Abstract: Background and Objective: Dillenia suffruticosa (Griff.) Martelli is abundantly thriving in various forest types in Brunei Darussalam, including the tropical Bornean heath forests. This study aimed to investigate the plant hormones that can be used to propagate stem cuttings of D. suffruticosa, which is a promising approach to supply plantlets to revegetate degraded tropical heath forests. Methodology: Stem cuttings of D. suffruticosa were treated using three single auxin hormone: 0.1% Indole Acetic Acid (IAA), 0.1% Indole Butyric Acid (IBA) and 0.2% Naphthalene Acetic Acid (NAA) and 5 commercially available rooting hormones: Yates, Clonex and Seradix 1, 2 and 3 and planted in a medium containing a mixture of equal parts black soil and perlite. The survival and rooting percentages, number and lengths of roots produced and number and surface areas of new leaves were assessed at 4-weekly intervals until week 16. Results: Results indicated that the cuttings treated with single auxin treatments (mainly 0.2% NAA and 0.1% IAA) had higher survival and rooting percentages, longer roots, larger leaf surface areas and produced more leaves than those cuttings treated with commercial rooting hormones. Conclusion: It is concluded that compared to commercial rooting hormones, 0.2% NAA and 0.1% IAA were the most effective auxin hormones in promoting survival and growth of Dillenia suffruticosa cuttings.
INTRODUCTION
Heath and Peat Swamp Forests of Brunei Darussalam, which cover 3,500 and 90,000 ha of the country’s total land area, respectively are unique in terms of their biodiversity and ecosystem features1. In the Belait District of Brunei Darussalam, two types of tropical heath forests exist: Kerangas (on higher and drier areas) and kerapah (on permanently waterlogged and lower areas)2,3. These forests are intermixed with the Peat Swamp Forests at Lumut and Badas and some of them occur along the Brunei Liquefied Natural Gas Company (BLNG) water pipelines that carry water from the Belait River to the BLNG plant in Lumut (pers. obs). In 2013, a 20 m wide fire corridor was created along the BLNG Pipeline Road as a preventive measure to reduce fire threats to the water pipelines (BLNG, pers. com.). However, the removal of vegetation cover has conversely increased the risk of fires to the dry and exposed peat within the fire corridor. As such, there is currently an urgent need to revegetate this fire corridor especially with increasing frequencies of dry spells and droughts that have increased fire incidences.
Cleared and degraded forests created from anthropogenic disturbances take a long time to recover naturally, thus supplementing their regeneration through rehabilitation and reforestation activities using tree planting methods are necessary4. Wildings and stem cuttings of native plant species have been widely used to revegetate degraded lands and forests5,6. A rapidly growing number of rooted stem cuttings of plants has been vegetatively propagated and used in many reforestation programmes7,8.
An investigation on the vegetative propagation of Dillenia suffruticosa (Griff.) Martelli, an abundant shrub that is native to Brunei Darussalam9 was conducted. Dillenia suffruticosa from the family Dilleniaceae was chosen because it is a potential candidate for rehabilitation of degraded lands due to its ability to grow in various habitats including roadsides, secondary forests, open sites in undisturbed forest, peat swamps, heath forests, white sands and also well drained and eroded soils10. Dillenia suffruticosa can be used to stabilise slopes in preventing landslides11. Since obtaining D. suffruticosa seedlings via seed propagation is difficult and almost impossible12, using auxins: Naphthalene Acetic Acid (NAA) and Indole Acetic Acid (IAA) in stem cuttings seem to be the best option to produce plantlets9.
This study investigated the different hormones (either using single auxin or commercially available plant rooting hormones) that could increase the survival and rooting percentages as well as root and leaf production of D. suffruticosa stem cuttings. Therefore, the main aim of the study was to determine which hormone could be used for the mass propagation of D. suffruticosa and thus, help to rapidly supply plantlets for the revegetation of degraded heath forests in Brunei Darussalam.
MATERIALS AND METHODS
Sampling of stem cuttings: A total of 150 softwood stem cuttings of Dillenia suffruticosa (Griff.) Martelli were collected from the heath forests within the Universiti Brunei Darussalam campus (4°58’41.5"N, 114°53’39.8"E) and along the Muara-Tutong highway (4°58’59.81"N, 114°54’20.34"E). A maximum of two leafy softwood stem cuttings (8-12 cm) were sampled per plant to reduce any damage to the wild stock plants.
Preparation of stem cuttings: Similar to Abidin and Metali9, all organs on the stem cuttings were removed to ensure they concentrate their energy in forming root and shoot instead of producing reproductive organs. A replicate of 10 cuttings were treated with single auxin hormones, 0.2% Naphthalene Acetic Acid (NAA), 0.1% Indole Acetic Acid (IAA) and 0.1% Indole Butyric Acid (IBA) and 20 cuttings with commercial rooting hormone powders, Seradix 1, 2, 3 and Yates and rooting hormone gel, Clonex, using a quick-dip method9,13. The auxins and their respective concentrations were selected and prepared by following Abidin and Metali9, whereas the commercial hormones were purchased from the local markets. The active constituent(s) of Seradix (unknown concentrations) and Clonex (0.3%) was IBA and Yates were 0.005% IAA and 0.002% NAA. Distilled water was used as a control treatment.
All stem cuttings were planted into square plastic pots containing autoclaved growth media of equal parts black soil and perlite. The pots were placed on the benches in a plant house with misted conditions and the benches with pots were covered with a PVC plastic cover to maintain high relative humidity. The temperatures, relative humidity and light intensities recorded in the plant house were between 22-29°C, 80-90% and 300 μmoL m–2 sec–1 (Apogee Quantum MQ-200 PAR Meter, Apogee Instruments, USA), respectively. After 6 weeks, the cuttings were re-potted into large pots with only black soils. The pots were subsequently transferred to a plant shade with temperatures between 22-34°C, relative humidity of 60-75% and light intensities of 450 μmol m–2 sec–1. Fertilizers (NPK, 15:15:15) were added into the potted soil every 4 weeks and watered twice a day.
Assessment of cuttings and statistical analysis: The cuttings were assessed every 4 weeks until the 16th week for their percentages of survival and rooting, number and lengths of new roots produced and number and surface areas of new leaves. All statistical analyses were conducted using R version 3.3.014. Differences in the number of roots, root lengths, number of leaves and leaf surface areas between the different hormones were determined using one-way ANOVA and Tukey’s HSD tests. Assumptions of normality and equal variances were checked during one-way ANOVA and were not violated.
RESULTS
Stem cuttings treated with single auxin treatments (i.e., 0.2% NAA, 0.1% IAA and 0.1% IBA) had 100% survival recorded throughout the 1st 12 weeks (Table 1). However, by week 16, only stem cuttings treated with 0.2% NAA remained at 100% survival. Stem cuttings treated with auxins produced roots faster with 100% rooting percentage achieved by week 4 when treated with 0.2% NAA and 100% rooting achieved by week 8 when treated with 0.1% IAA (Table 1). Cuttings treated with 0.1% IBA produced roots more slowly compared to the other hormone treatments, reaching only 90% rooting by week 16. In contrast, percentage survival for all stem cuttings treated with commercial rooting hormones fell considerably to 40% and below by week 4, except for cuttings treated with Yates (Table 1). Percentage survival for all commercial rooting hormones reached 0% survival by week 12, except Yates-treated stems. None of the cuttings treated with commercial hormones (except Yates) had successfully rooted (Table 1).
Cuttings treated with 0.2% NAA recorded significantly higher production of roots (Fig. 1a) and significantly longer root lengths (9.34±1.42 cm, Fig. 1b) than the other plant hormones. In contrast, cuttings treated with 0.1% IAA had significantly higher number of new leaves produced (10 new leaves, Fig. 1c) and significantly larger leaf surface area (166±48.6 cm2, Fig. 1d). Cuttings treated with distilled water (control) and all commercial rooting hormones (except for Yates) did not successfully produce any roots and leaf, thus no data for these treatments were recorded by week 16.
DISCUSSION
Stem cuttings survived and rooted better when they were treated with single auxin treatments than the control and commercial rooting hormone treatments. The present findings on survival and rooting percentages are consistent with a previous study by Abidin and Metali9, in which both studies reported that cuttings of D. suffruticosa treated with NAA and IAA produced more and longer roots, more new leaves and greater leaf areas, respectively than any other treatments. Proper root formation is important for survival and growth of cuttings9 and well-formed root systems allow plants to grow and compete for resources, like nutrients and water15. In this study, NAA (0.2%) and IAA (0.1%) showed significant effects on root and leaf productions, thus contributed to higher survival and rooting percentages of the cuttings. Similarly, Kebede et al.16 also reported very high survival (98-100%) of stem cuttings of two native tree species treated with NAA at concentrations of 0.2 and 0.4%. In addition, Usman and Akinyele17 recorded the highest number of roots and root length production of cuttings of a tree species treated with NAA. Hossain and Urbi18 reported that, other than NAA, IAA was also known to be very effective in promoting growth and root length production in stem cuttings of an herb.
| Table 1: | Survival percentages and rooting percentages of Dillenia suffruticosa stem cuttings treated with different hormones (0.2% Naphthalene Acetic Acid (NAA), 0.1% Indole Acetic Acid (IAA), 0.1% Indole Butyric Acid (IBA), Yates, Clonex, Seradix 1, 2 and 3) and distilled water as the control treatment at an interval of 4 weeks until week 16 |
| Fig. 1(a-d): | Variation in the (a) Number of roots produced, (b) Length of roots, (c) Number of new leaves produced and (d) Surface area of leaves of Dillenia suffruticosa stem cuttings treated with plant hormones (0.2% NAA, 0.1% IAA, 0.1% IBA, Yates, Clonex and Seradix 1, 2 and 3) and distilled water as the control treatment. Mean value is denoted by the diamond-shaped symbol. Different letters indicate significant difference between treatments (p<0.001). Dillenia suffruticosa stem cuttings treated with Clonex, Seradix and Control had zero mean values for all 4 parameters. Only data at week 16 were presented |
Higher percentage survival of cuttings treated with Yates compared to the other commercial rooting hormones used in this study could be due to the presence of both NAA and IAA as the active ingredients in Yates. However, due to its very low concentrations (0.005% IAA and 0.002% NAA), stem cuttings treated with Yates recorded lower rooting and survival percentages in week 16 and the effect was not clear-cut as those of single NAA and IAA hormone treatment. Kebede et al.16 also reported that at low concentrations of NAA and IBA hormones, root productions were also low. Various studies have reported that different concentrations of hormones differentially affected survival, rooting percentage, leaf and roots production of cuttings17-20. Copes and Mandel21 noted that a combination of NAA and IBA did not significantly increase root and leaf productions compared to the significant effects achieved with one hormone treatment, NAA or IBA, which was consistent with the present findings.
In this study, stem cuttings treated with Clonex rooting hormone gel which contained 0.3% IBA had negative results on survival, root and leaf production compared to 0.1% synthetic IBA treatment. This finding was similar to studies of stem cuttings propagation of three tropical species, whereby cuttings treated with commercial rooting hormone Clonex also showed lower rooting percentage compared to cuttings treated with synthetic IBA22 and this could possibly be due to high IBA concentrations in Clonex.
The use of single auxins (especially 0.2% NAA) will result in a high survival rate of stem cuttings. The stem cutting technique with plant hormone will produce plantlets that will be used in the revegetation of tropical degraded heath forests in Brunei Darussalam. Raising plantlets from vegetative propagation using plant hormones then consecutively using them to revegetate forests are not new7,8. There is a possibility that the revegetation programme in Lumut using D. suffruticosa will also be extended to the Peat Swamp forests as this species is well known to be grown in many forest habitats10.
It is concluded that Dillenia suffruticosa stem cuttings grow better when treated with single auxin treatment than using commercial root-promoting hormones. Synthetic auxins, in particular 0.2% NAA and 0.1% IAA are more suitable for use in the mass propagation of D. suffruticosa stem cuttings. D. suffruticosa plantlets can be rapidly mass produced using stem cutting techniques with a single auxin treatment (mainly NAA and IAA), which can then be transplanted to the degraded heath forests in Lumut. The suitability of this species in the revegetation and rehabilitation programme will be further assessed via field monitoring of planted stem cuttings.
SIGNIFICANCE STATEMENTS
| • | The effects of auxins on growth of Dillenia suffruticosa cuttings were investigated for 16 weeks |
| • | This study compared the effects of single auxin and commercial root-promoting hormone treatments on survival and rooting percentages and leaf and root production of cuttings |
| • | A higher survival and rooting percentages and significantly longer root and greater number of roots were reported in NAA-treated cuttings than the other hormone treatments and water |
| • | Based on the number and surface area of new leaves, IAA performed significantly better than other hormones |
| • | It is concluded that compared to the commercial root-promoting hormones, 0.2% NAA and 0.1% IAA were the most effective auxins in propagating Dillenia suffruticosa cuttings |
| • | The raised plantlets will be used in revegetation of degraded heath forests in Brunei Darussalam |
ACKNOWLEDGMENTS
This research study was funded by the ASEAN-Korea Environmental Cooperation Project (AKECOP) for Brunei Darussalam awarded to Universiti Brunei Darussalam. Many thanks to the Forestry Department, Ministry of Primary Resources and Tourism for entry and collection permit (Reference No. [70]/JPH/UND/17 PT. 1). The authors are grateful to Dr Jonathan Davies (Wetlands International Brunei) and Noralinda Ibrahim (Forestry Department) for useful advice and field support. We also thank BLNG for permission to access and conduct research at Lumut and for logistical support.