Stomatal Traits and Flowering Behavior as Selection Criteria in Apricots

Abstract

Significant genotypic variation with reference to altitudinal changes was found in stomatal frequency, size, and flowering characters of sixteen apricot genotypes collected from different altitudes and planted in one orchard. Stomatal frequency ranged from 167–343 stomata/mm², and stomatal size ranged from 65–102 μm. In genotypes collected from higher altitudes, delayed flowering, more time requirement to complete flowering, and ripening was found. A decrease in petal length and fruit weight was also found with increasing altitude. A difference in leaf stomatal characters and flowering behavior in apricot genotypes may be taken into consideration as suitable traits for selection of apricot to be grown at different altitudes.

Keywords:

• Prunus armeniaca
• apricot
• stomatal size
• stomatal frequency
• flowering

INTRODUCTION

Apricot (Prunus armeniaca L.) is one of the economically important fruit trees, which is particularly prone to erratic fruit production (Mehlenbacher et al., 1990). The cause for this behavior is yet not properly understood. Recent climatic changes (i.e., global warming) differently affect plant processes at different altitudes. Tree growth might be adversely affected by evaporation of water from the leaves due to increased heat load (Coder, 1996). The mechanism developed by plants as a response to water stress may be found in adaptations by roots, stems, leaves, and fruits; leaf modifications include change in size and number of stomata (Kozlowski, 1976).

Leaf surfaces are equipped with small openings called stomata, which allow carbon dioxide to enter into the leaf and oxygen to escape, facilitating photosynthesis. The cost to the plant for this vital exchange is water loss. Apricot trees bear large canopies, representing a great evaporative surface and show low levels of root and stem hydraulic conductivity (Alarcon et al., 2000). Under high day temperatures, these characteristics may result in a transpiration rate that exceeds the water absorption capacity resulting in water stress. Stomata on plant leaves are considered to have an important role for adaptability to varying environmental conditions (Salisbury and Ross, 1992). Stomatal frequency differs greatly from species to species (Ryugo, 1988). Stomatal frequency may also vary in response to the amount of annual rainfall in different regions (Misirli and Aksoy, 1994). With increasing elevation, solar irradiance, leaf to air temperature differences, and the diffusivity of water vapor may increase, while air and soil temperatures may decrease, leading to a complex array of potential elevational effects on plant transpiration (Smith and Geller, 1979).

Stomata regulate the exchange of gases between leaves and the atmosphere and, thus, control the water use efficiency of photosynthesis, i.e., the balance between water loss and CO₂ uptake. The development of stomata (about 400 million years ago) is, therefore, considered to be a key event in the evolution of advanced land plants (Hetherington and Woodward, 2003). Stomatal diffusion resistance and, hence, conductance is directly related to the size and spacing of stomata on the leaf surface (Jones, 1992) so guard cell length and stomatal density both can be considered important ecophysiological parameters (Beerling et al., 1993). Guard cell length and stomatal density both are sensitive to growth environment, and there is considerable genotypic variation and phenotypic plasticity for these two traits. Elevational changes and temperature variation affects flowering and fruiting characters also, of these number of days to complete flowering process, petal length, fruit weight, number of days required for ripening, and total soluble solids of fruit pulp are more vulnerable traits. But, in most of the selection studies, these traits have not been considered. Therefore, an understanding of the relationship between stomatal features, flowering characters, and environmental factors would be of considerable importance for the better fruit tree growth on the warming globe.

MATERIALS AND METHODS

Present investigation was conducted at apricot orchard of the National Bureau of Plant Genetic Resources Regional Station Bhowali, District Nainital, Uttarakhand (India), which is approximately ten years old and planted at a plant-to-plant and row-to-row distance of 5 × 5 m. The orchard is located at 29° 24′ 28.7″ N latitude and 79° 30′ 47.2″ E longitude at an altitude of 1,480 m above sea level. The annual precipitation average over the last two decades was 1,200–1,800 mm. The soil is sandy loam with medium organic matter content and numerous pebbles. Sixteen accessions of apricot are available in the orchard collected from different altitudes ranging from 1,450–2,000 m and grown with regular cultural practices.

Stomatal size and frequency in leaves of apricot genotypes present in the apricot orchard as well as at locations from where these accessions were collected was measured in the summer of 2009 and 2010. For stomata counting, the fourth fully expanded young leaves located on the tip of two branches facing two different directions (south and north) per tree were collected and readings from the abaxial side of the leaves were recorded. The epidermis layer of the abaxial side of leaf was painted with clear fingernail polish and a piece of transparency sheet (2 cm²) was pressed on it by fingers. Later, the imprints of stomata were obtained by striping the transparency sheet from the leaves (Schechter et al., 1992). Two imprints were taken from each side of the main vein of a leaf (total four imprints per leaf). By using a light microscope with a 40 × 10 magnification, in a 0.18 mm² vision field, stomata were counted on transparent sheets that were placed on a glass slide. The obtained values were calculated as stomata numbers per mm². The counting was replicated on three trees for each accession. Guard cell length (the size of the stomata) was measured on 15 different stomata for each leaf sample. Objectives of 20× and 40× (depending on the size and density of stomata) were used in conjunction with a 5× ocular.

The date on which 5% of the flower buds opened was recorded as date of the start of flowering and the date on which 85–90% of the flower buds opened as the end of flowering. The length from base to tip of petal was expressed as petal length. The number of days from the end of flowering to the date of harvest was recorded as days to ripening. Total soluble solids in the fruit pulp at the time of harvesting are expressed as TSS.

Data for each parameter were evaluated for statistical significance using two-way analysis of variance (ANOVA) to compare the means considering genotype and altitude as independent variables. The variation between two groups was assessed by computation of least significant difference taking ‘t’ values for error d.f. at 5% level of significance.

RESULTS AND DISCUSSION

Fruit tree studies are mainly focused on production and productivity of the plants at the experimental site in an environment where the experiments are set up. But usually trees are often exposed to different environmental conditions that may adversely affect both the plant development and yield. Even if planted in similar environments, conditions may vary in due course of time due to climate change as found in the present experiment (Figs. 1 and 2).

FIGURE 1 Weekly mean maximum temperature in January at the experiment orchard.

FIGURE 2 Weekly mean maximum temperature in June at the experiment orchard.

The leaf stomatal frequency significantly varied among the apricot accessions. The highest leaf stomata frequency was 343 stomata/mm² found in IC-319186 growing at higher altitude (i.e., 2,000 m) and lowest was 167 stomata/mm² in IC-360702 growing at lower altitude (i.e., 1,475 m) (Table 1). In earlier studies, Kundu and Tigerstedt (1998) found that stomatal density and guard cell length varied among different origins of plant and stomatal density correlated with leaf-level net photosynthesis and whole-plant dry weight. Other comparative studies have indicated that xeric species typically have higher stomatal densities than mesic species (Carpenter and Smith, 1975) and sun leaves typically have higher stomatal densities than shade leaves (Lichtenthaler, 1985). Plants growing at higher altitudes experience more water scarcity as compared to those growing at lower altitudes, and these plants avoid a water stress situation by modulating their stomatal frequency as found in the present investigation. Sun/shade plasticity for stomatal density also varies among species. In three Quercus spp., Ashton and Berlyn (1994) found that the capacity for phenotypic plasticity is correlated with drought tolerance. Stomatal density appears to be relatively plastic compared to stomatal length (either guard cell length or stomatal aperture length), which shows less variation (Richardson et al., 2001). There is a general negative correlation between guard cell length and stomatal density (Carpenter and Smith, 1975). In conformity with this, in the present investigation genotypes having maximum stomatal frequency, i.e., IC 319186 (343 stomata. mm⁻²), had minimum stomatal sizes, i.e., 65 μm and vice versa. Across functional groups and even including fossil plants, this relationship is almost exactly compensatory. Although stomatal densities ranging from 5 to 1,000 mm² have been observed, the concurrent changes in mean guard cell length result in a nearly constant stomatal conductance (Hetherington and Woodward, 2003). Earlier studies show a consistent inverse relationship between stomatal density and atmospheric CO₂ concentration, this is thought to be an adaptive response to maximize water-use efficiency (Woodward, 1987). By analysis of enrichment experiments, Woodward and Kelly (1995) have shown that stomatal density generally decreases by about 9% in response to enrichment from 350 to 700 ppm CO₂ and stomatal density has been shown to vary predictably in response to past changes in atmospheric CO₂ concentration at a range of time scales (hundreds, thousands, and tens of thousands of years) (Beerling et al., 1993; Körner, 1988). In accordance with these findings in the present study, genotypes naturally growing at lower altitudes have low stomatal frequency and more at higher altitudes. This may be due to presence of higher CO₂ concentrations at lower altitudes as compared to high altitudes.

TABLE 1 Altitudinal Variation in Leaf Stomatal Frequency and Size of Apricot Genotypes ^z

		Altitude		Stomatal frequency (number.mm^−2)		Stomatal size (μm)
S. No.	Genotype (accession number)	At native place	At orchard	At native place	At orchard	At native place	At orchard
1	IC360694	1,500	1,480	177 ± 1.2	175 ± 2.1	85 ± 1.2	86 ± 1.8
2	IC360696	1,460	1,480	176 ± 1.3	177 ± 2.2	102 ± 1.6	101 ± 1.6
3	IC360697	1,450	1,480	173 ± 1.5	174 ± 2.3	81 ± 1.5	80 ± 1.4
4	IC360699	1,475	1,480	167 ± 1.6 ^y	168 ± 2.5	89 ± 1.4	88 ± 1.5
5	IC360700	1,475	1,480	174 ± 2.3	151 ± 2.1	79 ± 1.5 ^y	78 ± 1.6
6	IC360701	1,600	1,480	212 ± 2.5	194 ± 2.4	77 ± 1.2	71 ± 1.5
7	IC360702	1,800	1,480	254 ± 2.7	231 ± 2.3	79 ± 1.3	85 ± 1.6
8	IC360692	1,650	1,480	233 ± 2.5 ^y	224 ± 1.9	74 ± 1.6	78 ± 1.2
9	IC247427	1,600	1,480	234 ± 2.1	222 ± 1.5	74 ± 1.5	79 ± 1.4
10	IC247428	1,600	1,480	245 ± 1.6	236 ± 1.7	71 ± 1.2	77 ± 1.8
11	IC247429	1,600	1,480	229 ± 1.5	220 ± 1.6	77 ± 1.1	81 ± 1.6
12	IC273865	1,950	1,480	286 ± 1.6	273 ± 1.8	72 ± 1.6	81 ± 1.5
13	IC360686	1,900	1,480	301 ± 1.5	287 ± 2.0	78 ± 2.1	86 ± 1.4
14	IC360687	1,900	1,480	322 ± 1.4 ^y	308 ± 2.1	74 ± 2.0	82 ± 2.1
15	IC360688	1,900	1,480	257 ± 1.3	241 ± 2.3	70 ± 2.3	79 ± 2.4
16	IC319186	2,000	1,480	343 ± 1.7	324 ± 2.1	65 ± 2.5 ^y	75 ± 2.4
^zValues presented are mean ± S.E.
^ySignificant at p ≤ 0.05.

Stomatal conductance is a numerical measure of the rate of passage of either water vapor or carbon dioxide through the stomata. Conductance plays an important role in the plant-atmosphere water exchange and, hence, it is a key parameter in many ecological models (Chen et al., 1999). Diffusion of CO₂ into the mesophyll of leaves and water vapor from the leaves to the atmosphere is mainly driven by the stomatal aperture, which is controlled by a complex system of plant physiological processes. Nearly all the water transpired by plants is lost through the stomata as water vapor diffusion via cuticle can be neglected. Investigations under natural conditions are strongly affected, for instance, by water vapor pressure deficit, soil moisture, general conditions of the plant (healthy/unhealthy), position of leaves or needles (shaded/exponated), and age of leaves or needles; therefore, it is difficult to compare published data. Stomatal conductance to water vapor diffusion varies with incident solar irradiance, leaf to air humidity difference, vapor pressure deficit of the air, and plant water status. In high altitudes, elevational effects differently affect plant transpiration as compared to plains. Stomatal diffusion resistance and, hence, conductance, is directly related to the size and spacing of stomata on the leaf surface. Generally, the accessions growing in areas with higher average air temperature have higher conductance values. In annual plants, stomatal conductance was found to be related to yield, e.g., Ulloa et al. (2000) confirmed that high stomatal conductance was associated with high cotton lint yields at supra-optimal temperatures for the cotton crop under irrigated environments. In the present investigation, adjustment in stomatal size and number might be a strategy of plants to maintain optimal stomatal conductance.

With an increase in altitude, air temperature decreases and the plant intercepts less heat units per unit leaf area, which delays the start of flowering and increases duration of the flowering phase; thus, more days are required to complete the flowering process (Table 2). Due to lower temperatures at high altitudes, environmental conditions for growth and development are not congenial. Therefore, a decrease in petal length was also found (Table 2). An increase in number of days required for ripening was also found in genotypes of high altitudes (Table 3). The decrease in temperature might be the reason for delay in ripening. Fruit ripening is a biochemical process in which optimal temperature is an important factor for its proper regulation. The decrease in fruit weight found in the present investigation (Table 3) might be either due to a decrease in net photosynthate production or translocation of photosynthate to the sink. Both these processes play a crucial role in increasing fruit productivity. Therefore, changes in photosynthate production and translocation with respect to altitudinal variation might be another area yet to be addressed properly in this particular fruit crop. Moreover, TSS, which is the measure of total soluble sugars translocated and stored in the fruit, was also found to be decreased with increasing elevation (Table 3); the reason might be adverse conditions for plant growth at higher elevations and expenditure of more chemical energy for survival rather than to be translocated to fruit for storage. In spite of regular cultural operations, these plants show a variation in flowering and fruiting behavior as compared to plants of the same accession grown at the native place. Therefore, for planting apricots at a different elevation, consideration of these characters might help growers in deciding their preference for selection of genotypes.

TABLE 2 Altitudinal Variations in Date of Start of Flowering, Number of Days to Complete Flowering Process, and Petal Length ^z

		Date of start of flowering		Number of days to complete flowering process		Petal length (cm)
S. No.	Genotype (accession number)	At native place	At orchard	At native place	At orchard	At native place	At orchard
1	IC360694	15-Feb	15-Feb	21 ± 1.0	18 ± 1.2	1.5 ± 0.073	1.3 ± 0.072
2	IC360696	13-Feb	13-Feb	29 ± 1.2	19 ± 1.1	1.6 ± 0.065	1.3 ± 0.063
3	IC360697	13-Feb	14-Feb	23 ± 1.1 ^y	18 ± 1.2	1.6 ± 0.066	1.4 ± 0.061
4	IC360699	14-Feb	14-Feb	26 ± 0.9	19 ± 1.1	1.5 ± 0.055	1.4 ± 0.058
5	IC360700	15-Feb	14-Feb	28 ± 1.5	19 ± 1.3	1.4 ± 0.053	1.5 ± 0.054
6	IC360701	18-Feb	15-Feb	28 ± 0.9	19 ± 1.4	1.3 ± 0.052	1.3 ± 0.049
7	IC360702	20-Feb	14-Feb	34 ± 1.1 ^y	19 ± 0.9	1.0 ± 0.051 ^y	1.5 ± 0.053
8	IC360692	18-Feb	14-Feb	35 ± 0.8	18 ± 0.9	1.0 ± 0.048	1.5 ± 0.046
9	IC247427	19-Feb	14-Feb	31 ± 1.2 ^y	18 ± 0.9	1.1 ± 0.049	1.3 ± 0.042
10	IC247428	20-Feb	14-Feb	33 ± 0.9	19 ± 1.1	1.2 ± 0.051	1.4 ± 0.054
11	IC247429	21-Feb	14-Feb	33 ± 0.9	19 ± 1.0	1.1 ± 0.046	1.4 ± 0.047
12	IC273865	26-Feb	15-Feb	37 ± 1.5	20 ± 1.0	0.9 ± 0.041 ^y	1.5 ± 0.038
13	IC360686	25-Feb	14-Feb	38 ± 1.4	20 ± 1.2	0.9 ± 0.039	1.3 ± 0.041
14	IC360687	25-Feb	15-Feb	37 ± 1.2	20 ± 1.3	0.8 ± 0.036	1.3 ± 0.031
15	IC360688	27-Feb	15-Feb	36 ± 1.3	20 ± 1.0	0.9 ± 0.043	1.5 ± 0.044
16	IC319186	28-Feb	15-Feb	39 ± 1.2 ^y	20 ± 0.9	0.7 ± 0.031 ^y	1.4 ± 0.035
^zValues presented are mean ± S.E.
^ySignificant at p ≤ 0.05.

TABLE 3 Altitudinal Variations in Fruit Weight, Days to Harvest/Ripening and Total Soluble Solids ^z

		Fruit weight (g)		Days to harvest/ripening		Total soluble solids
S. No.	Genotype (accession number)	At native place	At orchard	At native place	At orchard	At native place	At orchard
1	IC360694	44 ± 0.68	43 ± 0.61	44 ± 0.85	42 ± 0.78	13.03 ± 0.31	13.00 ± 0.28
2	IC360696	42 ± 0.62	41 ± 0.58	42 ± 0.82	43 ± 0.74	13.46 ± 0.32	13.01 ± 0.25
3	IC360697	42 ± 0.79	40 ± 0.59	39 ± 0.84	41 ± 0.72	13.51 ± 0.35	13.10 ± 0.26
4	IC360699	41 ± 0.68	43 ± 0.64	44 ± 0.82	44 ± 0.75	13.41 ± 0.36	13.02 ± 0.25
5	IC360700	46 ± 0.67	43 ± 0.62	45 ± 0.81	43 ± 0.67	13.32 ± 0.32	13.01 ± 0.25
6	IC360701	50 ± 0.73 ^y	45 ± 0.52	48 ± 0.89 ^y	45 ± 0.76	13.27 ± 0.31	13.04 ± 0.27
7	IC360702	53 ± 0.66	45 ± 0.54	44 ± 0.87	43 ± 0.75	12.23 ± 0.38 ^y	13.05 ± 0.29
8	IC360692	51 ± 0.67	46 ± 0.56	47 ± 0.86	46 ± 0.70	11.97 ± 0.39	13.06 ± 0.24
9	IC247427	54 ± 0.67	46 ± 0.55	51 ± 0.85	45 ± 0.66	12.21 ± 0.35	13.16 ± 0.21
10	IC247428	58 ± 0.72 ^y	47 ± 0.51	46 ± 0.87	46 ± 0.72	12.51 ± 0.36	13.22 ± 0.22
11	IC247429	60 ± 0.70	46 ± 0.49	49 ± 0.88	43 ± 0.73	12.58 ± 0.34	13.14 ± 0.23
12	IC273865	60 ± 0.75	47 ± 0.48	52 ± 0.89 ^y	44 ± 0.65	11.93 ± 0.36 ^y	13.25 ± 0.26
13	IC360686	62 ± 0.74	47 ± 0.51	57 ± 0.90	45 ± 0.76	11.47 ± 0.38	13.26 ± 0.25
14	IC360687	63 ± 0.76	48 ± 0.44	54 ± 0.85	46 ± 0.63	11.74 ± 0.37	13.17 ± 0.24
15	IC360688	63 ± 0.73	48 ± 0.47	55 ± 0.81	44 ± 0.65	11.17 ± 0.39	13.18 ± 0.23
16	IC319186	64 ± 0.75	48 ± 0.49	58 ± 0.82 ^y	46 ± 0.68	11.02 ± 0.40	13.21 ± 0.25
^zValues presented are mean ± S.E.
^ySignificant at p ≤ 0.05.

Elevational studies can be seen as “natural experiments” from which we can learn about plant adaptation or response to different environmental conditions (Körner, 1999). Global warming is having a significant impact on plant species around the world, but the most dramatic effects may not be felt for decades. The geographical range of numerous species had shifted polewards or to a higher elevation indicating that some plants are occupying areas that were previously not suitable for survival of these plants. Therefore, differences in stomatal characters and flowering behavior that existed among the genotypes may be taken into consideration as selection criteria for apricots to be grown in regions with higher average temperatures. An understanding of the relationship between stomatal features, flowering behavior, and environmental factors would be of considerable importance for the better fruit tree growth on the warming globe.

ACKNOWLEDGMENTS

The authors are thankful to the Director, National Bureau of Plant Genetic Resources, Pusa, New Delhi for providing necessary facilities for the work.