Effect of Different Disinfection Treatments and Bud Type on Growth Initiation of Pineapple cv. MD2 in In Vitro Conditions

Document Type : Research Paper

Authors

1 Assistant Professor, Date Palm and Tropical Fruit Research Center, Horticultural Sciences Research Institute, Agricultural Research, Education and Extension Organization, Ahwaz, Iran.

2 Assistant Professor, Field and Horticultural Crops Sciences Research Department, Baluchestan, Agricultural and Natural Resources Research and Education Center, Agricultural Research Education and Extension Organization, Iranshahr, Iran.

3 Research Officer, Date Palm and Tropical Fruits Research Center, Horticultural Sciences Research Institute, Agricultural Research, Education and Extension Organization, Ahwaz, Iran.

Abstract

The aim of this study was to determine the optimal conditions for in vitro culture initiation of pineapple cv. MD2 using tissue culture technique, in three different experiments. In the first experiment, the effect of sterilization treatment on contamination, mortality and development rate of explants was investigated. In the second experiment, the buds were divided into five groups based on their position on the pineapple stem to determine the highest rate of bud regeneration. In the third experiment, the effect of different concentrations of benzyl adenine (BA) (0, 0.1, 0.5 and 1 mgl-1) was studied alone or in combination with 0.01 mgl-1 of naphthalene acetic acid (NAA) concentrations on regeneration of buds. The results showed that sodium hypochlorite (2.5%) in 20 minutes was the most effective for sterilizing pineapple buds. All concentrations of BA promoted bud regeneration but the 3 mgl-1 was the optimum. Apical buds and the third section axillary buds had the highest regeneration rate at 93% and 92%, respectively.
 
Keywords: Pineapple, benzyl adenine, apical bud, tissue culture, sodium hypochlorite.
 
Introduction
Pineapple [Ananas comosus (L.) Merr.] is one of the most economically important tropical fruit that belongs to Bromeliaceae. Pineapple plants can be propagated using vegetative materials of slips, suckers, stem sections or crowns. However, in all these three cases plant material is often limited, thus in vitro culture is a commercial alternative approach. In vitro micropropagation of pineapple plantlets has many advantages over conventional methods of vegetative propagation. Tissue culture enhances rapid propagation of suckers to supplement conventional methods to produce pineapples to meet both local and international market demands (Shamim et al., 2016). However, tissue culture initiation, the most critical step towards rapid in vitro propagation, is challenged by microbial contamination, exogenous growth hormone requirements and bud type for growth (Vujovic et al., 2012). Therefore, the aim of this study was to determine optimal sterilization protocols and BA concentration to initiate pineapple in vitro culture using apical and axillary buds as sources of explants.
 
Materials and Methods
In this study the buds of suckers of pineapple cv. MD2 was used for in vitro culture in three different experiments. In the first experiment, explants were sterilized with sodium hypochlorite and ethanol at different concentrations, alone or in combination, and durations (2.5% sodium hypochlorite for 5, 10, 15 minutes, 70% ethanol for 3, 5, 7 minutes, 1% sodium hypochlorite for 10 minutes + 70% ethanol for 3 minutes and 2.5% sodium hypochlorite for 15 minutes + 70% ethanol for 1 minute), followed by three rinsing with sterilized distilled water under aseptic conditions. The explants were placed on solidified Murashige and Skoog (MS) based medium supplemented with 0.5 mgl-1 of benzyl adenine (BA) combined with 0.1 mgl-1 of naphthalene acetic acid (NAA). In the second experiment, buds were grouped based on their position on the pineapple stem (apical, 1st section, 2nd section, 3rd section, and 4th section) to determine the highest rate of bud regeneration. Following sterilization, different buds were transferred to MS medium containing 1 mgl-1 BA+ 0.1 mgl-1 NAA at pH = 5.8. In the third experiment, the effect of different concentrations of BA (0, 0.1, 0.5 and 1 mgl-1) was studied alone or in combination with 0.01 mgl-1 NAA concentrations on regeneration of buds.
 
Results and Discussion
The results showed that sodium hypochlorite (2.5%) in 20 minutes was the most effective in sterilizing pineapple buds. The effectiveness of sodium hypochlorite (2.5%) enhanced with increasing exposure time. The highest explant regeneration rate was observed in the disinfection treatment with 2.5% sodium hypochlorite for 20 minutes (84.49%) followed by 2.5% sodium hypochlorite treatment for 15 minutes + 70% ethanol for 1 minute (75.39%). Ethanol alone could not successfully disinfect pineapple buds. There was high level of harmful microbial contamination of plant tissues associated 70% ethanol treatment, hence the cultures deteriorated.
The apical bud followed by the third section buds were the first to show growth and had the highest rate of regeneration (95% after two weeks). The third section buds recorded (90%) regeneration rate within 1-3 weeks.
All concentrations of BA promoted bud regeneration. The highest regeneration rate was observed in the culture medium containing BA at concentration level of 3 mgl-1 and 0.1 mgl-1 NAA with an average of 75.75%. The lowest regeneration rate in the MS culture medium without growth regulators with an average of 42%. These results are in agreement with findings of Zuraida et al. (2011) and Usman et al, (2013).
 
References
Shamim, M., Kumar, M., Ranjan, T., Kumar, R. R., Pal, A. -K., Kumar, V., Jha, V. B. and Kumar, P. 2016. Importance of micropropagation in pineapple fordisease free plantlets and rapid multiplication. Journal of Pharmacognosy and Phytochemistry, 5(4), pp. 359-362.
Usman, I. S. 2013. Development of an efficient protocol for micro propagation of pineapple. African Journal of Agricultural Research, 8(18), pp.2053-2056. DOI: 10.5897/AJAR12.1763
Vujovic, T., Ruzic, D. J. and Cerovic, R. 2012. In vitro shoot multiplication as influenced by repeated subculturing of shoots of contemporary fruit rootstocks. Horticultural Science, 39, pp.101-107. DOI: 10.17221/208/2011-HORTSCI
Zurida, A. R., Shahanadz, A. H. N., Harteeni, Roowi, S., Che Radziah, C. M. Z. and Sreeramanan, S. 2011. A novel approach for rapid micropropagation of maspine pineapple shoots using liquid shake culture system. African Journal of Biotechnology, 10, pp.3859-3866.

Keywords

Main Subjects

References

Almeida, W., Santana, G., Rodriguez, A. and Costa, M.  2002. Optimization of a protocol for micropropagation of pineapples. Revista Brasileira de Fruticultura, 2, pp.296-300. DOI: 10.1590/S0100-29452002000200005
 
 
Al-Saif, A. M., Hossain, A. B. M. S. and Rosna, M. T. 2011. Effects of benzylaminopurine naphthalene acetic acid on proliferation and shoot growth of pineapple (Ananas comosus L. Merr) in vitro. African Journal of Biotechnology, 10(27), pp.5291-5295. DOI: 10.5897/AJB11.370
 
 
Atawia, A. R., Abd EL-Latif, F. M., EL-Gioushy, S. F., Sherif, S. S. and Kotb, O. M. 2016. Studies on micropropagation of pineapple (Ananas comosus L.). Middle East Journal of Agriculture Research, 5(2), pp.224-232.
 
 
Badou, B. T., Agbidinoukoun, A., Cacaï, G. T., Dossoukpèvi, R. C. and Ahanhanzo, C. 2018. Effects of system benzylaminopurine-adenine sulphate in combination with naphthalene acetic on in vitro regeneration and proliferation of pineapple (Ananas comosus (L.) Mill var. comosus). American Journal of Biotechnology and Bioscience, 2(9), pp.1-15.
 
 
Bhatia, P. and Ashwath, N. 2002. Development of rapid method for micro propagation of a new pineapple [Ananas comosus (L.) Merr.] clone, “Yeppoon gold”. Acta Horticultura. 575, pp.125-131. DOI: 10.17660/ActaHortic.2002.575.11
 
 
Cassells, A. C. 1991. Problems in tissue culture: culture contamination. Pp. 31– 44. In: Derbergh, P. C. and Zimmerman, R. H. (eds), Micropropagation: Technology and Application. Kluwer Publishers, Dordrecht. DOI: 10.1007/978-94-009-2075-0_3
 
 
Cunha, G. A. P. 2005. Applied aspects of pineapple flowering. Bragantia, 64, pp.499-516.  DOI: 10.1590/S0006-87052005000400001
 
 
Danso, K. E., Ayeh, K. O., Oduro, V., Amiteye, S. and Amoatey, H. M. 2008. Effect of 6-benzylaminopurine and α-naphthalene acetic acid on in vitro production of MD2 pineapple planting materials. World Applied Sciences Journal, 3, pp.614-619. DIO: 10.5897/AJB11.370
 
 
Daud, N. H., Jayaraman, S. and Mohamed, R. 2012. Methods paper: an improved surface sterilization technique for introducing leaf, nodal and seed explants of Aquilaria malaccensis from field sources into tissue culture. Asia-Pacific Journal of Molecular Biology and Biotechnology, 2, pp.55- 58.
 
 
Estrela, C., Ribeiro R. G., Estrela, C. R., Pécora J. D. and Sousa-Neto, M. D. 2003. Antimicrobial effect of 2% sodium hypochlorite and 2% chlorhexidine tested by different methods. Brazilian Dental Journal, 14(1), pp.58-62. DOI: 10.1590/s0103-64402003000100011
 
 
Farahani, F. 2014. Micropropagation and growth of in vitro pineapple (Ananas comosus L. merr) in Iran. Plant Archives, 14, pp.337-341.
 
 
Firoozabady, E. and Gutterson, N. 2003. Cost-effective in vitro propagation methods for pineapple. Plant Cell Reports, 21, pp.844-850. DOI: 10.1007/s00299-003-0577-x
 
 
Habiba, U., Reja, S., Saha, M. L. and Khan, M. R. 2002. Endogenous bacterial contamination during in vitro culture of table banana: identification and prevention. Plant Tissue Culture, 12(2), pp.117−124.
 
 
Khan, S., Nasib, A. and Saeed, B. A. 2004. Employment of in vitro technology for large scale multiplication of pineapples (Ananas comosus). Pakistan Journal of Botany, 36, pp.611-615.
 
 
Kyte, L. and Kleyn, J. G. 1996. Plants from test tubes: an introduction to micropropagation. 3rd edition. Timber Press, USA. 240 pp.
 
 
Ibrahim, M. A., Al-Taha, H. A. and Seheem, A. A. 2013. Effect of cytokinin type and concentration and source of explant on shoot multiplication of pineapple plant Ananas comosus Queen) in vitro. Acta Agriculturae Slovenic, 101, pp.15-20. DOI: 10.14720/aas.2013.101.1.14940
 
 
Nelson, B. J., Asare, P. A. and Arthur, R. 2015. In vitro growth and multiplication of pineapple under different duration of sterilization and different concentration of benzylaminopurine and sucrose. Biotechnology, 14(1), pp.35-40. DOI: 10.3923/biotech.2015.35.40
 
 
Moore, G. A., DeWald, M. G. and Evans, M. H. 1992. Micropropagation of pineapple (Ananas comosus L.). Pp.460-470. Bajaj, Y. P. S. (ed.) Biotechnology in Agriculture and Forestry, 18-High Tech and Micropropagation II.  Springer-Verlag Berlin Heidelberg.
 
 
Murashige, T. and Skoog, F. 1962. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plantarum, 15, pp.473-497.
 
 
Sen, S. K. 1990. Pineapple. Pp.  252-279. In: Bose, T. K. and Mitra, S. K (eds.) Fruits: Tropical and Subtropical. Naya Prokash, Calcutta, India.
 
 
Shamim, M., Kumar, M., Ranjan, T., Kumar, R. R., Pal, A.-K., Kumar, V., Jha, V. B. and Kumar, P. 2016. Importance of micropropagation in pineapple for disease free plantlets and rapid multiplication. Journal of Pharmacognosy and Phytochemistry, 5(4), pp.359-362.
 
 
Teixeira, S. L., Ribeiro, J. M. and Teixera, M. T. 2006. Influence of NaClO on nutrient medium sterilization and on pineapple (Ananas comosus cv. Smooth Cayenne) behavior. Plant Cell, Tissue and Organ Culture, 86, pp.375-378. DOI: 10.1007/s11240-006-9121-3
 
 
Usman, I. S. 2013. Development of an efficient protocol for micro propagation of pineapple. African Journal of Agricultural Research, 8(18), pp.2053-2056. DOI: 10.5897/AJAR12.1763
 
 
Vujovic, T., Ruzic, D. J. and Cerovic, R. 2012. In vitro shoot multiplication as influenced by repeated subculturing of shoots of contemporary fruit rootstocks. Horticultural Science, 39, pp.101-107. DOI: 10.17221/208/2011-HORTSCI
 
 
Zurida, A. R., Shahanadz, A. H. N., Harteeni, Roowi, S., Che Radziah, C. M. Z. and Sreeramanan, S. 2011. A novel approach for rapid micropropagation of maspine pineapple shoots using liquid shake culture system. African Journal of Biotechnology, 10, pp.3859-3866.
Seed and Plant Journal
Volume 40, Issue 2
June 2025
Pages 210-193

Files

Share

How to cite

Statistics