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Copyright: © 2018 Birhanu Gizaw et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Received: 30 November -0001 | Accepted: 01 January 1970 | Published: 23 January 2018
Keywords: Germination, Priming, Phosphate, Seedling vigor, PGPR.
Germination and seedling establishment are critical stages in the plant life cycle. The aim of this study was to evaluate the effect of phosphate solubilizing fungi on seed germination and seedling growth of Faba bean. Phosphorus is an essential macronutrient next to nitrogen required by plant for vital biosynthesis. Factorial experiment based a completely randomized block design with three replications were used. This experiment had two factors, the first factor with one level of bean cultivar. In the second factor, three levels of phosphate solubilizer fungi were applied as treatments. Single inoculation of each strain was done. The results revealed that seed treated withTrichosporon beigelii B, C. albidus var aurius, Phichia norvegensis and Control showed 780.43, 749.93, 618.23, 500.73 seedling vigor index respectively and improved germination rates up to 91%. The highest mean growth of plumule length (5.6 cm) and radicle length (3.42) within 15 days growth recorded by Trichosporon beigelii B, and C.albidus var aurius respectively. The results of this study suggest that T. begili and C. albidus var aurius have the potential to increase the growth and development of plumul size, radicle length, and fresh biomass of Faba bean seed. Bio-priming is a promising technique for vital application of using beneficial microbes to promote seed germination and seedling development through phytase enzyme production, phyto hormone production, phosphate solubilization and other plant growth promoting properties as well as bio control activities.
Faba bean (Vicia faba L.) is the fourth most important pulse crop in the world (Sainte, 2011). Ethiopia is the world’s second largest producer of Faba bean next to China; followed by, Egypt, Italy, and Morocco (Salunkhe and Kadam, 1989). Its share is only 6.96 % of world production and 40.5 % within Africa (Chopra et al., 1989). In Ethiopia, the average yield of Faba bean under small-holder farmers is not more than 1.6 t ha-1 (CSA, 2013). Faba bean ranks first in pulse crop in the total area coverage and the total production of Ethiopia. It accounts about 36% of the country’s pulse production (IFPRI, 2010). Currently, the total area, under cultivation with Faba bean in the country, is estimated to be about 0.54 million hectare and the total production is 696 million kilogram (MoARD, 2009). The productivity of Faba bean in Ethiopia is quite lower 15.2 qt/ha (CSA, 2011) than UK 30qt/ha (Winch, 2006). The country is considered as the secondary center of diversity and also one of the nine major agro-geographical production regions of Faba bean (Telaye, Bejiga et al. 1994). The production is mainly concentrated in the high-altitudes of Ethiopia ranging between altitudes1800-2400 m.a.s.l with annual rain fall ranges from 700 to 1100 mm. and has suitable environmental and soil conditions for highland pulse crops production (Telay, 1985). Faba bean (Vicia faba L.) is widely grown in many parts of the world as a source of protein, starch, cellulose and minerals from its mature seed for both human and animal nutrition (Crepon, 2010). It is also traditionally used as a cover crop to recover nitrogen content and prevent erosion of the soil, and is appreciated for its good agronomic characteristics (Kopke, 2010). In addition to its high contents of protein and carbohydrates, V. faba is also rich in fiber, vitamins and minerals, and has a hypocholesterolaemic effect (Ofuya .,2005). Most farmers in Ethiopia cultivate local Faba bean varieties (Thijssen et al., 2008). Local varieties are low yielding and susceptible to both biotic and abiotic factors. Which are highly susceptible to disease and low yielding (Samuel et al. (2008). Among biotic stresses, diseases have always been the major limiting factors for Faba bean cultivation. The major ones include ascochyta blight (Ascochyta fabae Speg.), rust (Uromyces viciae-fabae) and chocolate spot (Botrytis fabae Sard.) and black root rot Fusarium solani highly contribute to the low productivity of the crop in Ethiopia (Dereje1993), Chocolate spot is considered to be the most important and destructive in Ethiopia causing yield loss of up to 61% on susceptible cultivars (Dereje and Beniwal, 1987). Germination and early seedling establishment are critical stages in the plant life cycle and important for agricultural productivity. It determines plant density, uniformity and management options (Cheng et al., 1999; Hadas., 2004). Rapid and uniform seedling emergence leads to successful establishment as it produces a deep root system before the upper layers of soil dry out, harden, or reach supra-optimal Temperatures (Harris1996). Most of the leguminous plant seeds are attacked by certain soil borne pathogens i.e.,Fusarium solani Mart sacc., Rhizoctonia solani Kuhn, Fusarium oxysporum, Sclerotium rolfsii and Pythium spp before they germinate and seedling establishment which attack roots causing damping-off and root rot diseases. (Abdel-Kader, 1997; El-Mougy, 2001). Several root rot and wilt pathogens such as Rhizoctonia solani, Fusarium solani and Macrophomina phaseolina are reported to attack Faba bean roots and stem base causing serious losses in seed germination and plant stand as well (Abdel-Kader et al., 2011). These diseases cause substantial losses to beans crop, yield losses in severely infested areas may be as high as 50% (Estevez deJensen et al., 2001).A method to improve the rate and uniformity of germination is the priming or physiological advancement of the seed lot (Finch-Savage, 2004; Halmer, 2004). Seed treatment is an important process that provides insurance against seed-borne as well as soil-borne plant pathogens and insects (Gwary et al., 2007). It is a relatively cheap and effective way of controlling seed-borne plant diseases (Dawar & Ghaffar 1998). Bio priming of seeds with different bacterial strains particularly rhizobacteria have been shown to be effective in suppressing disease infection by inducing a resistance mechanism called ‘induced systemic resistance’ (ISR) in varied agronomic and horticultural crops (Van Loon, 1998). Among various bacterial genera, Bacillus and Pseudomonas spp. are ubiquitous rhizosphere inhabitant bacteria that are the most studied bio-priming agents reported as disease suppressing in plants (Weller 1988). Priming seeds of many crops with biological control agents (BCA), Bacillus subtillus and Pseudomonas fluorescens are the most effective approach for controlling seed and root rot pathogens (Begum, 2010, El-Mohamedy, 2013). Various seed priming techniques have been developed, including hydro-priming (soaking in water), halo-priming (soaking in inorganic salt solutions), osmo-priming (soaking in solutions of different organic osmotica), thermo-priming (treatment of seeds with low or high temperatures), solid matrix priming (treatment of seed with solid matrices) and bio-priming (treatment of seed using biological agent) (Ashraf and Foolad, 2005). Seed biopriming is a pre-sowing approach for influencing the seedling development by stimulating pre germination metabolic activities prior to the emergence of radicle and improvement in the germination rate and performance of plant (Taylor, 1998, Halmer
Materials And Methods:
The study was carried out at Ethiopian biodiversity institute microbial laboratory at 2017 GC.
Faba Bean SEED (Vicia faba) Collection
Faba bean seed purchased from Shola Market from AddisAbaba
Isolation of Microorganism:
Phosphate solubilizing fungi were isolated and identified from teff rhizosphere soil of crop fields collected from Gojam and North Showa. Soil sample (10 g) was mixed with 90 mL sterile distilled water from each collected soil sample. It was vigorously shaken and left to stand for 5 min. Homogeneous soil solution was serially diluted
Determination of Phosphate solubilization index (SI):
Phosphate solubilization index (SI) was calculated using the formula outlined in Premonoet al. (1996). A loopful of 24h old cultures was spotted at four points on Pikovskaya’s agar plate and incubated at 260C for 2 to 4 days .The diameter of colony and halo zone was measured using transparent ruler.
SI= Colony +Halo Zone diameter Colony diameter.
Faba bean Seed Disinfection:
Thirty six Faba bean seeds were disinfect using 70% ethanol alcohol for 15 minute and 3% hypochlorite for 10 minute and washed 6 times by distilled water. Finally disinfected bean seed were kept on sterilized Whiteman No1 filter paper inside petridish until germination bioassay carryout.
Bio-priming of Faba bean (Vicia faba) with plant growth promoting Microbes and its vigor index:
Faba bean seed germination Bioassay:
Twenty seven seeds of Faba bean were dipped into Malt extract broth culture for Trichosporon beigelii B, Cryptococcus albidus var aerius, Phichia norvegensis for 6 hour which contain the fungal suspension 90% inoculum density measured by Terbidiometer. While nine seeds were dipped in distilled water as control. Three seeds per plate of inoculated Faba beans from each strain in triplicate were plated in petri dishes with one layer of whatmanNo.1 filter paper. Both
Vigor index= (Mean radicle length+ mean plumule length) % Germination
Percentage of germination:
Percentage germination was calculated counting the number seed germinated divided for the total number of seed multiplied by hundred
GP= Seeds germinated/total seeds x 100
Fresh biomass determination:
At the end the experiments the root of Faba bean was carefully uprooted and washed to remove soil. Root separated from the base of stem and weighed. Fresh biomass of seedling stem, leaves was weighed and the fresh biomass result was recorded.
Statistical analysis:
The data of seed germination, fresh biomass, plumule length, radicle length, its vigour index were analyzed using descriptive statistics like percentage frequency, mean and Stata vere.13.
Isolation of phosphate solubilizing fungi:
A total of sixteen phosphate solubilizing fungi were isolated and identified from teff rhizosher soil and based on the SI and phosphate solubilizing efficiency three superior fungi species, Trichosporon beigelii B, Cryptococcus albidus var aerius, Phichia norvegensis were selected and evaluated for seed germination test. (Table 1, figure 1).
Plant growth promoting Fungi(PGPF)
Phosphate solubilizing index at 15 days
Trichosporon beigelii B
Cryptococcus albidus var aerius
Table.1. Phosphate solubilizing indexmeasure on Pikovskaya’s agar media
Figure1. Phosphate solubilizing fungi on Pikovskaya’s agar media . 1. Trichosporon beigelii B 2, Phichia norvegensis.3.Cryptococcus albidus var aerius.
Percentage of germination:
Among 36 seed sowed 33 seed were germinated and 3 fails to produce radicle and plumule. Their percentage germination was 91.6 % in this study.
Effect of Phosphate solubilizing fungi inoculation on seed germination and seedling growth
The results of Faba bean seed inoculation with Trichosporon beigelii B, Phichia norvegensis. Cryptococcus albidus var aerius.in the presence of Phosphate sources on growth parameters significantly increase height compared to control. Among all fungi inoculum Trichosporon beigelii B gave the maximum growth in plumule mean height (5.6 cm) followed by Cryptococcus albidus var aerius (4.2 cm), Phichia norvegensis (3.6 cm) and control (3.2 cm) within 15 days growth. C.albidusvar aurius showed the highest mean radicle growth (3.4cm) followed by Control (2.7 cm), Trichosporon beigelii(2.9 cm) and least radicle growth recorded by Phichia norvegensis (1.8 cm)in 15 days growth (Table2 and Figure 2).
Mean plumul growth(cm)
Mean radicle growth(cm)
Trichosporon beigelii B
Cryptococcus albidus var aerius
Table.2. The effect of plant growth microbs on plumul and radicle growth of Faba bean within 15 days*=Significant at p<0.05. Note; Mean in column followed by the same superscripts are not statistically different at P<0.05 according to Turkeys test
Vigor index result:
The number of germinated seeds inoculated by Trichosporon beigelii B, C.albidus var aurius , Phichia norvegensis and control was counted. Root and shoot length of individual seedling was measured to determine the vigour index. The result were summarized in Table 3
Plant growth promoting Fungi | Vigor index of Germinated seed |
Trichosporon beigelii B 780.43 | |
Cryptococcus albidus var aerius 500.73 |
Table 3. Vigor index of germinated seed treated by each plant growth promoting fungi.
It gives me a great pleasure to acknowledge Dr. Genene Tefera for his unreserved guidance and encouragement and support in providing and facilitating the necessary equipment. Finally goes to Ethiopian biodiversity institute, microbial directorate for every budget grant to carry out this study and its research team for their unreserved support at laboratory and field work especially, Endeshaw Abatneh, Endegena Aynalem, as part of research group in tireless effort in teff rhizospher soil sample collection. Lastly, I acknowledged Woyenshet Lule for her kindly support especially laboratory chemicals facilitation.
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