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Frontiers of Agricultural Science and Engineering

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Front. Agr. Sci. Eng.    2024, Vol. 11 Issue (3) : 499-513    https://doi.org/10.15302/J-FASE-2024559
Isolation, characterization and identification of plant growth-promoting rhizobacteria from the rhizosphere of Acacia mangium at sandy BRIS soil
Zakiah MUSTAPHA1(), Radziah OTHMAN2, Nik Nurnaeimah NIK MUHAMMAD NASIR1, Dhiya Dalila ZAWAWI1, Mohd Khairi CHE LAH3, Hafizan JUAHIR1()
1. School of Agriculture Science and Biotechnology, Faculty of Bioresources and Food Industry, Universiti Sultan Zainal Abidin, Besut Campus, 22200 Besut, Terengganu, Malaysia
2. Department of Land Management, Faculty of Agriculture, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia
3. Faculty of Plantation and Agrotechnology, Universiti Technologi MARA Cawangan Pahang, 26400 Jengka, Pahang, Malaysia
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Abstract

● Isolation of potential PGPR from rhizosphere sandy BRIS soil of Acacia mangium .

● The isolated rhizobacteria showed significantly varied growth in organic molasses medium supplemented with KNO3.

● The ability to fix atmospheric N2, solubilize P and K, produce IAA and siderophores varied differently for single and mixed strains of the isolated rhizobacteria.

● The single or mixed strains of rhizobacteria had a significant effect on corn phenology, growth and yield.

● Identification of the isolated rhizobacteria at the molecular level.

This study has isolated, characterized, and identified potential plant growth-promoting rhizobacteria (PGPR) with multiple PGP characteristics (N2-fixation, P- and K-solubilization, IAA, and siderophores production) from the rhizosphere BRIS soil of Acacia mangium. A total of 24 pure colonies were isolated and only 8 colonies were selected for further evaluation of the growth rate in 5% organic molasses medium supplemented with 2% KNO3. Based on the biochemical, potential PGP characteristics and growth performance, 3 superior PGPR strains were selected and identified as Paraburkholderia unamae (UA1), Bacillus amyloliquefaciens (UA6), and Enterobacter asburiae (UAA2) by partial sequencing of the 16S rRNA gene. The selected bacterial strains either in single or mixed (UA1 + UA6 + UAA2) cultures have shown a significant biochemical estimation of the PGP characteristics. Each strain has its own PGPR traits superiority with UA1 showing the best PGP characteristic followed by UA6 and UAA2. The use of mixed bacterial strains was beneficial as it showed the best performance in N2-fixation, siderophores production, and significant effect on corn phenology, growth and yield compared to using a single strain. These types of microbes showed potential to be used as biofertilizer and should be exploited more.
Keywords Biofertilizer      corn      organic molasses me- dium      PGP characteristic      phytohormone      potassium nitrate      siderophores     
Corresponding Author(s): Zakiah MUSTAPHA,Hafizan JUAHIR   
Just Accepted Date: 28 April 2024   Online First Date: 30 May 2024    Issue Date: 17 July 2024
 Cite this article:   
Zakiah MUSTAPHA,Radziah OTHMAN,Nik Nurnaeimah NIK MUHAMMAD NASIR, et al. Isolation, characterization and identification of plant growth-promoting rhizobacteria from the rhizosphere of Acacia mangium at sandy BRIS soil[J]. Front. Agr. Sci. Eng. , 2024, 11(3): 499-513.
 URL:  
https://academic.hep.com.cn/fase/EN/10.15302/J-FASE-2024559
https://academic.hep.com.cn/fase/EN/Y2024/V11/I3/499
Location Coordinate Colony count (CFU·g–1)
Aerobic Anaerobic
Point 1 5°45′26.4′′ N 102°37′21.3′′ E 1.06 × 106 ± 0.88c 1.16 × 106 ± 3.18c
Point 2 5°45′43.7′′ N 102°37′29.7′′ E 9.67 × 105 ± 0.58bc 1.01 × 106 ± 0.88bc
Point 3 5°45′32.1′′ N 102°8′18.4′′ E 8.00 × 105 ± 1.00ab 7.00 × 105 ± 0.58a
Tab.1  Bacterial count from three different rhizospheric soil of Acacia mangium at BRIS soil
Isolates Gram reaction Size Shape Color Catalase Nitrate reduction Indole Urease Ammonia Nitrogenfixation Phosphatesolubilization Potassiumsolubilization Siderophores production
UA1 – Pinpoint Round Creamy white + + + + + + +++ +++ ++
UA2 – Pinpoint Round Creamy white + + + + + + ++ ++ ++
UA3 + Moderate Rod Creamy white + + + + – + + + ++
UA4 + Large Rod Creamy white + + + + + + + + ++
UA5 + Large Rod Creamy white + + + + + + ++ ++ +++
UA6 + Large Rod Creamy white + + + + + + ++ ++ +++
UAA1 – Small Rod Red + + – – – + + ++ +++
UAA2 – Small Rod Creamy white + + – – + + ++ +++ +++
Tab.2  Morphological and biochemical characterization of selected eight isolates of BRIS soil rhizobacteria
Fig.1  Growth of bacterial isolates in 5% molasses medium supplemented with 2% KNO3 at 5 days after incubation. Mean with the same letters are not significantly different at p < 0.05 Tukey’s multiple comparisons, n = 3. Bar indicates the standard error of the treatment’s mean.
PGPR traits Unit Control UA1 UA6 UAA2 Mixed strain(UA1 + UA6 + UAA2)
BNF nmol·mL–1·h–1 0.0±0.00a 17.1±1.20b 18.6±0.48bc 17.0±0.65b 21.6±0.70c
P-solubilization pH (day 6) 5.9±0.00e 5.8±0.00c 5.9±0.02d 5.8±0.01c 5.9±0.00e
pH (day 12) 6.0±0.00e 5.4±0.01a 5.6±0.01b 5.0±0.01 5.5±0.00b
μg·mL–1 (day 6) 0.0±0.00a 45.8±2.08cd 29.1±2.65b 43.4±2.18abc 41.1±4.54abc
μg·mL–1 (day 12) 0.0±0.00a 54.4±4.23d 32.0±2.38bc 49.1±2.97d 43.9±3.72cd
K-solubilization mg·L–1 0.0±0.00a 17.5±0.49d 11.6±0.52b 12.3±0.66b 15.2±0.17c
IRCs (IAA) production μg·mL–1 0.0±0.00a 17.3±0.33e 13.5±0.24c 9.9±0.48b 15.8±0.25d
Siderophores production % 0.0±0.00a 14.7±2.32b 31.7±2.32b 20.3±1.34c 43.0±2.66d
Tab.3  Estimation of PGPR characteristics isolated from BRIS soil
Treatment Emergence(day) Tasseling(day) Maturity(day) Plant height(m) Leaf no. of plant Cob length(cm) Grain yield(t·ha–1) Total biomass(t·ha–1) Harvest index
T1 11.00b 57.10d 107.67a 2.21a 13.08a 20.34a 2.49a 4.36a 57.11a
T2 8.50a 53.00bc 115.08b 2.20a 14.33b 22.18b 2.97bc 4.74ab 62.64ab
T3 8.00a 51.75ab 121.50c 2.28a 14.83b 23.69c 3.09bc 4.88b 63.57b
T4 8.40a 53.67c 116.75b 2.19a 14.42b 21.79b 2.86b 4.83ab 59.25ab
T5 7.90a 50.91a 122.83c 2.21a 15.33b 24.49d 3.17c 5.02b 63.77b
Tab.4  Mean values of corn phenology, growth and yield parameters as affected by different treatments of BRIS soil PGPR
Fig.2  Phylogenetic tree of (a) UA1, (b) UA6, and (c) UAA2 based on the 16S rRNA sequences of the selected strains and related bacteria.
Isolates 16S rRNA fragment length (bp) Closest relatives in NCBI NCBI accession number Similarity (%)
UA 1 1337 Paraburkholderia unamae strain CATux-40 HQ023248.1 99
UA 6 1351 Bacillus amyloliquefaciens strain RD7-7 CP016913.1 100
UAA 2 1406 Enterobacter asburiae strain ENIPBJ-CG1 CP014993.1 99
Tab.5  Molecular bacterial identification using 16S rRNA gene sequences
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