The role of Ni- and Cd-resistant rhizobacteria in promoting the growth of rice seedlings and alleviating the combined phytotoxicity of Ni and Cd.

The problem of potentially toxic metal pollution is increasingly acute with the development of human society. In this study, we investigated the remediation of nickel (Ni) and cadmium (Cd) co-contamination through inoculating rice with three new-isolated Ni- and Cd-resistant plant growth-promoting rhizobacteria (PGPR) Y3, Y4, and Y5. These three strains possessed growth-promoting properties, including 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, the ability of indoleacetic acid (IAA) production, phosphate solubilization, siderophores production, and exopolysaccharide (EPS) development. According to 16S rDNA sequence homology, strains Y3, Y4, and Y5 were identified as Pseudomonas sp., Chryseobacterium sp., and Enterobacter sp., respectively. Based on the results of rice germination experiments conducted under combined toxicity, we set the contamination concentrations for Ni2+ at 20 μg mL-1 and Cd2+ at 40 μg mL-1. Then we conducted potting experiments at these concentration levels to study the effects of strains Y3, Y4, and Y5 on rice growth under synergistic Ni and Cd stress. The results indicated that the inoculated strains Y3, Y4, and Y5 were effective in promoting the growth of rice seedlings under the combined stress of Ni and Cd, and conferring tolerance to Ni and Cd by increasing the antioxidant enzyme activities of the seedlings. Among them, strain Y3 exhibited stronger ACC deaminase activity, IAA production capacity, and EPS production capacity, showing the most pronounced growth-promoting effect on rice. It was demonstrated that after inoculation with strain Y3, the germination rate of rice seeds increased by 43 %, the fresh weight of stems improved by 35 %, and the chlorophyll content enhanced by 70 % and other growth-promoting phenomena. Additionally, under Ni and Cd stress, strain Y5 performed better than strain Y4 in terms of IAA production capacity and its influence on rice root growth, suggesting that IAA production might play a specifically essential role in root growth under Ni and Cd stress.

Zhou W ，Yang J ，Qi L ，Wang G ，Guan C ，Li Q ... -  《-》

Bioaccumulation of cadmium by Enterobacter sp. and enhancement of rice seedling growth under cadmium stress.

Bacteria-mediated plant growth promotion and bioremediation of heavy metal containing soil is a widely accepted eco-friendly method. The present study is aimed to screen out cadmium resistant bacterial strain from metal contaminated rice rhizosphere and evaluate its effects on the growth of rice seedlings under cadmium stress. Among four different isolates (designated as S1, S2, S3 and S5), the S2 isolate was screened on the basis of different PGP traits and multi heavy metal resistance (minimum inhibitory concentration for cadmium, lead and arsenic were 3500, 2500 and 1050 µg/ml respectively). The selected S2 strain has ability to produce ACC deaminase (236.11 ng α-keto-butyrate/mg protein/h), IAA (726 µg/ml), solubilize phosphate (73.56 ppm) and fix nitrogen (4.4 µg of nitrogen fixed/h/mg protein). The selected strain was identified as Enterobacter sp. on the basis of phenotypic characterization, MALDI-TOF MS analysis of ribosomal proteins, FAME analysis and 16 S rDNA sequence homology. The high cadmium removal efficiency (> 95%) of this strain from the growth medium was measured by Atomic Absorption Spectrophotometer and it was due to intracellular cadmium accumulation evidenced by SEM-EDX-TEM-EDX study. SEM analysis also revealed no distortion of surface morphology of this strain even grown in the presence of high cadmium concentration (3000 µg/ml). Inoculation of this strain with rice seedlings significantly enhanced various morphological, biochemical characters of seedling growth compared with un-inoculated seedlings under Cd stress. The strain also exhibited alleviation of cadmium-induced oxidative stress, reduction of stress ethylene and decreased the accumulation of cadmium in seedlings as well that conferred cadmium tolerance to the plant. Thus the S2 strain could be considered as a potent heavy metal resistant PGPR applicable in heavy metal contaminated agricultural soil for bioremediation and plant growth promotion as well. A cadmium resistant plant growth promoting Enterobacter sp. was isolated that accumulated cadmium evidenced by SEM-TEM-EDX study. It reduced Cd uptake and enhanced growth in rice seedlings.

Mitra S ，Pramanik K ，Sarkar A ，Ghosh PK ，Soren T ，Maiti TK ... -  《-》

Characterization of Cd-resistant Klebsiella michiganensis MCC3089 and its potential for rice seedling growth promotion under Cd stress.

Application of heavy metal resistant plant growth promoting rhizobacteria has an important role as they help to evade metal-induced toxicity in plants on one hand and enhance plant growth on the other. The present study is therefore focused on the characterization of a cadmium resistant bacterial strain isolated from heavy metal contaminated rhizospheric soil designated as S8. This S8 strain was selected in terms of cadmium resistance and plant growth promoting traits. Moreover, it also showed resistance to lead and arsenic to a considerable extent. The selected strain S8 was identified as Klebsiella michiganensis by modern approaches of bacterial taxonomy. The plant growth promoting traits exhibited by the strain include 1-aminocyclopropane-1-carboxylic acid deaminase activity (58.33 ng α-keto butyrate/mg protein/h), Indole-3-acetic acid production (671 μg/ml), phosphate solubilization (71.98 ppm), nitrogen fixation (3.72 μg of nitrogen fixed/h/mg protein) etc. Besides, the strain also exhibited high cadmium removal efficiency (73-97%) from the medium and intracellular accumulation as well. Its efficiency to alleviate cadmium-induced toxicity was determined against a rice cultivar in terms of morphological and biochemical changes. Enhanced growth and reduced oxidative stress were detected in presence of the bacterium. On the basis of these results, it can be concluded that K. michiganensis strain S8 is cadmium accumulating plant growth promoting rhizobacterium that can be applied in cadmium contaminated agricultural soil to achieve better productivity of rice.

Mitra S ，Pramanik K ，Ghosh PK ，Soren T ，Sarkar A ，Dey RS ，Pandey S ，Maiti TK ... -  《-》

Alleviation of phytotoxic effects of cadmium on rice seedlings by cadmium resistant PGPR strain Enterobacter aerogenes MCC 3092.

Heavy metal resistant PGPR mediated bioremediation, phytostimulation and stress alleviation is an eco-friendly method for sustainable agriculture in the metal contaminated soil. The isolation of such PGPR is highly demanding to reduce heavy metals in contaminated cultivated fields for agricultural benefit. The present study was successful to isolate a potent multi-heavy metal resistant PGPR strain, identified as Enterobacter aerogenes strain K6 based on MALDI-TOF MS, FAME analysis and 16S rDNA sequence homology, from rice rhizosphere contaminated with a variety of heavy metals/metalloid near industrial area. The strain exhibited high degree of resistance to Cd2+, Pb2+ and As3+ upto 4000 μg/mL, 3800 μg/mL and 1500 μg/mL respectively. Intracellular Cd accumulation of this strain was evidenced by AAS-SEM-TEM-EDX-XRF studies. Moreover, it showed several important PGP traits like IAA production, nitrogen fixation, phosphate solubilization, ACC deaminase activity even under high Cd stress (upto 3000 μg/mL). The combined effect of Cd resistance and PGP activities of this strain was manifested to the significant (p < 0.05) growth promotion of rice seedling under Cd stress by reducing oxidative stress (through antioxidants), stress ethylene and Cd uptake in seedlings. Thus K6 strain conferred Cd-tolerance in rice seedlings and could be applied as PGPR in contaminated fields.

Pramanik K ，Mitra S ，Sarkar A ，Maiti TK ... -  《-》

Enhancement of growth and salt tolerance of rice seedlings by ACC deaminase-producing Burkholderia sp. MTCC 12259.

Increasing soil salinity is often associated with accelerated ethylene production in plants, leading to overall growth reduction. The salt-tolerant 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-producing PGPR may alleviate salt stress by reducing the production of stress ethylene. In this study, a salt-tolerant ACC deaminase-producing strain named P50 was isolated from a coastal rice field in Odisha, India, which enhanced the growth of rice seedlings under salt stress. The P50 strain was identified as Burkholderia sp. based on phenotypic characteristics, MALDI-TOF MS data for ribosomal proteins and 16S rDNA sequence-based homology. Various PGP traits of strain P50 were characterized, among which the ACC deaminase activity was optimized at different physical conditions and confirmed by enzyme assay, as well as FTIR. The IAA, EPS and proline production of this strain were estimated under increasing NaCl concentrations essential for plant growth promotion under salt stress. Finally, the P50 strain was utilized in a gnotobiotic assay using rice seedlings (cv. Swarnamasuri) under saline stress. Seedlings treated with the P50 strain showed improvement in various morphological and biochemical characteristics, ROS scavenging antioxidant enzymatic activities, and reduced amounts of stress ethylene compared to non-inoculated strains under salinity. In addition, isolation of the ACC deaminase mutant of this strain was not found to reduce stress ethylene, confirming that the P50 strain was associated with a reduction in stress ethylene. Strain P50 was also found to colonize the root surfaces of rice seedlings associated with the plant-microbe interaction process. Thus, as an effective salt-tolerant PGPR, strain P50 can be utilized in salt-affected agricultural fields to improve plant growth in a sustainable manner.

Sarkar A ，Pramanik K ，Mitra S ，Soren T ，Maiti TK ... -  《-》