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Research Article

Legume Research, volume 44 issue 5 (may 2021) : 593-601

Effect of seed biopriming with indigenous PGPR, Rhizobia and Trichoderma sp. on growth, seed yield and incidence of diseases in French bean (Phaseolus vulgaris L.)

Shivangi Negi1,*, Narender K. Bharat1, Manish Kumar1
1Department of Seed Science and Technology, Dr. Y.S. Parmar University of Horticulture and Forestry, Nauni, Solan-173 230, Himachal Pradesh, India.

  • Submitted20-03-2019|

  • Accepted24-05-2019|

  • First Online 14-08-2019|

  • doi 10.18805/LR-4135

Cite article:- Negi Shivangi, Bharat K. Narender, Kumar Manish (2019). Effect of seed biopriming with indigenous PGPR, Rhizobia and Trichoderma sp. on growth, seed yield and incidence of diseases in French bean (Phaseolus vulgaris L.) . Legume Research. 44(5): 593-601. doi: 10.18805/LR-4135.

ABSTRACT

The effect of seed biopriming with different bioagents including plant growth promoting rhizobacteria (PGPR-1), rhizobial biofertilizer (Rhizobium strain B1) and biological control agent (Trichoderma viride) was observed on plant growth, seed yield and incidence of diseases in French bean cv. Contender conducting a field experiment during kharif season in the year  2017 and 2018. Under field conditions in both the years, field emergence (95.18 %), plant height at 30 days after sowing (34.09 cm), plant height at final harvest (56.99 cm), days to pod harvest (52.33), harvest duration (18.67), pod length at final harvest (16.83 cm), number of pods per plant (20.17), dry pod weight (2.72 g), pod yield per plant (38.64 g), number of seeds per pod (7.17), seed yield per plant (20.76 g), seed yield per plot (875.33 g), seed yield per hectare (23.34 q), 100 seed weight (34.19 g), quality of harvested seeds were recorded significantly higher after seed biopriming with PGPR-1+ Rhizobium strain B1 (T4) as compared to carbendazim seed treatment and  untreated control. This treatment combination also reduced the incidence of major diseases like, Rhizoctonia root rot and Angular leaf spot significantly as compared to carbendazim seed treatments and untreated control. It can be concluded from the present investigation that seed biopriming of French bean cv. Contender with PGPR-1+Rhizobium strain B1 @ 109cfu/ml for 8 hours was an effective treatment which significantly improved plant growth, pod yield, seed yield, seed quality and seed vigour and reduced disease incidence as compared to seed treatment with carbendazim @ 0.2% as well as untreated control under field conditions.

INTRODUCTION

French bean (Phaseolus vulgaris L.) is one of the most important leguminous crops. It is a short duration crop and can be grown under different cropping patterns worldwide. In India it is largely grown in hilly areas of Himachal Pradesh, Uttarakhand, Jammu and Kashmir and North Eastern states during summer and as winter and autumn crop in parts of Uttar Pradesh, Maharashtra, Karnataka, Andhra Pradesh, Punjab, Haryana, Bihar, Gujarat, Madhya Pradesh and Tamil Nadu. It is grown in a total area of 230,000 hectares with a production of 2278,000 metric tons (NHB. 2016-2017). French bean is consumed as immature tender fruits and green grains as vegetables and mature seed as pulses (rajmash). It is very nutritive and per 100 g of green pod contains 1.7 g protein, 0.1 g fat, 4.5 g carbohydrate, 1.8 g fibre and is also rich in minerals and vitamins. Dry bean (seed) is an important source of protein, participating in human diet all over the world (Singh, 2001). Being a leguminous plant, French bean also play an important role in agriculture cropping system by virtue of its ability to fix atmospheric nitrogen with symbiotic association of Rhizobium in soil.
 
As bean is cultivated through seed the treatment of seeds is an important tool for ensuring good crop health, allowing growers to realize the genetic and physiological potential of crop variety. The quality of seed alone is known to account for at least 10 to 15 per cent increase in the crop productivity. Seed priming is a quality enhancement technique used for a rapid and uniform germination of seeds and optimum plant stand in the field. This technique is often used as a seed invigoration treatment for improving germination and vigour in low vigour seed lots. It appears to reverse the detrimental effects of seed deterioration (Srinivasan et al., 2009). Biopriming is a new technique of seed treatment that integrates biological and physiological method of improving plant growth and controlling disease. It is recently used as an alternative method for controlling many seed and soil borne pathogens. It is, thus considered as an advanced technique of seed treatment which is a combination of application of beneficial microorganism on seed surface and seed hydration (Singh et al., 2016). Biopriming has great promise for enhancing the efficacy, shelf life and consistent performance of bioagents (Callan et al., 1997). Plant growth promoting rhizobacteria (PGPR’s) and rhizobial biofertilizers are naturally occurring soil bacteria that aggressively colonize plant roots and benefit plants by providing growth promotion and disease suppression. The PGPR’s have been demonstrated to increase growth and productivity of many commercial crops (Saharan and Nehra, 2011). Effect on enhancement in plant growth by root-colonizing species of Rhizobium and Pseudomonas is well documented in literature (Sahni et al., 2008). Trichoderma has been exploited as biocontrol agent against a range of plant pathogenic fungi because it antagonizes a number of plant pathogens. Many strains of Trichoderma have been widely used as biological control agents as well as plant growth promoters (Harman, 2000). Keeping in view, the importance of the crop and the benefits of the seed biopriming with beneficial microbes, the present investigation were under taken which biopriming of bean seed was done and their effect on emergence, growth, health and yield parameters were worked out.

MATERIALS AND METHODS

The present study was carried out at experimental farm of Department of Seed Science and Technology, Dr Y.S. Parmar University of Horticulture and Forestry, Nauni, Solan (HP) during the kharif seasons of 2017 and 2018. The experimental farm is located at altitude of 1183 m above mean sea level with latitude of 30.51°N and longitude of 77.09° E in the mid- hill zone of Himachal Pradesh.
 
Physio-chemical properties of the soil of experimental field
 
The physic-chemical analysis of soil samples taken from experimental field was got done from the Department of Soil Science and Water Management, Dr. Y.S. Parmar University of Horticulture and Forestry, Nauni, Solan (HP). The physio-chemical properties of soil were as follows:
 

 
Meteorological data
 
The climate of the area is sub-tropical to sub-temperate and semi- humid. The area experiences three major seasons in a year. The winter season commences from October to March, summer season extends from April to June followed by the monsoon season from July to September. Generally, December and January months are the coldest while, May and June are the hottest months. During the crop seasons of 2017 and 2018, the meteorological data on rainfall, temperature and relative humidity during the course of investigation are presented in the following Table.


 
Procurement of the seed
 
The seeds of French bean cv. Contender were collected from the Department of Seed Science and Technology, Nauni, Solan (HP). The seeds were 6 months old and had been stored at 5°C.
 
Procurement of bioagents
 
The mother cultures of PGPR-1 and Rhizobium strain B1 were procured from the Soil Microbiology Laboratory, Department of Basic Sciences, whereas the culture of Trichoderma viride was obtained from the Biological Control Laboratory, Department of Plant Pathology, YSP UHF, Nauni, Solan (HP). These cultures were further maintained on the artificial growing media in the Department of Seed Science and Technology, Dr. YSP UHF Nauni, Solan (HP).
 
Experiment layout
 
The experiment was conducted in open field conditions using randomized complete block design (RCBD) with three replications.
 
Treatment combinations used
 
Ten treatment combinations were used viz., seed biopriming with PGPR-1 (T1), Rhizobium strain- B1 (T2), Trichoderma viride (T3), PGPR-1 + Rhizobium strain B1 (T4), PGPR-1 + Trichoderma viride (T5), Rhizobium strain B1 + Trichoderma viride (T6), PGPR-1 + Rhizobium strain B1 + Trichoderma viride (T7), Hydropriming (T8), Carbendazim seed treatment @ 0.2% (T9) and untreated control (T10).
 
Preparation of cultures and suspension of bioagents
 
Cultures of bioagents, PGPR-1 and Rhizobium strain-B1 were grown on the nutrient agar medium (NA) while that of Trichoderma viride were grown on potato dextrose agar medium (PDA). The compositions of NA and PDA are given below:
 
Nutrient Agar (NA)
Beef extract                                                  :                    3 g
Peptone                                                         :                    5 g
Agar agar                                                      :                    15g
Nacl                                                               :                    5 g
Distilled water                                                :                    1000 ml
Potato Dextrose Agar (PDA)
Peeled potato                                                :                    200 g
Dextrose                                                       :                    20 g
Agar -Agar                                                     :                    20 g
Distilled water                                                :                    1000 ml
 
Each bioagent was grown  in 9 cm Petriplate incubated in BOD incubator for 7 days at 25 ± 1°C for one week. After a week one ml of sterilized autoclaved double distilled water was added to each Petriplate containing fresh cultures and scraped with the help of a spade to produce slurry and then transferred to 250ml of sterilized distilled water to make a suspension. The concentration of the cell/ spore suspension was observed to be 109cfu/ml in case of PGPR and Rhizobium and 107cfu/ml in case of Trichoderma.
 
Biopriming of seed
 
The seeds of French bean with no cracks or other visible deformations were selected and surface sterilized with 1.5% sodium hypochlorite (NaOCl) solution for 5 minutes. Seeds were then rinsed thrice with sterilized distilled water and dried under laminar air flow on sterilized blotting paper (Jain et al., 2012). The surface sterilized and dried seeds were bioprimed by soaking in the spore/cell suspensions of PGPR-1 for 6 h, Rhizobium strain- B1 for 8 h, Trichoderma viride for 6 h, PGPR + Rhizobium strain B1, PGPR + Trichoderma viride, Rhizobium strain B1 + Trichoderma viride and PGPR + Rhizobium strain B1 + Trichoderma viride (T7) for 8 h separately. The bioprimed seeds were shade dried to bring down their moisture content to original moisture content.
 
Sowing of seed
 
The seeds were sown in a plot having size 2.00 m × 1.50 m (3 m2 area) accommodating 9 plants/ plot at the spacing of 45 × 15 cm. One seed per hill was dibbled at a depth of 2.0 to 2.5 cm. The standard cultural practices were adopted as recommended for growing French bean in the mid hills as included in the Package of Practices for Vegetables Crops, Dr YS Parmar UHF, Nauni, Solan (Anonymous, 2010).
 
Observations
 
The observations on growth parameters like field emergence, plant height at 30 DAS and at final harvest, days to pod harvest; yield parameters like number of pods per plant, dry pod weight (g), pod yield per plant (g), number of seeds per pod, seed yield per plant (g), seed yield per plot (g), seed yield per hectare (kg) and 100 seed weight (g) were taken following standard procedures. The observations on incidence of Rhizoctonia root rot and Angular leaf spot were recorded as per the methodology adopted by Mathew and Gupta (1996) and Mathew et al., (1998), respectively.
 
Statistical analysis
 
Before going for statistical analysis, all the observations in respect of field emergence (%) and incidence of diseases (%) were transformed into arc sine root transformation values. The data recorded were analyzed using MS-Excel, OPSTAT and SPAR 2.0 package as per the design of experiment. For working out the analysis of variance, the data analysis was done as suggested by Panse and Sukhatme (1967).

RESULTS AND DISSCUSSION

Field emergence and growth parameters
 
Different seed biopriming treatments had a significant effect on the field emergence and plant growth parameters like, plant height at 30 DAS, plant height at final harvest, days to pod harvest and duration of harvest as compared to the fungicide treatment and untreated control (Table 1 and Fig 1). Amongst various treatments, seed biopriming with PGPR-1 + Rhizobium strain B1 (T4) resulted in the highest percentage of field emergence (95.18%) followed by (93.70%) emergence in seed biopriming with Rhizobium strain B1 (T2). These two treatments were, however, at par with each other. Similar trend was observed w.r.t. the plant growth parameters and seed biopriming with PGPR-1 + Rhizobium strain B1 (T4) resulted in a significantly more plant height at 30 DAS (34.09 cm), plant height at final harvest (56.99 cm) and duration of harvest (18.67 days) than carbendazim seed treatment as well as untreated (control). The plant growth parameters observed after seed treatment with carbendazim viz., plant height at 30 DAS (27.47 cm), plant height at final harvest (48.37cm) and duration of harvest (13.83 days) were significantly less than the biopriming treatments.
 

Table 1: Effect of seed biopriming with PGPR, Rhizobia and Trichoderma on field emergence and growth parameters in French bean.


 

Fig 1: Effect of biopriming on field emergence and growth characters in French bean (Year 2017-18 pooled).


 
Under the present investigation, seed biopriming of French bean with a combination of PGPR-1 and Rhizobium strain B1 provided higher emergence and plant growth in field as compared to the rest of the treatments including fungicide treatment. These results are in accordance with the findings of Junges et al., (2016), Entesari et al., (2013), Yadav et al., (2013) and Ahmed et al., (2012) who have also reported enhancement in seedling emergence in bean, soybean and mungbean after biopriming the seeds with different bioagents. The effect of seed biopriming with the combination of PGPR and Rhizobium strain B1 on growth parameters was observed higher than that of biopriming with the individual bioagent under present study. Which might have happened because of the fact that Rhizobium and PGPR share a common microhabitat, the root soil inter surface, where interactions between different microbial groups were reported during root colonization. Co-inoculation of Rhizobium with PGPR enhanced nodulation and nitrogen fixations through the production of plant hormones like auxins, flavonoids, nod factor or enzymes in legumes (Bansal and Srivastava 2012). Junges et al., (2016) who have also observed that the combined inoculation of bioagents was more effective than their individual inoculations. Seed biopriming increased germination rate and uniformity in emergence through metabolic repair during imbibition and build-up of germination related physiological processes. According to Taylor and Harman (1990) initiation of all these germination related physiological processes helped in the establishment and proliferation of bioagents on the spermosphere. The conditions on the seeds during priming increased the potential for microbial proliferation and several studies have demonstrated increased ability of applied bioagents to establish on seeds during biopriming or pre-germination processes. Ahmed et al., (2012) have observed that increase in root and shoot growth of mungbean upon co-inoculation with Rhizobium and Pseudomonas was due to lowering of ethylene levels in germinating seeds by these strains. The increase in growth parameters could also be due to the fact that seed biopriming with bacterial antagonists increased the population of antagonists on the seeds and protected them from the attack of plant pathogens (Callan et al., 1990). Biopriming ensures the entire seed batch is at the same point in the germination process, so that once planted, bioprimed seed showed more rapid and uniform emergence than unprimed seed (Rowse, 1996).
 
Yield attributes
 
The data in Table 2 and Fig 2 revealed that seed biopriming with PGPR-1 + Rhizobium strain B1 (T4) also resulted in a significant improvement in yield attributes of French bean crop like, number of pods per plant, dry pod weight, pod yield per plant, number of seeds per pod, seed yield and 100 seed weight as compared to that of hydropriming, carbendazim seed treatment and untreated control. The number of pods per plant (20.17), dry pod weight (2.72 g), pod yield per plant (38.64 g), number of seeds per pod (7.17), seed yield (23.34 q/ha) and 100 seed weight (34.19 g) upon seed biopriming with PGPR-1 + Rhizobium strain B1 (T4) were at par with that of seed biopriming with Rhizobium strain B1 (T2) (Fig 3 and 4). The carbendazim seed treatment was found less effective than these biopriming treatments and resulted in significantly lesser number of pods per plant (11.50), dry pod weight (2.08 g), pod yield per plant (27.77g), number of seeds per pod (5.17), seed yield (17.98q/ ha) and 100 seed weight (30.44g), and it was at par w.r.t. some parameters with that of hydropriming and untreated (control).
 

Table 2: Effect of seed biopriming with PGPR, Rhizobia and Trichoderma on yield attribute in French bean.


 

Fig 2: Effect of biopriming on yield characters in French bean (Year 2017-18 Pooled).


 

Fig 3: Flowering and pod setting in PGPR-1 + Rhizobium strain B1.


 

Fig 4: Best treatments over control.


 
In the present investigation, higher seed yield was recorded when the seeds of French bean were bioprimed with PGPR-1 + Rhizobium strain B1 (T4). The present findings are in agreement with that of Yadegari and Rahmani (2010) who have also found that co-inoculation with PGPRs and Rhizobium increased seed yield in common bean as compared to the uninoculated (control). The increased seed yield was attributed to the increased physiological activities like synthesis of chlorophyll, carbohydrates, amino acids and translocation of photosynthates into developing pods and seeds in bioprimed plants. Abadeh et al., (2013) have also observed increased grain yield in red lentil after seed biopriming with PGPR as compared to the non-bioprimed seeds. Inoculation of seeds with PGPR and Rhizobium resulted in early and better nodulation which was translated into higher shoot nitrogen accumulation and there by increased higher seed yield. This increased seed yield in bioprimed plants was due to the increased uptake and availability of major and minor nutrients together with growth regulators, enzymes and amino acids at all the crucial stages of growth and development. These physiological changes caused more number of pods and bolder seeds in them (Bais et al., 2006). Rhizobacteria have also been found to induce phytoalexins in roots and these phytoalexins protect the roots against pathogens and help their development, mobilization of insoluble nutrients followed by enhancement of uptake by plants (Valverde et al., 2006).
 
Incidence of diseases
 
The data presented in Table 3 and Fig 5 revealed that the seed biopriming with bioagents including PGPR, Rhizobium and Trichoderma alone and their combinations significantly affected the incidence of Rhizoctonia root rot and angular leaf spot diseases in French bean. Amongst different treatments, minimum incidence of Rhizoctonia root rot (2.60%) as well as Angular leaf spot (2.97 %) was observed in the seed biopriming with PGPR-1 + Rhizobium strain B1 (T4) followed by 3.70 and 3.34 % in Rhizobium strain B1 (T2), 3.71 and 4.07 % in PGPR-1 + Rhizobium strain B1 + Trichoderma viride (T7), respectively. These three biopriming treatments were statistically at par with each other. The incidence of root rot and angular leaf spot in French bean plots grown after seed treatment with carbendazim @ 0.2% was observed to be 9.26 and 9.63% which was significantly higher than all the biopriming treatments. The disease incidence in untreated plots was observed maximum which was significantly higher than all the other treatments.
 

Table 3: Effect of seed biopriming with PGPR, Rhizobia and Trichoderma on incidence of major diseases in French bean.


 

Fig 5: Effect of biopriming on incidence of diseases in French bean (Year 2017-18 pooled).


       
In the present investigation, the seed biopriming with PGPR-1 + Rhizobium strain B1was found most effective treatment in reducing the diseases like Rhizoctonia root rot and Angular leaf spot in French bean. There existed no report in literature on the effect of seed biopriming in reduction of the root rot disease in French bean. Hence, this seems to be the first report in this regard. However, many workers have reported that seed biopriming with biological control agents reduced the incidence of various diseases in different leguminous crops such as faba bean (El-Mougy and Kader, 2008), soybean (Begum et al., 2010) and green gram (El-Mohamedy et al., 2015). Besides, growth promotion activities, PGPRs and Rhizobia also act as biocontrol agents against some plant pathogens and inhibit the progress of the pathogens following different mechanisms like, competition, lysis, antibiosis, siderophore production and hyper-parasitism. More importantly, the seed biopriming with PGPR, rhizobia and Trichoderma triggered the defence mechanisms during the initial germination process involving synthesis of salicylic acid and jasmonic acid and thus provided basal resistance against pathogens. These signals play important role in induction of induced systemic resistance (ISR) and systemic acquired resistance (SAR) and both are effective against a broad spectrum of pathogens (Mahmood et al., 2016). Application of these organisms through seed biopriming might have activated and followed the same mechanism and resulted in decreased incidence of Rhizoctonia root rot and angular leaf spot diseases. In addition to this these beneficial organisms also help in increased nutrient uptake and plant growth as discussed earlier and a healthy plant is less affected by the attack of pathogens.

CONCLUSION

In french bean cv. Contender, seed biopriming with a combination of plant growth promoting rhizobacteria (PGPR-1) + rhizobial biofertilizer (Rhizobium strain B1) @ 109cfu/ml for 8 hours or with Rhizobium strain B1alone @ 109cfu/ml for 6 h or with a combination of PGPR-1 + Rhizobium strain B1 + Trichoderma viride@ 109cfu/ml for 8 h before sowing was observed to be an effective treatment which improved plant growth, pod yield, seed yield, seed quality and seed vigour and reduced the incidence of major diseases like Rhizoctoniaroot rot and angular leaf spot significantly as compared to seed treatment with carbendazim @ 0.2% as well as untreated control under field conditions.

ACKNOWLEDGEMENT

The authors thank the authorities of Dr YS Parmar University of Horticulture and Forestry, Nauni, Solan (HP), India for providing all the facilities to conduct these investigations.

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