological control of Pythium aphanidermatum a pathogen of cucumber, by endophytic actinomycetes. Eur J Plant Pathol 2009;128(4):527-39.
10. Nimnoi P, Pongsilp N, Lumyong S. Endophytic actinomycetes isolated from Aquilaria crassna Pierre ex Lec and screening of plant growth promoters production. World J Microbiol Biotechnol 2010;26(2):193-203.
11. Ramesh KV, Garima M, Pradeep S, Jha KK, Khosa RL. Alpinia galanga - An important medicinal plant: A review. Pharma Sin 2011;2(1):142-54.
12. Ravichandra V, Hanumantharayappa B, Papasani VM. Evaluation of cardio protective activity of galangin against doxorubicin induced cardiomyopathy. Int J Pharm Pharm Sci 2014;6(9):86-90.
13. Barik BR, Kundu AB, Dey AK. Two phenolic constituents from Alpinia galanga rhizomes. Phytochemistry 1987;26(7):2126-7.
14. Das A, Santhy KS. Chemical characterisation of Alpinia galanga (L.) Willd by GC-MS, XRD, FTIR and UV-Vis spectroscopic methods. Int J Pharm Pharm Sci 2015;7(9):499-501.
15. Daubresse N, Francesch C, Mhamdi F, Rolando C. A mild synthesis of coumaryl, coniferyl, sinapyl aldehydes and alcohols. Synthesis 1994;4:369-71.
16. Lee SJ, Ando T. Optically active 1’-acetoxychavicol acetate and its positional isomers: Synthesis and repellent effect against adzuki bean weevil. J Pestic Sci 2001;26(1):76-81.
17. Loubinoux B, Miazimbakana J, Gerardin P. Reactivity of new precursors of quinone methides. Tetrahedron Lett 1989;30(15):1939-42.
18. Noro T, Sekiya T, Katoh M, Oda Y, Miyase T, Kuroyanagi M, et al. Inhibitors of xanthine oxidase from Alpinia galanga. Chem Pharm Bull 1988;36(1):244-8.
19. Goel MK, Kukreja AK, Bisht NS. In vitro manipulations in St. John’s wort (Hypericum perforatum L.) for incessant and scale up micropropagation using adventitious roots in liquid medium and assessment of clonal fidelity using RAPD analysis. Plant Cell Tissue Organ Cult 2009;96:1-9.
20. Jain SM. Tissue culture-derived variation in crop improvement. Euphytica 2001;118(2):153-66.
21. Amalraj VA, Velayudhan KC, Abraham Z. Threatened medicinal plants in western ghats. In: Karunakaran CK, editor. Proceedings Symposium on Rare Endangered Endemic Plants Western Ghats. Thiruvananthapuram, Kerala, India: Kerala Forest Department; 1991. p. 215-20.
22. Phillips R, Arnott SM, Kaplan SE. Antibiotics in plant tissue culture: Rifampicin effectively controls bacterial contaminants without affecting the growth of short-term explant cultures of Helianthus tuberosus. Plant Sci Lett 1981;21(3):235-40.
23. Murashige T, Skoog F. A revised medium for rapid growth and bio-assays with tobacco tissue cultures. Physiol Plant 1962;15(3):473-97.
24. Porebski S, Bailey LG, Baum BR. Modification of CTAB DNA extraction protocols for plants containing high polysaccharide and polyphenol components. Plant Mol Biol Rep 1997;15(1):8-15.
25. Abdelmageed AH, Faridah QZ, Norhana FM, Julia AA, Kadir MA. Micropropagation of Etlingera elatior (Zingiberaceae) by using axillary bud explants. J Med Plants Res 2011;5(18):4465-9.
26. Rakkimuthu R, Jacob J, Aravinthan K. In vitro micropropagation of Alpinia zerumbet variegate, an important medicinal plant, through rhizome bud explants. Res Biotechnol 2011;2(1):7-10.
27. Wilkins WL. Disinfection, Sterilization, and Preservation. 5th ed. Philadelphia, PA: Lea & Febiger; 2000.
28. Kiso T, Fujita K, Ping X, Tanaka T, Taniguchi M. Screening for microtubule-disrupting antifungal agents by using a mitotic-arrest mutant of Aspergillus nidulans and novel action of phenylalanine derivatives accompanying tubulin loss. Antimicrob Agents Chemother 2004;48(5):1739-48.
29. Campbell EA, Korzheva N, Mustaev A, Murakami K, Nair S, Goldfarb A, et al. Structural mechanism for rifampicin inhibition of bacterial rna polymerase. Cell 2001;104(6):901-12.
30. Nasher MA, Hay RJ. Synergy of antibiotics against Streptomyces somaliensis isolates in vitro. J Antimicrob Chemother 1998;41(2):281-4.
31. Kochuthressia KP, Britto SJ, Raj LJ, Jaseentha MO, Senthilkumar SR. Efficient regeneration of Alpinia purpurata (Vieill.) K. Schum. Plantlets from rhizome bud explants. Int Res J Plant Sci 2010;1(2):43-7.
32. Tefera W, Wannakrairoj S. A micropropagation method for korarima (Aframomum corrorima [Braun] Jansen). ScienceAsia 2004;30:1-7.
33. Baskaran P, Jayabalan N. Effect of growth regulators on rapid micropropagation and psoralen production in Psoralea corylifolia L. Acta Physiol Plant 2008;30:345-51.
34. Yusuf NA, Annuar MM, Khalid N. Rapid micropropagation of Boesenbergia rotunda (L.) Mansf. Kulturpfl. (A valuable medicinal plant) from shoot bud explants. Afr J Biotechnol 2011;10(7):1194-9.
35. Rani V, Raina SN. Genetic fidelity of organized meristem-derived micropropagated plants: A critical reappraisal. In Vitro Cell Dev Biol Plant 2000;36(5):319-30.
36. Mohanty S, Parida R, Sahoo S, Nayak S. In vitro conservation of nine medicinally and economically important species of zingiberaceae from Eastern India. Proc Natl Acad Sci India Sect B Biol Sci 2014;84(3):799-803.
37. Rajasekharan PE, Abdul-Kareem VK, Ravish BS, Mini S. Analysis of genetic diversity in Alpinia galanga using ISSR markers. Indian J Plant Genet Resour 2016;29(2):194-8.
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How to Cite
Rg, B., R. Mv, and S. K. T. “CURBING ACTINOMYCETES AND THIDIAZURON ENHANCED MICROPROPAGATION IN THE RARE ALPINIA GALANGA - A MEDICINAL ZINGIBER”. Asian Journal of Pharmaceutical and Clinical Research, Vol. 10, no. 7, July 2017, pp. 153-8, doi:10.22159/ajpcr.2017.v10i7.17734.
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