Evaluation of Arsenic-Tolerant Plant Growth-Promoting Rhizobacteria from Manipur for Mitigating Arsenic Translocation and Enhancing Growth in Rice (Oryza sativa)

Evaluation of Arsenic-Tolerant Plant Growth-Promoting Rhizobacteria from Manipur for Mitigating Arsenic Translocation and Enhancing Growth in Rice (Oryza sativa) | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Evaluation of Arsenic-Tolerant Plant Growth-Promoting Rhizobacteria from Manipur for Mitigating Arsenic Translocation and Enhancing Growth in Rice (Oryza sativa) Nirmala Akoijam, Santa Ram Joshi This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7415469/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 26 Dec, 2025 Read the published version in Scientific Reports → Version 1 posted 9 You are reading this latest preprint version Abstract Arsenic (As) contamination in paddy fields poses a serious risk to food safety by promoting arsenic accumulation in rice. This study evaluates the bioremediation potential of two arsenic-tolerant plant growth-promoting rhizobacteria (PGPR)— Bacillus paramycoides TNCB-27 and Pseudomonas shirazica TNB-16—isolated from agricultural soils in Thoubal, Manipur, India. Greenhouse experiments were conducted to assess their effects on arsenic uptake, translocation, and rice plant growth under arsenite [As(III)]- and arsenate [As(V)]- spiked conditions. Inoculated plants showed significantly reduced arsenic levels in shoots, likely due to enhanced root sequestration and microbial transformation of arsenic, as indicated by lower translocation factors. Morphological alterations in bacterial cells post-arsenic exposure were observed via scanning and transmission electron microscopy (SEM, TEM). Fourier transform infrared spectroscopy (FTIR) revealed changes in bacterial functional groups and exopolysaccharides, suggesting their role in arsenic binding. This is the first report on PGPR from Manipur demonstrating both arsenic remediation and plant growth-promoting abilities, offering a sustainable microbial approach to reduce arsenic bioavailability and accumulation in rice agroecosystems. Biological sciences/Biotechnology Earth and environmental sciences/Environmental sciences Biological sciences/Microbiology Biological sciences/Plant sciences Arsenic Bioaccumulation Bacteria Bioremediation Rice Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Introduction Bioaccumulation of arsenic (As) in rice is an alarming issue as rice forms a major constituent in diets across the world 1 . Consumption of rice containing inorganic arsenic is a widespread concern as countries like Mexico, Bangladesh, Thailand and a few others have reported the presence of arsenic in concentrations extremely close to the threshold safety limits of World Health Organisation (WHO) and Food and Agriculture Organisation (FAO) 2 – 4 . The health effects of arsenic exposure in humans are numerous which manifests into different types of cancer 5 . Usually, rice cultivation involves a flooded anaerobic environment that favours the prevalence of arsenite species in the soil, while arsenate species are more common in aerobic conditions. Several other factors like pH and redox conditions also play significant role in the interconversion between arsenic species 6 , 7 . Additionally, seasonal variations in arsenic accumulation in rice grains have also been a common phenomenon 8 – 10 . Since arsenite shares a chemical analogue with silicate, its uptake by the plant via specialised silicate transporters is common 11 . Similarly, arsenate is a phosphate analogue and transporter proteins that would otherwise assist in the uptake of phosphate have been reported to transport arsenate efficiently 12 . Recent reports have demonstrated the role of different silicate transporters in translocation of arsenite from roots to shoots and ultimately to rice grains, which makes it necessary to obstruct the process of arsenic translocation from roots to shoots in order to reduce arsenic accumulation in the grains 13 , 14 . There have been studies in recent times which have used gene editing technologies like CRISPR-CAS9 to modify expression of the silicate transporter genes in the root thereby limiting arsenite uptake without affecting the plant’s silicon requirements 15 . Although this approach seems to precisely address the problem, however, there are multiple challenges that arise while using this technology e.g. off-target effects due to inefficient delivery of the CRISPR components, possible shrinking of genetic pool with consistent use of this technology which could lead to plant cultivars being susceptible to pests and diseases 16 . In this regard, bioremediation techniques which usually do not involve manipulating or modifying the genetic organisation of plants are options to help mitigate the problem of arsenic contamination and accumulation in rice grains. The past few decades have seen tremendous development in bioremediation strategies in order to implement environmentally friendly approaches that can safely restore the target sites. Bioremediation makes use of biological processes in organisms like bacteria, fungi, algae and plants to remediate sites contaminated by heavy metals, hydrocarbons and other toxic wastes 17 . Some commonly used techniques which employ the use of either microbes or plants for bioremediating contaminated sites are biopile, windrows, bioreactors, bioventing, biosparging, bioslurping and phytoremediation 18 . Microbes found in metal-contaminated sites face selective pressure and have gradually evolved various mechanisms of heavy metal resistance consisting of extracellular barrier, intra- and extra-cellular sequestration, efflux (active transport) of metal ions, detoxification by cellular enzymes and reduction of metal ions 19 . Changes that arsenic brings about in bacterial cell morphology and shape while also examining their arsenic removal efficiency from a growth medium were considered as methods of assessment in the present study. Additionally, plant growth-promoting potential as well as arsenic translocation of select isolates for rice plants grown in soil spiked with arsenic were also assessed in the study. These results would shed light on those bacterial isolates which can withstand the toxic effects of arsenic as well as enhance growth of rice plants while simultaneously reducing arsenic translocation and accumulation in the rice grains. This was designed to develop biofertilizers containing As- tolerant and remediating bacteria which could be used for mitigation of arsenic toxicity in agricultural fields. Furthermore, while microbial bioremediation studies often emphasize arsenic reduction or plant growth enhancement separately, our study integrates both aspects in a greenhouse setup and couples this with mechanistic investigations using SEM, TEM, and FTIR. To our knowledge, this is the first report that evaluates the potential of B. paramycoides TNCB-27 and P. shirazica TNB-16 from Manipur for arsenic mitigation in rice, supported by both physiological, biochemical and ultrastructural evidence. Results Arsenic removal from growth medium The total arsenic concentration in the culture supernatant was measured using ICP-OES after 48 hours of incubation with the bacterial isolates under four treatment conditions: arsenite- and arsenate-amended media inoculated with Bacillus paramycoides TNCB-27 and Pseudomonas shirazica TNB-16 (Table 1 ). Since ICP-OES does not differentiate between arsenic species, the results reflect the residual total arsenic remaining in the medium post-incubation. Arsenic removal was higher in arsenate-amended media (~ 36%) than in arsenite-amended media (~ 28%). Isolate TNCB-27 showed greater reduction in arsenate-containing media, whereas TNB-16 performed marginally better in arsenite-containing media. Nonetheless, both isolates demonstrated comparable total arsenic removal efficiencies across the treatments. Table 1 Removal of arsenic from culture medium by bacterial isolates Arsenite-amended Arsenate-amended C i (ppm) C f (ppm) R% C i (ppm) C f (ppm) R% TNB-16 100 77.9 28.4 100 73.5 36.0 TNCB-27 100 78.1 28.0 100 73.2 36.6 FTIR Analysis FTIR spectra of the control and arsenic-treated samples were recorded and are provided as supplementary information (Figs. S1- S6). Tables 2 and 3 show the FTIR bands of TNB-16 and TNCB-27 (both control and arsenic-treated) and the shifts in peaks that occurred after arsenic-treatment. It was evident from the data that in arsenic-free or control cells there were a number of peaks which were otherwise not observed in the treated samples. Similarly, the treated samples in case of TNB-16 also showed new peaks in addition to some common ones. In TNCB-27 samples, there were a number of unique peaks observed for both control and treated cells. Some of these did not have any assigned functional groups that corresponded to wave numbers in the database. This implied that arsenic-binding on the cell surface had induced changes on the chemical moieties of the cell indicating morphological changes and membrane fluidity. Table 2 FTIR absorption bands and their subsequent assignments in the cells of the isolate TNB-16 As-free As³⁺-loaded As⁵⁺-loaded Frequency Frequency Frequency 540.65 666.71 619.17 621.97 699.71 P-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate) 656.43 655.92 982.7 697.72 702.04 1091.77 873.62 P-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate) 773.64 1155.09 897.45 897.67 1234.28 C-N and C-O stretching amide 938.94 934.29 1317.24 981.86 1083.21 P-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate) 1341.99 1087.58 1226.91 Combination of C-N stretching and N-H bending 1401.73 Alkanes C-H bending 1235.25 Aliphatic C-N stretching 1342.75 1452.14 1320.01 Aromatic C-N stretching 1402.44 Alcohols and phenols O-H bending 1548.45 1341.06 1451.96 Aromatic C = C stretching 1650.54 Amides N-H bending 1402.13 Alkanes C-H bending 1554.99 2099.05 1452.04 Aromatic C = C stretching 1643.61 Amides C = O stretching 2323.76 1549.1 2113.06 2356.34 1647.77 Amides C = O stretching 2931.44 Carboxylic acids O-H stretching 2877.79 Carboxylic acids O-H stretching 2100.58 2962.62 2930.37 2879.6 Carboxylic acids O-H stretching 3421.23 Alcohols and phenols -OH stretching 2962.59 2932.46 3298.78 Alcohols and phenols -OH stretching 2962.98 3416.35 3422.08 Alcohols and phenols -OH stretching 3724.96 3758.48 Table 3 FTIR absorption bands and their assignments in the cells of the isolate TNCB-27 As-free As³⁺-loaded As⁵⁺-loaded Frequency Frequency Frequency 542.36 662.57 622.61 618.22 701.6 660.1 660.2 895.6 P-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate) 701.07 701.49 940.26 896.25 P-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate) 778.55 1079.93 Aliphatic C-N stretching 1078.28 Aliphatic C-N stretching 1081.4 Alkenes C-H bending 1241.35 1154.66 1156.74 1313.94 Aromatic C-N stretching 1241.66 1237.39 1401.95 Alkane C-H bending 1314.27 Aromatic C-N stretching 1312.27 Aromatic C-N stretching 1447.05 1402.08 Alkane C-H bending 1401.22 Alkane C-H bending 1551.97 Aromatic C = C stretching 1449 1450.32 Aromatic C = C stretching 1646.95 Amides C = O stretching 1549.93 Aromatic C = C stretching 1554.88 2109.81 1648.39 Alkene C = C stretching 1643.25 Amides C = O stretching 2339.47 2104.28 2104.9 2965.43 Carboxylic acids O-H stretching 2880.51 Carboxylic acids O-H stretching 2320.79 3448.4 Alcohols and phenols -OH stretching 2933.02 2356.01 2963.29 2929.55 Alkane C-H stretching 3297.32 Alcohols and phenols -OH stretching 2961.87 Carboxylic acids O-H stretching 3414.42 3423 Alcohols and phenols -OH stretching 3702.56 3725.46 3764.73 3795.14 3814.05 Scanning Electron Microscopy visualized cells SEM visualization of isolates was carried out at 50000X and 10000X for treated and untreated cells. Untreated (TNB-16) cells [Fig. 1 (a) & (b)] showed whole cell morphology with an approximate cell size of ~ 1µm. In the arsenite-treated cells, a few cells showed elongation with a size of ~ 5µm [Fig. 1 (c)] as well as cell disruption [Fig. 1 (d)]. In the case of arsenate-treated cells, cell elongation was observed in most cells and other changes like cell disruption and cell rupture was less prominent [Fig. 1 (e) & (f)]. This is in accordance to the difference in toxicity levels with arsenite being more toxic than arsenate, hence the damaging effects observed in arsenite-treated cells are more pronounced than the arsenate-treated cells. The effects of arsenic treatment were more pronounced in the isolate TNCB-27 cells where cell elongation was a common occurrence in the presence of both arsenite and arsenate. The control or untreated cells were observed to be more or less smooth in appearance with an average cell size of ~ 2 µm [Fig. 2 (a) & (b)] whereas cells with arsenite treatment demonstrated elongation upto several folds which is a significant increase in cell size [Fig. 2 (c) & (d)]. In arsenate-treated cells, cell size remained more or less the same as compared to the control cells, however there were appearances of indentations on the cell surface which could be in response to the toxicity and concentration of the arsenate treatment [Fig. 2 (e) & (f)]. Transmission Electron Microscopy visualized cells In addition to the morphological changes observed with the help of scanning electron microscopy in the arsenic-treated cells, transmission electron microscopy provided an insight into the internal changes that occur simultaneously. In the case of the isolate TNB-16, the cells demonstrated size elongation in response to arsenite treatment, whereas the arsenate-treated cells showed cytoplasmic shrinkage, cell distortion and cell rupture [Fig. 3 (a), (b) and (c)]. Similarly, TNCB-27 cells treated with arsenite showed ruptured cells and cytoplasmic shrinkage, whereas cells treated with the less-toxic arsenate displayed cell distortion [Fig. 4 (a), (b) and (c)]. Arsenic bioremediation by the isolates After 90 days of growth in arsenite-spiked soil, total arsenic was estimated in the soil, roots, and shoots of control and treated plants, revealing variation in arsenic content across treatments (Figs. 5 & 6 ). In arsenite-spiked soil, total arsenic concentrations in soil for both control and treated plants were similar (~ 100–120 ppm). Roots accumulated substantially higher arsenic levels (> 1000 ppm) compared to soil and shoots. Notably, shoots of plants treated with Pseudomonas shirazica TNB-16 and Bacillus paramycoides TNCB-27 exhibited significantly lower arsenic accumulation compared to control plants. A similar pattern was observed in arsenate-spiked soils, with treated plants showing reduced shoot arsenic levels, underscoring the bioremediation potential of both isolates. Statistical analysis via two-way ANOVA followed by Dunnett’s multiple comparisons test confirmed the significance of these observations (P < 0.05), with highly significant differences (**** P < 0.0001) between controls and bacterial treatments in arsenate conditions. To further quantify arsenic mobility within the plants, the translocation factor (TF)—calculated as the ratio of arsenic concentration in shoots to roots—is presented in Fig. 7 . Although bacterial inoculation resulted in a non-significant reduction of TF under arsenite treatment, a significant decrease in TF was observed under arsenate treatment with both isolates. This indicates that the bacteria effectively limited arsenic translocation from roots to shoots in arsenate conditions. Notably, Bacillus paramycoides TNCB-27 exhibited the lowest TF under arsenate stress, suggesting enhanced sequestration of arsenic in the roots and reduced movement to aerial parts. Statistical analysis using two-way ANOVA followed by Tukey’s multiple comparisons test confirmed these differences as highly significant (**** P < 0.0001) between control and treated plants under arsenate stress. Discussion The arsenic-removal assay performed in the present study revealed the bioremediation potential of the two test isolates, Pseudomonas shirazica TNB-16 and Bacillus paramycoides TNCB-27 with similar removal percentages of about 28 and 36% for arsenite and arsenate respectively. Other studies have even reported higher removal efficiency by Bacillus sp. isolated from agricultural fields 20 . Another study also demonstrated the arsenic removal capacity of bacteria belonging to several genera including Pseudomonas sp. 21 . This disparity could be attributed to the type of media used e.g. Luria-Bertani broth which was used in our experiments which contains trace amounts of phosphate ions that may have altered or interfered with arsenate removal due to the similar chemical structure of the two molecules 22 . A comparison of the FTIR studies between the arsenic-treated and untreated bacterial cells revealed the chemical changes that occur between arsenic and the various bacterial cell wall components like polysaccharides. In both TNB-16 and TNCB-27, the IR spectra of the As-treated cells displayed new peaks which are otherwise not detected in the untreated cells, implicating a shift in peaks owing to the binding or interaction of arsenic with the functional groups of the cell wall components. This could be better described as an interaction of the negatively charged groups on the bacterial cell surface with the positively charged arsenite or arsenate by mechanisms such as van der Waals forces, electrostatic interaction, or covalent bonding 23 . Peaks were observed around 700–900 cm − 1 for both isolates, signifying an interaction of arsenic with oxygen-containing groups, likely from phosphates, hydroxyl groups, or other cell wall components. Additionally, shifts in peaks were observed after arsenic treatment, for instance in P. shirazica TNB-16, shifting of the peaks from 1226 cm − 1 to 1234 cm − 1 and 1235 cm − 1 in arsenite- and arsenate-treated could be attributed to aliphatic C-N stretching. Similarly, in B. paramycoides TNCB-27, the shift from 3423 cm − 1 to 3448 cm − 1 and 3414 cm − 1 indicated an O-H stretch of hydroxyl groups 24 . Scanning Electron Microscopy studies offer insights into morphological changes when bacterial cells interact with arsenite and arsenate. The transformation after arsenic treatment was evident in both B. paramycoides TNCB-27 and P. shirazica TNB-16 cells; while untreated cells appeared plump and smooth, treated cells appeared distorted and even elongated which was also reported in Exiguobacterium sp. and Bacillus sp. wherein a fourfold increase in cell volume was observed in comparison to the untreated cells, suggesting an accumulation of arsenic in the cells which was further verified by TEM and EDX analysis 25 . This phenomenon of cell volume increase has been described as a defence mechanism of bacteria when exposed to stress factors like heavy metals in its environment 26 . Morphological changes in the bacterial cells observed in the present study in response to arsenic treatment could thus be a similar defence mechanism that allows them to survive and thrive despite the toxic effects of arsenic. It was determined that in the presence of toxic compounds, bacterial cells decrease the ratio between surface and volume of the cells, thereby reducing their relative surfaces to lessen the effects of the toxic compound 27 , 28 . Transmission Electron Microscopy further elucidated the effects of arsenic on bacterial cells by providing an insight of the cell interior whereby the treated cells were characterized by increased cell volume and also cytoplasm shrinkage, which is also said to be a common observance in metal-treated bacteria 29 . Although, EDX analysis would have verified the accumulation of arsenic within the cells, however, electron-dense regions (which appear as dark spots) were observed in arsenite-treated TNB-16 and TNCB-27 suggesting the possibility of arsenite accumulation. Untreated TNCB-27 appears to be more electron-dense than that of untreated TNB-16 owing to the presence of a thicker cell wall in Gram-positive bacilli 30 . The bioremediation study yielded significant results where the test isolates were able to not only promote plant growth in the presence of arsenic salts in the soils 31 but also reduce the translocation of arsenic into the shoots and edible parts of the plant by immobilizing the arsenic in the roots of the plant. While initial soil amendments contained defined arsenic species (arsenite or arsenate), speciation was not monitored during the 90-day greenhouse experiment. Inter-conversion between arsenite and arsenate may have occurred under soil conditions and microbial activity, reflecting natural biogeochemical processes. This limitation is consistent with previous studies and highlights the need for future investigations incorporating arsenic speciation techniques to clarify these transformations. Notably, no grain formation was observed in any of the treatment groups, including the control. This is likely due to the high arsenic concentrations used in the experiment, which may have induced phytotoxic effects severe enough to hinder reproductive development in rice plants. Arsenic toxicity has been previously reported to interfere with flowering and grain filling stages, potentially by disrupting hormonal balance, nutrient uptake, or photosynthetic efficiency. As a result, the assessment of arsenic accumulation was limited to the vegetative tissues—roots and shoots. Future studies may consider adjusting arsenic concentrations or extending the growth period to evaluate the full impact on grain development under more moderate stress conditions. The observed decrease in arsenic accumulation in the shoots following inoculation with B. paramycoides TNCB-27 corroborates findings from studies with another Bacillus spp. 28 . The notably high arsenic concentrations in roots, especially under arsenite treatment, are in line with earlier reports documenting preferential accumulation of As(III) in rice roots 32 . This is plausible considering that As(III) enters root cells primarily through aquaporin channels and forms complexes with thiol groups, leading to its retention 33 . However, the contribution of surface-bound arsenic to total root arsenic concentrations cannot be excluded, warranting future work focused on root surface analysis and arsenic speciation. Plant growth-promoting rhizobacteria such as Bacillus cereus PMM6 have demonstrated growth promotion of rice seedlings in arsenic stress 34 . Collectively, this study reiterates and highlights the potential in utilising these arsenic-tolerant, plant growth-promoting rhizobacteria as a sustainable strategy to reduce arsenic accumulation in rice grains and to remediate contaminated agricultural soils. Methods Bacterial Isolation and Identification The bacteria used in this study Bacillus paramycoides TNCB-27 and Pseudomonas shirazica TNB-16 were previously isolated from agricultural soil samples collected in Thoubal, Manipur, India and identified in an earlier study 31 . The isolation was performed using Luria-Bertani (LB) agar, and pure colonies were obtained using the same medium. The bacterium was identified based on 16S rRNA sequencing and its sequence was deposited in GenBank under accession number OL455722 and OL455721 respectively. The two isolates selected for this study are evaluated for parameters which have not been previously studied. Arsenic removal analysis using Inductively coupled plasma-optical emission spectrometry (ICP-OES) To assess the arsenic removal efficiency of the bacterial isolates, an in vitro assay was conducted using Luria-Bertani (LB) broth supplemented with either sodium arsenite or sodium arsenate at a concentration of 100 ppm. A 100 mL aliquot of the supplemented medium was inoculated with bacterial isolates previously selected based on their plant growth-promoting potential (adjusted to 0.5 McFarland standard). The cultures were incubated at 37°C for 48 hours under shaking conditions. Post-incubation, the cultures were centrifuged at 10,000 rpm for 5 minutes to separate the biomass, and the supernatants were collected for arsenic quantification. Total arsenic concentration in the supernatant (i.e., residual arsenic in the growth medium) was measured using Inductively Coupled Plasma–Optical Emission Spectrometry (ICP-OES; Thermo Scientific™ iCAP™ 7600) at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India. Quantification was based on external calibration with certified multi-element standards and internal standard correction. The instrument had a detection limit (LOD) of 0.0044 ppm and a background equivalent concentration (BEC) of 0.002 ppm. The calibration curve demonstrated strong linearity with an R² value of 0.9985. LB broth without arsenic supplementation served as the control. The percentage of arsenic removed from the medium was calculated using the following equation: % Removal = \(\:\frac{\mathbf{C}\mathbf{i}-\mathbf{C}\mathbf{f}}{\mathbf{C}\mathbf{f}}\:\mathbf{x}\:100\) where, C i is the initial concentration and, C f is the final concentration. This method evaluates the capacity of the bacterial isolates to reduce soluble arsenic in the medium, thereby indicating potential for bioremediation 35 . Analysis of bacterial surface moieties using Fourier Transform Infrared Spectroscopy (FTIR) The FTIR technique is used to identify chemical bonds or functional groups and detect changes in the membrane surface moieties in bacteria which could be responsible for biosorption of arsenic. The pellet obtained after the centrifugation step in the previous methodology was dried overnight in a hot air oven followed by mixing with spectrochemical grade potassium bromide (KBr) in the ratio of 1:100. The FTIR spectra of the samples were then observed over a range of 500 to 4000 cm − 1 at a resolution of 4 cm − 1 using the Perkin Elmer Spectrum 400 FTIR. For control samples, pellets of bacterial isolates grown in normal LB media with no arsenic supplementation were used 36 . Scanning Electron Microscopy (SEM) for assessment of bacteria in response to As Sample preparation for SEM was done following standard techniques 37 involves fixing the bacterial cell pellets (both control and test) in 3% glutaraldehyde solution overnight. These pellets were then washed with 0.2M of cacodylate buffer for 3 changes with a 15 min interval at 4℃. Following this, dehydration of the samples is carried out by washing at increasing concentrations of acetone from 30–100% for 3 changes with a 15 min interval at 4℃ at each step. The final step involves drying the samples using a critical point drying fluid like tetramethylsilane (TMS) and allowing its complete evaporation. Before sample viewing, the dried samples are then mounted on a metal stub using adhesive tape and coated with a thin layer of conductive material like gold using a sputter coater. The prepared specimens were examined under a Scanning Electron Microscope (JSM–6360, Jeol) at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India. Transmission Electron Microscopy (TEM) for assessment of bacteria in response to As Similar to SEM sample preparation, bacterial pellets were obtained by centrifugation and carefully washed with 0.1 M phosphate buffer (PBS). These were then fixed with Karnovsky’s fixative constituted of 0.2M cacodylate buffer, 10% formaldehyde and 25% glutaraldehyde solution at 4℃ overnight. These pellets were then washed with 0.1M of cacodylate buffer for 3 changes with a 10 min interval at 4℃. Secondary fixation of the cells was with 1% osmium tetroxide (1% OsO 4 ) for 2 h at 4℃. Post-fixation, the cell pellets were serially dehydrated using 30%, 50%, 70%, 80%, 90% and 95% acetone for 2 changes with a 15 min interval at 4℃ at each step. This was followed by another washing step with propylene oxide for 30 min with two changes in room temperature. Following dehydration, the samples are then infiltrated and embedded in a mixture of embedding medium: propylene oxide in a 1:3 ratio (overnight) and gradually increasing the ratio to 3:1 (1h) then finally in embedding medium at 50℃ for 12h. The mixture is then cured into a hard block at 60℃ for 24-48h. Sectioning of the block is done using a microtome to produce 60-90nm thin sections which then undergo post-staining with uranyl acetate for 30–120 min at room temperature. The stained sections were mounted on specimen grids and viewed under a Transmission Electron Microscope (TEM, JEM-2100 PLUS (HR), Jeol) at a magnification of 3000X 38 . The TEM analysis was carried out at the Sophisticated Analytical Instrument Centre (SAIC), IASST, Guwahati. Arsenic bioremediation by the bacterial isolates Rice seeds of the local Balum variety were surface sterilized and then bacterized with the isolates, TNB-16 and TNCB-27. The seeds were allowed to germinate as per previous protocol 39 . The germinated seedlings were then transplanted into pots of 16x18cm dimensions filled with 3 kg of autoclaved soil in spiked with 50 mg/kg (or ppm) arsenite and 50 mg/kg (or ppm) arsenate respectively. Sterile or unbacterized seedlings grown in the same soil conditions served as control in this study. The pots were maintained under greenhouse conditions with daytime and nighttime temperatures of 28°C and 22°C, respectively. No fertilizers were applied throughout the experiment, and the plants were irrigated with distilled water on alternate days. After harvest, arsenic translocation from roots to shoots was determined by estimating arsenic content in the soil, roots, and shoots. The three sample types were dried overnight at approximately 65℃ followed by digestion with aqua regia (3:1 HCl: HNO 3 ) by volume) for estimation of the total arsenic content using Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES) analysis (iCAP 7000 Series Duo, Thermo Fisher Scientific, USA) 40 at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India. The glasswares were prewashed with 10% nitric acid and rinsed with distilled water to avoid external metal contamination. The analysis employed an internal standard correction and was calibrated using multi-element standards. The instrument detection limit (LOD) for arsenic was 0.0045 ppm, and the background equivalent concentration (BEC) was 0.012 ppm for solid samples while the calibration curve showed linearity with an R² value of 0.9994. Root samples were washed thoroughly with distilled water to remove adhering soil particles. However, we recognize that this may not completely eliminate surface-bound arsenic. Therefore, the root arsenic concentrations reported here may include both internalized and adsorbed forms. The translocation factor (TF) was calculated as the ratio of arsenic concentration in the shoot to that in the root (TF = As concentration in shoot / As concentration in root), and was used to evaluate the efficiency of arsenic translocation from roots to aerial parts under different bacterial treatments and arsenic species. Statistical Analysis All experimental data were analyzed using two-way analysis of variance (ANOVA) to assess the effects of treatments on arsenic accumulation and translocation. Post-hoc comparisons were performed using Tukey’s or Dunnett’s multiple comparison tests to identify significant differences between groups. Statistical significance was set at P < 0.05. Analyses were conducted using GraphPad Prism 9 (version 9.5.1). Declarations Author Contributions All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by NA. The first draft of the manuscript was written by NA and SRJ and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Data availability The 16S rRNA sequences of Bacillus paramycoides TNCB-27 and Pseudomonas shirazica TNB-16 has been deposited and can be accessed at GenBank with accession nos. OL455722 and OL455721 respectively. The FTIR spectra can be accessed in the supplementary file provided. Competing Interests The authors declare no competing interests. Acknowledgments and Funding NA acknowledges the financial support received from Council of Scientific & Industrial Research [09/0347(12424)/2021-EMR-I] as fellowship to carry out the present study. References Rahman, M. A., Hasegawa, H., Rahman, M. M., Miah, M. A. M. & Tasmin, A. Arsenic accumulation in rice (Oryza sativa L.): human exposure through food chain. Ecotoxicol Environ Saf 69 , 317–324 (2008). Hossain, M. F. Arsenic contamination in Bangladesh—an overview. Agric Ecosyst Environ 113 , 1–16 (2006). Nookabkaew, S., Rangkadilok, N., Mahidol, C., Promsuk, G. & Satayavivad, J. Determination of arsenic species in rice from Thailand and other Asian countries using simple extraction and HPLC-ICP-MS analysis. J Agric Food Chem 61 , 6991–6998 (2013). García-Rico, L., Valenzuela-Rodríguez, M. P., Meza-Montenegro, M. M. & Lopez-Duarte, A. L. 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[CE- Cell elongation, CD- Cell disruption]\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/cd61a8e6d4c05761d30b6f19.png"},{"id":92102589,"identity":"4725bddd-79af-49b5-9bed-9b2499712d90","added_by":"auto","created_at":"2025-09-24 15:56:21","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":831827,"visible":true,"origin":"","legend":"\u003cp\u003eSEM images of the isolate TNCB-27 (a) Control at 5000X magnification (b) Control at 10000X magnification (c) 100 ppm As³\u003csup\u003e⁺\u003c/sup\u003e treated at 5000X magnification (d) 100 ppm As³\u003csup\u003e⁺\u003c/sup\u003e treated at 10000X (e) 100 ppm As⁵\u003csup\u003e⁺\u003c/sup\u003e treated at 5000X (f) 100 ppm As⁵\u003csup\u003e⁺\u003c/sup\u003e treated at 10000X [CE- Cell elongation, CI- Cell indentation]\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/6c3a6ca80c02902e79ffcd00.png"},{"id":92101607,"identity":"56a787b8-0beb-4d96-99dd-5f5632ecb42d","added_by":"auto","created_at":"2025-09-24 15:48:21","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":550186,"visible":true,"origin":"","legend":"\u003cp\u003eTEM images of the isolate TNB-16 (a) Control at 3000X magnification (b) 100 ppm As³\u003csup\u003e⁺\u003c/sup\u003e treated at 3000X magnification (c) 100 ppm As⁵\u003csup\u003e⁺\u003c/sup\u003e treated at 3000X magnification. [CE- Cell Elongation, CS- Cytoplasm Shrinkage, CD- Cell Distortion, CR- Cell Rupture]\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/393b3425d672b0b552e019ee.png"},{"id":92101612,"identity":"6a6223d5-056f-461b-9d0b-6d6cf0dcb1a0","added_by":"auto","created_at":"2025-09-24 15:48:21","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":493545,"visible":true,"origin":"","legend":"\u003cp\u003eTEM images of the isolate TNCB-27 (a) Control at 3000X magnification (b) 100 ppm As³\u003csup\u003e⁺\u003c/sup\u003e treated at 3000X magnification (c) 100 ppm As⁵\u003csup\u003e⁺\u003c/sup\u003e treated at 3000X magnification. [CS- Cytoplasm Shrinkage, CD- Cell Distortion, CR- Cell Rupture]\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/259245041bc0cf7c54bcdc3b.png"},{"id":92101611,"identity":"8553abd9-25d6-4fae-bf87-054677fcf33f","added_by":"auto","created_at":"2025-09-24 15:48:21","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":191980,"visible":true,"origin":"","legend":"\u003cp\u003eArsenic content in plant biomass of control and treated plants after arsenite treatment\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/e51c50d99a376049d4c9366b.png"},{"id":92101608,"identity":"bcb1b426-43d4-4dd0-8f61-fb9f8c41dcae","added_by":"auto","created_at":"2025-09-24 15:48:21","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":123371,"visible":true,"origin":"","legend":"\u003cp\u003eArsenic content in plant biomass of control and treated plants after arsenate treatment\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/8044e96953d8f29dff1c18d6.png"},{"id":92101133,"identity":"7160dcbf-c103-4957-8562-f29f3ca5af3a","added_by":"auto","created_at":"2025-09-24 15:40:21","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":89926,"visible":true,"origin":"","legend":"\u003cp\u003eTranslocation factor (TF) of rice plants grown in arsenite- and arsenate-spiked soils under different bacterial treatments.\u003c/p\u003e","description":"","filename":"floatimage7.png","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/360cb3bfea60ba8704bf4a72.png"},{"id":99172217,"identity":"46874e5e-4980-4fff-b365-d97d08763289","added_by":"auto","created_at":"2025-12-29 16:02:28","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4073306,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/8579cbdc-a50e-4b69-a86f-9e3d59633f88.pdf"},{"id":92102590,"identity":"8eae1e5b-d3af-4d11-b483-1fa3c65cef0a","added_by":"auto","created_at":"2025-09-24 15:56:21","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":558425,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryInformation.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7415469/v1/3b556d2f49eeb70b60c4c5e6.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Evaluation of Arsenic-Tolerant Plant Growth-Promoting Rhizobacteria from Manipur for Mitigating Arsenic Translocation and Enhancing Growth in Rice (Oryza sativa)","fulltext":[{"header":"Introduction","content":"\u003cp\u003eBioaccumulation of arsenic (As) in rice is an alarming issue as rice forms a major constituent in diets across the world \u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Consumption of rice containing inorganic arsenic is a widespread concern as countries like Mexico, Bangladesh, Thailand and a few others have reported the presence of arsenic in concentrations extremely close to the threshold safety limits of World Health Organisation (WHO) and Food and Agriculture Organisation (FAO) \u003csup\u003e\u003cspan additionalcitationids=\"CR3\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. The health effects of arsenic exposure in humans are numerous which manifests into different types of cancer \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e. Usually, rice cultivation involves a flooded anaerobic environment that favours the prevalence of arsenite species in the soil, while arsenate species are more common in aerobic conditions. Several other factors like pH and redox conditions also play significant role in the interconversion between arsenic species \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. Additionally, seasonal variations in arsenic accumulation in rice grains have also been a common phenomenon \u003csup\u003e\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e. Since arsenite shares a chemical analogue with silicate, its uptake by the plant via specialised silicate transporters is common \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. Similarly, arsenate is a phosphate analogue and transporter proteins that would otherwise assist in the uptake of phosphate have been reported to transport arsenate efficiently \u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eRecent reports have demonstrated the role of different silicate transporters in translocation of arsenite from roots to shoots and ultimately to rice grains, which makes it necessary to obstruct the process of arsenic translocation from roots to shoots in order to reduce arsenic accumulation in the grains \u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e,\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. There have been studies in recent times which have used gene editing technologies like CRISPR-CAS9 to modify expression of the silicate transporter genes in the root thereby limiting arsenite uptake without affecting the plant\u0026rsquo;s silicon requirements \u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. Although this approach seems to precisely address the problem, however, there are multiple challenges that arise while using this technology e.g. off-target effects due to inefficient delivery of the CRISPR components, possible shrinking of genetic pool with consistent use of this technology which could lead to plant cultivars being susceptible to pests and diseases \u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eIn this regard, bioremediation techniques which usually do not involve manipulating or modifying the genetic organisation of plants are options to help mitigate the problem of arsenic contamination and accumulation in rice grains. The past few decades have seen tremendous development in bioremediation strategies in order to implement environmentally friendly approaches that can safely restore the target sites. Bioremediation makes use of biological processes in organisms like bacteria, fungi, algae and plants to remediate sites contaminated by heavy metals, hydrocarbons and other toxic wastes \u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e. Some commonly used techniques which employ the use of either microbes or plants for bioremediating contaminated sites are biopile, windrows, bioreactors, bioventing, biosparging, bioslurping and phytoremediation \u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eMicrobes found in metal-contaminated sites face selective pressure and have gradually evolved various mechanisms of heavy metal resistance consisting of extracellular barrier, intra- and extra-cellular sequestration, efflux (active transport) of metal ions, detoxification by cellular enzymes and reduction of metal ions \u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e. Changes that arsenic brings about in bacterial cell morphology and shape while also examining their arsenic removal efficiency from a growth medium were considered as methods of assessment in the present study. Additionally, plant growth-promoting potential as well as arsenic translocation of select isolates for rice plants grown in soil spiked with arsenic were also assessed in the study. These results would shed light on those bacterial isolates which can withstand the toxic effects of arsenic as well as enhance growth of rice plants while simultaneously reducing arsenic translocation and accumulation in the rice grains. This was designed to develop biofertilizers containing As- tolerant and remediating bacteria which could be used for mitigation of arsenic toxicity in agricultural fields. Furthermore, while microbial bioremediation studies often emphasize arsenic reduction or plant growth enhancement separately, our study integrates both aspects in a greenhouse setup and couples this with mechanistic investigations using SEM, TEM, and FTIR. To our knowledge, this is the first report that evaluates the potential of \u003cem\u003eB. paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003eP. shirazica\u003c/em\u003e TNB-16 from Manipur for arsenic mitigation in rice, supported by both physiological, biochemical and ultrastructural evidence.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eArsenic removal from growth medium\u003c/h2\u003e\u003cp\u003eThe total arsenic concentration in the culture supernatant was measured using ICP-OES after 48 hours of incubation with the bacterial isolates under four treatment conditions: arsenite- and arsenate-amended media inoculated with \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Since ICP-OES does not differentiate between arsenic species, the results reflect the residual total arsenic remaining in the medium post-incubation. Arsenic removal was higher in arsenate-amended media (~\u0026thinsp;36%) than in arsenite-amended media (~\u0026thinsp;28%). Isolate TNCB-27 showed greater reduction in arsenate-containing media, whereas TNB-16 performed marginally better in arsenite-containing media. Nonetheless, both isolates demonstrated comparable total arsenic removal efficiencies across the treatments.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eRemoval of arsenic from culture medium by bacterial isolates\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e\u003cp\u003eArsenite-amended\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c7\" namest=\"c5\"\u003e\u003cp\u003eArsenate-amended\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eC\u003csub\u003ei\u003c/sub\u003e\u003c/p\u003e\u003cp\u003e(ppm)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eC\u003csub\u003ef\u003c/sub\u003e\u003c/p\u003e\u003cp\u003e(ppm)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eR%\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eC\u003csub\u003ei\u003c/sub\u003e\u003c/p\u003e\u003cp\u003e(ppm)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eC\u003csub\u003ef\u003c/sub\u003e\u003c/p\u003e\u003cp\u003e(ppm)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eR%\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTNB-16\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003e100\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003e77.9\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003e28.4\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003e73.5\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003e36.0\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eTNCB-27\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e\u003cb\u003e100\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e\u003cb\u003e78.1\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e\u003cb\u003e28.0\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e\u003cb\u003e100\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e\u003cb\u003e73.2\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e\u003cb\u003e36.6\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eFTIR Analysis\u003c/h3\u003e\n\u003cp\u003eFTIR spectra of the control and arsenic-treated samples were recorded and are provided as supplementary information (Figs. S1- S6). Tables\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e and \u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e show the FTIR bands of TNB-16 and TNCB-27 (both control and arsenic-treated) and the shifts in peaks that occurred after arsenic-treatment. It was evident from the data that in arsenic-free or control cells there were a number of peaks which were otherwise not observed in the treated samples. Similarly, the treated samples in case of TNB-16 also showed new peaks in addition to some common ones. In TNCB-27 samples, there were a number of unique peaks observed for both control and treated cells. Some of these did not have any assigned functional groups that corresponded to wave numbers in the database. This implied that arsenic-binding on the cell surface had induced changes on the chemical moieties of the cell indicating morphological changes and membrane fluidity.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eFTIR absorption bands and their subsequent assignments in the cells of the isolate TNB-16\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAs-free\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAs\u0026sup3;⁺-loaded\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAs⁵⁺-loaded\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003e540.65\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003e666.71\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003e619.17\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e621.97\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e699.71\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eP-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e\u003cb\u003e656.43\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e655.92\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e982.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e\u003cb\u003e697.72\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e702.04\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1091.77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e873.62\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"3\" rowspan=\"4\"\u003e\u003cp\u003eP-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e773.64\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1155.09\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e897.45\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e897.67\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1234.28\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eC-N and C-O stretching amide\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e938.94\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e934.29\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1317.24\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e981.86\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1083.21\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eP-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1341.99\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1087.58\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1226.91\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eCombination of C-N stretching and N-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1401.73\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlkanes C-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1235.25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAliphatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1342.75\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1452.14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1320.01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAromatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1402.44\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlcohols and phenols O-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1548.45\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1341.06\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1451.96\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAromatic C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1650.54\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAmides N-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1402.13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAlkanes C-H bending\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1554.99\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2099.05\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1452.04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAromatic C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1643.61\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAmides C\u0026thinsp;=\u0026thinsp;O stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2323.76\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1549.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2113.06\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2356.34\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1647.77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAmides C\u0026thinsp;=\u0026thinsp;O stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2931.44\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2877.79\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"2\" rowspan=\"3\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e2100.58\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2962.62\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2930.37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e2879.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"2\" rowspan=\"3\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3421.23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2962.59\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e2932.46\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e3298.78\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e2962.98\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e3416.35\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e3422.08\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e3724.96\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e3758.48\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eFTIR absorption bands and their assignments in the cells of the isolate TNCB-27\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAs-free\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAs\u0026sup3;⁺-loaded\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAs⁵⁺-loaded\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eFrequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003e542.36\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003e662.57\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003e622.61\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e618.22\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e\u003cb\u003e701.6\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e\u003cb\u003e660.1\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e660.2\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e895.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eP-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e\u003cb\u003e701.07\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e701.49\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e940.26\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e896.25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eP-O-P stretching (phospholipids, ribose phosphate chain pyrophosphate)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e778.55\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1079.93\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAliphatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1078.28\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"2\" rowspan=\"3\"\u003e\u003cp\u003eAliphatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1081.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlkenes C-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1241.35\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1154.66\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1156.74\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1313.94\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAromatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1241.66\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1237.39\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1401.95\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlkane C-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1314.27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAromatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1312.27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAromatic C-N stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1447.05\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1402.08\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlkane C-H bending\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1401.22\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlkane C-H bending\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1551.97\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAromatic C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1449\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1450.32\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAromatic C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e1646.95\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAmides C\u0026thinsp;=\u0026thinsp;O stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1549.93\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAromatic C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1554.88\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2109.81\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1648.39\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAlkene C\u0026thinsp;=\u0026thinsp;C stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1643.25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAmides C\u0026thinsp;=\u0026thinsp;O stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2339.47\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2104.28\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2104.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e2965.43\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2880.51\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"2\" rowspan=\"3\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2320.79\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e3448.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2933.02\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2356.01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e2963.29\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2929.55\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlkane C-H stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3297.32\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2961.87\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eCarboxylic acids O-H stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3414.42\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3423\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlcohols and phenols -OH stretching\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e3702.56\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e3725.46\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e3764.73\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e3795.14\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003e3814.05\u003c/b\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\n\u003ch3\u003eScanning Electron Microscopy visualized cells\u003c/h3\u003e\n\u003cp\u003eSEM visualization of isolates was carried out at 50000X and 10000X for treated and untreated cells. Untreated (TNB-16) cells [Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (a) \u0026amp; (b)] showed whole cell morphology with an approximate cell size of ~\u0026thinsp;1\u0026micro;m. In the arsenite-treated cells, a few cells showed elongation with a size of ~\u0026thinsp;5\u0026micro;m [Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (c)] as well as cell disruption [Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (d)]. In the case of arsenate-treated cells, cell elongation was observed in most cells and other changes like cell disruption and cell rupture was less prominent [Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (e) \u0026amp; (f)]. This is in accordance to the difference in toxicity levels with arsenite being more toxic than arsenate, hence the damaging effects observed in arsenite-treated cells are more pronounced than the arsenate-treated cells.\u003c/p\u003e\u003cp\u003eThe effects of arsenic treatment were more pronounced in the isolate TNCB-27 cells where cell elongation was a common occurrence in the presence of both arsenite and arsenate. The control or untreated cells were observed to be more or less smooth in appearance with an average cell size of ~\u0026thinsp;2 \u0026micro;m [Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e (a) \u0026amp; (b)] whereas cells with arsenite treatment demonstrated elongation upto several folds which is a significant increase in cell size [Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e (c) \u0026amp; (d)]. In arsenate-treated cells, cell size remained more or less the same as compared to the control cells, however there were appearances of indentations on the cell surface which could be in response to the toxicity and concentration of the arsenate treatment [Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e (e) \u0026amp; (f)].\u003c/p\u003e\n\u003ch3\u003eTransmission Electron Microscopy visualized cells\u003c/h3\u003e\n\u003cp\u003eIn addition to the morphological changes observed with the help of scanning electron microscopy in the arsenic-treated cells, transmission electron microscopy provided an insight into the internal changes that occur simultaneously. In the case of the isolate TNB-16, the cells demonstrated size elongation in response to arsenite treatment, whereas the arsenate-treated cells showed cytoplasmic shrinkage, cell distortion and cell rupture [Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e (a), (b) and (c)]. Similarly, TNCB-27 cells treated with arsenite showed ruptured cells and cytoplasmic shrinkage, whereas cells treated with the less-toxic arsenate displayed cell distortion [Fig. \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e (a), (b) and (c)].\u003c/p\u003e\n\u003ch3\u003eArsenic bioremediation by the isolates\u003c/h3\u003e\n\u003cp\u003eAfter 90 days of growth in arsenite-spiked soil, total arsenic was estimated in the soil, roots, and shoots of control and treated plants, revealing variation in arsenic content across treatments (Figs. \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e \u0026amp; \u003cspan class=\"InternalRef\"\u003e6\u003c/span\u003e). In arsenite-spiked soil, total arsenic concentrations in soil for both control and treated plants were similar (~\u0026thinsp;100\u0026ndash;120 ppm). Roots accumulated substantially higher arsenic levels (\u0026gt;\u0026thinsp;1000 ppm) compared to soil and shoots. Notably, shoots of plants treated with \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16 and \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 exhibited significantly lower arsenic accumulation compared to control plants. A similar pattern was observed in arsenate-spiked soils, with treated plants showing reduced shoot arsenic levels, underscoring the bioremediation potential of both isolates. Statistical analysis via two-way ANOVA followed by Dunnett\u0026rsquo;s multiple comparisons test confirmed the significance of these observations (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05), with highly significant differences (**** P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) between controls and bacterial treatments in arsenate conditions.\u003c/p\u003e\n\u003cp\u003eTo further quantify arsenic mobility within the plants, the translocation factor (TF)\u0026mdash;calculated as the ratio of arsenic concentration in shoots to roots\u0026mdash;is presented in Fig. \u003cspan class=\"InternalRef\"\u003e7\u003c/span\u003e. Although bacterial inoculation resulted in a non-significant reduction of TF under arsenite treatment, a significant decrease in TF was observed under arsenate treatment with both isolates. This indicates that the bacteria effectively limited arsenic translocation from roots to shoots in arsenate conditions. Notably, \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 exhibited the lowest TF under arsenate stress, suggesting enhanced sequestration of arsenic in the roots and reduced movement to aerial parts. Statistical analysis using two-way ANOVA followed by Tukey\u0026rsquo;s multiple comparisons test confirmed these differences as highly significant (**** P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) between control and treated plants under arsenate stress.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe arsenic-removal assay performed in the present study revealed the bioremediation potential of the two test isolates, \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16 and \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 with similar removal percentages of about 28 and 36% for arsenite and arsenate respectively. Other studies have even reported higher removal efficiency by \u003cem\u003eBacillus\u003c/em\u003e sp. isolated from agricultural fields \u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. Another study also demonstrated the arsenic removal capacity of bacteria belonging to several genera including \u003cem\u003ePseudomonas\u003c/em\u003e sp. \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e. This disparity could be attributed to the type of media used e.g. Luria-Bertani broth which was used in our experiments which contains trace amounts of phosphate ions that may have altered or interfered with arsenate removal due to the similar chemical structure of the two molecules \u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eA comparison of the FTIR studies between the arsenic-treated and untreated bacterial cells revealed the chemical changes that occur between arsenic and the various bacterial cell wall components like polysaccharides. In both TNB-16 and TNCB-27, the IR spectra of the As-treated cells displayed new peaks which are otherwise not detected in the untreated cells, implicating a shift in peaks owing to the binding or interaction of arsenic with the functional groups of the cell wall components. This could be better described as an interaction of the negatively charged groups on the bacterial cell surface with the positively charged arsenite or arsenate by mechanisms such as van der Waals forces, electrostatic interaction, or covalent bonding \u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. Peaks were observed around 700\u0026ndash;900 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for both isolates, signifying an interaction of arsenic with oxygen-containing groups, likely from phosphates, hydroxyl groups, or other cell wall components. Additionally, shifts in peaks were observed after arsenic treatment, for instance in \u003cem\u003eP. shirazica\u003c/em\u003e TNB-16, shifting of the peaks from 1226 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e to 1234 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and 1235 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in arsenite- and arsenate-treated could be attributed to aliphatic C-N stretching. Similarly, in \u003cem\u003eB. paramycoides\u003c/em\u003e TNCB-27, the shift from 3423 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e to 3448 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and 3414 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e indicated an O-H stretch of hydroxyl groups \u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e .\u003c/p\u003e\u003cp\u003eScanning Electron Microscopy studies offer insights into morphological changes when bacterial cells interact with arsenite and arsenate. The transformation after arsenic treatment was evident in both \u003cem\u003eB. paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003eP. shirazica\u003c/em\u003e TNB-16 cells; while untreated cells appeared plump and smooth, treated cells appeared distorted and even elongated which was also reported in \u003cem\u003eExiguobacterium\u003c/em\u003e sp. and \u003cem\u003eBacillus\u003c/em\u003e sp. wherein a fourfold increase in cell volume was observed in comparison to the untreated cells, suggesting an accumulation of arsenic in the cells which was further verified by TEM and EDX analysis \u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. This phenomenon of cell volume increase has been described as a defence mechanism of bacteria when exposed to stress factors like heavy metals in its environment \u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e. Morphological changes in the bacterial cells observed in the present study in response to arsenic treatment could thus be a similar defence mechanism that allows them to survive and thrive despite the toxic effects of arsenic. It was determined that in the presence of toxic compounds, bacterial cells decrease the ratio between surface and volume of the cells, thereby reducing their relative surfaces to lessen the effects of the toxic compound\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e,\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eTransmission Electron Microscopy further elucidated the effects of arsenic on bacterial cells by providing an insight of the cell interior whereby the treated cells were characterized by increased cell volume and also cytoplasm shrinkage, which is also said to be a common observance in metal-treated bacteria \u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e. Although, EDX analysis would have verified the accumulation of arsenic within the cells, however, electron-dense regions (which appear as dark spots) were observed in arsenite-treated TNB-16 and TNCB-27 suggesting the possibility of arsenite accumulation. Untreated TNCB-27 appears to be more electron-dense than that of untreated TNB-16 owing to the presence of a thicker cell wall in Gram-positive bacilli \u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eThe bioremediation study yielded significant results where the test isolates were able to not only promote plant growth in the presence of arsenic salts in the soils \u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e but also reduce the translocation of arsenic into the shoots and edible parts of the plant by immobilizing the arsenic in the roots of the plant. While initial soil amendments contained defined arsenic species (arsenite or arsenate), speciation was not monitored during the 90-day greenhouse experiment. Inter-conversion between arsenite and arsenate may have occurred under soil conditions and microbial activity, reflecting natural biogeochemical processes. This limitation is consistent with previous studies and highlights the need for future investigations incorporating arsenic speciation techniques to clarify these transformations. Notably, no grain formation was observed in any of the treatment groups, including the control. This is likely due to the high arsenic concentrations used in the experiment, which may have induced phytotoxic effects severe enough to hinder reproductive development in rice plants. Arsenic toxicity has been previously reported to interfere with flowering and grain filling stages, potentially by disrupting hormonal balance, nutrient uptake, or photosynthetic efficiency. As a result, the assessment of arsenic accumulation was limited to the vegetative tissues\u0026mdash;roots and shoots. Future studies may consider adjusting arsenic concentrations or extending the growth period to evaluate the full impact on grain development under more moderate stress conditions.\u003c/p\u003e\u003cp\u003eThe observed decrease in arsenic accumulation in the shoots following inoculation with \u003cem\u003eB. paramycoides\u003c/em\u003e TNCB-27 corroborates findings from studies with another \u003cem\u003eBacillus\u003c/em\u003e spp. \u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e. The notably high arsenic concentrations in roots, especially under arsenite treatment, are in line with earlier reports documenting preferential accumulation of As(III) in rice roots \u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e. This is plausible considering that As(III) enters root cells primarily through aquaporin channels and forms complexes with thiol groups, leading to its retention \u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e. However, the contribution of surface-bound arsenic to total root arsenic concentrations cannot be excluded, warranting future work focused on root surface analysis and arsenic speciation. Plant growth-promoting rhizobacteria such as \u003cem\u003eBacillus cereus\u003c/em\u003e PMM6 have demonstrated growth promotion of rice seedlings in arsenic stress \u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e. Collectively, this study reiterates and highlights the potential in utilising these arsenic-tolerant, plant growth-promoting rhizobacteria as a sustainable strategy to reduce arsenic accumulation in rice grains and to remediate contaminated agricultural soils.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec10\" class=\"Section2\"\u003e\u003ch2\u003eBacterial Isolation and Identification\u003c/h2\u003e\u003cp\u003eThe bacteria used in this study \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16 were previously isolated from agricultural soil samples collected in Thoubal, Manipur, India and identified in an earlier study \u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e. The isolation was performed using Luria-Bertani (LB) agar, and pure colonies were obtained using the same medium. The bacterium was identified based on 16S rRNA sequencing and its sequence was deposited in GenBank under accession number OL455722 and OL455721 respectively. The two isolates selected for this study are evaluated for parameters which have not been previously studied.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003eArsenic removal analysis using Inductively coupled plasma-optical emission spectrometry (ICP-OES)\u003c/h2\u003e\u003cp\u003eTo assess the arsenic removal efficiency of the bacterial isolates, an in vitro assay was conducted using Luria-Bertani (LB) broth supplemented with either sodium arsenite or sodium arsenate at a concentration of 100 ppm. A 100 mL aliquot of the supplemented medium was inoculated with bacterial isolates previously selected based on their plant growth-promoting potential (adjusted to 0.5 McFarland standard). The cultures were incubated at 37\u0026deg;C for 48 hours under shaking conditions. Post-incubation, the cultures were centrifuged at 10,000 rpm for 5 minutes to separate the biomass, and the supernatants were collected for arsenic quantification. Total arsenic concentration in the supernatant (i.e., residual arsenic in the growth medium) was measured using Inductively Coupled Plasma\u0026ndash;Optical Emission Spectrometry (ICP-OES; Thermo Scientific\u0026trade; iCAP\u0026trade; 7600) at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India.\u003c/p\u003e\u003cp\u003eQuantification was based on external calibration with certified multi-element standards and internal standard correction. The instrument had a detection limit (LOD) of 0.0044 ppm and a background equivalent concentration (BEC) of 0.002 ppm. The calibration curve demonstrated strong linearity with an R\u0026sup2; value of 0.9985. LB broth without arsenic supplementation served as the control.\u003c/p\u003e\u003cp\u003eThe percentage of arsenic removed from the medium was calculated using the following equation:\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003e% Removal = \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\frac{\\mathbf{C}\\mathbf{i}-\\mathbf{C}\\mathbf{f}}{\\mathbf{C}\\mathbf{f}}\\:\\mathbf{x}\\:100\\)\u003c/span\u003e\u003c/span\u003e\u003c/h2\u003e\u003cp\u003ewhere, C\u003csub\u003ei\u003c/sub\u003e is the initial concentration\u003c/p\u003e\u003cp\u003eand, C\u003csub\u003ef\u003c/sub\u003e is the final concentration.\u003c/p\u003e\u003cp\u003eThis method evaluates the capacity of the bacterial isolates to reduce soluble arsenic in the medium, thereby indicating potential for bioremediation \u003csup\u003e\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\u003ch2\u003eAnalysis of bacterial surface moieties using Fourier Transform Infrared Spectroscopy (FTIR)\u003c/h2\u003e\u003cp\u003eThe FTIR technique is used to identify chemical bonds or functional groups and detect changes in the membrane surface moieties in bacteria which could be responsible for biosorption of arsenic. The pellet obtained after the centrifugation step in the previous methodology was dried overnight in a hot air oven followed by mixing with spectrochemical grade potassium bromide (KBr) in the ratio of 1:100. The FTIR spectra of the samples were then observed over a range of 500 to 4000 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e at a resolution of 4 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e using the Perkin Elmer Spectrum 400 FTIR. For control samples, pellets of bacterial isolates grown in normal LB media with no arsenic supplementation were used \u003csup\u003e\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\u003ch2\u003eScanning Electron Microscopy (SEM) for assessment of bacteria in response to As\u003c/h2\u003e\u003cp\u003eSample preparation for SEM was done following standard techniques \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e involves fixing the bacterial cell pellets (both control and test) in 3% glutaraldehyde solution overnight. These pellets were then washed with 0.2M of cacodylate buffer for 3 changes with a 15 min interval at 4℃. Following this, dehydration of the samples is carried out by washing at increasing concentrations of acetone from 30\u0026ndash;100% for 3 changes with a 15 min interval at 4℃ at each step. The final step involves drying the samples using a critical point drying fluid like tetramethylsilane (TMS) and allowing its complete evaporation. Before sample viewing, the dried samples are then mounted on a metal stub using adhesive tape and coated with a thin layer of conductive material like gold using a sputter coater. The prepared specimens were examined under a Scanning Electron Microscope (JSM\u0026ndash;6360, Jeol) at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\u003ch2\u003eTransmission Electron Microscopy (TEM) for assessment of bacteria in response to As\u003c/h2\u003e\u003cp\u003eSimilar to SEM sample preparation, bacterial pellets were obtained by centrifugation and carefully washed with 0.1 M phosphate buffer (PBS). These were then fixed with Karnovsky\u0026rsquo;s fixative constituted of 0.2M cacodylate buffer, 10% formaldehyde and 25% glutaraldehyde solution at 4℃ overnight. These pellets were then washed with 0.1M of cacodylate buffer for 3 changes with a 10 min interval at 4℃. Secondary fixation of the cells was with 1% osmium tetroxide (1% OsO\u003csub\u003e4\u003c/sub\u003e) for 2 h at 4℃. Post-fixation, the cell pellets were serially dehydrated using 30%, 50%, 70%, 80%, 90% and 95% acetone for 2 changes with a 15 min interval at 4℃ at each step. This was followed by another washing step with propylene oxide for 30 min with two changes in room temperature. Following dehydration, the samples are then infiltrated and embedded in a mixture of embedding medium: propylene oxide in a 1:3 ratio (overnight) and gradually increasing the ratio to 3:1 (1h) then finally in embedding medium at 50℃ for 12h. The mixture is then cured into a hard block at 60℃ for 24-48h. Sectioning of the block is done using a microtome to produce 60-90nm thin sections which then undergo post-staining with uranyl acetate for 30\u0026ndash;120 min at room temperature. The stained sections were mounted on specimen grids and viewed under a Transmission Electron Microscope (TEM, JEM-2100 PLUS (HR), Jeol) at a magnification of 3000X \u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e. The TEM analysis was carried out at the Sophisticated Analytical Instrument Centre (SAIC), IASST, Guwahati.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003eArsenic bioremediation by the bacterial isolates\u003c/h2\u003e\u003cp\u003eRice seeds of the local \u003cem\u003eBalum\u003c/em\u003e variety were surface sterilized and then bacterized with the isolates, TNB-16 and TNCB-27. The seeds were allowed to germinate as per previous protocol \u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e. The germinated seedlings were then transplanted into pots of 16x18cm dimensions filled with 3 kg of autoclaved soil in spiked with 50 mg/kg (or ppm) arsenite and 50 mg/kg (or ppm) arsenate respectively. Sterile or unbacterized seedlings grown in the same soil conditions served as control in this study. The pots were maintained under greenhouse conditions with daytime and nighttime temperatures of 28\u0026deg;C and 22\u0026deg;C, respectively. No fertilizers were applied throughout the experiment, and the plants were irrigated with distilled water on alternate days.\u003c/p\u003e\u003cp\u003eAfter harvest, arsenic translocation from roots to shoots was determined by estimating arsenic content in the soil, roots, and shoots. The three sample types were dried overnight at approximately 65℃ followed by digestion with aqua regia (3:1 HCl: HNO\u003csub\u003e3\u003c/sub\u003e) by volume) for estimation of the total arsenic content using Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES) analysis (iCAP 7000 Series Duo, Thermo Fisher Scientific, USA) \u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e at the Sophisticated Analytical Instrumentation Facility (SAIF), North-Eastern Hill University (NEHU), Shillong, India. The glasswares were prewashed with 10% nitric acid and rinsed with distilled water to avoid external metal contamination. The analysis employed an internal standard correction and was calibrated using multi-element standards. The instrument detection limit (LOD) for arsenic was 0.0045 ppm, and the background equivalent concentration (BEC) was 0.012 ppm for solid samples while the calibration curve showed linearity with an R\u0026sup2; value of 0.9994. Root samples were washed thoroughly with distilled water to remove adhering soil particles. However, we recognize that this may not completely eliminate surface-bound arsenic. Therefore, the root arsenic concentrations reported here may include both internalized and adsorbed forms.\u003c/p\u003e\u003cp\u003eThe translocation factor (TF) was calculated as the ratio of arsenic concentration in the shoot to that in the root (TF\u0026thinsp;=\u0026thinsp;As concentration in shoot / As concentration in root), and was used to evaluate the efficiency of arsenic translocation from roots to aerial parts under different bacterial treatments and arsenic species.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\u003ch2\u003eStatistical Analysis\u003c/h2\u003e\u003cp\u003eAll experimental data were analyzed using two-way analysis of variance (ANOVA) to assess the effects of treatments on arsenic accumulation and translocation. Post-hoc comparisons were performed using Tukey\u0026rsquo;s or Dunnett\u0026rsquo;s multiple comparison tests to identify significant differences between groups. Statistical significance was set at \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05. Analyses were conducted using GraphPad Prism 9 (version 9.5.1).\u003c/p\u003e\u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by NA. The first draft of the manuscript was written by NA and SRJ and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe 16S rRNA sequences of \u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16 has been deposited and can be accessed at GenBank with accession nos. OL455722 and OL455721 respectively. The FTIR spectra can be accessed in the supplementary file provided.\u003c/p\u003e\n\n\u003cp\u003e\u003cstrong\u003eCompeting Interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\n\u003cp\u003e\u003cstrong\u003eAcknowledgments and Funding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNA acknowledges the financial support received from Council of Scientific \u0026amp; Industrial Research [09/0347(12424)/2021-EMR-I] as fellowship to carry out the present study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eRahman, M. A., Hasegawa, H., Rahman, M. M., Miah, M. A. M. \u0026amp; Tasmin, A. Arsenic accumulation in rice (Oryza sativa L.): human exposure through food chain. \u003cem\u003eEcotoxicol Environ Saf\u003c/em\u003e \u003cstrong\u003e69\u003c/strong\u003e, 317\u0026ndash;324 (2008).\u003c/li\u003e\n\u003cli\u003eHossain, M. F. 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Determination of heavy metal concentrations in plants exposed to different degrees of pollution using ICP-AES. \u003cem\u003eFresenius J Anal Chem\u003c/em\u003e \u003cstrong\u003e354\u003c/strong\u003e, 648\u0026ndash;652 (1996).\u003c/li\u003e\n\u003c/ol\u003e "}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Arsenic Bioaccumulation, Bacteria, Bioremediation, Rice","lastPublishedDoi":"10.21203/rs.3.rs-7415469/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7415469/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eArsenic (As) contamination in paddy fields poses a serious risk to food safety by promoting arsenic accumulation in rice. This study evaluates the bioremediation potential of two arsenic-tolerant plant growth-promoting rhizobacteria (PGPR)\u0026mdash;\u003cem\u003eBacillus paramycoides\u003c/em\u003e TNCB-27 and \u003cem\u003ePseudomonas shirazica\u003c/em\u003e TNB-16\u0026mdash;isolated from agricultural soils in Thoubal, Manipur, India. Greenhouse experiments were conducted to assess their effects on arsenic uptake, translocation, and rice plant growth under arsenite [As(III)]- and arsenate [As(V)]- spiked conditions. Inoculated plants showed significantly reduced arsenic levels in shoots, likely due to enhanced root sequestration and microbial transformation of arsenic, as indicated by lower translocation factors. Morphological alterations in bacterial cells post-arsenic exposure were observed via scanning and transmission electron microscopy (SEM, TEM). Fourier transform infrared spectroscopy (FTIR) revealed changes in bacterial functional groups and exopolysaccharides, suggesting their role in arsenic binding. This is the first report on PGPR from Manipur demonstrating both arsenic remediation and plant growth-promoting abilities, offering a sustainable microbial approach to reduce arsenic bioavailability and accumulation in rice agroecosystems.\u003c/p\u003e","manuscriptTitle":"Evaluation of Arsenic-Tolerant Plant Growth-Promoting Rhizobacteria from Manipur for Mitigating Arsenic Translocation and Enhancing Growth in Rice (Oryza sativa)","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-24 15:40:16","doi":"10.21203/rs.3.rs-7415469/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-19T20:13:27+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-17T15:25:31+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"269525834251384153623645440988855630676","date":"2025-10-07T16:01:11+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-09-23T10:26:00+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"145233463385725590719278218466888482860","date":"2025-09-16T11:24:32+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-09-16T08:56:27+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-02T12:40:40+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-08-27T12:57:52+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-08-27T12:54:09+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"bb7cceef-c96a-4cb7-8124-71249527719c","owner":[],"postedDate":"September 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":55185888,"name":"Biological sciences/Biotechnology"},{"id":55185889,"name":"Earth and environmental sciences/Environmental sciences"},{"id":55185890,"name":"Biological sciences/Microbiology"},{"id":55185891,"name":"Biological sciences/Plant sciences"}],"tags":[],"updatedAt":"2025-12-29T15:58:59+00:00","versionOfRecord":{"articleIdentity":"rs-7415469","link":"https://doi.org/10.1038/s41598-025-30386-7","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2025-12-26 15:57:00","publishedOnDateReadable":"December 26th, 2025"},"versionCreatedAt":"2025-09-24 15:40:16","video":"","vorDoi":"10.1038/s41598-025-30386-7","vorDoiUrl":"https://doi.org/10.1038/s41598-025-30386-7","workflowStages":[]},"version":"v1","identity":"rs-7415469","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7415469","identity":"rs-7415469","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}