The Impact of Seed Size and Aging on Physiological Performance of Lentil under Water Stress

Introduction

Crop yields are restricted by water shortages in many parts of the world (Austin 1989). The physiological responses of plants to water stress and their relative importance for crop productivity vary with species, soil type, nutrients and climate. Decreasing water content is accompanied by loss of turgor and wilting, cessation of cell enlargement, closure of stomata, reduction in photosynthesis and interference with many other basic metabolic processes (Kramer and Boyer 1995). Relative water content is a good sign for water status in plants and spots it better than water potential (Patakas et al. 2002). Membrane stability index (MSI) and relative water content (RWC) have been decreased as a result of water deficit (Bayoumi et al. 2008). Tolerant cultivars have more RWC in comparison with sensitive cultivars under drought stress (Patakas et al. 2002).

Plants can partly protect themselves against mild drought stress by accumulating osmolytes. Proline is one of the most common compatible osmolytes in drought stressed plants. For example, proline content of pea increased under drought stress (Alexieva et al. 2001). Other stresses such as salinity can also induce proline accumulation (Sairam et al. 2002; Ghassemi-Golezani et al. 2011). Proline metabolism in plants, however, has mainly been studied in response to osmotic stress (Verbruggen and Hermans, 2008). The accumulation of proline in plant tissue is, therefore, a clear marker for drought and salt stresses. Proline accumulation may also be part of the stress signal influencing adaptive responses (Maggio et al. 2002). Thus, increasing proline concentration can be used as an evaluating parameter for irrigation scheduling and for screening drought resistant varieties (Bates et al. 1973; Gunes et al. 2008).

Maximum seed quality is obtained at or slightly after mass maturity and thereafter seeds begin to deteriorate on mother plant (Ghassemi-Golezani and Mazloomi-Oskooyi 2008; Ghassemi-Golezani and Hosseinzadeh-Mahootchy 2009) and during storage, loose vigor and viability (Ellis and Roberts 1981). Several biochemical and physiological changes have been observed in seeds during aging, resulting in a progressive decline in seed quality (Marcos-Filho and McDonald 1998). When seed aging increases, germination rate and uniformity and tolerance to environmental stresses and consequently seedling emergence and post emergence seedling growth decrease (Khan et al. 2003).

Seed size is an important characteristic of many plant life histories (Harper et al. 1970), since it is generally proportional to the amount of food reserves that will be destined to the embryo (Lloret et al. 1999). Larger seeds have a better performance than small seeds, especially under competitive conditions (Moles and Westoby 2004). Results of some researchers clarified that seed size notably affected seedling establishment, plant height, seed weight and number of seeds per spike in wheat (Royo et al. 2006). Also, larger seeds with well-developed root systems of seedlings may gain an advantage by allowing to reach the soil moisture at deeper levels (Leishman and Westoby 1994). Since the interaction of seed size and aging on field performance of lentil is not documented, this research was carried out to investigate these interactive effects on some physiological characteristics and yield of lentil under different irrigation treatments.

Materials and Methods

Seeds of lentil (Lens culinaris Medik. cv. Kimia) were obtained from Dryland Agricultural Research Center, Kermanshah, Iran. A sub-sample of the seeds was kept as bulk (S₁) with 1000 grain weight of 42 g. The other seeds were separated by a sieve with four millimeters diameter. The seeds that remained on the sieve were considered as large (S₂) with 1000 grain weight of 50 g and those passed the sieve were considered as small (S₃) seeds with 1000 grain weight of 35 g. Seeds of each size were divided into three sub-samples. A sub-sample was kept as control or high vigor seed lot with 97.2% normal germination (A₁). The two other sub-samples with about 20% moisture content were artificially aged, using controlled deterioration test (ISTA 2010) at 40°C for 2 and 4 days, reducing germination to 90.8% and 82.3% (A₂ and A₃, respectively). So, three seed lots with different levels of aging were provided for laboratory tests and field experiment.

The field experiment was conducted at the Research Farm of the University of Tabriz (Latitude 38˚05’ N, Lon­gitude 46˚17’ E, Altitude 1360 m above the sea level) in 2011. All the seeds were treated with Benomyl at a rate of 2 g kg^-1 before sowing. Seeds were hand sown in about 5 cm depth with a density of 100 seeds m^-2 on 5th May 2011. Each plot consisted of 6 rows with 4 m length, spaced 25 cm apart. The experiment was arranged as split plot factorial, based on randomized complete block design with three replications. All plots were irrigated immediately after sowing and subsequent irrigations were carried out after 70 (I₁), 120 (I₂) and 170 (I₃) mm evaporation from class A pan. Weeds were removed by hand during crop growth and development.

Relative water content was determined according to Barr and Weatherley (1962). Fresh weight of the youngest fully expanded leaves of five plants from each plot was determined. Then, leaves were soaked in distilled water for 24 h. After that, the water on the leaves was quickly and carefully removed by tissue paper and subsequently turgid weight was recorded. Leaf dry weight was obtained after drying the samples for 48 h at 75°C. Relative water content was calculated by the following equation:

RWC = [(fresh weight – dry weight) / (turgid weight – dry weight)] × 100

Leaf temperature (ºC) was directly measured by an infra-red thermometer (TES-1327) at the flowering stage. Fresh leaf samples (0.1 g) from each plot were taken in 10 ml double-distilled water in glass vials and kept at 40ºC for 10 min. Initial conductivity (C₁) was recorded with a conductivity meter after transferring the sample to 25ºC. The samples were kept at 100ºC for 30 min and cooled at 25ºC. Final conductivity (C₂) was measured according to Sairam (1994). The membrane stability index (MSI) was calculated as:

MSI = [1 – (C₁/C₂)] × 100

Proline content was determined according to Bates et al. (1973). Ground cover was measured every week by viewing the canopy through wooden frame (50 × 50 cm dimensions) divided into 100 equal sections. The sections were counted when more than half filled with crop green area. Finally, plants of 1 m² in the middle part of each plot were harvested and grain yield per unit area was determined. Analyses of variance of the data based on the experimental de­sign and comparison of means at P≤0.05 were carried out, using MSTATC software.

Results

Analyses of variance of the data showed significant effects of irrigation treatments on RWC, leaf temperature, proline, ground cover and grain yield per unit area. Ground cover and grain yield per unit area significantly affected by seed size and relative water content (RWC), leaf temperature, membrane stability index (MSI), ground cover and grain yield were significantly influenced by seed aging (Table 1). Interactions of irrigation × seed size for ground cover and grain yield per unit area, irrigation × seed aging for relative water content (RWC) and membrane stability index (MSI) and seed size × seed aging for ground cover were also significant (Table 1).

           Table 1. Analysis of variance of the effects of seed size and aging on some

           physiological traits of lentil under different irrigation treatments

             *, **: Statistically significant at p≤0.05 and p≤0.01, respectively

             RWC: Relative water content, MSI: Membrane stability index

Relative water content (RWC), ground cover and grain yield per unit area significantly decreased with decreasing water availability, but mean leaf temperature and proline content increased as water stress severed. The highest RWC, MSI, ground cover and grain yield were recorded for plants from the non-aged seed lot (A₁) (Table 2).

                    Table 2. Means of physiological traits and grain yield of lentil  affected by irrigation

                     treatments and seed aging

Different letters in each column for each treatment indicate significant difference at p≤ 0.05

I₁, I₂ and I₃: Irrigations after 70,120 and 170 mm evaporation from class A pan, respectively

A₁, A₂ and A₃: Control and aged seed lots of lentil for 2 and 4 days at 40°C, respectively

RWC: Relative water content, MSI: Membrane stability index

Seed aging decreased RWC and MSI of the resultant plants under all irrigation treatments. This decline for the A₃ plants under severe water limitation was larger than that under other irrigation treatments (Figure 1).

Figure1. Means of relative water content (RWC) and membrane stability index (MSI) for plants from differentially aged seed lots of lentil under different irrigation treatments

Different letters indicate significant difference at p≤ 0.05

 I₁, I₂ and I₃: Irrigation after 70, 120 and 170 mm evaporation from class A pan, respectively

A₁, A₂ and A₃: Control and aged seed lots of lentil for 2 and 4 days at 40°C, respectively

Ground cover was reduced with increasing seed aging; the highest reduction was recorded for plants from the most aged small seeds (Figure 2a). Plants from large seeds were superior in ground cover and grain yield per unit area under limited irrigation conditions (I₂ and I₃). However, these traits for the plants from bulk and large seeds under well watering were statistically similar. Plants from small seeds produced the lowest ground cover and grain yield under all irrigation treatments (Figures 2b, 2c).

Discussion

The decrease in leaf RWC as a result of water stress (Table 2) could be related to low water uptake and more transpiration rate under stress conditions (Lourtie et al. 1995). Lower RWC of plants from most aged seeds (A₃) due to water deficit (Figure 1) was the result of delayed seedling emergence and less rooting. However, vigorous plants can tolerate drought stress, because of expanded rooting system and better water uptake (Ghassemi-Golezani et al. 2012).

Figure 2. Means of ground green cover for lentil plants affected by seed size × aging (a) and

seed size × irrigation treatments (b) and grain yield of plants from

different seed sizes grown under different water supply (c)

Different letters indicate significant difference at p≤ 0.05

I₁, I₂ and I₃: Irrigations after 70, 120 and 170 mm evaporation from class A pan, respectively

S₁, S₂ and S₃: Bulk, large and small seeds of lentil, respectively

A₁, A₂ and A₃: Control and aged seed lots of lentil for 2 and 4 days at 40°C, respectively