Effect of species, cutting position and exogenous rooting substances on rooting of honeybush (Cyclopia spp.) cuttings

Abstract

Effect of the cutting position, terminal and subterminal, and rooting substance on the rooting of cuttings of two Cyclopia species (Cyclopia genistoides and C. intermedia) taken in winter, spring or summer was studied under controlled glasshouse conditions. Cuttings taken in summer, irrespective of the species, rooted better than cuttings taken either in winter or spring. This poor rooting performance of cuttings in winter and spring resulted in no significant treatment effects and for this reason only results of cuttings taken in summer are discussed. Cyclopia genistoides rooted significantly better than C. intermedia as measured by rooting percentage, number of roots per cutting, length of longest root and mean root length. Terminal cuttings from C. genistoides also increased rooting percentage and number of roots compared to subterminal cuttings, but not so in C. intermedia. Longest roots were produced by terminal cuttings irrespective of species. Treatment with different exogenous rooting substances as a main factor did not have any significant effect on rooting in this study.

Keywords:

• Cyclopia genistoides
• Cyclopia intermedia
• honeybush
• subterminal cutting
• terminal cutting

Cyclopia, which grows in the coastal districts of the Western and Eastern Cape provinces of South Africa, is a very distinct genus of the Podalyrieae and is classified as a member of the Fabaceae family (Schutte 1995). The stems and leaves of Cyclopia species are used to manufacture a sweetish herbal infusion known as honeybush tea (du Toit et al. 1998). The bushes are normally found on the shady and cooler southern slopes of mountain ranges and are about 1.5 m in height, but can reach up to 3 m (Bond and Goldblatt 1984).

Currently, commercial plantations of Cyclopia species are mainly established from seed and this has been indicated to result in variation in the quality of tea. Seedlings from indiscriminately collected seed from the wild or from commercial plantations resulted in large variation within plantations (Joubert 2010). This prompted a need to find alternative means of propagation other than by seed. Vegetative propagation by cuttings can be used as a tool for improving the quality of commercially produced species of honeybush (Joubert et al. 2011). Vegetative plant propagation is the most important method used in clonal regeneration of many horticultural crops such as ornamentals, fruits, nuts and vegetables (Nanda and Kochhar 1987, Hartmann et al. 2002). Establishment and growth rate of stem cuttings, however, depends on many factors, such as seasonal and age variation, portion and diameter of the stem, growing media, moisture level, nutrient status and temperature, amongst others (Hartman et al. 2002). Auxins, a plant growth substance, have an essential role in coordination of many growth and behavioural patterns in the plant life cycle (Hobbie 1998, Delker et al. 2008). Indole-3-acetic acid (IAA), α-naphthalene acetic acid (NAA) and indole-3-butyric acid (IBA) are typically the principal auxins that are available commercially and can be applied in liquid (liquid formulation) or in talc (powder formulation) for rooting and sprouting of stem cuttings (Ercìslí and Guleruz 1999, Hopkins and Hüner 2009). This study was conducted to evaluate the effect of rooting substances (IBA solution, a combination of IBA and NAA solutions, and IBA in talc) and cutting position, both terminal and subterminal, on rooting of two Cyclopia species (C. genistoides and C. intermedia) over different seasons.

Experiments were conducted in a glasshouse (33°56′47.32″ S, 18°15′59.16″ E; altitude 130 m above sea level [masl]) at the University of Stellenbosch. Mother stock plants of C. genistoides and C. intermedia were collected randomly in different seasons from a seven-year-old plantation on the experimental farm of ARC–Infruitec/Nietvoorbij (33°54′53.84″ S, 18°51′40.09″ E; altitude 156 masl) situated in Stellenbosch, Western Cape, South Africa. The stock plants were collected in the morning while they were still turgid and stored in a cold-room (4–6 °C) within an hour, to prevent moisture loss and to decrease biochemical activity. The cuttings were removed from the cold-room within a few hours and prepared for uniformity (length and stem size) and treated immediately thereafter.

Cuttings with a stem diameter of ±4 mm and length of ±8 cm with bottom leaves stripped were treated with Seradix^® 2 powder (active ingredient 4-indole-3-butyric acid at 3 g kg^-1), Dip & Root^™, a commercial rooting stimulator (active ingredient IBA at 10 g l^-1 and NAA at 5 g l^-1), and 2 and 4 g l^-1 IBA solutions. Cuttings were dipped in distilled water as a control treatment. Cuttings that were taken from the upper ±10 cm of the terminal portion of the branches were regarded as terminal cuttings, whereas any other cutting obtained after removing the terminal portion of the shoot was referred to as a subterminal cutting. Immediately after application of the treatments, the cuttings were planted in polystyrene trays filled with a moist growing medium of river sand and fine pine bark (1:1) as a rooting substrate. The trays were placed in a controlled environment in a glasshouse, with daily misting scheduled to irrigate for 30 s every 30 min. The glasshouse was set at a 20/25 °C night/ day temperature and no bottom heating was applied.

The experiment was laid out as a split plot (Snedecor and Cochran 1967) with rooting substances allocated to main plots, and species and cutting position as subplots. All treatment combinations were replicated five times. Each experimental unit consisted of 10 cuttings. Rooting percentage, number of roots per cutting, length of longest root and mean root length were assessed for analysis after 62 d of growth.

Analysis of variance (ANOVA) was performed using the GLM (General Linear Model) procedure of SAS statistical software version 9.1 (SAS Institute 2000). The Shapiro-Wilk test was performed to test for normality (Shapiro and Wilk 1965). Student's t-least significant difference was calculated at the 5% confidence level to compare treatment means (Ott 1998). Data for different rooting seasons were analysed separately.

Although experiments were replicated in spring, summer and winter, generally very poor rooting was experienced with winter and spring cuttings with no significant differences because of the treatments applied. For this reason only results from the summer cuttings will be presented and discussed. The highest rooting percentage (86%) was obtained from terminal cuttings of C. genistoides when treated with 2 g l^-1 IBA. However, no significant differences were found between the four rooting treatments (including the control) in terms of rooting percentage, number of roots, length of longest root or the mean root length (Table 1). This may be an indication that cuttings of these two Cyclopia species have the potential to root with or without treatment with rooting-promoting substances during the summer season.

Rooting percentage, number of roots, length of longest root per cutting and mean root length were highly significantly affected by species, but both rooting percentage and number of roots per cutting showed a sigificant interaction with cutting position (Table 1). In general, in comparison with C. intermedia, C. genistoides had a higher rooting percentage (57.8 vs 28.6), produced more roots (2.01 vs 0.71) and had both the longest root (21.5 vs 7.59 mm) and longest mean root length (13.73 vs 5.63 mm) (Table 2). In this study, cuttings of C. intermedia had fewer thick roots in comparison with the many, but thin roots in C. genistoides (MMM and EYR pers. obs.).

Table 1 : Significant levels of the main factors on rooting of honeybush (Cyclopia spp.) cuttings during summer

	Pr > F
Source	Rooting percentage	Number of roots	Length of longest root	Mean root length
Hormone treatment (T)	ns	ns	ns	Ns
Species (S)	<0.0001	<0.0001	<0.0001	<0.0001
Cutting position (P)	<0.0001	<0.0001	0.0360	ns
SxP	0.001	0.0016	ns	ns
TxS	ns	ns	ns	ns
TxP	ns	ns	ns	ns
TxSxP	ns	ns	ns	ns

Table 2 : Effect of species on rooting of honeybush (Cyclopia spp.) cuttings during summer. Means within a column with the same letter are not significantly different (LSD = 0.05)

	Per cutting
Species	Rooting percentage of roots	Number of roots	Length of longest root (mm)	Mean root length (mm)
C. genistoides	57.80^a	2.01^a	21.51^a	13.73^a
C. intermedia	28.60^b	0.71^b	7.59^b	5.63^b

Cutting position had a significant effect on rooting percentage, number of roots and lengths of longest root per cutting but, as already mentioned, significant interactions between species and cutting position occurred for rooting percentage and number of roots per cutting (Table 1). From Table 3 it is, however, clear that the interactions are because of the absence of significant differences between rooting percentage and number of roots from terminal and subterminal cuttings of C. intermedia. Terminal cuttings from C. genistoides showed 74.8% rooting compared to 40.8% from subterminal cuttings and 2.89 roots per cutting compared to 1.13 roots from subterminal cuttings. Cyclopia intermedia, in comparison, showed only 32.8% and 24.4% rooting and 0.98 and 0.44 roots from terminal and subterminal cuttings, respectively.

Table 3 : Interactive effect of species and cutting position on rooting of honeybush (Cyclopia spp.) cuttings during summer. Means within a column with the same letter are not significantly different (LSD = 0.05). T = Terminal cutting position, ST = subterminal cutting position

	Per cutting
Species	Cutting position	Rooting percentage (%)	Number of roots	Length of longest root (mm)	Mean root length (mm)
C. genistoides	T	74.80^a	2.89^a	25.32^a	15.33^a
C. intermedia	T	32.80^bc	0.98^b	7.80^c	6.15^b
C. genistoides	ST	40.80^b	1.13^b	17.70^ab	12.12^a
C. intermedia	ST	24.40^c	0.44^bc	7.39^c	5.10^b

On average, terminal cuttings had a higher rooting percentage (53.8%) compared to subterminal cuttings (32.6%) and produced on average 1.94 roots per cutting with a longest root of 16.56 mm compared to 0.79 roots and 12.55 mm, respectively, for subterminal cuttings (Table 3). Terminal cuttings are generally believed to give the highest rooting percentage and are usually recommended for propagation of most cuttings, although some commercial nurseries also use subterminal cuttings with good results (Brits 1986, Harré 1988, Malan 1992). From this study, it was clear that terminal cuttings rooted better than subterminal cuttings and, thus, would be ideal to use for rooting purposes.

Results of this study indicated that stem cuttings of C. genistoides and C. intermedia taken in summer can be rooted more easily than in winter or spring. Although the highest rooting percentage (86%) was obtained from terminal cuttings of C. genistoides when treated with 2 g l^-1 IBA during summer, responses to rooting treatments were not significant. Terminal cuttings of C. genistoides rooted better than subterminal cuttings of this species and significantly better than terminal or subterminal cuttings of C. intermedia. More research is, however, needed before specific treatments can be recommended.

Acknowledgements

The authors acknowledge the Agricultural Research Council (ARC), Agribusiness in Natural African Plant Products (ASNAPP) and the National Department of Agriculture for funding this study.