Volume 11, Issue 22 (11-2023)
Ecol Iran For 2023, 11(22): 13-23 |
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Ahmadi A, Moslehi Jouybari M, Shabani S, Zavvar S, Maghsudloo M K, Rabbani Nasab H. (2023). The Effect of Some Disinfectants on the Contamination Control of Box Tree Explants under in Vitro Culture. Ecol Iran For. 11(22), 13-23. doi:10.61186/ifej.11.22.12
URL: http://ifej.sanru.ac.ir/article-1-467-en.html
URL: http://ifej.sanru.ac.ir/article-1-467-en.html
Akram Ahmadi *1 
, Maryam Moslehi Jouybari2, Saeid Shabani3, Sepideh Zavvar1, Mohammad Karim Maghsudloo4, Hojjatollah Rabbani Nasab5
, Maryam Moslehi Jouybari2, Saeid Shabani3, Sepideh Zavvar1, Mohammad Karim Maghsudloo4, Hojjatollah Rabbani Nasab5
1- Research Department of Natural Resources, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
2- Research Assistant, Research Department of Natural Resources, Hormozgan Agricultural and Natural Resources Research and Education Center, AREEO, Bandar Abbas, Iran
3- Research Assistant, Research Department of Natural Resources, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
4- Research Expert, Research Department of Natural Resources, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
5- Research Associate, Plant Protection Research Department, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
2- Research Assistant, Research Department of Natural Resources, Hormozgan Agricultural and Natural Resources Research and Education Center, AREEO, Bandar Abbas, Iran
3- Research Assistant, Research Department of Natural Resources, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
4- Research Expert, Research Department of Natural Resources, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
5- Research Associate, Plant Protection Research Department, Golestan Agriculture and Natural Resources Research and Education Center, AREEO, Gorgan, Iran
Abstract: (2102 Views)
Extended Abstract
Background: Hyrcanian boxwood (Buxus hyrcana Pojark.) is an evergreen tree native to the Hyrcanian forests in northern Iran. This species is considered one of the endangered plant species due to habitat loss, overexploitation, and environmental changes. In vitro propagation of this valuable species offers a viable solution to prevent its extinction and promote conservation efforts. However, one of the significant challenges faced in the in vitro culture of Buxus hyrcana is the presence of fungal and bacterial contamination. Such contamination not only affects the successful propagation of this species but also leads to increased costs and resource wastage. The objective of the present study is to investigate the efficacy of various disinfectants, including 15% Percidine, Aqua Percidine, Super Percidine, and Percidine Plus, in removing contamination from B. hyrcana explants under in vitro culture conditions. These treatments were compared to the conventional method of using sodium hypochlorite, which is commonly employed for sterilization in plant tissue culture. By evaluating the effectiveness of these disinfectants, we aim to identify a reliable method for reducing contamination and improving the success rates of in vitro propagation for Hyrcanian boxwood.
Methods: The experiment was conducted using a factorial arrangement as a completely randomized design, with three replications to ensure statistical validity. Initially, explants, which included both twigs and seeds, were thoroughly washed with ordinary water combined with a few drops of dishwashing liquid for five minutes. This step was crucial to eliminate any surface contaminants that could interfere with subsequent sterilization processes. Following this, the explants were treated with a 1% benomyl fungicide for ten minutes to further reduce fungal presence. After the pre-treatment, the sterilization of explants was carried out under laminar airflow conditions using the experimental disinfectant treatments. The treatments included varying concentrations of Percidine (15%), Aqua Percidine, Super Percidine, and Percidine Plus at concentrations of 0%, 1%, 2%, and 4%. These treatments were compared against the standard sodium hypochlorite method, which served as the control. Once the sterilization process was complete, the plant materials were placed on Murashige and Skoog (MS) medium, a widely used nutrient medium for plant tissue culture that provides essential nutrients for growth. To assess contamination levels, the cultures were monitored under controlled growth conditions at a temperature of 25±1 °C, with a light intensity ranging from 1000 to 1500 lux, following a photoperiod of 16 hours of light and 8 hours of darkness. This setup ensured optimal growth conditions for the explants while allowing for accurate evaluation of contamination rates.
Results: The results of the study indicated a significant difference in fungal and bacterial contamination levels between the seed and twig explants. Specifically, contamination in the seed explants was considerably reduced compared to the twig explants. This finding suggests that the type of explant used plays a critical role in the success of sterilization protocols. Among the disinfectants tested, Super Percidine demonstrated the highest efficiency, achieving a contamination reduction rate of 63% in twig explants. This result highlights the potential of Super Percidine as an effective sterilizing agent for this particular type of explant. In addition to Super Percidine, Aqua Percidine also showed promising results in seed explants, with a contamination reduction rate of 33%. This indicates that both disinfectants can effectively reduce contamination on the surface of seeds, making them suitable choices for in vitro propagation efforts. The comparative analysis of the various treatments revealed that while sodium hypochlorite is commonly used, the alternative disinfectants tested in this study may offer improved outcomes, particularly for specific explant types.
Conclusion: In conclusion, the findings of this research suggest that the use of Super Percidine at a concentration of 2% is particularly beneficial for sterilizing twig explants of Hyrcanian boxwood. Furthermore, both Super Percidine (2%) and Aqua Percidine (2%) are recommended for use with seed explants, as they effectively reduce contamination rates and enhance the likelihood of successful in vitro propagation. These results contribute valuable insights into the optimization of sterilization protocols for endangered plant species, ultimately aiding in conservation efforts and the sustainable management of Hyrcanian boxwood populations.Future studies should explore the long-term effects of these sterilization methods on the growth and development of Buxus hyrcana in vitro, as well as the potential for scaling up these techniques for broader conservation applications. By improving our understanding of effective propagation methods, we can better support the preservation of this important species and its habitat.
Background: Hyrcanian boxwood (Buxus hyrcana Pojark.) is an evergreen tree native to the Hyrcanian forests in northern Iran. This species is considered one of the endangered plant species due to habitat loss, overexploitation, and environmental changes. In vitro propagation of this valuable species offers a viable solution to prevent its extinction and promote conservation efforts. However, one of the significant challenges faced in the in vitro culture of Buxus hyrcana is the presence of fungal and bacterial contamination. Such contamination not only affects the successful propagation of this species but also leads to increased costs and resource wastage. The objective of the present study is to investigate the efficacy of various disinfectants, including 15% Percidine, Aqua Percidine, Super Percidine, and Percidine Plus, in removing contamination from B. hyrcana explants under in vitro culture conditions. These treatments were compared to the conventional method of using sodium hypochlorite, which is commonly employed for sterilization in plant tissue culture. By evaluating the effectiveness of these disinfectants, we aim to identify a reliable method for reducing contamination and improving the success rates of in vitro propagation for Hyrcanian boxwood.
Methods: The experiment was conducted using a factorial arrangement as a completely randomized design, with three replications to ensure statistical validity. Initially, explants, which included both twigs and seeds, were thoroughly washed with ordinary water combined with a few drops of dishwashing liquid for five minutes. This step was crucial to eliminate any surface contaminants that could interfere with subsequent sterilization processes. Following this, the explants were treated with a 1% benomyl fungicide for ten minutes to further reduce fungal presence. After the pre-treatment, the sterilization of explants was carried out under laminar airflow conditions using the experimental disinfectant treatments. The treatments included varying concentrations of Percidine (15%), Aqua Percidine, Super Percidine, and Percidine Plus at concentrations of 0%, 1%, 2%, and 4%. These treatments were compared against the standard sodium hypochlorite method, which served as the control. Once the sterilization process was complete, the plant materials were placed on Murashige and Skoog (MS) medium, a widely used nutrient medium for plant tissue culture that provides essential nutrients for growth. To assess contamination levels, the cultures were monitored under controlled growth conditions at a temperature of 25±1 °C, with a light intensity ranging from 1000 to 1500 lux, following a photoperiod of 16 hours of light and 8 hours of darkness. This setup ensured optimal growth conditions for the explants while allowing for accurate evaluation of contamination rates.
Results: The results of the study indicated a significant difference in fungal and bacterial contamination levels between the seed and twig explants. Specifically, contamination in the seed explants was considerably reduced compared to the twig explants. This finding suggests that the type of explant used plays a critical role in the success of sterilization protocols. Among the disinfectants tested, Super Percidine demonstrated the highest efficiency, achieving a contamination reduction rate of 63% in twig explants. This result highlights the potential of Super Percidine as an effective sterilizing agent for this particular type of explant. In addition to Super Percidine, Aqua Percidine also showed promising results in seed explants, with a contamination reduction rate of 33%. This indicates that both disinfectants can effectively reduce contamination on the surface of seeds, making them suitable choices for in vitro propagation efforts. The comparative analysis of the various treatments revealed that while sodium hypochlorite is commonly used, the alternative disinfectants tested in this study may offer improved outcomes, particularly for specific explant types.
Conclusion: In conclusion, the findings of this research suggest that the use of Super Percidine at a concentration of 2% is particularly beneficial for sterilizing twig explants of Hyrcanian boxwood. Furthermore, both Super Percidine (2%) and Aqua Percidine (2%) are recommended for use with seed explants, as they effectively reduce contamination rates and enhance the likelihood of successful in vitro propagation. These results contribute valuable insights into the optimization of sterilization protocols for endangered plant species, ultimately aiding in conservation efforts and the sustainable management of Hyrcanian boxwood populations.Future studies should explore the long-term effects of these sterilization methods on the growth and development of Buxus hyrcana in vitro, as well as the potential for scaling up these techniques for broader conservation applications. By improving our understanding of effective propagation methods, we can better support the preservation of this important species and its habitat.
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