EFFECT OF BRADYRHIZOBIUM INOCULUM AND LEVELS OF PHOSPHORUS ON THE YIELD AND QUALITY OF SOYBEAN

Background: Soybean is one of the major oil seed crops in Bangladesh. Combine application of Rhizobium inoculation and phosphorus fertilizer could be an important factor to get maximum yield and quality of soybean. Objective: To evaluate the effect of Bradyrhizobium inoculum and levels of phosphorus on yield and quality of soybean varieties. Methodology : The experiment evaluated three factors namely, varieties, Bradyrhizobium inoculum and phosphorus levels. The three varieties were: i) Binasoybean-1 ii) Binasoybean-2 and iii) BARI Soybean-6. Bradyrhizobium inoculum levels were: i) control (no inoculum), ii) 50% of recommended dose (RD) (25 g kg -1 seed) iii) 100% of RD (50 g kg -1 seed). The phosphorus levels were: i) control (no phosphorus) ii) 50% of RD (18 kg ha -1 ), iii) 100% of RD (36 kg ha -1 ). The experiment was laid out in a randomized complete block design with three replications. Data were recorded on yield, yield contributing characters and quality parameters of seeds. Results: Binasoybean-1 performed superiorly in terms of all the yield and yield contributing characters. Application of 100% of RD of Bradyrhizobium inoculum and application of 100% of RD of phosphorus also showed superior performance. Considering the interaction effect, the findings of the study indicate that Binasoybean-1 responded well to the application of 100% of RD of phosphorus and 100% of RD of Bradyrhizobium and gave maximum seed yield. The highest protein content (39.21%) was found in Binasoybean-2, 50% of RD of phosphorus with no Bradyrhizobium inoculum and the highest oil content was found (19.33%) in Binasoybean-2, no phosphorus and 50% of RD of Bradyrhizobium inoculum. Implication: Binasoybean-1 with the application of 100% of RD of phosphorus and 100% of RD of Bradyrhizobium might be recommended to obtain higher yield soybean in Bangladesh. Conclusion: From this study it may be concluded that recommended dose of phosphorus and Bradyrhizobium is beneficial for a higher yield of Binasoybean-1.


INTRODUCTION
Soybean (Glycine max L) is an important economic crop among grain legumes, mostly grown in a wide range of environments all over the world.It covers 120.30 million ha of land worldwide with the production of 333.67 million tons (FAO, 2021).Protein and oil are two significant components of soybeans that contribute to their overall quality.It contains oil 20-22%, protein 42-45%, carbohydrates 30-35% and total sugar 10-12% and also high amount of the amino acid, thiamin, vitamins, niacin, riboflavin, phosphorus, calcium and iron (Wahhab et al., 2001).It contributes 25% of the global edible oil production (Jaybhay et al., 2021).The multipurpose use of soybean is gradually increasing day by day in Bangladesh.About 0.717 lac hectares of land is under soybean cultivation and annual production is approximately 1.35 lac metric tons with an average yield of 1.717 t ha -1 (DAE, 2021).But the supply of soybean is very lower than the demand (Rashid et al., 2023).Considering the ever-increasing demand of edible oil of our country, it is extremely needed to increase the total production of oil crops by fitting the existing cropping patterns by replacing the high yielding variety (HYV) with low yielding varieties through improving management practices as well as increasing the area of cultivation wherever possible (Salam and Kamruzzaman, 2015).Proper nutrient management is one of the cultivation techniques that are expected to contribute substantially in increasing soybean production (Bagale, 2021).
Varietal differences in response to nutrient levels and plant efficiencies for nutrient uptake have been reported for many species (Pal et al., 2016;Paul et al., 2018;Jha et al., 2023).The search for efficient plants in nutrient uptake and use has been stimulated since large genetic variability was reported for these characters within germplasm of several species (Furlani et al., 2002;Paul et al., 2019).Variety has been reported to affect the yield and quality of soybean.Bangladesh Agricultural University, Bangladesh Agricultural Research Institute and Bangladesh Institute of Nuclear Agriculture have developed different high yielding soybean varieties (Nimu et al., 2020;Das et al., 2022;Rabbani et al., 2023).Different varieties can respond differently with different fertilizer levels.Also, studies elsewhere show that low native soil phosphorus availability coupled with poor utilization efficiency of added nutrients is a major constraint limiting the productivity of soybean.In soybean, phosphate fertilizer shows a significant effect in stimulating the development of roots, so the plants will be more resistant on drought, accelerate the harvest and add nutritional value from seeds.Phosphorus has been demonstrated to increase root nodule weight and quantity as well as to improve seed yield of soybean (Khanam et al., 2016;Barman et al., 2023) and faba bean (Yasmin et al., 2020).
Legume plants particularly soybean have the ability to fix nitrogen from atmosphere by symbiotic relationship with Rhizobial bacteria (Coskan and Dogan, 2011).These bacteria are located around root hair and fixes atmospheric nitrogen using particular enzyme called nitrogenase.When this mutualistic symbiosis established, rhizobia use plant resources for their own reproduction whereas fixed atmospheric nitrogen is used to meet nitrogen requirement of both itself and the host plants.Supply of nitrogen through biological nitrogen fixation has ecological and economic benefits (Ndakidemi et al., 2006).The symbiotic relationship between the soybean root and Rhizobial root colonies and subsequent symbiotic nitrogen fixation is one of the most important physiological processes, which occurs in the growth, and development of the soybean plant.Research done by Bambara and Ndakidemi, (2009) concluded that Rhizobium sp.inoculation in legumes stimulated growth and is an alternative source to the expensive commercial nitrogen fertilizers.Nitrogen is highly needed for all enzymatic reactions in a plant, also is a major part of the chlorophyll molecules and plays a necessary role in photosynthesis and is a major component of several vitamins (Uchida, 2000).In legumes and other leafy vegetables, nitrogen improves the quality and quantity of dry matter and protein (Uchida, 2000).
In soybean production, phosphorus and inoculation with the appropriate Bradyrhizobium strains have quite prominent effects on yield parameters (Kumaga and Ofori, 2004).Inoculation is an activity of transferring microorganisms in the form of bacteria and fungi from the place or source of origin to the new medium.Rhizobium inoculation on soybean plants has a long been known as one of the biological fertilizers.In soybean plants to produce 1 kg of seeds, plants absorb 70-80 grams of nitrogen from the soil so that if the yield of 1.5 tons/ha it will absorb 105-120 nitrogen from the soil (Purwaningsih et al., 2015).The factors which control the amount of nitrogen fixed include available soil nitrogen, genetic determinants of compatibility in both symbiotic partners and lack of other yield-limiting factors like edaphic factors associated with phosphorus deficiency (Harold et al., 1992).The absence of the required Rhizobia species and optimal phosphorus levels limit legume production in different parts of the world.Inoculation with compatible and suitable Rhizobia with optimum phosphorus levels may be essential where a low population of native Rhizobial strains prevail and is one of the key components of which grain legume farmers can use to optimize yields and seed quality.Therefore, there is a scope to work on effect of Bradyrhizobium inoculum and phosphorus fertilization on yield and quality of soybean varieties.

Features of the experimental location
To find out the effect of Bradyrhizobium inoculum and levels of phosphorus on yield and quality of soybean varieties an experiment was carried out at the experimental field of Farm Management Section, Bangladesh Agricultural University (90º 50' E and 24º 75' N and at an altitude of 18 meter) during the Rabi (winter) season of 2019.The experimental area belongs to the non-calcareous dark grey floodplain soil under Old Brahmaputra Floodplain Agro-ecological Zone (AEZ-9).The land was well drained medium high with silt-loam textured soil.The soil almost neutral in reaction (pH 6.8), low in organic matter content (1.27%) and the general fertility level of the soil was low (1.1% total N, 25 ppm available P and 0.16 me % exchangeable K).The experimental area characterized by subtropical monsoon climate with a humid environment.

Experimental treatments and design
The experiment consisted of three factors namely, varieties, Bradyrhizobium inoculum (Brdyrhizobium japonicum) and phosphorus levels.Three varieties included i) Binasoybean-1 ii) Binasoybean-2 and iii) BARI Soybean-6.While three different Bradyrhizobium inoculum were i) control (no Bradyrhizobium inoculum), ii) 50% of RD (25 g kg -1 seed) iii) 100% of RD (50 g kg -1 seed) and phosphorus levels were i) control (no phosphorus) ii) 50% of RD (18 kg ha -1 ), iii) 100% of RD (36 kg ha -1 ).Triple super phosphate was used as a source of phosphorus.The recommended dose (RD) of Brdyrhizobium japonicum was 50 g per kg soybean seed (BINA, 2020).The experiment was laid out in a Randomized Complete Block Design with three replications.The unit plot size was 4.0 m × 2.5 m.

Field preparation and fertilizer application
The piece of land selected for carrying out the experiment was opened with a power tiller and was exposed to the sun for a week after which the land was harrowed, ploughed and cross-ploughed several times followed by laddering to obtain a good tilth.Manures and fertilizers were applied by following Fertilization Recommendation Guide-2012(FRG, 2012).Well decomposed cowdung at the rate of 20 t ha -1 was applied during final land preparation.Urea, muriate of potash (MoP), gypsum and boric acid were applied at the rate of 60, 120, 110 and 10 kg per ha, respectively without triple superphosphate (TSP).Entire amount of urea, MoP, gypsum and boric acid were applied at the time of final land preparation.Triple superphosphate was applied as per experimental treatment specification.The recommended dose of TSP was 36 kg per ha.

Seed inoculation application
The seeds were inoculated with commercial Bradyrhizobium japonicum inoculant (Legumefix) as per treatment specification before sowing.The soybean seeds were put in a plastic bucket and moistened with ordinary tap water, stirred uniformly with a wooden spatula.The inoculants were added to the moistened seeds, stirred gently and uniformly, until the seeds were evenly coated.The seeds were then spread on a sheet of canvas material under a shade for at least one hour to allow the inoculants adequately adhere to the surface of the seeds.The sowing was done early in the morning to avoid exposing the inoculants to direct sunrays, which might affect the quality of the inoculants.

Agronomic management of the crop
Furrows were made for sowing seeds when the land was in proper moisture condition and seeds were sown.The seed rate was 80 kg ha -1 .During seed emergence period, weeding and thinning were done on the 25 days after the emergence (DAE).Keeping only the vigorous seedling, the rest of the seedlings were removed.Two irrigations were applied in the experimental plots during the growing period.The first irrigation was applied on the 4 th week after emergence and the second irrigation was applied on the 8 th week after emergence by flood irrigation method.The crop was harvested at 80-85% pod maturity of the terminal raceme.Prior to harvest five plants were selected randomly from each unit plot and uprooted to record data on branches per plant (no.), length of pod (cm), effective pods per plant (no.), seeds per pod (no.), 1000-seed weight (g), protein (%) and oil (%) content of soybean seeds.The harvesting was done at different dates, as the maturity period of the genotypes was not same.The plants were sun dried properly.Seeds were separated from pods and finally seed and stover yields were recorded and converted to ton per ha.

Measurement quality parameters
Protein content was computed by multiplying N content in soybean seed determined by Microkjeldahl assay by a conventional factor of 6.25 (Jackson, 1973).The oil content of soybean seed was extracted by Folsch method (Folsch et al., 1957) by using chloroform:methanol in 2:1 ratio in a beaker with stirring.The extractant was removed by heating and oil obtained was expressed in percentage.

Data analysis
Data were compiled and analyzed to find out the significance of variation resulting from the experimental treatments.All the collected data were analyzed following three way factorial analysis of variance (ANOVA) technique and mean differences were adjudged by the Duncan Multiple Range Test (Gomez and Gomez, 1984) using the program MSTAT-C (Russel, 1986).

Effect of variety on yield, yield contributing and quality parameters
All the yield and yield contributing characters were significantly influenced by varietal effect except seeds per pod, 100-seed weight and harvest index.The maximum number of branches per plant (2.88), pods per plant (44.08), seed yield (2.43 t ha -1 ) and stover yield (4.37 t ha -1 ) were produced by Binasoybean-1.The longest pod (3.36 cm) was found in Binasoybean-2.The minimum number of branches per plant (2.68), pod length (3.21 cm), pods per plant (40.48), seed yield (2.21 t ha -1 ) and stover yield (3.96 t ha -1 ) were found in BARI Soybean-6.The protein and oil content of soybean were significantly influenced by the variety.From the Table 1, it is observed that the difference between protein content and oil content was huge.The highest protein content (37.48%) was found in Binasoybean-1 and oil content was found (17.76%) in the same variety.The lowest protein content (36.77%) was found in Binasoybean-2 and lowest oil content (16.84%) was recorded in the BARI Soybean-6 (Table 1).

Effect of Phosphorus on yield, yield contributing and quality parameters
Application of different level of phosphorus had significant (p<0.01)effect on all the yield and yield contributing characters except branches per plant, seeds per pod, 100-seed weight and harvest index.The maximum number of pods per plant (43.78), seeds per pod (2.98) were observed at 100% of RD of phosphorus.Maximum pod length (3.31 cm), seed yield (2.51 t ha -1 ) and stover yield (4.49 t ha -1 ) were recorded in 100% of RD of phosphorus.On the other hand, minimum no. of pods per plant (40.07), seeds per pod (2.56), seed yield (2.15 t ha -1 ) and stover yield (3.87 t ha -1 ) were documented in control treatment.The protein and oil content of soybean were not significantly influenced by the effect of phosphorus (Table 2).3).

Interaction effect of variety and phosphorus on yield, yield contributing and quality parameters
The interaction effect of variety and phosphorus inoculum was significant (p<0.01) on all the yield and yield contributing characters except 100-seed weight and harvest index.) and stover yield (3.84 t ha -1 ) was produced in Binasoybean-1 with control treatment.The protein and oil content of soybean were not significantly influenced by the interaction effect of variety and phosphorus (Table 4).

Interaction effect of variety and Bradyrhizobium inoculum on yield, yield contributing and quality parameters
Effect of interaction of variety and Bradyrhizobium inoculum on number of branches per plant was significant (p<0.01).The interaction of Binasoybean-1 and 50%   ) and I2= 100% of RD (50 g kg -1 ) The highest number of pods per plant (60.00) was found in Binasoybena-1, 100% of RD of phosphorus and 50% of RD of Bradyrhizobium inoculum and the lowest one (31.76)was found in Binasoybena-1, 50% of RD of phosphorus with no Bradyrhizobium inoculum and BARI Soybean-6, 50% of RD of phosphorus with 50% of RD of Bradyrhizobium inoculum.The interaction of variety, phosphorus and Rhizobium inoculum exerted significant in respect of seed yield.The highest seed yield (2.90 t ha -1 ) and stover yield (4.94 t ha -1 ) wasobtained in Binasoybean-1, 100% of RD of phosphorus with 100% of RD of Bradyrhizobium inoculum and the lowest seed yield (1.89 t ha -1 ) and stover yield (1.89 tha -1 ) was obtained from BARI Soybean-6, no phosphorus and no Bradyrhizobium inoculum.The protein and oil content of soybean were significantly influenced by the interaction effect of variety, phosphorus and Bradyrhizobium.The highest protein content (39.21%) was found in Binasoybean-1, 50% of RD of phosphorus with no Bradyrhizobium inoculum and BARI Soybean-6, 50% of RD of phosphorus with no Bradyrhizobium inoculum.The highest oil content (19.33%) was found in Binasoybean-1, no phosphorus and 50% of RD of Bradyrhizobium inoculum.On the other hand, the lowest protein content (33.00%) was found in Binasoybean-2, 100% of RD of phosphorus with 50% of RD of Bradyrhizobium inoculum and the lowest oil content (15.18%) was recorded in BARI Soybean-6, no phosphorus with 50% of RD of Bradyrhizobium inoculum (Table 7).

DISCUSSION
In Bangladesh, soybean has great potential as it is the crop with the richest source of protein (Salam and Kamruzzam, 2015), and it requires less fertilizer, reducing the input cost.Previously neglected, soybean cultivation is now gaining popularity due to the availability of high-yielding, short-duration varieties and a suitable climatic condition (Islam et al., 2022).In this experiment, various soybean cultivars responded differentially to phosphorus and Bradyrhizobium inoculation levels.The varietal effect had a considerable impact on all yield and yield contributing factors.In comparison to the other two varieties, the Binasoybean-1 variety produced the highest yield.Seed yield differences among the three varieties may be due to their variant genetic make-up.Seed yield variation of soybean in different varieties was reported by previous studies (Khan et al., 2015;Nimu et al., 2020;Das et al., 2022;Rabbani et al., 2023).Protein and oil content are affected by the variety since they are a property of that variety, even if the oil and protein content might alter due to changes in agronomic techniques, the soil, or the climatic conditions in which the varieties are cultivated (Araujo et al., 2008).) and I2= 100% of RD (50 g kg -1 ) Among the various elements that can contribute to the success of soybean cultivation, phosphorus has a considerable impact on yield and yield qualities (Kumaga and Ofori, 2004).Phosphorus-treated plants in this experiment demonstrated a much higher soybean yield.Utilizing 100% of the recommended dose of phosphorus produced better results in terms of yield and factors leading to yield.The outcome is consistent with those made by Pauline et al. (2010) and Aise et al. (2011), who found a similar conclusion on soybean seed yield under the assumption of the right P application.Because nutritional deficit affected the growth and development of soybean, the decrease in seed production at the lower P level was probablycaused by this (Xiang et al., 2012).Phosphorus is also an essential ingredient for  (2003) and Tomar et al. (2004).They showed that both inoculation and increasing levels of phosphorus have significant effects on protein contents of soybean.
Bradyrhizobium inoculation can boost soybean output, but its effectiveness depends on a number of factors, including the genotype of the crop.In this investigation, the greatest yield was achieved when Binasoybean-1 interacted with 100% RD of phosphorus and 100% RD of Bradyrhizobium.When variety coupled with phosphorus and variety coupled with Bradyrhizobium inoculum, it is observed that Binasoybean-1 with 100% of the RD of P, or Binasoybean-1 with 100% of RD of Bradyrhizobium inoculum produced the highest seed yield which was statistically identical to Binasoybean-2 with 100% of RD of Bradyrhizobium inoculum.In the case of combination of P and Bradyrhizobium inoculum, 100% of the RD of P with 100% of RD of Bradyrhizobium inoculum also produced the highest seed yield of soybean.In the case of combination of variety, phosphorus and Bradyrhizobium inoculum, Binasoybean-1 with combination of 100% of RD of phosphorus and 100% of RD of Bradyrhizobium inoculum produced the highest seed yield of soybean.Begum et al. (2015) studied the effect of phosphorus on the performance of soybean and reported that application of P at 54 kg ha -1 showed the highest number of effective pods per plant.Ashraf et al. (2002) reported a specific combination of soybean genotype with Bradyrhizobium strains resulting in many folds increase in the amount of N2 fixed and grain yield of soybean.Ghasem et al. (2015) reported RI inoculated treatment increased 23% seed in pod compared with non-inoculated treatment.Few reports also stated that RI increased the number of nodule and yield in soybean (Majid et al. 2009).Several scientists reported that soybean growth, 100-grain weight and seed yield is significantly increased by the adding of 90 and 100 kg P2O5 ha -1 (Taj et al., 1986).Matusso et al. (2015) reported that the combination of RI and P significantly increased seed yield in soybean.Abbasi et al. (2008) used the combination of Rhizobium inoculum and P and reported that there was a positive effect of Rhizobium inoculum and P on growth, nodulation and yield of soybean.Few reports showed that application of Rhizobium inoculum and P significantly increased the number of pods per plant (Matusso et al., 2015).Akter et al. (2021) revealed that use of Rhizobium inoculum at 50 g kg -1 seed in combination with 54 kg ha -1 P appeared as the best practice for producing the highest grain yield of Binasoybean-1.

CONCLUSION
The findings of the study indicate that Binasoybean-1 responded well to the application of 100% of RD of phosphorus and 100% of RD of Bradyrhizobium and gave maximum seed yield with highest protein content.Finally, it may conclude that Binasoybean-1 with the application of 100% of RD of phosphorus and 100% of RD of Bradyrhizobium might be recommended to obtain optimum yield and quality of soybean.However, further trial with the treatment combinations on different agro-ecological zones of Bangladesh will be useful to confirm the result of the present study.

Funding.
Bangabandhu Science and Technology fellowship Bangabandhu Science and Technology Fellowship Trust, Ministry of Science and Technology, Government of the People's Republic of Bangladesh

Table 1 . Effect of variety on yield, yield contributing and quality parameters of soybean.
*= Significant at 5% level of probability, ** =Significant at 1% level of probability, NS = Non significant

Table 3 . Effect of level Bradyrhizobium on yield, yield contributing and quality parameters of soybean.
*In a column, figures with the same letter (s) or without letter do not differ significantly whereas figures with dissimilar letter differ significantly (as per DMRT) ** =Significant at 1% level of probability, NS = Non significant