Medicinal Plant Research 2025, Vol.15, No.2, 80-87 http://hortherbpublisher.com/index.php/mpr 84 degradation. In addition, substrates that are compatible with sustainable cultivation manners assist in the conservation of this endangered species by reducing pressure on natural populations (Shao et al., 2014). The use of natural and biodegradable materials in substrate formulation is also within environmentally sustainable cultivation manners, which promote biodiversity and soil fertility. 6 Technical Challenges in Application and Practice 6.1 Difficulties in applying substrates to large-scale cultivation Among the key issues in large-scale culturing of Anoectochilus roxburghii via substrates is the variable nature of substrate composition affecting the growth and seedling survival rate of the plants. Different substrate compositions, such as peat, river sand, and peanut shells, have been shown to significantly affect the seedling survival rate and growth quality of A. roxburghii (Mechergui et al., 2024). Optimum substrate ratios obtained from laboratory experiments may not be directly scalable due to differences in environmental conditions and resource availability (Han et al., 2022). It may also be challenging to maintain consistent substrate quality across large areas, which would lead to varying survival rates and growth rates. 6.2 Issues of cost, resource availability, and sustainability of substrates Cost and availability of substrates are relevant concerns with large-scale culture of A. roxburghii. Substrates like peat and bark, though suitable, will not be readily available, sustainable, and cost-effective for commercial use as they are limited and expensive (Jim, 2024). Their extraction and use also involve environmental costs, which have questioned sustainability. Alternatives like utilizing locally obtained materials or creating artificial substrates can be looked into so as to overcome these drawbacks (Yu et al., 2022). 6.3 Directions for improving transplanting techniques for tissue culture seedlings To improve the transplanting techniques of Anoectochilus roxburghii tissue culture seedlings, research shows improving substrate composition and environmental conditions. Experiments have proved working with particular combinations of substrates, e.g., peat, muck, and vermiculite, under controlled conditions significantly enhances survival rates and quality of growth (Jin et al., 2017). Besides, integration of the endophytic fungi technology has been reported to enhance significantly the seedling survival and vigor and been proven to be a potential option for transplanting success improvement. It is clear from studies that the role of endophytic fungi in the improvement of A. roxburghii seedling survival and vigor of growth is considerable. These fungi have the capability of releasing plant growth hormones, such as indole-3-acetic acid (IAA), to stimulate root branching and elongation, thereby increasing the nutrient and water uptake potential of the seedlings from the medium. This is of extreme significance during A. roxburghii tissue culture seedling transplantation as they are extremely sensitive to environmental differences, and a healthy root system has the capability to make them survive (Pascual et al., 2018; Ye et al., 2020) (Figure 2). Figure 2 The Impact of Endophytic Fungi on the Transplantation of Anoectochilus roxburghii (Adopted from Ye et al., 2020)
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