ACC Deaminase Gene Research Articles

Isolation, Cloning and Sequence Analysis of 1-Aminocyclopropane-1-Carboxylate Deaminase Gene from Native Sinorhizobium meliloti

Background: Many plant growth-promoting bacteria including Rhizobia contain the 1-aminocyclopropane-1-carboxylate (ACC) deaminase enzyme that can leave ACC, and thereby lower the level of ethylene in stressed plants. Drought and salinity are the most common environmental stress factors for plants in Iran. Objectives: The main aim of this research was development of bio-fertilizers containing ACC deaminase enzyme which is very important in conditions of stressed drought and salinity. Materials and Methods: In this research 168 isolates of native Sinorhizobium meliloti were evaluated for ACC deaminase activity. These isolates were classified in four groups based on growth rate on ACC containing medium and enzyme activity. One isolate from each group was selected for molecular characterization. The nucleotide sequence of 16S rRNA gene of the selected isolates were determined. The ACC deaminase genes (acdS) on total and chromosomal DNA of S. meliloti KYA40, and KYA71 strains were isolated and cloned in pTZ57R/T vector and the obtained recombinant plasmids were used for sequence analysis. Results: The sequence of acdS genes from strains KYA71 and KYA40 and corresponding proteins were analyzed with respect to available sequences in NCBI database. The 16S rRNA gene sequences of S. meliloti strains submitted to the GeneBank/NCBI database. The acdS gene of KYA71 may be located on chromosomal DNA and in KYA40 it is located on one of the mega plasmids. These two genes have 99% similarity with three nucleotide differences which only lead to a change in one amino acid 48, threonine in KYA40 acdS gene and methionine in KYA71. Conclusions: The comparison of amino acid sequences of KYA40 and KYA71 with other sequences in the database showed that the amino acids 37 to 58 in almost all strains were similar. Therefore, it was concluded that it was a conserved region in this location of acdS genes and any changes in this region may cause change in ACC deaminase activity.

New insights into 1-aminocyclopropane-1-carboxylate (ACC) deaminase phylogeny, evolution and ecological significance.

The main objective of this work is the study of the phylogeny, evolution and ecological importance of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, the activity of which represents one of the most important and studied mechanisms used by plant growth–promoting microorganisms. The ACC deaminase gene and its regulatory elements presence in completely sequenced organisms was verified by multiple searches in diverse databases, and based on the data obtained a comprehensive analysis was conducted. Strain habitat, origin and ACC deaminase activity were taken into account when analyzing the results. In order to unveil ACC deaminase origin, evolution and relationships with other closely related pyridoxal phosphate (PLP) dependent enzymes a phylogenetic analysis was also performed. The data obtained show that ACC deaminase is mostly prevalent in some Bacteria, Fungi and members of Stramenopiles. Contrary to previous reports, we show that ACC deaminase genes are predominantly vertically inherited in various bacterial and fungal classes. Still, results suggest a considerable degree of horizontal gene transfer events, including interkingdom transfer events. A model for ACC deaminase origin and evolution is also proposed. This study also confirms the previous reports suggesting that the Lrp-like regulatory protein AcdR is a common mechanism regulating ACC deaminase expression in Proteobacteria, however, we also show that other regulatory mechanisms may be present in some Proteobacteria and other bacterial phyla. In this study we provide a more complete view of the role for ACC deaminase than was previously available. The results show that ACC deaminase may not only be related to plant growth promotion abilities, but may also play multiple roles in microorganism's developmental processes. Hence, exploring the origin and functioning of this enzyme may be the key in a variety of important agricultural and biotechnological applications.

Recent developments in use of 1-aminocyclopropane-1-carboxylate (ACC) deaminase for conferring tolerance to biotic and abiotic stress

Ethylene is an essential plant hormone also known as a stress hormone because its synthesis is accelerated by induction of a variety of biotic and abiotic stress. The plant growth promoting bacteria containing the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase enhances plant growth by decreasing plant ethylene levels under stress conditions. The expression of ACC deaminase (acdS) gene in transgenic plants is an alternative approach to overcome the ethylene-induced stress. Several transgenic plants have been engineered to express both bacterial/plant acdS genes which then lowers the stress-induced ethylene levels, thus efficiently combating the deleterious effects of environmental stresses. This review summarizes the current knowledge of various transgenic plants overexpressing microbial and plant acdS genes and their potential under diverse biotic and abiotic stresses. Transcription regulation mechanism of acdS gene from different bacteria, with special emphasis to nitrogen fixing bacteria is also discussed in this review.

Isolation and characterization of drought-tolerant ACC deaminase and exopolysaccharide-producing fluorescent Pseudomonas sp.

The enzyme 1-aminocyclopropane-1-carboxylate deaminase catalyzes the degradation of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of the plant hormone ethylene, into α-ketobutyrate and ammonia. The enzyme has been detected in a limited number of bacteria and plays a significant role in sustaining plant growth and development under biotic and abiotic stress conditions by reducing stress-induced ethylene production in plants. We have screened 32 fluorescent Pseudomonas sp. isolated from rhizosphere and non-rhizosphere soils of different crop production systems for drought tolerance using polyethylene glycol 6000 (PEG 6000). Nine of these isolates were tolerant to a substrate metric potential of −0.30 MPa (15 % PEG 6000) and therefore considered to be drought-tolerant. All of these drought-tolerant isolates were screened for ACC deaminase activity using ACC as the sole nitrogen source, and one (SorgP4) was found to be positive for ACC, producing 3.71 ± 0.025 and 1.42 ± 0.039 μM/mg protein/h of α-ketobutyrate under the non-stress and drought stress condition, respectively. The isolate SorgP4 also showed other plant growth-promoting traits, such as indole acetic acid production, phosphate solubilization, siderophore and hydrogen cyanide production. The ACC deaminase gene (acdS) from the isolate SorgP4 was amplified, and the nucleotide sequence alignment of the acdS gene showed significant homology with acdS genes of NCBI Genbank. The 16S rRNA gene sequencing analysis identified the isolate as Pseudomonas fluorescens. Both sequences have been submitted to the NCBI GenBank under the accession numbers JX885767 and KC192771 respectively.

ACC deaminase genes are conserved amongMesorhizobiumspecies able to nodulate the same host plant

Rhizobia strains expressing the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase have been reported to display an augmented symbiotic performance as a consequence of lowering the plant ethylene levels that inhibit the nodulation process. Genes encoding ACC deaminase (acdS) have been studied in Rhizobium spp.; however, not much is known about the presence of acdS genes in Mesorhizobium spp. The aim of this study was to assess the prevalence and phylogeny of acdS genes in Mesorhizobium strains including a collection of chickpea-nodulating mesorhizobia from Portugal. ACC deaminase genes were detected in 10 of 12 mesorhizobia type strains as well as in 18 of 18 chickpea Mesorhizobium isolates studied in this work. No ACC deaminase activity was detected in any Mesorhizobium strain tested under free-living conditions. Despite the lack of ACC deaminase activity, it was possible to demonstrate that in Mesorhizobium ciceri UPM-Ca7(T) , the acdS gene is transcribed under symbiotic conditions. Phylogenetic analysis indicates that strains belonging to different species of Mesorhizobium, but nodulating the same host plant, have similar acdS genes, suggesting that acdS genes are horizontally acquired by transfer of the symbiosis island. This data, together with analysis of the symbiosis islands from completely sequenced Mesorhizobium genomes, suggest the presence of the acdS gene in a Mesorhizobium common ancestor that possessed this gene in a unique symbiosis island.

Disparate role of rhizobial ACC deaminase in root-nodule symbioses

The enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase converts ACC, a precursor of the plant hormone ethylene, into ammonia and α-ketobutyrate. ACC deaminase is widespread among the rhizobia in which it might play a crucial role in protecting rhizobia against inhibitory effects of ethylene synthesized by the host plant in response to the nodulation process. The beneficial action of this enzyme was demonstrated in several rhizobia such as Mesorhizobium loti and Rhizobium leguminosarum where knock-out mutants of the ACC deaminase gene showed nodulation defects. The genome of the slow-growing rhizobial species Bradyrhizobium japonicum also carries an annotated gene for a putative ACC deaminase (blr0241). Here, we tested the possible importance of this enzyme in B. japonicum by constructing an insertion mutant of blr0241 and studying its phenotype. First, the activity of ACC deaminase itself was measured. Unlike the B. japonicum wild type, the blr0241 mutant did not show any enzymatic activity. By contrast, the mutant was not impaired in its ability to nodulate soybean, cowpea, siratro, and mungbean. Likewise, symbiotic nitrogen fixation activity remained unaffected. Furthermore, a co-inoculation assay with the B. japonicum wild type and the blr0241 mutant for soybean and siratro nodulation revealed that the mutant was not affected in its competitiveness for nodulation and nodule occupation. The results show that the role previously ascribed to ACC deaminase in the rhizobia cannot be generalized, and species-specific differences may exist.

Plant Growth Promoting Characterization of Indigenous Azotobacteria Isolated from Soils in Iran

It has been well known that the bacteria of the genus Azotobacter, in addition to the beneficial N(2)-fixing activity, are able to improve plant growth by a number of direct and indirect mechanisms. To identify this potential in indigenous azotobacteria, the efficiency of 17 isolates of Azotobacter from the rhizosphere of wheat and barley plants cultivated in salt- and/or drought-affected soils in Iran were evaluated for their ability to dissolve inorganic and organic phosphates, siderophore secretion, indole acetic acid (IAA) production; and protease, chitinase, and ACC deaminase (ACCD) activities. First, they were biochemically characterized and one isolate (strain) was identified by 16S rDNA sequencing. Eight isolates were designated as Azotobacter vinelandii and the remaining isolates were identified as A. chroococcum. All isolates hydrolyzed the organic and inorganic phosphate compounds and effectively produced IAA. Fifteen isolates produced siderophore, but only one isolate showed protease activity which is being reported for the first time in relation to Azotobacter. None of the 17 isolates was capable of producing ACCD or chitinase. However, polymerase chain reaction amplification of the ACCD coding genes, by the use of the gene-specific primers, indicated that not all contain the ACCD gene. The standard screening methods with slight modifications, especially in the case of ACCD assay, were applied. The results showed that the use of specific screening methods, modified according to bacterial nutritional requirements, are the efficient methods for precise evaluation of the plant growth promoting rhizobacteria activity.

Enhanced chickpea growth-promotion ability of a Mesorhizobium strain expressing an exogenous ACC deaminase gene

Aims The main goal of the study reported herein was to assess the nodulation performance of a Mesorhizobium strain transformed with an exogenous ACC deaminase gene (acdS), and its subsequent ability to increase chickpea plant growth under normal and waterlogged conditions. Methods The Mesorhizobium ciceri strain LMS-1 was transformed with the acdS gene of Pseudomonas putida UW4 by triparental conjugation using plasmid pRKACC. A plant growth assay was conducted to verify the plant growth promotion ability of the LMS1 (pRKACC) transformed strain under normal and waterlogging conditions. Bacterial ACC deaminase and nitrogenase activity was measured. Results By expressing the exogenous acdS gene, the transformed strain LMS-1 showed a 127% increased ability to nodulate chickpea and a 125% promotion of the growth of chickpea compared to the wild-type strain, under normal conditions. Plants inoculated with the LMS-1 wild-type strain showed a higher nodule number under waterlogging stress than under control conditions, suggesting that waterlogging increases nodulation in chickpea. No significant relationship was found between ACC deaminase and nitrogenase activity. Conclusions The results obtained in this study show that the use of rhizobial strains with improved ACC deaminase activity might be very important for developing microbial inocula for agricultural purposes.

Metagenomic analysis of the 1-aminocyclopropane-1-carboxylate deaminase gene (acdS) operon of an uncultured bacterial endophyte colonizing Solanum tuberosum L

Deamination of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) is a key plant-beneficial trait found in many plant growth-promoting bacteria. In this study, we analysed ACC deaminase genes (acdS) of bacterial endophytes colonizing field-grown potato plants. PCR analysis revealed the presence of two types of acdS genes, the dominant one showing high homology to an acdS gene derived from Pseudomonas fluorescens. Construction, functional screening and sequence analysis of metagenomic libraries revealed clones containing the acdS gene identified in the PCR library. Sequence analysis of one metagenomic clone identified the entire acdS operon of an uncultivated endophyte and revealed that the acdS gene is coupled upstream with an acdR transcriptional regulator gene as previously found in P. putida strain UW4 (Grichko and Glick 2000). However, in-silico analysis of 195 fully sequenced, acdS-containing bacterial genomes revealed that the majority of strains, including numerous strains belonging to the genus Pseudomonas, do not contain an acdR regulatory gene in the vicinity of the acdS gene or elsewhere in the genome. The acdR (+)-acdS (+) operon was exclusively found in several Alpha- and Betaproteobacteria most prominently in the genus Burkholderia.

The Vicia sativa spp. nigra - Rhizobium leguminosarum bv. viciae symbiotic interaction is improved by Azospirillum brasilense

This study aimed to examine the effects of inoculation with wild type (Sp7) and mutant strains of Azospirillum brasilense on the Vicia sativa spp. nigra (vetch)-Rhizobium leguminosarum bv. viciae (Rlv) symbiosis. The A. brasilense mutants were ipdC- and napA-, impaired in indole pyruvate decarboxylase and periplasmic nitrate reductase, respectively; and acdS+, carrying the ACC deaminase gene. Inoculations were done in pots, pouches and hydroponics and we measured shoot and root weight parameters as well as effects on root morphology and nod gene induction ability by roots. In pots, wild type Sp7 and the acdS+ strain, but ipdC- and napA- mutants, lead to an increase in root hair density, 3–4 cm above the root tip. In pouches, combined inoculation with Rlv and strains Sp7, acdS+ or ipdC-, but napA-, increased shoot dry matter and nodulation relative to Rlv alone. In a hydroponic system, co-inoculation with strains Sp7 or acdS+, but with ipdC- and napA- mutants, enhanced root secretion of nod gene-inducing flavonoids in comparison with Rlv-inoculated plants. These results support that auxin production by A. brasilense has a positive effect on root secretion of nod gene-inducing flavonoids and auxin absorption activity by the plant.

Characterization of ACC deaminase from the biocontrol and plant growth-promoting agent Trichoderma asperellum T203

1-aminocyclopropane-1-carboxylate (ACC) deaminase activity was evaluated in the biocontrol and plant growth-promoting fungus Trichoderma asperellum T203. Fungal cultures grown with ACC as the sole nitrogen source showed high enzymatic activity. The enzyme encoding gene (Tas-acdS) was isolated, and an average 3.5-fold induction of the gene by 3 mM ACC was detected by real-time PCR. Escherichia coli bacteria carrying the intron-free cDNA of Tas-acdS cloned into the vector pAlter-EX1 under the control of the tac promoter revealed specific ACC deaminase (ACCD) activity and the ability to promote canola (Brassica napus) root elongation in pouch assays. RNAi silencing of the ACCD gene in T. asperellum showed decreased ability of the mutants to promote root elongation of canola seedlings. These data suggest a role for ACCD in the plant root growth-promotion effect by T. asperellum.

Cloning and Characterization of a Plasmid Encoded ACC Deaminase from an Indigenous Pseudomonas fluorescens FY32

In addition to the characterized mechanisms responsible for many direct effects of plant growth promoting bacteria (PGPB) on plants, it has been suggested that a number of PGPB contain the enzyme ACC deaminase that catalyzes degradation of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of ethylene, into alpha-ketobutyrate and ammonia. As part of an effort to obtain an ACC deaminase encoding gene from a collection of soil samples, only one bacterial isolate, Pseudomonas fluorescens FY32 was capable of growing on ACC as a sole source of nitrogen. The ACC deaminase gene was amplified from the above isolate by polymerase chain reaction (PCR) giving an expected DNA fragment, 1017 bp. Sequence analysis of the fragment showed that it was highly homologous (94% and 98% identities at nucleotide and amino acid levels, respectively) to the previously characterized acdS gene from Pseudomonas sp. 6G5. Furthermore, fusion of the ACC deaminase ORF with lacZ gene resulted in the expression of active enzyme in Escherichia coli. In addition, further analyses revealed that the acdS gene was plasmid-encoded so that a large plasmid (pFY32) with almost 50 kb in size was identified from this bacterium. Furthermore, transfer of pFY32 into E. coli DH5alpha proved its ACC deaminase activity. This result was in accordance with previous reports suggesting horizontal transfer of the acdS gene. However, it needs more investigation to identify whether this pFY32 plasmid has undergone lateral gene transfer during the evolutionary process.

Detection and Characterization of ACC Deaminase in Plant Growth Promoting Rhizobacteria

The enzyme 1-aminocyclopropane-1-carboxylate deaminase converts ACC, the precursor of the plant hormone ethylene to α-ketobutyrate and ammonium. The enzyme has been identified in few soil bacteria, and is proposed to play a key role in plant growth promotion. In this study, the isolates of plant growth promoting rhizobacteria were screened for ACC deaminase activity based on their ability to grow on ACC as a sole nitrogen source. The selected isolates showed the presence of other plant growth promoting characteristics such as IAA production, phosphate solubilization and siderophore production. The role of ACC deaminase in lowering ethylene production under cadmium stress condition was also studied by measuring in vitro ethylene evolution by wheat seedlings treated with ACC deaminase positive isolates. Nucleic acid hybridization confirmed the presence of ACC deaminase gene (acdS) in the bacterial isolates.

Isolation and Characterization of ACC Deaminase Gene from Two Plant Growth-Promoting Rhizobacteria

Lowering of plant ethylene by deamination of its immediate precursor 1-aminocyclopropane-1-carboxylate (ACC) is a key trait found in many rhizobacteria. We isolated and screened bacteria from the rhizosphere of wheat for their ACC-degrading ability. The ACC deaminase gene (acdS) isolated from two bacterial isolates through PCR amplification was cloned and sequenced. Nucleotide sequence alignment of these genes with previously reported genes of Pseudomonas sp. strain ACP and Enterobacter cloacae strain UW4 showed variation in their sequences. In the phylogenetic analysis, distinctness of these two genes was observed as a separate cluster. 16S rDNA sequencing of two isolates identified them to be Achromobacter sp. and Pseudomonas stutzeri.

Screening of plant growth promoting traits ofBacillus thuringiensis

This study aimed to evaluate the plant growth promoting (PGP) potential ofBacillus thuringiensis. In this context, several genetic determinants of factors implicated in PGP potential were investigated by polymerase chain reaction (PCR) in 16B. thuringiensis strains of different origin and belonging to different subspecies. PCR screening was performed on acid phosphatase, phytase, siderophore biosynthesis protein, 1-aminocyclopropane-1-carboxylate (ACC) deaminase and indolpyruvate decarboxylase (ipdC). Production of indol acetic acid (IAA)-like compounds and of ACC deaminase, and capability of solubilising mineral phosphate were investigated by phenotypic tests. All the strains were PCR positive for the presence of the siderophore biosynthesis protein, ACC deaminase and acid phosphatase genes. Five and seven strains gave an amplicon with the expected length for the phytase andipdC genes respectively. All the strains produced IAA compounds and seven had a high capacity to solubilise inorganic phosphorous. Qualitative phenotypic test for ACC deaminase activity showed that seven strains are able to grow on salt minimal medium containing ACC as sole nitrogen source, indicating the expression of theaccd genes. Our screening results in thirteen strains having more than one PGP trait and showed thatB. thuringiensis harbours and expresses several PGP determinants that could be very interesting in field application to enhance the plant growth. To our knowledge, this is the first report on the multiple plant growth promoting potential ofB. thuringiensis.

Transcriptional regulation of ACC deaminase gene expression inPseudomonas putidaUW4

One of the major mechanisms that plant growth-promoting bacteria use to facilitate plant growth is through the lowering of plant ethylene levels by the bacterial enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase. Many of the bacterial ACC deaminase genes (acdS) that have been examined to date are under the transcriptional control of a leucine-responsive regulatory protein, Lrp, encoded by acdR and referred to here as AcdR. The work presented here is focused on how AcdR and the newly discovered AcdB protein from Pseudomonas putida UW4 are involved in the regulation of acdS expression. First, the results of gel retardation experiments showed that AcdR binds to the acdS regulatory region, and this binding activity in vitro is not affected by the addition of 2 mmol x L-1 ACC but can be eliminated by addition of 20 microg x mL-1 leucine. Second, a potential regulatory protein, AcdB, involved in the regulation of acdS expression, was identified through both yeast 2-hybrid screen and coimmunoprecipitation based on its ability to bind to AcdR; subsequently, its binding to the acdS regulatory region in the presence of ACC was shown by gel retardation experiments. The data are interpreted in terms of a model in which AcdR and AcdB co-regulate the expression of the acdS gene.

ACC deaminase from plant growth-promoting bacteria affects crown gall development

In addition to the well-known roles of indoleacetic acid and cytokinin in crown gall formation, the plant hormone ethylene also plays an important role in this process. Many plant growth-promoting bacteria (PGPB) encode the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, which can degrade ACC, the immediate precursor of ethylene in plants, to alpha-ketobutyrate and ammonia and thereby lower plant ethylene levels. To study the effect of ACC deaminase on crown gall development, an ACC deaminase gene from the PGPB Pseudomonas putida UW4 was introduced into Agrobacterium tumefaciens C58, so that the effect of ACC deaminase activity on tumour formation in tomato and castor bean plants could be assessed. Plants were also coinoculated with A. tumefaciens C58 and P. putida UW4 or P. putida UW4-acdS- (an ACC deaminase minus mutant strain). In both types of experiments, it was observed that the presence of ACC deaminase generally inhibited tumour development on both tomato and castor bean plants.

Light quality regulation of endogenous levels of auxin, abscisic acid and ethylene production in petioles and leaves of wild type and ACC deaminase transgenic Brassica napus seedlings

Brassica napus L. seedlings responded to low red to far-red (R/FR) ratio by elongating petioles and decreasing leaf expansion. These typical shade avoidance traits were correlated with significantly decreased endogenous indole-3-acetic acid (IAA) levels and significantly increased endogenous abscisic acid (ABA) levels and ethylene production. The transgenic (T) B. napus line bearing the bacterial ACC deaminase gene, did not respond to low R/FR ratio with altered petiole and leaf growth and less ethylene (especially by petioles) was produced. As with WT seedlings, T seedlings had significantly lower IAA levels in both petioles and leaves under low R/FR ratio. However, ABA levels of low R/FR ratio-grown T seedlings either increased (petioles) or were unaltered (leaves). Our results further suggest that low R/FR ratio regulates endogenous IAA levels independently of ethylene, but there may be an interaction between ABA and ethylene in leaf development.

Do low-ethylene-producing transgenic canola (Brassica napus) plants expressing the ACC deaminase gene differ from wild-type plants in response to UVB radiation?

Few studies have considered ethylene involvement in plant responses to ultraviolet-B (UVB) radiation. We studied the responses to UVB radiation of one wild-type (WT, ‘Westar’) canola (Brassica napus L.) with normal ethylene evolution and two transgenic (C1, C2) lines with lower ethylene evolution. Canola plants were grown under biologically effective levels of UVB (UVBBE) radiation: 0.03 (low), 4.88 (medium), and 9.78 (high) kJ·m–2·d–1 in controlled-environment growth chambers. The growth and physiological parameters of the plants were measured. Of the two transgenic lines, C1 demonstrated higher ethylene evolution than C2 but lower than WT. The lowest aboveground and belowground biomass was found with exposure to high UVB radiation. WT produced more biomass than C2. Net CO2 assimilation and transpiration did not vary among plant lines or UVB treatments. Water-use efficiency was lower under high UVB radiation than under low UVB. The quantum yield of photosystem II was higher for C2 than for either WT or C1. WT did not differ from transgenic plants in respect to photosynthetic pigments and UV-screening compounds. Photosynthetic pigment concentration decreased, but concentration of UV-screening compounds, thickness of epicuticular wax, and the rate of root hydraulic conductance were increased by exposure to UVB radiation. While there appears to be a lack of ethylene involvement in some of the measured physiological parameters, the transgenic plants exhibited differential sensitivity to UVB in a few key measured parameters.

ACC deaminase from Pseudomonas fluorescens mediated saline resistance in groundnut (Arachis hypogea) plants

To study the effect of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase from Pseudomonas fluorescens against saline stress under in vitro and field conditions in groundnut (Arachis hypogea) plants. Four plant growth-promoting rhizobacteria (PGPR) strains were used in this study to evaluate their efficacy in groundnut plants against saline stress under in vitro. Among the four PGPR strains used, Ps. fluorescens strain TDK1 showed greater performance in improving the plant growth parameters of groundnut seedlings in vitro. PCR amplification using Pseudomonas-specific 16S-23S rRNA internal transcribed spacers (ITS) primers revealed that all the four strains belonged to the group of fluorescent pseudomonads. ITS region of Ps. fluorescens strain TDK1 was cloned and sequenced. ACC deaminase activity using biochemical and molecular (PCR) analysis revealed that among all the four strains, Ps. fluorescens strain TDK1 showed greater amount of ACC deaminase activity and positive reaction to PCR amplification. ACC deaminase gene from Ps. fluorescens strain TDK1 was isolated, cloned and sequenced. Pseudomonas bioformulations were developed and they were tested in groundnut plants under saline-affected soils. The results indicated the superior performance by Ps. fluorescens strain TDK1 possessing ACC deaminase activity in improving yield parameters in groundnut plants despite salinity. Pseudomonas fluorescens strain TDK1 possessing ACC deaminase activity enhanced the saline resistance in groundnut plants, which in turn resulted in increased yield when compared with the groundnuts treated with Pseudomonas strains not having ACC deaminase activity. The promising role of ACC deaminase from Ps. fluorescens strain TDK1 in alleviating saline stress has been concluded in groundnut plants. This study will be useful for exploiting the activity of ACC deaminase from microbial strains against various biotic and abiotic stresses wherever ACC accumulated as precursor for ethylene biosynthesis.