STUDY OF GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN (NIGELLA SATIVA L.) TREATED BY HYDRO AND OSMOPRIMING UNDER SALT STRESS CONDITIONS
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GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN UNDER SALT STRESS CONDITIONS
Cercetări Agronomice în Moldova
Vol. XLVIII , No. 2 (162) / 2015
STUDY OF GERMINATION AND SEEDLING GROWTH
OF BLACK CUMIN (NIGELLA SATIVA L.) TREATED
BY HYDRO AND OSMOPRIMING UNDER SALT
STRESS CONDITIONS
A. AHMADIAN1*, Y. SHIRI2, M. FROOZANDEH2
*E-mail: Aahmadian59@gmail.com
Received October 20, 2014
ABSTRACT. The objective of the study weight, shoot and root length were higher
was to determine the responsible factors for but mean germination time and abnormal
germination and early seedling growth due germination percentage were lower than
to salt toxicity or osmotic effect and to Na2SO4, at the same water potential. The
optimize the best priming treatment for root / shoot weight and R/S length enhanced
these stress conditions. To study the effect with increase of osmotic potential in both
of osmopriming and hydropriming on NaCl and Na2SO4 solutions. NaCl had less
germination and seedling growth of black inhibitor effect on seedling growth than the
cumin (Nigella sativa L.) under salt stress germination. It was concluded that
conditions this experiment was conducted at inhibition of germination at the same water
Torbat-Heydariyeh University, Torbat, Iran. potential of NaCl and Na2SO4 resulted from
The treated seeds (control, hydropriming salt toxicity rather than osmotic effect. The
and ZnSO4) of black cumin were evaluated findings of this experiment can be useful
at germination and seedling growth for and applied to achieve best germination and
tolerance to salt (NaCl and Na2SO4) uniform emergence under field conditions
conditions at the same water potentials of for farmers of medicinal plants.
0.0, -0.3, -0.6, -0.9 and -1.2MPa. Electrical
conductivity (EC) values of the NaCl Key words: Black cumin (Nigella sativa
solutions were 0.0, 6.5, 12.7, 18.4 and 23.5 L.); Salt stress; ZnSO4; Priming; Seedling.
dSm-1, respectively. Results showed that
hydropriming increased germination and INTRODUCTION
seedling growth under salt stress.
Germination delayed in both solutions, The black cumin (Nigella sativa
having variable germination with different L.), belonging to the family
priming treatments. In NaCl treatment, Ranunculaceae is an ancient crop
germination percentage, root and shoot which is originated in the East-
1 Department of Medicinal Plants, Saffron Institute, University of Torbat-e Heydariyeh, Iran
2 Agriculture Research Center, University of Zabol, Iran
69A. AHMADIAN, Y. SHIRI, M. FROOZANDEH
southern Europe. It is particularly with NaCl and Na2SO4 solutions
grown in southern Europe (The increased germination percentage and
Balkans), northern Africa and rate and primary seedling growth
Hindustan. The black cumin is under salt stress. Seed priming has
generally short-lived annual, typical been successfully demonstrated to
of disturbed soils or natural improve germination and emergence
communities of semiarid regions, with in seeds of many crops, particularly
a dominance of therophytes. In the seeds of vegetables and small seeded
natural form, flowers are bluish, with grasses (Heydecker and Coolbaer,
a variable number of multi-ovule 1977; Bradford, 1986). The beneficial
carpels, developing into a follicle after effects of priming have also been
pollination, with single fruits, partially demonstrated for many field crops
connected to form a capsule structure. such as wheat, sugar beet, maize,
Seeds, of generally small size (1-5 soybean and sunflower (Parera and
mg), dark grey or black color and with Cantliffe, 1994; Singh, 1995; Khajeh-
corrugated integuments, represent the Hosseini et al., 2003; Sadeghian and
useful product (Michel, 1983; El- Yavari, 2004). Dharmalingam and
Fouly, 1983). Basu (1990) reported beneficial effect
A major constraint to seed of a hydration-dehydration seed
germination is soil salinity, a common treatment on germination of
problem in irrigated areas of Iran, sunflower. Rao et al. (1987) reports
with low rainfall (Kaya et al., 2003; that primed Brassica seeds may
Kafi, 2002; Kizil et al., 2003; Mehra et reduce the risk of poor stand
al., 2003; Mwale et al., 2003). Soil establishment in cold and moist soils.
salinity may affect the germination of However, Singh and Rao (1993)
seeds either by creating an osmotic stress that KNO3 effectively improved
potential external to the seed, germination, seedling growth and
preventing water uptake, or through seedling vigour index of the seeds of
the toxic effects of Na+ and Cl- ions sunflower varieties with low
on the germinating seed (Khajeh- germination.
Hosseini et al., 2003). Salt and The aims of the present study
osmotic stresses are responsible for were to determine the responsible
both inhibition or delayed seed factors for failure of germination of
germination and seedling black cumin seeds under saline
establishment (Almansouri et al., conditions due to the toxic effects of
2001; Angadi et al., 2002). Under NaCl and Na2SO4 by comparing seed
these stresses there is a decrease in germination under a range of osmotic
water uptake during imbibition and, potentials due to NaCl and Na2SO4.
furthermore, salt stress may cause Furthermore, the study examined the
excessive uptake of ions (Murillo- possibilities to overcome salt stress by
Amador et al., 2002). seed treatments with hydropriming or
Ahmadian et al. (2009) reported treatment with ZnSO4.
that hydropriming and osmopriming
70GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN UNDER SALT STRESS CONDITIONS
MATERIALS AND METHODS 1996). The rolled paper with seeds was
put into sealed plastic bags to avoid
This experiment was carried out at moisture loss. Seeds were allowed to
the Department of Medicinal plants, germinate at 25±1°C, in the dark, for 7
Faculty of Agriculture, University of days. Germination was considered to have
Torbat-e Heydariyeh, Iran in 2012. occurred when the radicles were 2 mm
Germination and early seedling growth (7 long. Germination percentage was
days) of the cultivar were studied using recorded every 24 h, for 7 days. To
distilled water (control) and under determine the toxic effects of the
osmotic potentials of -0.3, -0.6, -0.9 and - solutions on germination, non-germinated
1.2MPa, for NaCl (Coons et al., 1990) or seeds in each treatment were transferred
polyethylene glycol (PEG 6000) (Michel to distilled water and counted 3 days later.
and Kaufmann, 1973). NaCl Mean germination time (MGT) was
concentrations had the electrical calculated to assess the rate of
conductivity (EC) values of 6.5, 12.7, germination (Ellis and Roberts, 1980).
18.4 and 23.5 dSm-1, respectively. The seedlings with short, thick and spiral
formed hypocotyls and stunted primary
Seed treatments root were considered as abnormally
For hydropriming, cumin seeds germinated (ISTA, 2003).
(4.4% seed moisture) were immersed in Root length, shoot length and
distilled water at 25°C, for 18 h, under seedling fresh weights were measured
dark conditions. The hydropriming after the 7th day. Three grams of the
duration was determined by controlling seeds from each seed treatment were
seed imbibition during germination. For placed in Petri dishes, containing distilled
ZnSO4 treatment, the seeds were water to determine water uptake of seeds
immersed in 500 ppm ZnSO4 solution at necessary for germination. The water
25°C, for 2 h, in the dark (Singh and Rao, uptake was expressed as the percentage
1993). Thereafter, the seeds were rinsed increase in moisture content on fresh
with tap water three times. The treated weight basis.
seeds were surface-dried and dried back
to their original moisture content at room Experimental design
temperature (about 22°C, 45% relative The experimental design was three
humidity) determined by changes in seed factors factorial (3×2×5), arranged in a
weight. Moisture content of untreated complete randomized design with three
seeds (control, 4.4% moisture content), replications and 25 seeds per replication.
hydroprimed and ZnSO4 treated seeds was The first factor was seed treatment
equilibrated at room temperature for 2 (control, hydropriming and ZnSO4), the
days. second, solutions of NaCl and Na2SO4
and the third was osmotic potential level
Germination tests (0, -0.3, -0.6, -0.9 and -1.2 MPa). Data for
Three replicates of 25 seeds were germination percentage were subjected to
germinated between double layered rolled arcsine transformation, and analysis of
Anchor germination papers with 10 ml of variance was made using MSTAT-C
respective test solutions. The papers were program (Michigan State University). The
replaced every 2 days to prevent differences between the means were
accumulation of salts (Rehman et al., compared using LSD values (P < 0.05).
71A. AHMADIAN, Y. SHIRI, M. FROOZANDEH
Germination percentage was
RESULTS influenced by salt stresses, but
inhibition was greater in Na2SO4
A significant three-way (Table 2). Hydropriming showed
interaction (seed treatment, solution maximum germination under all
and stress) was found (P < 0.05, 60 osmotic potential of NaCl solutions.
d.f.) for all investigated characters. Germination percentage drastically
The germination rate decreased with declined and delayed with increase of
decrease in osmotic potential in both osmotic stress due to Na2SO4 in MPa
NaCl and Na2SO4 solution; however, lower than -0.6 and NaCl in MPa
Na2SO4 delayed it more compared to lower than -0.9. Considering seed
NaCl (Table 1). Both priming treatments, hydropriming gave higher
treatments increased the rate of seed germination percentage in Na2SO4
germination. Hydropriming resulted solution. Transfer of non-germinated
in the accelerated germination for seeds from PEG solution to the
both NaCl and Na2SO4, especially distilled water resulted in 100%
under low osmotic potential. Water germination regardless of osmotic
uptake of primed seeds did not change potential (data not shown). ZnSO4 and
significantly (P < 0.05) (data not hydropriming diminished the
shown), while the time to seed abnormal germination in NaCl and
germination for hydropriming, ZnSO4 Na2SO4. Increased water stress was
and control was delayed by 12, 18 and accompanied with increase of
38 h, respectively (data not shown). abnormal germination in both
solutions potential (data not shown).
Table 1 - Germination rate (seed/days) of black cumin seeds treated with ZnSO4,
hydropriming and control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa Control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 7.76 7.76 10.13 10.13 7 7
-0.3 6.53 5.8 9.36 7.33 6.15 5.1
-0.6 5.4 3.73 8.3 6.3 4.73 3.33
-0.9 3.76 2.5 7.2 4.63 4 2.17
-1.2 2.33 1.37 6 3.13 2.47 1.29
Although root length was root from the seed. Greater reduction
affected due to salt stress, significant in shoot length due to Na2SO4,
and higher inhibition due to Na2SO4 compared to NaCl, was very evident
was very evident (P < 0.05; Table 3). (P < 0.05), with no recorded shoot
At -1.2 MPa, radicle growth stopped growth at -0.9 MPa of Na2SO4 (Table
after emergence of radicle or primary 4). However, hydropriming enhanced
72GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN UNDER SALT STRESS CONDITIONS
shoot growth at -0.9 MPa of Na2SO4. shoot and root weights were recorded
Also, hydropriming exhibited higher from NaCl, compared to osmotic
shoot growth due to all concentrations stress at -0.6 MPa due to Na2SO4 and
of NaCl. Depending on decrease in above. The root / shoot weight and
shoot and root length, shoot and root R/S length increased with increase in
weight gradually declined with the osmotic potential in both NaCl and
decreasing osmotic potential of Na2SO4 solution (Fig. 1).
solutions (Tabs. 5 and 6). Higher
Table 2 - Germination percentage of seeds treated with ZnSO4, hydropriming and
control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 56 56 69 69 59 59
-0.3 51 46 66 66 58 57
-0.6 37 29 52 49 47 45
-0.9 28 9 44 19 36 16
-1.2 10 6 16 13 13 12
Table 3 - Root length (cm) of black cumin seedlings treated with ZnSO4,
hydropriming and control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa Control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 1.87 1.87 1.83 1.83 1.80 1.80
-0.3 1.45 1.42 1.78 1.58 1.82 1.74
-0.6 1.22 1.12 1.52 1.28 1.68 1.41
-0.9 0.84 0.52 1.00 0.72 1.44 0.67
-1.2 0.54 0.00 0.66 0.00 0.69 0.00
Table 4 - Shoot length (cm) of black cumin seedlings treated with ZnSO4,
hydropriming and control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa Control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 1.54 1.54 1.38 1.38 1.46 1.46
-0.3 1.36 0.96 1.13 1.05 1.18 1.12
-0.6 0.87 0.68 0.79 0.68 0.86 0.67
-0.9 0.59 0.32 0.38 0.29 0.46 0.21
-1.2 0.20 0.00 0.23 0.00 0.23 0.00
73A. AHMADIAN, Y. SHIRI, M. FROOZANDEH
Table 5 - Root weight (mg plant-1) of black cumin seedlings treated with ZnSO4,
hydropriming and control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa Control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 0.69 0.69 0.89 0.89 0.77 0.77
-0.3 0.56 0.45 0.82 0.78 0.67 0.65
-0.6 0.55 0.32 0.66 0.62 0.55 0.52
-0.9 0.22 0.2 0.53 0.47 0.32 0.33
-1.2 0.13 0.00 0.18 0.00 0.16 0.00
Table 6 - Shoot weight (mg plant-1) of cumin seedlings treated with ZnSO4,
hydropriming and control (untreated) at salt stress of NaCl and Na2SO4
Seed treatments
MPa Control (untreated) Hydropriming ZnSO4
NaCl Na2SO4 NaCl Na2SO4 NaCl Na2SO4
0 0.44 0.44 0.64 0.64 0.52 0.52
-0.3 0.31 0.2 0.57 0.53 0.42 0.4
-0.6 0.3 0.17 0.41 0.37 0.3 0.27
-0.9 0.16 0.11 0.28 0.22 0.07 0.08
-1.2 0.12 0.00 0.17 0.00 0.06 0.00
DISCUSSION increased by seed priming, but stress
conditions delayed it considerably.
Both seed treatments showed Compared to Na2SO4, germination
enhanced performance under stress rate for NaCl was more at equivalent
conditions. Germination rate was osmotic potential. This could be
74GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN UNDER SALT STRESS CONDITIONS
explained by more rapid water uptake germination without protrusion of
in hydroprimed seeds because radicle. Akinola et al. (2000) reported
germination for hydropriming, ZnSO4 that higher duration of exposure to
and control started at 12, 18 and 38 h, seed treatment resulted in higher
respectively (data not shown). It cumulative germination in wild
supports that hydropriming caused sunflower and Caseiro et al. (2004)
more rapid water uptake than the found that hydropriming was the most
amount of water for germination. The effective method for improving seed
results are in line with the findings of germination of onion, especially when
Ahmadian et al. (2009) in cumin. the seeds were hydrated for 96 h,
Sung and Chiu (1995) observed that compared to 48 h. The beneficial
mean germination time was effects of hydropriming on
accelerated by hydropriming without germination were found in this study.
changing amount of water uptake in ZnSO4 shortened MGT, however,
watermelon. final germination was higher from
Hydropriming clearly improved hydropriming, suggesting toxicity of
both rate of germination and mean ZnSO4 due to ion accumulation in the
germination time both under salt embryo (Demir and Van De Venter,
stress conditions. Furthermore, 1999).
hydropriming resulted in increase of Seeds always germinated better
normal germination. The results are in in NaCl than Na2SO4 at the equivalent
line with the findings of Thornton and water potential in line with earlier
Powell (1992) in Brassica, Srinivasan observations made for soybean by
et al. (1999) in mustard, and Khajeh-Hosseini et al. (2003), and for
Ahmadian et al. (2009) in cumin. cumin by Ahmadian et al. (2009).
Fujikura et al. (1993) indicated the This may be due to the uptake of Na+
beneficial effects of hydropriming on and Cl- ions by the seed, maintaining
aged seeds, with respect to a water potential gradient allowing
germination and percentage of normal water uptake during seed germination.
seedlings, in cauliflower. Lower germination percentage
Furthermore, Sadeghian and Yavari obtained from Na2SO4 compared with
(2004) reported that increasing NaCl, at equivalent water potential in
drought stress resulted in increasing each priming method suggest that
abnormal seedlings in sugar beet. It is adverse effects of Na2SO4 on
concluded that superiority of germination were due to specific ion
hydropriming on germination could accumulation rather than osmotic
be due to soaking time effects rather effect. These results agree with
than ZnSO4 treatment. Because Murillo-Amador et al. (2002) in
hydroprimed seeds compared to cowpea, Demir and Van De Venter
ZnSO4 treated seeds were allowed to (1999) in watermelon, they affirmed
imbibe water for a longer time and that drought or salinity may influence
went through the first stage of germination by decreasing the water
75A. AHMADIAN, Y. SHIRI, M. FROOZANDEH
uptake and toxicity of ions. Under salt the risk of poor stand establishment
stress, Na+, Cl- and SO4- may be under drought and salt stress and
taken up by the seed and toxic effect permit more uniform growth under
of NaCl and Na2SO4 might appear. conditions of irregular rainfall and
However, our findings at high salinity drought on saline soils.
concentration of 23.5 dSm-1 showed Parera and Cantliffe (1994) and
that decrease in germination McDonald (1999) emphasize that
percentage was significant. hydropriming is the simplest approach
The main effect of seed to hydrating seeds and minimizes the
treatments was an increase in use of chemicals. However, if the
germination rate; however, post- seeds are not accurately hydrated, the
germination growth was also rate of hydration cannot be exactly
increased. Hydropriming improved controlled. It was observed that
seedling fresh weight, under osmotic hydropriming practically ensured
stress. Considering both seed rapid and uniform germination
treatments, it was concluded that accompanied with low abnormal
hydropriming improved root growth seedling percentage in line with Singh
and gave the highest root length in (1995) and Shivankar et al. (2003).
both solutions. El-Midaoui et al. They stress that it has high potential
(2003) reports that root and shoot in improving field emergence and
growth significantly decreased by ensures early flowering and harvest
osmotic stress, at -0.6MPa, and above under stress conditions especially in
induced by PEG 6000. Murillo- dry areas.
Amador et al. (2002) found that
seedling growth of cowpea was CONCLUSIONS
inhibited by both NaCl and PEG, but
higher inhibition occurred due to Our findings revealed that
PEG. Sung and Chiu (1995) proposed inhibition of germination at equivalent
that emergence force and seedling water potential of NaCl and Na2SO4
growth were strengthened by resulted from salt toxicity rather than
hydropriming in watermelon. osmotic effect. Both seed treatments
Seedling growth severely diminished gave better performance than control
with increased drought stress and (untreated) under salt stress with clear
genetic differences were found in effectiveness of hydropriming in
sugar beet (Sadeghian and Yavari, improving the germination percentage
2004). at low water potential. In commercial
In many coated seeds, production of black cumin in Iran,
germination and subsequent seedling germination of cumin seeds is a major
growth can be inhibited by problem to planting. To achieve a
mechanical restriction exerted by the uniform plant density in case of
seed coat (Sung and Chiu, 1995). drought, growers tend to sow 25 kg
Priming may be helpful in reducing ha-1 of seed while only 4.5 kg ha-1 is
76GERMINATION AND SEEDLING GROWTH OF BLACK CUMIN UNDER SALT STRESS CONDITIONS
needed. Hydrated seeds with higher cultivars to high temperature
germination percentage under salt combined with NaCl during
germination. J. Am. Soc. Hort. Sci.,
stress or micronutrient application 115, 1004-1007.
increased tolerance of seeds to salt Demir I., Van De Venter H.A., 1999 - The
stress. In addition, reported protocol is effect of priming treatments on the
simple, cheap and does not require performance of watermelon (Citrillus
lanatus (Thunb.) Matsum. & Nakai)
expensive chemicals and sophisticated seeds under temperature and
equipment. The protocol has practical osmotic stress. Seed Sci. Technol.,
importance and could be 27, 871-875.
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higher germination and uniform Maintance of viability and vigour in
sunflower. Seed Res., 18, 15-24.
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