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Characterization and Selection of Phosphate Solubilizing Bacteria and Their Use as Biofertilizer for the Growth and Yield of Rice and Wheat

Md. Azizul Haque
Senior Scientific Officer,
Soil Science Division, BINA, Mymensingh

Abstract/ Executive Summary:

Laboratory, pot and field experiments were conducted to select the efficient phosphate solubilizing bacterial inoculants for wheat and rice cultivation in Bangladesh. Forty phosphate solubilizing bacteria (PSB) were isolated on tricalcium phosphate (TCP) based solid medium from rhizosphere of rice, wheat, lentil and chickpea from four agroecological zones (AEZs) of Bangladesh and eight PSB were selected on the basis of P solubilization in liquid culture. The selected isolates widely varied in terms of decreasing pH, increasing P solubilization and acid phosphatase activity in liquid media with five insoluble P substrates viz. tricalcium phosphate (TCP), ferric phosphate (Fe-P), aluminium phosphate (Al-P), rock phosphate (RP) and calcium phytate (Ca-phytate). The PSB isolate MR1 showed the highest P solubilization (45.1%) and acid phosphatase activity (136.3 µg pNP mL^-1 h^-1) from the Al-P and Ca-phytate substrates, respectively followed by the isolate MW1. Morphological, biochemical and molecular characterization indicated that all the eight PSB isolates belong to the genus Pseudomonas and were identified as P. aeruginosa (IL1, MR1, RC1, IC2 and MdR3), P. fluorescens (IW1 and MyR2) and P. putida (MW1). Combined restriction fragment length polymorphism (RFLP) analyses revealed that all the eight PSB isolates depicted in four distinct groups which varied from phenotypic and biochemical point of view but indicated that they were genetically related. Six PSB inoculants with an uninoculant (I0, MR1, IL1, IW1, IC2, RC1 and MW1) were tested with four levels of P fertilizer to investigate their performance on wheat and T. aman rice in pot culture using Mymensingh, Madhupur and Ishurdi soils. The PSB inoculant MR1 and MW1 showed better results in terms of available P, phosphatase activity and PSB population in soils resulting increased grain and straw yields of wheat (Triticum aestivum) and T. aman rice (Oryza sativa) in pot culture with all three soils. The field experiments were conducted to investigate the effects of two PSB inoculants (MR1 and MW1) and their mixture with inorganic or organic sources of P on T. aman rice and wheat at Mymensingh, Madhupur and Ishurdi. Fourteen treatments were comprised with all possible combinations with two PSB inoculants, their mixture and inorganic (50 or 100% P from triple super phosphate) or organic sources of P (50% P from cowdung) for both the crops and the experiments were conducted following RCB design with four replications for each treatment. Application of single or mixed PSB inoculants with 50% P from triple super phosphate (TSP) or cowdung (CD) increased the PSB population and phosphatases activity in rhizosphere of T. aman rice and wheat. Strong significant correlation (P0.01) was observed between these two variables resulted in higher availabile soil P due to inoculation of PSB with T. aman rice and wheat which were also comparable to the 100% P from TSP. Consequences of higher availability of soil P due to PSB inoculation attributed to increase the P uptakes and grain yields of T. aman rice (5.7 to 34.1, 2.6 to 21.4 and 0.9 to 15.2% yield increase) and wheat (35.1 to 92.4, 21.3 to 61.9 and 4.6 to 26.1% yield increase) over P0 control treatment which were also comparable to 100% P from TSP (28.1, 14.5, and 12.3% yield increase for T. aman rice and 77.4, 51.4 and 18.0% yield increase for wheat) at Mymensingh, Madhupur and Ishurdi, respectively. The PSB inoculant MR1 showed better performance at Mymensingh and Ishurdi while mixed inoculants (MR1 + MW1) performed better at Madhupur in both the crops. Thus, 50% P can be saved in the cultivation of T. aman rice and wheat by the integrated use of PSB inoculants with inorganic or organic sources of P. Survival of two PSB cultures (MR1 and MW1) and its mixtures were tested for long duration in six carrier materials against two temperature viz. air condition and room temperature. Two PSB inoculants (MR1 and MW1) and their mixtures survived with desired population (≥106 cfu g-1 carrier) for longer time in air condition (360 days) than that at room temperature (270 days) with all carrier materials. Peat soil was found the best carrier for survival of PSB inoculants followed by either its mixture with pond slurry or pressmud at the ratio of 1:1. The results revealed that mixture of peat soil either with pond slurry or pressmud can also be used as an alternate carrier for production of phosphatic biofertilizer than peat soil alone.

Keywords: Phosphate solubilizing bacteria, PSB, Biofertilizer, Characterization, Carrier materials, Rice yield, Wheat yield

Location: Dept. of Soil Science, BAU, Mymensingh and Soil Science Division, BINA, Myemensingh

Start Date: 00-00-2006

End Date: 00-00-2012

Program Area: Crop-Soil-Water Management

Commodity: Bio fertilizer

Objectives:

i) Isolation and characterization of phosphate solubilizing bacteria (PSB) from diverse soils of Bangladesh.
ii) To determine the phosphate solubilizing capacity and phosphatase activity of PSB isolates.
iii) To evaluate the effects of PSB as biofertilizer on wheat and rice under pot culture and diverse field conditions.
iv) To select a suitable carrier material for the production of phosphatic biofertilizer for wheat and rice.

Methodology:

Mainly, three types such as laboratory, pot and field experiments were conducted. Isolation of phosphate solubilizing bacteria (PSB), solubilization of phosphorus from different inorganic and organic compounds by PSB isolates, screening of highly efficient PSB isolates and their morphological, biochemical and molecular characterization were done in the laboratory. The selected PSB isolates were tested with three soils collected from different agroecological zones (AEZs) on wheat and rice in pot culture. Further, field experiments were conducted in order to evaluate the effects of PSB inoculants on rice and wheat crops in two consecutive years at three AEZs viz. AEZ 9 (BAU farm, Mymensingh), AEZ 28 (BADC farm, Madhupur) and AEZ 11 (BINA substation, Ishurdi). Finally, survival of selected PSB inoclants on different carrier materials at various temperatures were conducted in laboratory to select the carrier materials for long time storage of the PSB inoculants for use as biofertilizer.

Isolation
Collection of rhizosphere soil samples
Sixty rhizosphere soils (Table 2.1) of four crops viz. rice, wheat, lentil and chickpea were collected from four agro-ecological zones (AEZs) of Bangladesh (AEZs 9, 11, 28 and 26) for isolation of phosphate solubilizing bacteria.

Experiment-I: Selection and characterization of phosphate solubilizing bacteria
Isolation of phosphate solubilizing bacteria (PSB) from rhizosphere soils:
PSB were isolated from different rhizospere soils by serial dilution techniques (Subba Rao, 1999). The serial dilutions of soils were made from 10-1 to10-5. Phosphate solubilizing bacteria were isolated using Pikovskaya’s solid medium (Pikovskaya, 1948) containing tricalcium phosphate (TCP). The pH of the medium was adjusted to 7.2 prior to mix the agar. The medium was sterilized at 1210C temperature at 15 PSI pressure for 20 minutes. Using sterile technique 0.1 mL of aliquot of 10-3, 10-4 and 10-5 dilutions were spread on Pikovskaya’s solid medium. All the plates were incubated at 30 ⁰C for 7 days. The colonies that appeared with discrete clear zones (halo zones) around them indicated the dissolution of TCP, was assumed to be phosphate solubilizers. These colonies were picked up and further purified by repeated streaking on Pikovskaya’s medium. Thus the phosphate solubilizing bacterial (PSB) isolates were maintained on Pikovskaya’s medium slants which were stored in a refrigerator at 4⁰C for subsequent study.

Source of Information: Ph. D. Dessertation. Dept. of Soil Science, BAU, Mymensingh. year 2012.

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

• Finally it may be concluded that PSB inoculants MR1 or mixed inoculants (MR1 +MW1) can be use as phosphatic biofertilizer for T. aman rice and wheat cultivation. Integrated use of these biofertilizer with inorganic or organic sources of P can reduce the uses of 50% P from inorganic or organic sources of P in rice and wheat cultivation in Bangladesh.

• Among the carrier materials, single peat soil was the best carrier for survival of all the PSB inoculants followed by either its mixer with pond slurry or pressmud at the ratio of 1:1. However, mixtures of peat soil either with pond slurry or pressmud (1:1) also can be use as carrier for PSB inoculants as an alternate of single peat soils. All the PSB cultures might be survived with good population up to 270 DAI in room temperature and 360 DAI in air condition in different carrier materials.

Knowledge Management: