Introduction

Abiotic stressors that affect plants include nutritional deficiency, UV radiation, salt, high and
low temperatures, and drought (Mittler et al., 2006). The main stressor among these is
salinity, which restricts the geographic range and crop productivity (Zhu et al., 2016). While
salt stress generates both hyperosmotic and ionic effects on plants, which is the predominant
indicator of drought (Zhu et al., 2002). In addition to metabolic inefficiency in plants, salt
may cause oxidative damage and membrane lipid peroxidation (Munns et al., 2008; Datta et
al., 2012; Amoah et al., 2019). Soil salinity is the most detrimental environmental factor
affecting crop productivity globally among abiotic stressors (Chung et al., 2019). Around the
world, it damages millions of hectares of land, and each year it results in considerable
economic losses (Yoon and Humayun et al., 2009). A high salt content already affects 62
million hectares (20 percent) of watered land, and by 2050, it is predicted that over half of the
arable land would be salinized (Jamil et al., 2011, Humayun et al., 2010). It is well
recognized that soil salinity inhibits plant growth at first by osmotic stress, then ion toxicity
(Rahnama et al., 2010; James et al., 2011). Reactive oxygen species (ROS) produced by salt
stress could cause oxidative damage and significantly impede plant growth and development
(Khattab, 2007; Singh & Jha, 2016). The principal morphological lesions that have been
noticed under salinity stress include scorching of leaves and stems, abscission and senescence
of leaves, as well as suppression of root and shoot growth (Mayak et al., 2004; Singh & Jha,
2016). In addition, biochemical alterations can affect how cells oxidize, lead to nutritional
imbalances, alter normal metabolism, and cause chlorophyll to degrade (Yue et al., 2007;
Khan et al., 2019a). By limiting root growth, nutrient uptake, and metabolic activities, soil
salinity reduces crop output (Chung et al., 2019). As a result of poor root growth and
development, the active rhizosphere zone is diminished, affecting nutrient uptake
effectiveness. Salinity stress also has an impact on physiological, morphological, and
biochemical processes, resulting in a reduction in crop biomass and production (Yoon and
Humayun et al., 2009). By competing with K ions, more Na and Cl ions cause ionic
imbalances and ion toxicity. Chlorosis and necrosis are caused by ion toxicity, which disrupts
a variety of physiological processes in plants (Almeida et al., 2017). By competing with K for
binding sites and interfering with K homeostasis, Na suppresses crucial cellular processes and
enzymatic activities that depend on K for proper operation. It has been proposed that
maintaining low Na concentrations in the cytoplasm and cytosol and maintaining high
cytosolic K concentrations are necessary for plant life under salinity stress. All growth
phases, such as germination, seedling, vegetative, and mature stages, have shown
morphological changes in response to saline stress (Tavakkoli et al., 2010). Salt stress alters a
number of biochemical processes, including the modulation of phytohormones (increases in
the defence hormone salicylic acid [SA] and decreases in the stress hormone abscisic acid
[ABA]), changes in ion uptake (absorption or removal of ions), activation of antioxidant
enzymes, generation and accumulation of reactive oxygen species (ROS), and disruption of
the photosynthetic pathway. Salinity stress in plants also influences gene regulation at the
molecular level. Additionally, the effects of salt stress on chlorophyll, carotenoids, and
reduced PSII activity hinder photosynthesis (Demetriou et al., 2007, Lu et al., 2002, Misra et
al., 2001). A decrease in photosynthesis will eventually exhaust energy supplies, causing
plants to starve, expand their leaves, and age. Food security is now a significant concern due
to the steadily expanding global population. The 2020 Global Agricultural Productivity
(GAP) Report estimates that to satisfy the needs of 10 billion people for food in 2050, the
GAP must rise by 1.73 percent per year (GAP Report 2020). In contrast, soil quality has
significantly decreased as a result of climate change and human activity, which has reduced
agricultural productivity. Soil salinization is one of the utmost detrimental elements that
contribute to the deterioration of arable land among the different risks to soil functions
(Ilangumaran and Smith 2017; Otlewska et al., 2020). According to estimates, salinity now
affects more than 20% of all agricultural land globally, and by 2050, that percentage will rise
to 50%. (Otlewska et al., 2020). Low water potential caused by excessive salt build-up in the
soil hinders plant growth and development by generating ion toxicity, nutritional shortage,
and inhibition of photosynthesis. Additionally, the presence of salt stress can increase the
creation of reactive oxygen species (ROS), which are highly reactive with cellular
components and result in programmed cell death (Kushwaha et al., 2020; Otlewska et al.,
2020). Salt stress may affect almost all farmed crops. There have been significant attempts
undertaken to increase food yield in salty soils, including the development of salt-resistant
crops by traditional breeding and genetic engineering techniques. Although these tactics have
had some success, the entire procedure is time-consuming, technically challenging, and
expensive (Shokat and Großkinsky 2019). In addition, there may be certain regulatory
concerns associated with the creation and cultivation of genetically modified crops (Kumar et
al. 2020). An alternate technique is to use beneficial bacteria to improve crop tolerance to
salt, which is both cost-effective and environmentally favorable. Tomato is a commonly
grown vegetable crop that is consumed in many ways. After potatoes, lettuce, and onions, it
is the fourth most popular fresh-market vegetable. It is nutrient-rich and contains 37 minerals,
a lot of vitamins, water, proteins, fiber, carbs, and calories (Chew et al. 2014). It functions as
an antioxidant, lowering blood pressure, decreasing kidney stone risk, limiting cancer and eye
disease risks, and supporting healthy weight loss and skin (Cogswell et al. 2012). Recently,
scientists have developed an interest in employing endophytic bacteria that stimulate plant
development to mitigate the effects of salt stress (PGPEB). Endophytic colonization, in which
PGPEB dwell inside plant tissues and foster the growth and development of their host plant
without transmitting any illnesses, is an indication of a healthy plant system (Taghavi et al.,
2010). These naturally occurring bacterial endophytes not only aid in the development of
plants but also initiate systemic resistance to numerous stressors. Furthermore, bacterial
species are being employed to counteract the negative effects of salt stress and boost plant
development by creating a variety of bioactive secondary metabolites and phytohormones
(Mayak et al., 2004; Nadeem et al., 2010; Khan et al., 2019). Endophytic microorganisms can
mitigate the negative impacts of stress on plant development through morphological,
biochemical, and physiological adaptation. The bacteria can also influence the growth of the
host plant by modifying the availability of various soil nutrients through the following
mechanisms: (1) reducing root width; (2) lengthening root hairs; and (3) producing various
biochemicals, including phenolic compounds. Due to the sequestration of aluminum on the
root surfaces, these substances can also have an impact on the development of roots in acidic
environments (Malinowski and Belesky 2000; Barac et al. 2004; Bauer and Mathesius 2004).
A sustainable agricultural system preserves and enhances human health, rewards farmers and
consumers economically and spiritually, safeguards the environment and generates enough
food to feed a growing global population. The prevalent abiotic stress levels in the
environment are one of the most significant barriers to agricultural output worldwide.
Microbes that are found in plants can significantly contribute to the development of abiotic
stress tolerance. These organisms might include rhizoplane, rhizosphere, and endophytic
bacteria, as well as symbiotic fungi, and they work in several ways, such as initiating osmotic
responses, giving growth hormones and nutrients, functioning as biocontrol agents, and
inducing new genes in plants. Genetic engineering and plant breeding are critical to the
development of stress-tolerant crop varieties, but microbial inoculation to alleviate stresses in
plants could be a more cost-effective, environmentally friendly option. Taking the present
leads available, focused further study in this area is required, notably on field assessment and
deployment of prospective species as biofertilizers in stressed soil. Soil salinization is a
worldwide scourge on agricultural productivity. Crops planted on salty soils suffer from
severe osmotic stress, nutritional diseases and toxicities, poor soil physical conditions, and
decreased crop output. The current study focuses on increasing plant yield under stressful
conditions and increasing plant resilience to salt stress through the use of endophytic bacteria.

Material Method
Isolation of endophytes
A total of 52 bacterial endophytes were recovered from four distinct Halophytic plants
cultivated in farmers' fields in four separate areas of the Indo-Gangetic plains. The hypocotyl
and root portions were separated and cleaned under running water. Furthermore, the surface
sterilisation was carried out by the technique given by(Zinniel et al.2002).Using a sterile
pestle and mortar, surface sterilized plant tissue was pulverized in 12.5 mM potassium
phosphate buffer. The dilutions 10-2 and 10-3 were prepared and inoculated on a nutrient agar
medium. In addition, anatomically different colonies were collected and purified for future
use.
In vitro bio-efficacy
Three soil-borne tomato pathogens, Rhizoctonia solani (NAIMCC-F-02899), Sclerotium
rolfsii (NAIMCC-F-03053), and Villosciclava virens (NAIMCC), were investigated for
antagonistic effects of the bacterial endophytes. An entire one-week-old mother plate that was
fully colonised was taken out, and a 7-mm circular disc of the fungus was put upside down in
the centre of the PDA plate. Endophyte cultures that had been actively developed for 24
hours were streaked equidistantly on both sides of the pathogen disc. S. rolfsii was incubated
for 4 days, V. virens for 5 days and R. solani for 5 days. As a control, fungus-inoculated
plates lacking bacterial endophyte were used. This experiment included three replications and
was performed twice to ensure uniformity in the outcomes. The decrease in mycelium
growing radially was monitored, and the percentage inhibition was calculated.
Taxonomic diversity of bacterial endophytes

Following the manufacturer's instructions and using the AxyPrep Bacterial Genomic DNA
Miniprep Kit, DNA was extracted from pure bacterial colonies (Axygen Biosciences). PCR
was performed on isolated DNA samples using the conditions described in and bacteria-
specific primers (27F and 1492R) (Thomas et al., 2008).The Sanger technique was used to
perform 16S rRNA partial sequencing on all 52 isolates. Sequencing data were identified and
submitted to GeneBank using EzTaxon and NCBI databases (NCBI). The primer pairs listed
in supplementary data 2 were used to test bacterial endophytes for the presence of the
antimicrobial-producing genes NRPS, pks, bmyB, phz, ituD, ituC, srfA, and chiA. These
verified genera-specific primers were chosen since they were found in the Bacillus and other
G-ve genera.
Functional diversity: PGP traits
Indole acetic acid
Luria Bertani (LB) broth with and without 5g/mL tryptophan was used to study the synthesis
of indole acetic acid (IAA). Cultures were centrifuged for 10 minutes at 5000 RPM after 5
days of incubation. Two drops of orthophosphoric acid and 5 mL of Salkowski's reagent were
added to 10 ml of supernatant. When the pink color developed to a high degree, IAA
generation was suggested (Patten and Glick 1996), and absorbance was measured at 530 nm.
Ammonia production test
The production of ammonia by endophyte isolates was assessed as previously described
(Cappuccino and Sherman 1992). Peptone water was supplemented with endophyte cultures,
and the mixture was maintained at 30 °C for 48–72 hours for growth. Once the sample had
been incubated, 0.5 mL of Nessler's reagent was added, and the presence of a yellow-brown
tint was deemed positive.
Hydrogen cyanide production
The generation of hydrogen cyanide (HCN) was evaluated in accordance with Ahmad et al
(2008). In nutrient agar plates that had 4.4 g L-1 glycine added, cultures were streaked. The
upper half of a Petri plate was covered with Whatman No. 1 filter paper that had been soaked
in a solution of 2 percent sodium carbonate and 0.5 percent picric acid. The plates were
covered and incubated for 6 days at 30 °C. The HCN production for 6 days was demonstrated
by the color shift from yellow to orange-brown. HCN production was identified by the color
shift from yellow to orange-brown.
Phosphate solubilization
Using Pikovskaya's agar medium, the endophytic isolates' phosphorus solubilization was
evaluated (Pikovskaya 1948). Using Pikovskaya's agar medium, the endophytic isolates'
phosphorus solubilization was evaluated (Pikovskaya 1948). The existence of a clean zone
surrounding the colonies is indicative of successful phosphate solubilization.
Zinc solubilization
First, sterile zinc oxide plates were made, and the center of each plate was spot-inoculated
with one loopful of pure active bacterial culture. After securely sealing the plates with
parafilm, they were placed in an incubator and maintained there for 48 hours for bacterial
cultures. Following incubation, the diameter of bacterial growth and the zone of clearance
were measured and the following formula was used to calculate the results:
Solubilization efficiency=solubilization diameter/diameter of colony growth × 100

Siderophore production test

The modified CAS agar technique was used to test siderophore production (Chrome Azurol
S; Schwyn and Neilands 1987).After growing for 24 hours, endophyte cultures were spotted
onto CAS agar plates and incubated for three days at 30 °C. The siderophore production was
evident by the development of orange hallo around the colonies.
Exopolysaccharide production
By using the serial dilution approach, the bacteria from the sample were extracted. A stock
solution was created by taking a 1ml sample of the water and aseptically suspending it in
10ml of sterile physiological saline water. 1ml of the stock solution was added to 9ml of
sterile saline solution, and the mixture was carefully vortexed. Following serial dilution to a
concentration of 10-6 or 10-7, the sample was distributed by inoculating 0.1 ml of each
diluent suspension onto Nutrient Agar plates, which were suspended in 2% sucrose and
incubated for 48 hours to count the number of bacteria. The isolates that produced dense
mucoidal colonies were then chosen and purified using the streak plate technique on a
recently prepared agar medium to produce unique colonies.
Assessment of salt tolerance of Endophytes
The capacity of the chosen Endophytes to tolerate NaCl was further assessed. The isolates
were grown/cultured in NB media supplemented with various sodium chloride (NaCl) levels
(0, 400, 800, 1200, 1600, and 2000 mM) for this, then incubated at 28 ± 2 ◦C in a shaking
incubator (at 150 rpm) for two to three days. Following incubation, metabolically active cells
were examined using the viable count technique, and the strain displaying a greater amount
of salt was referred to as a salt-tolerant (halotolerant) PGPR strain.
Production of VOCs against R. solani
100μL of overnight grown broth culture was inoculated into beef extract peptone-containing
Petri dishes. These plates were incubated for 24 hours at 30°C. An infected petri dish
containing R. solani fermentation broth (10µL) was placed on top of a Petri dish containing a
PDA medium infected with R. solani. A control Petri dish without inoculation bacteria was
used at the same time. Two bottom dishes were sealed and incubated in the dark at 30°C. A
control R. solani colony was measured before reaching the edge of the Petri dish.
The inhibition rate was calculated:
Inhibition rate = [(colony diameter of control R. solani − colony diameter of treated R.
solani)/colony diameter of control R. solani] × 100%.

Plant Assay

Tomato seeds of the Pusa Ruby variety were surface disinfected by being washed in sterile
water, then dipped for 30 seconds in 70 percent ethanol, and then washed with sterile distilled
water. The seeds were then immersed in 2 percent sodium hypochlorite (NaOCl) thrice for 1
minute with sterile distilled water, and then submerged in 2 percent sodium thiosulfate for 10
minutes. Seeds that had been surface-sterilized were given the appropriate bacterial
endophyte inoculum treatment (24h old, 0.2 OD suspension). Each of the five treatments had
ten replications. The treatment detail were as follows: 4H1, 2H2, 2H18, 2H20, and control. In
98-well seedling trays with sterilized sand: soil combination, the seeds were planted (1:3).
Through the use of sterile Hoagland solution, nutrients were given (1X). Before collecting the
roots for the colonization research, the tomato plants were allowed to develop for a duration
of three weeks.

Visualization of Endophytic Colonization

The roots were carefully cleaned three times in sterilized distilled water before being stained
with LIVE/DEADTM BacLightTM bacterial viability stain (Invitrogen, USA) that contains
SYTO9 and propidium iodide (Stiefel et al., 2015). For 30 minutes in the dark, the roots were
dyed with a 35-M concentration of SYTO9 and a 400-M concentration of propidium iodide.
Phosphate buffered saline was used to wash away the extra fluorescent dyes. The stained
roots were mounted on glass slides and viewed using the 488 nm, 543 nm, and TD channels
of a confocal scanning laser microscope (CSLM; Nikon, Japan). The NIS element 3.2.3
software was used to process the photos (Nikon, Japan).

In vivo Assessment of Salinity Tolerance in Tomato

Soil Preparation and Nursery

Five kilograms of autoclaved soil were put into the pots. Row and column randomization
eliminated the variation within the pots. The nutrients were combined using urea,
diammonium phosphate, and muriate of potash at a ratio of 100:50:50 kg NPK per acre.
Three soaking and drying cycles were used to naturally compress the soil in these pots. The
pots were watered to 80% of the field's capacity, and 150 mM NaCl was used to maintain
salinity. In a planting tray containing soil, sand, and cocopit mixture (1:1:1), tomato seeds of
the variety Pusa Ruby were planted and maintained wet by misting them with water. The
roots were gently rinsed and treated with the appropriate endophytic inoculant by root
dipping after the plants had been removed after 30 days. The roots were soaked in 0.5 percent
carboxy methyl cellulose for 30 minutes before being transplanted into the pots with the
inoculum (5 ml/L; 24-hour-old 0.2 OD culture of respective endophytes). The treatment
information was the same as that utilized in the three-replication seedling tray experiment.
The results of this experiment were double-checked for consistency using the same
experimental design. Data was collected 45 days after transplantation (DAT), However, the
plant's root architecture, length, and dry weight were measured at 75 DAT.

Modulation in Gene Expression

RNA Extraction and cDNA Preparation

The plant sample (100 mg of a root, shoot, and leaf) was combined with 1 ml TRIzol reagent
and 10µl mercaptoethanol and incubated for 5 minutes at room temperature before grinding
in liquid nitrogen and processing according to the manufacturer's instructions for utilizing
PureLinkTM RNA mini-Kit (Invitrogen, United States). The total RNA extracted was
converted to cDNA using the High Capacity RNA-to-cDNA kit (Thermo Fisher Scientific,
USA) according to the manufacturer's instructions. The cDNA was then employed in real-
time quantitative PCR (RT-qPCR) experiments.

RT-qPCR Assay

The RT-qPCR-based expression studies of salt stress-responsive genes were performed using
the RT-qPCR Detection System (Bio-Rad, United States) and the Eva Green SYBR Green
Supermix Kit (Bio-Rad, United States), with three technical replicates of each sample. The
final concentration of gene-specific primers was kept constant at 10 pmol/µl, and internal
controls were used to calculate and normalize mRNA transcript levels. The reaction mixture
was kept to a final volume of 10 µl [diluted cDNA samples (50 ng/ml), 2 µl; forward and
reverse primers (10 µM), 1.5 µl each; and real-time master mix, 5 µl]. The RT-qPCR
procedure included denaturation at 95 °C for 2 min, 40 repetitions at 95 °C for 30 s, 60 °C for
30 s, and 72 °C for 30 s. The two endogenous genes actin and glyceraldehyde 3-phosphate
dehydrogenase's mean CT values were utilized to normalize the data using the 211CT
technique. The mean CT values of the three technical replicates from each biological
replication with control were used to compare transcript fold accumulation (Livak and
Schmittgen, 2001).

Total chlorophyll and carotenoid content

The total chlorophyll and carotenoid concentrations were calculated by homogenizing 0.5g of
fresh leaves in 80 percent acetone. Using a spectrophotometer, the extract's absorbance was
determined at wavelengths 645, 663, and 470 nm. The concentration of total chlorophyll and
carotenoid content was determined using the previously reported technique (Lichtenthaler
and Buschmann, 2001).

Total polyphenolic content (TPC)

For TPC analysis, methanolic extracts (1ml) from tomato leaf samples were combined with
water: methanol (1:1, 1ml, v/v), distilled water (3ml), and newly produced Folin-Ciocalteau
reagent (0.5ml) followed by vigorous mixing (Zheng and Shetty, 2000). 5 minutes of
incubation were followed by the addition of 1 ml of 5 percent sodium carbonate. After 30
minutes, the absorbance was measured at 725 nm. TPC was determined as gallic acid
equivalents (GAE) g-1 FW.

Total flavonoid content

According to the technique outlined by Irshad et al., 2012 the total flavonoid concentration in
the leaf extract was evaluated. Leaf extract (0.5 ml), distilled water (4 ml), and NaNO2 made
up the reaction mixture (50 percent , 0.3ml). AlCl3 (10%, 0.3ml) was well incorporated into
the mixture after 5 minutes of incubation, and NaOH (1M, 2ml) was added after another 5
minutes. With ethanol, the volume was adjusted to 10ml (95 percent ). TFC was reported as
mg quercetin equivalents g-1 FW and the absorbance was measured at 510 nm using
quercetin as the reference component.

Extraction for enzyme assays

Fresh leaves (0.5g) were crushed in a mortar and pestle that was stored in the refrigerator
using liquid nitrogen. In 12 ml of ice-cold phosphate buffer (100 mM; pH 7.0; 0.5 mM
EDTA) containing 1.4 mmol l-1 -mercaptoethanol, fine powder was homogenised. The
homogenate was centrifuged at 16,000 g for 15 min at room temperature to separate the
supernatant, which was then utilized to conduct enzyme tests.

Phenylalanine ammonia-lyase (PAL) assay

The reaction mixture was made by adding 0.1 mol of L-1 phenylalanine (pH 8.7), 0.2 mol of
L-1 phosphate buffer (pH 8.7), 0.2 mol of enzyme extract (0.2 ml), and distilled water
(1.3ml). The reaction was stopped by adding 0.5ml of trichloroacetic acid (1 mol l-1) after 30
minutes of incubation at 32°C. At 290 nm, the absorbance was measured, and the activity was
measured in mol of t-cinnamic acid (TCA) per g of formic acid (FW) (Brueske, 1980).

Superoxide dismutase (SOD)

The capacity of enzyme extract to stop the photochemical reduction of nitroblue tetrazolium
(NBT) chloride was used to evaluate the enzyme activity of SOD (EC 1.15.1.1). (Fridovich,
1974). The reaction mixture contains 200µl of enzyme extract combined with 100 mM
phosphate buffer (pH 7.8), 3 mM EDTA, 1.5 m sodium carbonate, 2.25 m NBT, and 3 mM
EDTA in a final volume of 3 ml. After adding 400µl of riboflavin (2 mol l-1), the process
was initiated by putting the tubes under two 15 W fluorescent lights for 15 min. After putting
the tube in complete darkness to stop the process, the absorbance at 560 nm was measured.
One unit of the enzyme extract lowered the absorbance by 50% in contrast to the control
group which did not contain any enzyme extract.

Peroxidase (PO) assay

PO (EC 1.11.1.7) activity was measured in a reaction mixture containing 500µl H2O2 (1
percent v v), 1.5ml pyrogallol (0.05mol l-1), and 50µl of an enzyme (Hammerschmidt et al.
1982). At 420 nm, the change in absorbance was noted at regular intervals of 30 s for three
minutes. U min-1g-1 FW was used to express the enzyme activity.

Ascorbate peroxidase (APX)

In the test combination of ascorbic acid (0.25mM), H2O2 (1.0mM), EDTA (0.1mM), and
phosphate buffer (25mM, pH 7.0), APX activity was calculated (Nakano and Asada, 1981).
Enzyme extract (0.2 ml) was added to the mixture to start the reaction, and after 60 seconds, a
decrease in absorbance at 290 nm was observed. It was discovered that the enzyme activity
was U min-1g-1FW.

Catalase (CAT)
Phosphate buffer (300 M, pH 7.2), enzyme extract (1 ml), and CAT (EC 1.11.1.6) activity
were measured in the reaction mixture (Aebi, 1984). When H2O2 (100 mM) was introduced
to start the process, it broke down enzymatically after being in the dark for one minute,
releasing O2. The O2 was determined as the reduction of H2O2 at 240nm (the extinction
coefficient of H2O2 is 0.036mM-1cm-1) and enzyme activity was defined as M H2O2
oxidized min-1g-1 FW.

Glutathione reductase (GR)

In the reaction mixture made by adding Tris-HCl buffer (50 mM, pH 7.6), oxidised
glutathione (1 mM GSSG, 100 l), MgCl2 (3 mM), NADPH (0.15 mM, 10 ml), and enzyme
extract (0.3 ml), GR (EC 1.6.4.2) was measured (Anderson, 1996). NADPH absorbance at
340 nm was gradually decreased to determine the GR activity, which was expressed in terms
of U (nmol oxidized NADPH) min-1 mg-1 FW.

Guaiacol peroxidase (GPX)

By observing the change in the reaction mixture containing sodium phosphate (10mM, pH
6.0), tetraguaiacol (1 percent, v/v), H2O2 (0.3 percent, v/v), and enzyme extract (0.3ml) at
470nm, GPX (EC 1.11.1.7) was determined (Zheng and Van Huystee, 1992). In terms of U
min1 mg-1 FW, the enzyme activity was expressed. The amount of enzyme required to
catalyze the oxidation of 1 mol of guaiacol per minute is represented by one unit of an
enzyme.

Non-enzymatic antioxidant assay

The leaf sample (1g) was crushed in a mortar and pestle with 10ml of 80% methanol before
being left to ferment at room temperature for a whole night. The supernatant from the
centrifugation of the extract at 4 °C for 15 min at 16000 g was utilized in non-enzymatic
antioxidant experiments (Wojdyo et al. 2007).

ABTS activity

The ABTS•+ decolorization technique was used to assess the ABTS activity in leaf extract
(Re et al. 1999). The stock solution was made by combining equal volumes of potassium
persulfate (2.45mM) and ABTS (7mM) and storing the mixture in the dark for 18 hours. The
stock solution was then further diluted using potassium phosphate buffer (0.1M, pH 7.4) to
achieve an absorbance of 0.70 (0.02) at 734 nm. 2.95mL of the ABTS radical working
solution was added to the sample (50µl) and properly blended. The absorbance was measured
at 734 nm, and the activity's percent inhibition was reported.

Reducing power (RP)

With quercetin as the reference component, the Reducing power of leaf extracts was
calculated using an adjusted ferric reducing antioxidant power test (Apati et al., 2003).
500ml of plant extract, 1ml of MeOH, 2.5 ml of phosphate buffer (pH 6.6), and 1 % w/v of
potassium ferrocyanide made up the reaction mixture. To stop the reaction, TCA (10% w/v,
2.5ml) was added after the mixture had been incubated at 50°C for 20 minutes in a water
bath. FeCl3 (0.1 percent w/v, 0.5ml) was added after the mixture had been further diluted
with deionized water (2.5ml). A 700 nm absorbance measurement revealed a percent increase
in Fe reduction activity.

Proline estimation

By crushing 0.5 g of leaf sample in 10 ml of 3 percent aqueous sulphosalicylic acid, the

proline concentration was determined. The glacial acetic acid and ninhydrin solutions were
added to 2 ml of the filtrate and heated in a water bath for 1 hour. Toluene (4 ml) was
combined and maintained at room temperature after the reaction was finished to separate the
toluene layer. Red color absorbance was measured at 520 nm using the top layer, which was
taken. As mentioned by (Sadasivam and Manickam, 1996) a standard curve was used for
calculation.

Determination of lipid peroxidation

Malondialdehyde (MDA) measurements were used to calculate the quantity of lipid

peroxidation (Heath and Packer 1968). The root tissues from the arsenic-treated and control
groups were divided into tiny pieces and crushed by adding 1 mL of 0.1 percent cold TCA.
The samples were then put into new tubes and centrifuged at 10,000 g for 20 minutes at room
temperature. A fresh tube was filled with 1 mL of supernatant, 1 mL of 20% TCA that
included 0.5 percent thiobarbituric acid, and 0.01 mL of a 4 percent butylated hydroxyl
toluene solution in ethanol. The tube was then incubated at 96 C for 35 min. The tubes were
then put in an ice bath and left there for 5 minutes before being centrifuged at 10,000 g for 5
minutes. The supernatant's absorbance was measured at 532 nm, and the absorbance at 600
nm was subtracted to adjust for non-specific turbidity. The amount of MDA was determined
based on its molar extinction coefficient, which was 156 mmol L 1 cm 1, or mmol L 1 g 1
FW.

Electrolyte Leakage

To measure the electrical conductivity, 10 leaf discs were put in 25ml of distilled water and
left at room temperature for 4 hours. To measure the electrical conductivity, the same was
autoclaved at 121°C for 30 minutes. The electrolyte leakage was calculated using the method
from Khare et al (2010).

Statistical Analysis

Three separate experimental sets (n=3) were used in the studies, which were carried out in a
completely randomized block design (CRBD). The mean value of the replicates combined
with the standard errors (SE) was used to express the results for all experiments that
underwent One Way ANOVA.

Discussion

In this work, we have identified a halotolerant endophytic bacterium that may enhance plant

development in salt-stress situations. The best candidates for testing whether endophytic

halotolerant bacteria may reduce the symptoms of salt stress in crop species are known to

have the potential to promote plant development. (Akram et al., 2016; Bharti et al., 2016;

Ilangumaran and Smith, 2017; Shahid et al., 2019; Bakka and Challabathula, 2020; Kumar et

al., 2020). Plant development and growth under various abiotic stress conditions were

demonstrated to be significantly benefited by the isolation and use of efficient endophytic

halotolerant bacteria. (Palacio-Rodríguez et al., 2017; Sapre et al., 2018; Vives-Peris et al.,

2018; Prittesh et al., 2020). Several endophytes have been reported to have the innate ability

to deal with salinity stress and enhance stress resilience in plant growth when inoculated with

plants in saline conditions. (Egamberdiever et al., 2019, Kear et al., 2019). Here, we

identified 52 salt-tolerant endophytes linked to the roots of halophytic plants. One of the

important findings of this study is the screening of IAA-producing, phosphate-solubilizing

(analyzed in vitro), with the hope that they can stimulate the growth of plants in fields under

saline conditions if provided in the form of biofertilizer. The primary auxin in plants, known

as IAA, is responsible for maintaining the growth and developmental phases of plants,

including tissue elongation and cell division, reactions to light, gravity, and pathogens,

among other things (Glick, 2012). Studies revealed that increased salt stress had an impact on

plants' xylem and phloem concentrations of IAA (Junghans et al., 2006). Moreover, IAA is

crucial for plant-microbe interactions, which may become unstable under various abiotic

conditions (such as salt stress) (Spaepen and Vanderleyden, 2011). Thus, by creating more

IAA under saline conditions and supplying the rhizosphere, salt-tolerant endophytes with

increased IAA production might promote growth-cornered plants. The PGP characteristics of

bacteria have been researched to identify those with a high potential for usage as

biofertilizers. These studies are essential because they find bacteria that are more

advantageous for plants before putting them to the test for specific qualities in the field

(ALKahtani et al., 2020). Among the several processes used by bacteria to promote plant,

development is the production of ammonia, IAA, and P. (Biasolo et al,. 2017). Most of the

endophytic bacterial isolates in this situation could produce varying amounts of ammonia.

Ammonia-producing bacteria are frequently reported to be able to provide ammonia as a

nitrogen source for plant development. Through the generation of ammonia from the

hydrolysis of urea into ammonia and carbon dioxide, bacterial endophytes can promote plant

development (Li et al.,2016). Most of the isolated endophytes had a varying ability to

solubilize phosphate, according to P-solubilization. According to Rodrigues et al., 47% of the

sugarcane-isolated bacterial endophytes showed poor P-solubilizing indices. Inoculating P-

solubilizing endophytic bacteria into the soil with limited phosphate availability results in

improved plant growth performance.

By chelating Fe+3 from complex materials and making it accessible to plants for growth

promotion, strains that generate siderophores may help iron-deficient plants (Khan and Doty

2009). According to He et al. (2010), phosphate solubilizing bacteria (PSB) are common in

the rhizosphere and may be employed as microbial inoculants in agriculture and forestry.

All of the isolated endophytic strains now exhibit PSB behavior by solubilizing phosphate,

aiding in the usage of phosphate stocks, fixed soil phosphorus, and phosphates used to

increase crop yields (Khan et al. 2006). The ability of the isolated strains to create ammonia

(NH3) as a secondary metabolite, which might be considered a key characteristic of plant

growth-promoting rhizobacteria (PGPR) capable of indirectly influencing plant development,

is essential (Wani et al. 2007). Endophytic bacteria concentrate, synthesize, and provide

nitrogen to the host plant when exposed to complex nitrogen sources. They also encourage

root and shoot elongation, which increases plant biomass (Marques et al. 2010).

Additionally, bacterial strains produced hydrogen cyanide (HCN), a desirable component of

PGPR. Some rhizobacteria prevent the growth of plant diseases and boost the host's disease

resistance system by producing HCN (Schippers et al. 1990; Whipps 2001).

In agriculture, a key issue that impacts soil health and crop productivity is zinc deficiency.

The solubilization of zinc in the soil is mostly a function of microorganisms. It had

previously been observed that such zinc compound solubilization, mediated by the synthesis

of organic acids, would lead to zinc release into the environment and bioaccumulation of Zn

inside the cells of bacterial species (Fasim et al., 2002). The main component of plants, zinc,

is essential to their growth. The most prevalent micronutrient shortage in crops globally is

zinc insufficiency, which significantly reduces agricultural output. It may not be cost-

effective to use zinc fertilizers to reduce zinc deficiency and boost crop output. Under salt

stress conditions, the synthesis of EPS is crucial and forms a chelate with ionic metals (Na+,
Cl-). It relieves the stress caused by salinity on plants by directly binding with cations (Na+)

and immobilizing them. One of the key areas for plant development because of the rice

plant's decreased Na+ absorption might be increased EPS synthesis by endophytic strains in

the inoculated plant (Upadhyay et al., 2011). The sizeable amount of EPS generation

improved biofilm development and affected soil aggregation. For instance, Upadhyay et al.

(2011) showed that wheat treated with bacterial culture under salt conditions significantly

improved its growth properties. On the other side, agricultural land has a greater phosphorus

shortage, and fertilizers are heavily used to increase crop yield. However, plants only

consume a small portion of the phosphorus given to agricultural soil. The use of phosphate

solubilizing bacteria (PSB) improves the effectiveness and availability of phosphorus to

plants (Wang et al., 2018).

Proline is a key osmoprotectant that shields bacterial cells from the osmotic stress caused by

salinity stress (Zhang et al., 2018). Thus, it is an essential solute that accumulates in

microorganisms and plants to reestablish normal activities.

The current study's findings clearly show a considerable increase in proline content and

antioxidant enzyme activity in plants treated with endophytic strains. Other researchers have

observed similar findings that bacterial-treated plants have higher concentrations of

antioxidant enzymes (CAT, SOD) than untreated plants under salt stress tolerance (Habib et

al., 2016; Shobana et al., 2020).

In tomatoes, the increase of NaCl is closely connected with an increase in antioxidant enzyme

activity (Habib et al., 2016; Al Kharusi et al., 2019). To limit the harm caused by salt stress,

plants have evolved an enzyme defense mechanism.

The findings imply that more antioxidant activity is necessary at higher NaCl doses to protect

plants from oxidative damage brought on by salinity (Li et al., 2020). One of the most

prevalent alterations under salt stress is proline accumulation in plant cells.

It is a significant solute that accumulates in plants and is considerably more abundant in

stressed plants than in healthy ones. This might help to explain why certain physiological

systems in plants are activated in salinity stress situations (Chun et al., 2018; Numan et al.,

2018).

The effect of the bacterial strain's inoculation is indicated in the current study by the greater

concentration of proline content in inoculated plants compared to non-inoculated plants. This

study revealed that plants may be able to reduce oxidative damage brought on by salt through

the action of bacteria (Ansari et al., 2019)

According to Liu et al. (2008), peroxidases contribute to several defensive mechanisms that

reduce the frequency of oxidative bursts during defensive responses (Lamb et al.,1997).

Although there was no appreciable difference in the peroxidase enzyme activity between the

treatments and the control in the tissues of the tomato leaf.

One of the enzymes that help plants resist pathogens is peroxoydase (Seevers et al.,1971)

Depending on the stage of development and the environmental stimulation, plant peroxidase

activity appears to be under rigorous regulation (Gadea et al., 1999). According to Maksimov

et al. (2003), fruit's defensive reactions to pathogens are connected with the activity of

peroxidase. Following pathogen and insect damage, tomato plants produce peroxidases.

To reduce the risk of disease transmission, peroxidases are engaged in the synthesis and

polymerization of phenolics, lignification, and hypersensitive reactions ( Bowles 1990).

Between the various treatments and the control, PAL activity in tomato tissues varied

dramatically.
There was no discernible change in PAL activity between the treatments and control in

tomato leaf tissues. In tomato leaf tissues, there was a substantial difference in PPO activity

between treatments and the control, but not in the leaves of tomato plants, where there was no

significant change. Regarding the amount of host resistance, defense-related enzyme levels

(PAL, POX, PPO, etc.) are critical (Ralph et al., 2006). By boosting antimicrobial activity,

they play a crucial role in the defensive mechanism against infections and may even have a

direct role in regulating the growth of diseases ( Melo et al., 2006). The severity of the

disease is also reduced by an increase in these defense-related enzymes (Eckardt et al.,2004,

Cao et al.,2006, Tian et al., 2006)

Important markers of soil health and agricultural yield include soil nutrients and organic

matter. An important measure of soil salinity and physicochemical characteristics is electrical

conductivity (EC). EC was reduced in non-inocula when soil salinity increased. The interior

tissues of plants offer bacterial endophytes a magnificent niche, which favorably triggers

modulations in host physiology and improves the health and production of the plants

(Amaresan et al., 2012). In the current study, 52 endophytic bacterial strains that were

isolated from various halophytic plants were examined for their effects. Endophytes, or

bacteria that live inside eukaryotic hosts, face challenging conditions because of a lack of

available nutrients, space, and control over cellular growth from their hosts. The endophytic

cell may experience simultaneous bursts of oxidation and the generation of ROS under such

circumstances. Increased catalase activity may be essential in these circumstances for

reducing oxidative stress. Endophyte survival within the host is thereby made possible.

Various physiological and morphological factors that promote plant development were

examined after the incubation of plants with each of these isolates to identify the most

efficient endophyte.
However, the vigour index was chosen as the main criterion for selecting the prospective

endophyte among all the factors examined here. The real justification for choosing the vigour

index is that it directly depends on three important growth characteristics, namely shoot

length, root length, and germination percentage (Abdul-Baki and Anderson, 1973). In our

investigation, it was shown that plants infected with endophytes 2H2, 2H18, 2H20, and 4H1

had vigour indexes that were around 97% and 85% higher than the control, which was the

highest. Therefore, it was determined that the best endophytic isolates were 2H2, 2H18,

2H20, and 4H1.

The increase in total seedling length (the sum of the shoot and root lengths) and germination

percentage in the plants infected with 2H2, 2H18, 2H20, and 4H1 may account for the much

higher vigour index. These plants showed 100% germination combined with significantly

longer seedling lengths.

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Microb. 10:1849. doi: 10.3389/fmicb.2019.01849

Sultana, S., Alam, S., and Karim, M. M. (2021). Screening of siderophore-producing salt-

tolerant rhizobacteria suitable for supporting plant growth in saline soils with iron

limitation. J. Agricult. Food Res. 4:100150. doi: 10.1016/j.jafr.2021.100150

Glick, B. R. (2012). Plant growth-promoting bacteria: mechanisms and

applications. Scientifica 2012:963401. doi: 10.6064/2012/963401

Junghans, U., Polle, A., Düchting, P., Weiler, E., Kuhlman, B., Gruber, F., et al. (2006).

Adaptation to high salinity in poplar involves changes in xylem anatomy and auxin

physiology. Plant, Cell Environ. 29, 1519–1531. doi: 10.1111/j.1365-3040.2006.01529.x

Spaepen, S., and Vanderleyden, J. (2011). Auxin and plant-microbe interactions. Cold Spring

Harbor Perspect. Biol. 3:a001438. doi: 10.1101/cshperspect.a001438

Sharma, S., Kulkarni, J., and Jha, B. (2016). Halotolerant rhizobacteria promote growth and

enhance salinity tolerance in peanut. Front. Microbiol. 7:1600. doi:

10.3389/fmicb.2016.01600

ALKahtani, M. D., Fouda, A., Attia, K. A., Al-Otaibi, F., Eid, A. M., Ewais, E. E. D., ... &

Abdelaal, K. A. (2020). Isolation and characterization of plant growth promoting endophytic

bacteria from desert plants and their application as bioinoculants for sustainable

agriculture. Agronomy, 10(9), 1325.

Biasolo, G.; Kucmanski, D.A.; Salamoni, S.P.; Gardin, J.P.; Minotto, E.; Baratto, C.M.

Isolation, characterization and selection of bacteria that promote plant growth in grapevines

(Vitis sp.). J. Agric. Sci. 2017, 9, 184–194. [CrossRef]

 Li, X.; Geng, X.; Xie, R.; Fu, L.; Jiang, J.; Gao, L.; Sun, J. The endophytic bacteria isolated

from elephant grass (Pennisetum purpureum Schumach) promote plant growth and enhance

salt tolerance of hybrid Pennisetum. Biotech. Biofuels 2016, 9, 190. [CrossRef] [PubMed]

Banik, A.; Dash, G.K.; Swain, P.; Kumar, U.; Mukhopadhyay, S.K.; Dangar, T.K.

Application of rice (Oryza sativa L.) root endophytic diazotrophic Azotobacter sp. strain

Avi2 (MCC 3432) can increase rice yield under green house and field condition. Microbiol.

Res. 2019, 219, 56–65. [CrossRef] [PubMed]

Khan, N.; Bano, A.; Curá, J.A. Role of Beneficial Microorganisms and Salicylic Acid in

Improving Rainfed Agriculture and Future Food Safety. Microorganisms 2020, 8, 1018.

[CrossRef] [PubMed]

Rodrigues, A.A.; Forzani, M.V.; Soares, R.; Sibov, S.T.; Vieira, J. Isolation and selection of

plant growth-promoting bacteria associated with sugarcane. Agric. Res. Tropics 2016, 46,

149–158. [CrossRef]

Lin, L.; Zhengyi, L.; Chunjin, H.; Zhang, X.; Chang, S.; Yang, L.; Yangrui, L.; Qianli, A.

Plant growth-promoting nitrogen-fixing Enterobacteria are in association with sugarcane

plants growing in Guangxi, China. Microbes Environ. 2012, 27, 391–398

Khan Z, Doty SL (2009) Characterization of bacterial endophytes of sweet potato plants.

Plant Soil 322:197–207

He CQ, Tan GE, Liang X, Du W, Chen YL, Zhi GY, Zhu Y (2010) Effect of Zn-tolerant

bacterial strains on growth and Zn accumulation in Orychophragmus violaceus. Appl Soil

Ecol 44:1–5

Khan MS, Zaidi A, Wani PA (2006) Role of phosphate solubilizing microorganisms in

sustainable agriculture - A review. Agron Sustain Dev 26:1–15

Wani PA, Khan MS, Zaidi A (2007) Effect of metal tolerant plant growth

promoting Bradyrhizobium sp. (vigna) on growth, symbiosis, seed yield and metal uptake by

green gram plants. Chemosphere 70:36–45

Marques APGC, Pires C, Moreira H, Rangel AOSS, Castro PML (2010) Assessment of the

plant growth promotion abilities of six bacterial isolates using Zea mays as indicator plant.

Soil Biol Biochem 42:1229–1235

Schippers B, Bakker AW, Bakker R, van Peer R (1990) Beneficial and deleterious effects of

HCN-producing Pseudomonads on rhizosphere interactions. Plant Soil 129:75–83

Whipps JM (2001) Microbial Interactions and biocontrol in the rhizophere. J Exp Bot

52:487–511

Fasim, F., Ahmed, N., Parsons, R., Gadd, G. M., 2002. Solubilization of zinc salts by

bacterium isolated from the air environment of tannery. FEMS Microbiol. Lett. 213: 1-6

Khan, R., Gurmani, A. R., Khan, M. S., and Gurmani, A. H. (2009). Residual, direct and

cumulative effect of zinc application on wheat and rice yield under rice wheat system. Soil

Environ. 28, 24–28.

Upadhyay, S.K., Singh, J.S., Singh, D.P., 2011. Exopolysaccharide producing plant

growthpromoting-rhizobacteria under salinity condition. Pedosphere. 21, 214–222.

doi.org/10.1371/journal.pone.0222302.

Wang, W., Wu, Z., He, Y., Huang, Y., Li, X., Ye, B. C., 2018. Plant growth promotion and

alleviation of salinity stress in Capsicum annuum L. by Bacillus isolated from saline soil in

Xinjiang. Ecotoxicol. Environ. Saf. 164, 520-529. https://doi.10.1016/j.ecoenv.2018.08.070.

Ansari, F. A., Ahmad, I., Pichtel, J., 2019. Growth stimulation and alleviation of salinity

stress to wheat by the biofilm forming Bacillus pumilus strain FAB10. Appl. Soil Ecol. 143,

45-54. https://doi.org/10.1016/j.apsoil.2019.05.023

Khan, M. S., Hemalatha, S., 2016. Biochemical and molecular changes induced by salinity

stressin Oryza sativaL. Acta. Physiol. Plant. 38(7), 167.https:// doi.10.1007/s11738-016-

2185-8

Bal, H. B., Nayak, L., Das, S., Adhya, T. K. 2013. Isolation of ACC deaminase producing

PGPR from rice rhizosphere and evaluating their plant growth promoting activity under salt

stress. Plant and soil, 366(1-2), 93-105. https://doi.10.1007/s11104-012-1402-5

Jaleel, C. A., Sankar, B., Sridharan, R., Panneerselvam, R. 2008. Soil salinity alters growth,

chlorophyll content, and secondary metabolite accumulation in Catharanthus roseus. Turkish

Journal of Biology, 32(2), 79-83

Zhang, S., Fan, C., Wang, Y., Xia, Y., Xiao, W., Cui, X. 2018. Salt-tolerant and plantgrowth-

promoting bacteria isolated from high-yield paddy soil. Canadian journal of microbiology,

64(12), 968-978. https://doi.org/10.1139/cjm-2017-0571

Samaddar, S., Chatterjee, P., Choudhury, A. R., Ahmed, S., Sa, T. 2019. Interactions between

Pseudomonas spp. and their role in improving the red pepper plant growth under salinity

stress. Microbiol. Res.219, 66-73. https://doi.org/10.1016/j.micres.2018.11.005.

Misra, S., Chauhan, P. S. 2020. ACC deaminase-producing rhizosphere competent Bacillus

spp. mitigate salt stress and promote Zea mays growth by modulating ethylene metabolism. 3

Biotech, 10(3), 1-14. https://doi.org/10.1007/s13205-020-2104-y

Habib, S. H., Kausar, H., Saud, H. M. 2016. Plant growth-promoting rhizobacteria enhance

salinity stress tolerance in okra through ROS-scavenging enzymes. BioMed research

international. https://doi.org/10.1155/2016/6284547.

Shobana, N., Thangappan, S., Uthandi, S. 2020. Plant Growth-Promoting Bacillus sp.

Cahoots Moisture Stress Alleviation in Rice Genotypes by Triggering Antioxidant Defense

System. Microbiol. Res. 126518. https://doi.org/10.1016/j.micres.2020.126518.

Chun, S. C., Paramasivan, M., Chandrasekaran, M. 2018. Proline accumulation influenced by

osmotic stress in arbuscular mycorrhizal symbiotic plants. Front. Microbiol.9,

2525.https://doi.org/10.3389/fmicb.2018.02525.

Samaddar, S., Chatterjee, P., Choudhury, A. R., Ahmed, S., Sa, T. 2019. Interactions between

Pseudomonas spp. and their role in improving the red pepper plant growth under salinity

stress. Microbiol. Res.219, 66-73. https://doi.org/10.1016/j.micres.2018.11.005

Numan, M., Bashir, S., Khan, Y., Mumtaz, R., Shinwari, Z. K., Khan, A. L., ... Ahmed, A. H.

2018. Plant growth promoting bacteria as an alternative strategy for salt tolerance in plants: a

review. Microbiol. Res.209, 21-32. https://doi.org/10.1016/j.micres.2018.02.003

Al Kharusi, L., Al Yahyai, R., Yaish, M. W. 2019. Antioxidant response to salinity in

salttolerant and salt-susceptible cultivars of date palm. Agriculture, 9(1),

8.https://doi.org/10.3390/agriculture9010008.

Li, X., Sun, P., Zhang, Y., Jin, C., Guan, C. 2020. A novel PGPR strain Kocuria rhizophila

Y1 enhances salt stress tolerance in maize by regulating phytohormone levels, nutrient

acquisition, redox potential, ion homeostasis, photosynthetic capacity and stress-responsive

genes expression. Environ. Exp. Bot. 104023.

https://doi.org/10.1016/j.envexpbot.2020.104023
Ansari, F. A., Ahmad, I., Pichtel, J., 2019. Growth stimulation and alleviation of salinity

stress to wheat by the biofilm forming Bacillus pumilus strain FAB10. Appl. Soil Ecol. 143,

45-54. https://doi.org/10.1016/j.apsoil.2019.05.023

Eckardt, N. A. (2004). Amino transferases confer "Enzymatic resistance" to downy mildew in

melon. Plant Cell, 16, 1-3.

Bowles, D. J. (1990). Defense-related proteins in higher plants. Annu. Rev. Biochem., 59,

873-907.

Lamb, C., and Dixon, R. A. (1997). The oxidative burst in plant disease resistance. Annual

Review of Plant Physiology and Plant Molecular Biology, 48, 251-75.

Liu, Y., Pan, Q. H., Yang, H. R., Liu, Y. Y. and Huang, W. D. (2008). Relationship between

H2O2 and Jasmonic acid in Pea leaf wounding response. Russ. J. Plant Physiol., 55(6), 851-

862.

Gadea, J., Conejero, V., Vera, P. (1999). Mol. Gen. Genet., 262, 212.

Seevers, P. M., Catedral, F. F. and Daly, J. M. (1971). The role of peroxidase iso enzymes in

resistance to wheat stem rust disease. Plant physiol., 48, 353-360.

Ralph, S., Park, J. Y., Bohlman, N. J. and Mansfield, S. D. (2006). Dirigent proteins in

conifer defense: gene discovery, phylogeny, and differential wound and insect-induced

expression of a family of DIR and DIR-like genes in spruce (Piceaspp.). Plant Mol. Biol., 60,

21-40.

[10] Cao, J., Zeng, K. and Jiang, W. (2006). Enhancement of post-harvest disease resistance

in Yalipear (Pyrus bretschneideri) fruit by salicylic acid sprays on the trees during fruit

growth. Eur. J. Plant Pathol., 114, 363-370.

[36] Tian, S., Wan, Y., Qin, G. and Xu, Y. (2006). Induction of defense responses against

Alternaria rot by different elicitors in harvested pear fruit. Appl. Microbiol. Biotechnol., 70,

729- 734.

Maksimov, I. V, Cherepanova, E. A. and Khairullin, R. M. (2003). Chitin-specific

peroxidases in plants. Biochemistry, 68, 111-115.

Akram, M. S., Shahid, M., Tariq, M., Azeem, M., Javed, M. T., Saleem, S., et al. (2016).
Deciphering Staphylococcus sciuri SAT-17 mediated anti-oxidative defense mechanisms and
growth modulations in salt stressed maize (Zea mays L.). Front. Microbiol. 7:867. doi:
10.3389/fmicb.2016.00867

Bharti, N., Pandey, S. S., Barnawal, D., Patel, V. K., and Kalra, A. (2016). Plant growth
promoting Rhizobacteria Dietzia natronolimnaea modulates the expression of stress
responsive genes providing protection of wheat from salinity stress. Sci. Rep. 6:34768

Ilangumaran, G., and Smith, D. L. (2017). Plant growth promoting Rhizobacteria in

amelioration of salinity stress: a systems biology perspective. Front. Plant Sci. 8:1768. doi:
10.3389/fpls.2017.01768

Shahid, M., Ahmed, T., Noman, M., Javed, M. T., Javed, M. R., Tahir, M., et al. (2019). Non-
pathogenic Staphylococcus strains augmented the maize growth through oxidative stress
management and nutrient supply under induced salt stress. Ann. Microbiol. 69, 727–739.
doi: 10.1007/s13213-019-01464-9

Bakka, K., and Challabathula, D. (2020). “Amelioration of salt stress tolerance in plants by
plant growth promoting Rhizobacteria: insights from ‘Omics’ approaches,” in Plant Microbe
Symbiosis, ed. A. Varma (Cham: Springer), 303–330. doi: 10.1007/978-3-030-36248-5_16

Kumar, A., Singh, S., Gaurav, A. K., Srivastava, S., and Verma, J. P. (2020). Plant growth
promoting bacteria: biological tools for the mitigation of salinity stress in plants. Front.
Microbiol. 11:1216. doi: 10.3389/fmicb.2020.01216
Egamberdieva, D., Wirth, S., Bellingrath-kimura, S. D., Mishra, J., and Arora, N. K. (2019).
Salt-tolerant plant growth promoting rhizobacteria for enhancing crop productivity of saline
soils. Front. Microbiol. 10, 2791. doi: 10.3389/fmicb.2019.02791

Kearl, J., McNary, C., Lowman, J. S., Mei, C., Aanderud, Z. T., Smith, S. T., et al. (2019).
Salt-tolerant halophyte rhizosphere bacteria stimulate growth of alfalfa in salty soil. Front.
Microb. 10:1849. doi: 10.3389/fmicb.2019.01849

Sultana, S., Alam, S., and Karim, M. M. (2021). Screening of siderophore-producing salt-
tolerant rhizobacteria suitable for supporting plant growth in saline soils with iron
limitation. J. Agricult. Food Res. 4:100150. doi: 10.1016/j.jafr.2021.100150

Glick, B. R. (2012). Plant growth-promoting bacteria: mechanisms and

applications. Scientifica 2012:963401. doi: 10.6064/2012/963401

Junghans, U., Polle, A., Düchting, P., Weiler, E., Kuhlman, B., Gruber, F., et al. (2006).
Adaptation to high salinity in poplar involves changes in xylem anatomy and auxin
physiology. Plant, Cell Environ. 29, 1519–1531. doi: 10.1111/j.1365-3040.2006.01529.x

Spaepen, S., and Vanderleyden, J. (2011). Auxin and plant-microbe interactions. Cold Spring
Harbor Perspect. Biol. 3:a001438. doi: 10.1101/cshperspect.a001438

Sharma, S., Kulkarni, J., and Jha, B. (2016). Halotolerant rhizobacteria promote growth and
enhance salinity tolerance in peanut. Front. Microbiol. 7:1600. doi:
10.3389/fmicb.2016.01600

ALKahtani, M. D., Fouda, A., Attia, K. A., Al-Otaibi, F., Eid, A. M., Ewais, E. E. D., ...
& Abdelaal, K. A. (2020). Isolation and characterization of plant growth promoting
endophytic bacteria from desert plants and their application as bioinoculants for
sustainable agriculture. Agronomy, 10(9), 1325.

Biasolo, G.; Kucmanski, D.A.; Salamoni, S.P.; Gardin, J.P.; Minotto, E.; Baratto, C.M.
Isolation, characterization and selection of bacteria that promote plant growth in grapevines
(Vitis sp.). J. Agric. Sci. 2017, 9, 184–194. [CrossRef]

Li, X.; Geng, X.; Xie, R.; Fu, L.; Jiang, J.; Gao, L.; Sun, J. The endophytic bacteria isolated
from elephant grass (Pennisetum purpureum Schumach) promote plant growth and enhance
salt tolerance of hybrid Pennisetum. Biotech. Biofuels 2016, 9, 190. [CrossRef] [PubMed]
 Banik, A.; Dash, G.K.; Swain, P.; Kumar, U.; Mukhopadhyay, S.K.; Dangar, T.K.
Application of rice (Oryza sativa L.) root endophytic diazotrophic Azotobacter sp. strain
Avi2 (MCC 3432) can increase rice yield under green house and field condition. Microbiol.
Res. 2019, 219, 56–65. [CrossRef] [PubMed]

Khan, N.; Bano, A.; Curá, J.A. Role of Beneficial Microorganisms and Salicylic Acid in
Improving Rainfed Agriculture and Future Food Safety. Microorganisms 2020, 8, 1018.
[CrossRef] [PubMed]

Rodrigues, A.A.; Forzani, M.V.; Soares, R.; Sibov, S.T.; Vieira, J. Isolation and selection of
plant growth-promoting bacteria associated with sugarcane. Agric. Res. Tropics 2016, 46,
149–158. [CrossRef]

Lin, L.; Zhengyi, L.; Chunjin, H.; Zhang, X.; Chang, S.; Yang, L.; Yangrui, L.; Qianli, A.
Plant growth-promoting nitrogen-fixing Enterobacteria are in association with sugarcane
plants growing in Guangxi, China. Microbes Environ. 2012, 27, 391–398

Khan Z, Doty SL (2009) Characterization of bacterial endophytes of sweet potato plants.

Plant Soil 322:197–207

He CQ, Tan GE, Liang X, Du W, Chen YL, Zhi GY, Zhu Y (2010) Effect of Zn-tolerant
bacterial strains on growth and Zn accumulation in Orychophragmus violaceus. Appl Soil
Ecol 44:1–5

Khan MS, Zaidi A, Wani PA (2006) Role of phosphate solubilizing microorganisms in

sustainable agriculture - A review. Agron Sustain Dev 26:1–15

Wani PA, Khan MS, Zaidi A (2007) Effect of metal tolerant plant growth
promoting Bradyrhizobium sp. (vigna) on growth, symbiosis, seed yield and metal uptake by
green gram plants. Chemosphere 70:36–45

Marques APGC, Pires C, Moreira H, Rangel AOSS, Castro PML (2010) Assessment of the
plant growth promotion abilities of six bacterial isolates using Zea mays as indicator plant.
Soil Biol Biochem 42:1229–1235

Schippers B, Bakker AW, Bakker R, van Peer R (1990) Beneficial and deleterious effects of
HCN-producing Pseudomonads on rhizosphere interactions. Plant Soil 129:75–83

Whipps JM (2001) Microbial Interactions and biocontrol in the rhizophere. J Exp Bot
52:487–511
Fasim, F., Ahmed, N., Parsons, R., Gadd, G. M., 2002. Solubilization of zinc salts by
bacterium isolated from the air environment of tannery. FEMS Microbiol. Lett. 213: 1-6
Khan, R., Gurmani, A. R., Khan, M. S., and Gurmani, A. H. (2009). Residual, direct and
cumulative effect of zinc application on wheat and rice yield under rice wheat system. Soil
Environ. 28, 24–28.

Upadhyay, S.K., Singh, J.S., Singh, D.P., 2011. Exopolysaccharide producing plant
growthpromoting-rhizobacteria under salinity condition. Pedosphere. 21, 214–222.
doi.org/10.1371/journal.pone.0222302.

Wang, W., Wu, Z., He, Y., Huang, Y., Li, X., Ye, B. C., 2018. Plant growth promotion and
alleviation of salinity stress in Capsicum annuum L. by Bacillus isolated from saline soil in
Xinjiang. Ecotoxicol. Environ. Saf. 164, 520-529. https://doi.10.1016/j.ecoenv.2018.08.070.

Ansari, F. A., Ahmad, I., Pichtel, J., 2019. Growth stimulation and alleviation of salinity stress
to wheat by the biofilm forming Bacillus pumilus strain FAB10. Appl. Soil Ecol. 143, 45-54.
https://doi.org/10.1016/j.apsoil.2019.05.023

Khan, M. S., Hemalatha, S., 2016. Biochemical and molecular changes induced by salinity
stressin Oryza sativaL. Acta. Physiol. Plant. 38(7), 167.https:// doi.10.1007/s11738-016-
2185-8

Bal, H. B., Nayak, L., Das, S., Adhya, T. K. 2013. Isolation of ACC deaminase producing PGPR
from rice rhizosphere and evaluating their plant growth promoting activity under salt stress.
Plant and soil, 366(1-2), 93-105. https://doi.10.1007/s11104-012-1402-5

Jaleel, C. A., Sankar, B., Sridharan, R., Panneerselvam, R. 2008. Soil salinity alters growth,
chlorophyll content, and secondary metabolite accumulation in Catharanthus roseus.
Turkish Journal of Biology, 32(2), 79-83

Zhang, S., Fan, C., Wang, Y., Xia, Y., Xiao, W., Cui, X. 2018. Salt-tolerant and plantgrowth-
promoting bacteria isolated from high-yield paddy soil. Canadian journal of microbiology,
64(12), 968-978. https://doi.org/10.1139/cjm-2017-0571

Samaddar, S., Chatterjee, P., Choudhury, A. R., Ahmed, S., Sa, T. 2019. Interactions between
Pseudomonas spp. and their role in improving the red pepper plant growth under salinity
stress. Microbiol. Res.219, 66-73. https://doi.org/10.1016/j.micres.2018.11.005.
Misra, S., Chauhan, P. S. 2020. ACC deaminase-producing rhizosphere competent Bacillus
spp. mitigate salt stress and promote Zea mays growth by modulating ethylene metabolism.
3 Biotech, 10(3), 1-14. https://doi.org/10.1007/s13205-020-2104-y

Habib, S. H., Kausar, H., Saud, H. M. 2016. Plant growth-promoting rhizobacteria enhance
salinity stress tolerance in okra through ROS-scavenging enzymes. BioMed research
international. https://doi.org/10.1155/2016/6284547.

Shobana, N., Thangappan, S., Uthandi, S. 2020. Plant Growth-Promoting Bacillus sp. Cahoots
Moisture Stress Alleviation in Rice Genotypes by Triggering Antioxidant Defense System.
Microbiol. Res. 126518. https://doi.org/10.1016/j.micres.2020.126518.

Chun, S. C., Paramasivan, M., Chandrasekaran, M. 2018. Proline accumulation influenced by

osmotic stress in arbuscular mycorrhizal symbiotic plants. Front. Microbiol.9,
2525.https://doi.org/10.3389/fmicb.2018.02525.

Samaddar, S., Chatterjee, P., Choudhury, A. R., Ahmed, S., Sa, T. 2019. Interactions between
Pseudomonas spp. and their role in improving the red pepper plant growth under salinity
stress. Microbiol. Res.219, 66-73. https://doi.org/10.1016/j.micres.2018.11.005

Numan, M., Bashir, S., Khan, Y., Mumtaz, R., Shinwari, Z. K., Khan, A. L., ... Ahmed, A. H.
2018. Plant growth promoting bacteria as an alternative strategy for salt tolerance in plants:
a review. Microbiol. Res.209, 21-32. https://doi.org/10.1016/j.micres.2018.02.003

Al Kharusi, L., Al Yahyai, R., Yaish, M. W. 2019. Antioxidant response to salinity in

salttolerant and salt-susceptible cultivars of date palm. Agriculture, 9(1),
8.https://doi.org/10.3390/agriculture9010008.

Li, X., Sun, P., Zhang, Y., Jin, C., Guan, C. 2020. A novel PGPR strain Kocuria rhizophila Y1
enhances salt stress tolerance in maize by regulating phytohormone levels, nutrient
acquisition, redox potential, ion homeostasis, photosynthetic capacity and stress-responsive
genes expression. Environ. Exp. Bot. 104023.
https://doi.org/10.1016/j.envexpbot.2020.104023
Ansari, F. A., Ahmad, I., Pichtel, J., 2019. Growth stimulation and alleviation of salinity stress
to wheat by the biofilm forming Bacillus pumilus strain FAB10. Appl. Soil Ecol. 143, 45-54.
https://doi.org/10.1016/j.apsoil.2019.05.023

Eckardt, N. A. (2004). Amino transferases confer "Enzymatic resistance" to downy mildew in

melon. Plant Cell, 16, 1-3.

Bowles, D. J. (1990). Defense-related proteins in higher plants. Annu. Rev. Biochem., 59,
873-907.

Lamb, C., and Dixon, R. A. (1997). The oxidative burst in plant disease resistance. Annual
Review of Plant Physiology and Plant Molecular Biology, 48, 251-75.

Liu, Y., Pan, Q. H., Yang, H. R., Liu, Y. Y. and Huang, W. D. (2008). Relationship between H2O2
and Jasmonic acid in Pea leaf wounding response. Russ. J. Plant Physiol., 55(6), 851- 862.

Gadea, J., Conejero, V., Vera, P. (1999). Mol. Gen. Genet., 262, 212.

Seevers, P. M., Catedral, F. F. and Daly, J. M. (1971). The role of peroxidase iso enzymes in
resistance to wheat stem rust disease. Plant physiol., 48, 353-360.

Ralph, S., Park, J. Y., Bohlman, N. J. and Mansfield, S. D. (2006). Dirigent proteins in conifer
defense: gene discovery, phylogeny, and differential wound and insect-induced expression
of a family of DIR and DIR-like genes in spruce (Piceaspp.). Plant Mol. Biol., 60, 21-40.

[10] Cao, J., Zeng, K. and Jiang, W. (2006). Enhancement of post-harvest disease resistance
in Yalipear (Pyrus bretschneideri) fruit by salicylic acid sprays on the trees during fruit
growth. Eur. J. Plant Pathol., 114, 363-370.

[36] Tian, S., Wan, Y., Qin, G. and Xu, Y. (2006). Induction of defense responses against
Alternaria rot by different elicitors in harvested pear fruit. Appl. Microbiol. Biotechnol., 70,
729- 734.

Maksimov, I. V, Cherepanova, E. A. and Khairullin, R. M. (2003). Chitin-specific peroxidases

in plants. Biochemistry, 68, 111-115.