Livestock Research for Rural Development 30 (1) 2018 Guide for preparation of papers LRRD Newsletter

Citation of this paper

Mechanical scarification and hot water treatments enhance germination of Leucaena leucocephala (Lam.) seeds

M Koobonye, B V Maule and K Mogotsi

Animal Production and Range Research Division, Department of Agricultural Research, Ministry of Agricultural Development and Food Security, P O Box 10275, Francistown, Botswana
kbmogotsi@gov.bw

Abstract

Seed coat-imposed dormancy in seeds of the leguminous forage Leucaena leucocephala is an important impediment to rapid, uniform and high germination rates. Thus to test the effectiveness of several presowing treatments on germinability of L. leucocephala seeds, the following treatments were imposed under a Completely Randomized Design with 4 replications of 100 seeds each; (1) mechanical chipping with a nail clip, (2) soaking in hot water at 80oC for 5 minutes, (3) preheating at 600C for 150 minutes in an air-circulating oven, (4) immersion in concentrated sulphuric acid for 5 minutes as well as (5) untreated seeds as control.

Final germination percentage was highest for mechanical clipping (97.8%) and the hot water (84.3%) treatments. The untreated seeds (control) attained only 2% final germination with low values for dry heat (22.8%) and sulphuric acid (5%) treatments. Mechanical scarification greatly reduced germination time as most seeds (87.5%) sprouted in the first 4 days and germination was complete by day 7 whilst the hot water, dry heat and sulphuric acid treatments had 14, 5 and 3.25% germination count respectively by day 4. It is therefore concluded that, while mechanical scarification gave the highest final germination percentage and rate, the hot water treatment was still highly effective in overcoming physical dormancy in L. leucocephala seeds when compared to untreated seeds. Practically, this is especially important when considering the labour intensiveness of mechanical scarification when bulk pretreatment of seeds is required.

Keywords: germination percentage, germination rate, physical dormancy, seed pre-treatment


Introduction

Leucaena leucocephala (Lam.) is a multipurpose perennial legume tree species used as livestock feed due to its high nutritive value and palatability (Jones 1979), to improve soil characteristics and in soil erosion control, in agroforestry systems as shade crops or windbreaks and as fuel wood (Shelton and Brewbaker 1998; Graham and Vance 2003). Because of its multiplicity of uses, L. leucocephala can be of importance especially to rural agricultural systems in arid and semi arid regions of southern Africa such as Botswana (APRU 1980; Aganga and Tshwenyane 2003), even though the plant’s biology and ecology predisposes it to invasion (Olckers 2011). That notwithstanding, the planting and successful adoption of this drought tolerant forage is limited to some extent by water impermeability of the seed coat or ‘hardseededness’, caused by one or more water-impermeable layers of palisade cells in the seed or fruit coat (Baskin et al 2000). Apart from it being impermeable to water and/or oxygen, the seed coat can also exert germination-restrictive action by its mechanical resistance to radicle protrusion as well as harbouring inhibitors to suppress seed germination. In nature, this seed coat dormancy may be overcome by passage through the digestive tract of animals. For example, Gardener et al (1993) found that seeds of L. leucocephala germinated to about 90% after 240-hour residence in rumen of cattle. Other natural factors observed to overcome seed physical dormancy include abrasion, fire, drying, freezing/thawing, high or widely fluctuating soil temperatures and microbial activity (Baskin and Baskin 1998; Owens et al 1995). From an ecological viewpoint, this physical dormancy occurring in 15 plant families of Angiosperms (Baskin et al 2000) is an important survival strategy as seeds time their germination to coincide with favourable natural conditions to maximize chances of successful establishment of seedlings.

However, it is this very same ecological adaptation that is disadvantageous when rapid, uniform and high germination rate of L. leucocephala seed is required. Thus several techniques to break seed coat-imposed dormancy have been investigated over the years including; soaking in hot water or acid (Gray 1962; Jones 1970; Quinlivan 1971; Rolston 1978; Oakes 1984; Shelton and Brewbaker 1998; Tadros et al 2011), mechanical scarification/nicking (Babeley and Kandya 1985; Padma et al 1994; Gosling et al 1995) as well as varying levels of exposure to dry heat treatments (Mott et al 1982).

Furthermore, dormancy of seeds can also be influenced by the diversity of climates and habitats in which seeds are produced as well as time of harvest (Finch-Savage and Leubner-Metzger 2006), thus the need for this study to investigate germination ecology of L. leucocephala seed when exposed to pre-sowing treatments in semi arid Botswana. Such information could enable successful planting, establishment and utilization of the species particularly as livestock feed to augment existing protein sources like Lablab purpureus (Madzonga and Mogotsi 2014) and indigenous forage trees (Moleele 1998; Dambe et al 2015) during drought years as well as benefit potential forage seed industry in the country and elsewhere.


Materials and methods

Seeds of L. leucocephala harvested from Impala Research Station (2108’ - 2111’ S, 2135’ - 2737’ E, 470mm annual rainfall, altitude of 1020m) near Francistown, Botswana were used in the study. Mature seeds were collected from dried pods just before dehiscence of the pod. Seeds were removed from pods manually and separated out to leave only intact, filled seeds.

Seeds were subjected to the following treatments; (1) mechanical chipping with a nail clip, (2) soaking in hot water at 80oC for 5 minutes, (3) preheating at 600C for 150 minutes in an air-circulating oven, (4) immersion in concentrated sulphuric acid (H 2SO4 98%) for 5 minutes followed by rinse with tap water as well as (5) untreated seeds as control. The treatments were arranged in a Completely Randomized Design (CRD) with 4 replications consisting of 100 seeds each. Treated seeds were placed between two layers of moist paper towel (Between Paper method) following ISTA (2009; 2013), after which the rolled towels were placed upright in transparent sealed containers in a growth chamber. A constant temperature of 30C was maintained. Seeds were considered germinated following the rupture of the seed coat and emergence of the radicle. Germinated seeds were discarded after counting. The germination period lasted for 10 days, with germination counts done at days 4, 7, 9 and 10. Normal and abnormal seedlings as well as hard, fresh and dead seeds were classified as described by ISTA (2009).

Data were subjected to one-way analysis of variance (ANOVA) using SAS statistical software package. Treatment means were separated using the Least Significant Difference (LSD) test at p<0.05.


Results and discussion

Germination of seeds of L. leucocephala was influenced by the different treatment techniques applied (Table 1). These actions break dormancy by disrupting or softening the seed coat or by fracturing specialized tissues in the seed coat to allow water penetration (Owens et al 1995). Final germination percentage was highest for mechanical clipping (97.8%) and the hot water (84.3%) treatments. The dry heat (22.8%) and sulphuric acid treatments (5%) attained relatively low values while untreated seeds (control) attained only 2% germination count. Mechanical scarification of the seed testa greatly reduced germination time as most seeds (87.5%) sprouted in the first 4 days and germination was complete by day 7 whilst the hot water, dry heat and sulphuric acid treatments had 14, 5 and 3.25% germination count respectively by day 4.Similar results were obtained by Padma et al (1994) who found that germination of L. leucocephala seeds could be improved by grindstone scarification, nicking or by soaking the seeds in 80C water for 5 minutes while Tadros et al (2011) found that seeds soaked in 70C water for 20 minutes and then soaked for 24, 48 or 72 hours in water at room temperature had germination rates above 97%. However, caution should be exercised as prolonged exposure to excessive heat may damage the embryo and thus negatively affect germination rates. Gosling et al (1995) also reported L. leucocephala seeds’ sensitivity to hot water treatments and observed significant reduction in germination capacity with increasing temperatures. In the same study by Tadros et al (2011), blade scarification treatment of L. leucocephala resulted in 97% seed germination which was just as effective as the soaking treatments, whereas sandpaper scarification was not enough to overcome the physical barrier to allow germination and thus failed to facilitate water imbibition or permeability of the seed coat to water and oxygen. In the present study, most seeds germinated within the first 4 days of count under the mechanical clipping treatment, while none went beyond 7 days (Figure 1). This was closely followed by the hot water treatment where the majority of germination was observed at the day 7 count. This is worth considering since longer periods of germination can expose seeds to deterioration and infection by pathogens.

Table 1. Percentage of normal seedlings, hard, fresh and dead seeds following different pre-sowing treatments of L. leucocephala seeds

Treatment

Normal
seedling (%)

Hard
(%)

Fresh
(%)

Dead
(%)

Mechanical scarification (nail clip)

97.8a

0.00a

0.00a

2.25a

Soaking in 80oC H2O for 5 min

84.3b

9.5b

2.00b

4.25a

Heating in 600C oven for 150 min

22.8c

73.8c

1.75b

1.75a

Immersion in 98% H2SO4 for 5 min

5.00d

93.5d

0.00a

1.50a

Control (untreated)

2.00d

95.5d

1.00ab

1.50a

SEM

5.61

9.71

1.11

4.95

p

0.0001

0.0001

0.0444

0.399

abcd Means in the same column with significantly different letters are different at p<0.05



Figure 1. Germination response of L. leucocephala seeds under different pre-sowing treatments as observed at days 4, 7 and 10

The use of dry heat treatment (analogous to heating by vegetation fire) did slightly increase germination of L. leucocephala seeds (with final germination percentage at 22.8%) compared to the control but this result was still lower vis--vis the clipping and hot water treatments. In contrast, Mott et al (1982) found optimum heat treatment to be 140C for 30 seconds whereas seeds of the same species from another lot gave optimum germination rates when exposed to 140C for 60 seconds. Teketay (1996) also found that dry heat overcame dormancy in L. leucocephala when seeds were exposed to 80C for 60 minutes. Dry heat treatments have also been used successfully to overcome water-impermeability of seeds of other legume species such as Stylosanthes (Mott 1979), as heat causes cracks on the seed coat and the palisade layer of the strophiole to split, thereby speeding up the imbibition process (Baskin and Baskin 1998). For plant species with heat-stimulated germination, some of these conducive conditions may be potentially encountered by seeds at or near the soil surface on open sites or during fires (Keeley 1987). Just like with wet heat treatments, increased severity of dry heat has the potential to increase number of seeds killed and thus practicability of this scarification technique may require that seeds be exposed to temperatures less than the optimum to provide some margin of safety (Mott et al 1982).

The least effective treatment was the use of 98% sulphuric acid for 5 minutes, with only 5% final seed germination percentage attained. Egley (1989) reported that the disintegration of the seed coat as well as the micropylar plug allows increased imbibition and subsequent germination of seeds. Even though germination percentage was low in the current study, the use of concentrated sulphuric acid (soaking for 5-60 minutes) has been successfully demonstrated to increase germination rates in Leucaena species (Teketay 1996; Shelton and Brewbaker 1998) and several other tree legumes like Acacia and Prosopis species (e.g. Idu et al 2002; Hanaoka et al 2014). So it is possible that the observed poor germination could have been in part attributed to the less severity of the treatment i.e. short duration of immersion in the acid. When working with Corchorus species in Botswana, Emongor et al (2004) argue that the use of hot water to break dormancy may be the better option as some farmers may not have access to or do not know how to handle sulphuric acid.

In conclusion, the present study demonstrated that mechanical scarification with a nail clip as well as soaking in hot water (80C for 5 minutes) respectively are highly effective in overcoming coat-imposed seed dormancy in L. leucocephala when compared to untreated seeds – resulting in prompt and high germination which can aid in successful field establishment and/or maintenance of readily-available plant stock in nurseries. That notwithstanding, it is worth considering the practicality of such techniques when bulk pretreatment of seeds is required. Mechanical scarification is laborious and time consuming and can lead to considerable seed damage. It is thus recommended that hot water treatment, if mastered to improve its reliability, be used as an alternative by the farming community in Botswana as it is more convenient.


Acknowledgement

The Department of Agricultural Research provided resources for the study. The authors would like to thank the Seed Testing Laboratory staff at Sebele, especially O T Ndubo and M Lefhoko for technical assistance in implementation of experiments M I Ithuteng of the Biometry Unit at Sebele is appreciated for statistical analysis.


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Received 9 November 2017; Accepted 16 November 2017; Published 1 January 2018

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