Impact of Azo Dyes and Ibuprofen on the Proteome of Serratia nematodiphila sp. MB307.

BACKGROUND: Micropollutants comprise organic/mineral substances that cause an undesirable impact on the environment, by affecting life at all scales. In this study, we explored the changes they impart on the global proteome of a soil bacterium Serratia nematodiphila MB307, for two classes of pollutants, i.e., Azo dyes (Methyl orange, Congo red) and a pharmaceutical (Ibuprofen). METHODS: The 100 µg pollutant supplemented alteration of pure S. nematodiphila MB307 culture after 24 hours of incubation at 37 °C and its control was analyzed using a differential proteomics approach. MaxQuant software with the Perseus package was used for data analysis purposes. RESULTS: Prominently, ribosomal proteins and chaperones were up or downregulated in the whole cell and membranous fraction. CONCLUSION: This illustrates dynamic protein production adaptation of bacteria, to cope with stress and cell growth/division trade-off for survival. A collective pattern of survival under stress or pollution resistance could not be decrypted for all classes of pollutants, portraying dissimilar mechanisms of coping with differently structured pollutant moieties.

Exploring the potential of a newly constructed manganese peroxidase-producing yeast consortium for tolerating lignin degradation inhibitors while simultaneously decolorizing and detoxifying textile azo dye wastewater.

MnP-YC4, a newly constructed manganese peroxidase-producing yeast consortium, has been developed to withstand lignin degradation inhibitors while degrading and detoxifying azo dye. MnP-YC4 tolerance to major biomass-derived inhibitors was promising. MnP induced by lignin was found to be highly related to dye decolorization by MnP-YC4. Simulated azo dye-containing wastewater supplemented with a lignin co-substrate (3,5-Dimethoxy-4-hydroxybenzaldehyde) decolorized up to 100, 91, and 76% at final concentrations of 20, 40, and 60%, respectively. MnP-YC4 effectively decolorized the real textile wastewater sample, reaching up to 91.4%, and the COD value decreased significantly during the decolorization, reaching 7160 mg/l within 7 days. A possible dye biodegradation pathway was proposed based on the degradation products identified by UV-vis, FTIR, GC/MS, and HPLC techniques, beginning with azo bond cleavage and eventually mineralized to CO2 and H2O. When compared to the phytotoxic original dye, the phytotoxicity of MnP-YC4 treated dye-containing wastewater samples confirmed the nontoxic nature.

Photocatalytic dye degradation and antimicrobial activities of Pure and Ag-doped ZnO using Cannabis sativa leaf extract.

A facile green route has been employed for the synthesis of ZnO and Ag-doped ZnO using Cannabis sativa as a reducing and stabilizing agent. The as-synthesized nanoparticles were characterized and tested for photocatalytic dye degradation and antimicrobial activity. The results suggested that nanoparticles have shown antimicrobial activity against different human pathogenic bacteria (Escherichia coli, Klebsiella pneumonia, MRSA, Pseudomonas aeruginosa, Salmonella typhi, Staphylococcus aureus) and fungal strains (Fusarium spp. and Rosellinia necatrix). Ag-doped nanoparticles comparatively have shown better removal Congo red and methyl orange under visible light. Therefore, green synthesized nanoparticles could have beneficial applications in environmental science and biological field.

Fabrication and characterisation of silver nanoparticles using bract extract of Musa paradisiaca for its synergistic combating effect on phytopathogens, free radical scavenging activity, and catalytic efficiency.

This work explores the rapid synthesis of silver nanoparticles (AgNPs) from Musa paradisiaca (M. paradisiaca) bract extract. The bio-reduction of Ag+ ion was recorded using ultraviolet-visible spectroscopy by a surface plasmon resonance extinction peak with an absorbance at 420 nm. The phytoconstituents responsible for the reduction of AgNPs was probed using Fourier transform infrared spectroscopy. The X-ray diffraction pattern confirmed the formation of crystalline AgNPs that were analogous to selected area electron diffraction patterns. Morphological studies showed that the obtained AgNPs were monodispersed with an average size of 15 nm. The biologically synthesised AgNPs showed higher obstruction against tested phytopathogens. The synthesised AgNPs exhibited higher inhibitory zone against fungal pathogen Alternaria alternata and bacterial pathogen Pseudomonas syringae. Free radical scavenging potential of AgNPs was investigated using 1,1-diphenyl-2-picryl hydroxyl and 2,2-azinobis (3-ethylbenzothiazoline)-6-sulphonic acid assays which revealed that the synthesised AgNPs act as a potent radical scavenger. The catalytic efficiency of the synthesised AgNPs was investigated for azo dyes, methyl orange (MO), methylene blue (MB) and reduction of o-nitrophenol to o-aminophenol. The results portrayed that AgNPs act as an effective nanocatalyst to degrade MO to hydrazine derivatives, MB to leucomethylene blue, and o-nitro phenol to o-amino phenol.

Carpogenic ZnO nanoparticles: amplified nanophotocatalytic and antimicrobial action.

This investigation has for the first time utilised environmental resource Prunus cerasifera seed extract phytochemicals for the green synthesis of carpogenic ZnO nanoparticles (NPs). Spherical morphology and size range of 56.57-107.70 nm at variable calcination temperatures without the use of any external reducing agent was obtained. The synthesised NPs exhibited hexagonal wurtzite geometry with an average crystal size 5.62 nm and a band gap of 3.4 eV. Carpogenic NPs were investigated for optical, compositional, morphological, and phytochemical make up via ultraviolet spectroscopy (UV-Vis), Fourier transform infrared analysis, X-ray powder diffraction, scanning electron microscopy, and gas chromatography and mass spectrometry. Carpogenic NPs degraded methyl red up to 83% with pseudo-first-order degradation kinetics (R2 = 0.88) in 18 min signifying their remediation role in environment in conformity with all principles of green chemistry. Photocatalytic assays were performed in direct solar irradiance. Nine pathogens of biomedical and agricultural significance having multi-drug resistance were inhibited in vitro via the Kirby-Bauer disc diffusion assay. The enhanced photocatalytic and antimicrobial inhibition not only makes carpogenic ZnO NPs a future photo-degradative candidate for environmental remediation but also a nanofertiliser, nanofungicide, and nanobactericide synthesised via bioinspired, biomimetic, green, and unprecedented route.

Continuous degradation of Direct Red 23 by calcium pectate-bound Ziziphus mauritiana peroxidase: identification of metabolites and degradation routes.

In the present study, oxido-reductive degradation of diazo dye, Direct Red 23, has been carried out by Ziziphus mauritiana peroxidases (specific activity 17.6 U mg-1). Peroxidases have been immobilized via simple adsorption and cross-linking by glutaraldehyde; adsorbed and cross-linked enzyme retained 94.28% and 91.23% of original activity, respectively. The stability of peroxidases was enhanced significantly upon immobilization; a marked widening in both pH and temperature activity profiles were observed. Adsorbed peroxidases exhibited similar pH and temperature optima as reported for the free enzyme. Thermal stability was significantly enhanced in case of cross-linked enzyme which showed 80.52% activity even after 2 h of incubation at 60 °C. Packed bed reactors containing adsorbed and cross-linked peroxidases were run over a period of 4 weeks; adsorbed peroxidases retained 52.86% activity whereas cross-linked peroxidases maintained over 77% dye decolorization ability at the end of the fourth week of its continuous operation. Gas chromatography coupled with mass spectrometry was used to analyze the degradation products; it showed the presence of four major metabolites. Degradation of dye starts with the 1-Hydroxybenzotriazole radical attack on the carbon atom of the phenolic ring bearing azo linkage, converting it into cation radical which underwent nucleophilic attack by a water molecule and results in cleavage of chromophore via symmetric and asymmetric cleavage pathways. Intermediates undergo spontaneous removal of nitrogen, deamination, and oxidation reactions to produce maleic acid as the final degradation product. Graphical abstract.

Optimization of Direct Blue-14 dye degradation by Bacillus fermus (Kx898362) an alkaliphilic plant endophyte and assessment of degraded metabolite toxicity.

Alkaliphilic bacteria possesses the ability to survive in the extreme conditions with high salt concentrations. The adaptability of alkaliphilic bacteria to extreme conditions has made them predominant degrader in the field of biodegradation. A moderately alkaliphilic endophyte was isolated from Centella asiatica with a potential to degrade a di-azo dye Direct Blue-14(DB-14). The isolate was identified as Bacillus fermus with 97% similarity strain Xmb064. On optimization, maximum of 92.76% biodegradation was attained with dye concentration at 68.78 ppm supplemented with 1 g of sucrose and 2.5% (v/v) of inoculum for 72 h incubation. Characterization of the biodegraded product carried out using UV-vis spectrophotometry, FT-IR and LC-MS confirmed the destabilization of di-azo bond followed with the degradation of DB-14. Cytogenotoxicity studies revealed the biodegraded products to be less toxic. The current study is the first report on the optimization, biotransformation and cytogenotoxicity of DB-14 by B. fermus strain Centella.

Asparagus densiflorus in a vertical subsurface flow phytoreactor for treatment of real textile effluent: A lab to land approach for in situ soil remediation.

This study explores the potential of Asparagus densiflorus to treat disperse Rubin GFL (RGFL) dye and a real textile effluent in constructed vertical subsurface flow (VSbF) phytoreactor; its field cultivation for soil remediation offers a real green and economic way of environmental management. A. densiflorus decolorized RGFL (40 gm L-1) up to 91% within 48 h. VSbF phytoreactor successfully reduced American dye manufacture institute (ADMI), BOD, COD, Total Dissolved Solids (TDS) and Total Suspended Solids (TSS) of real textile effluent by 65%, 61%, 66%, 48% and 66%, respectively within 6 d. Oxidoreductive enzymes such as laccase (138%), lignin peroxidase (129%), riboflavin reductase (111%) were significantly expressed during RGFL degradation in A. densiflorus roots, while effluent transformation caused noteworthy induction of enzymes like, tyrosinase (205%), laccase (178%), veratryl oxidase (52%). Based on enzyme activities, UV-vis spectroscopy, FTIR and GC-MS results; RGFL was proposed to be transformed to 4-amino-3- methylphenyl (hydroxy) oxoammonium and N, N-diethyl aniline. Anatomical study of the advanced root tissue of A. densiflorus exhibited the progressive dye accumulation and removal during phytoremediation. HepG2 cell line and phytotoxicity study demonstrated reduced toxicity of biotransformed RGFL and treated effluent by A. densiflorus, respectively. On field remediation study revealed a noteworthy removal (67%) from polluted soil within 30 d.

Biodegradation of Azure-B dye by Serratia liquefaciens and its validation by phytotoxicity, genotoxicity and cytotoxicity studies.

The azo dyes in textile industry are a major source of environmental pollution and cause serious threat to aquatic flora and fauna. The present study aims to evaluate the potential of previously isolated lignin peroxidase (LiP) enzyme producing Serratia liquefaciens in degradation of Azure-B (AB) dye. S. liquefaciens showed rapid decolourisation of AB dye (100 mg L-1) in mineral salt medium (MSM) supplemented with 0.2% glucose and yeast extract, and more than 90% dye decolourisation was observed at 48 h when incubated at 30 °C. Decolourisation conditions were optimized by Response Surface Methodology (RSM) using Box-Behnken Designs (BBD). The dye degradation was further confirmed by ATR-FTIR and GC-MS analysis. Toxicological studies of untreated (UT) and bacterial treated (BT) AB dye solutions were studied by using phytotoxicity, genotoxicity and cytotoxicity endpoints. Phytotoxicity assay using Vigna radiata indicated that bacterial treatment led to detoxification of AB dye. Genotoxicity assay with Allium cepa showed that pure AB dye solutions significantly reduced mitotic index (MI) and induced various chromosomal abnormalities (CAs) like c-mitosis, stickiness, chromosome break, anaphase bridges, vagrant chromosomes and binucleated and micronucleated cell in the root tip cells, whereas, bacterial treated solutions induced relatively less genotoxicity in nature. Improved cell survivability (%) was also noted in kidney cell line (NRK-52E) after S. liquefaciens treated dye solutions than the pure dye solutions. The findings suggest that S. liquefaciens could be a potential bacterium for azo dye degradation, as it is effective in lowering of toxic effects of AB dye.

Optimization of conditions for decolorization of azo-based textile dyes by multiple fungal species.

Wastewater from textile industries contains azo dye residues that negatively affect most environmental systems. The biological treatment of these wastes is the best option due to safety and cost concerns. Here we isolated and identified 19 azo dye-degrading fungi and optimized conditions resulting in enhanced degradation. The fungi belonged to five species of Aspergillus and a single Lichtheimia sp. All fungi were evaluated for their ability to decolorize 20 azo dyes. While the most easily transformable azo dye was direct violet (decolorization ranged from 71.1 to 93.3%), the most resistant to decolorization was fast green azo dye. The greatest degradation potential of azo dyes (direct violet and methyl red) was optimized using the most promising four fungal strains and changing media glucose concentration, nitrogen source, and micronutrients. Biomass, lignin peroxidase, and laccases production were also determined in the optimization studies. The decolorization of both azo dyes by the four fungal strains was greatly enhanced by glucose supplementation. The fungal strains were not able to produce lignin peroxidases in the absence of organic nitrogen source. Both yeast extract and casamino acid supplementation enhanced decolorization of direct violet and methyl red dyes and production of lignin peroxidase by the fungal strains. In contrast, the laccases were absent in the similar medium enriched with the same organic nitrogen sources.

Isolation of quercetin from the methanolic extract of Lagerstroemia speciosa by HPLC technique, its cytotoxicity against MCF-7 cells and photocatalytic activity.

The flavonoids present in the leaves of Lagerstroemia speciosa were extracted, characterized by spectral methods and studied for its cytotoxicity activity against MCF-cell lines and photocatalytic activity against azo dye. Direct and sequential soxhlet extraction was performed and its concentrated crude extract was subjected to high performance liquid chromatography. The yield obtained by the isolated compound (MEI-quercetin) from leaves of L. speciosa was found to be 1.8g from the methanolic extract. The phytochemical analysis and the Rf value of the isolated flavonoid was found to be 3.59. The isolated compound was characterized by Infrared Spectroscopy, NMR and Mass. Based on the characterization, the structure was elucidated as quercetin - a flavonoid. The isolated compound showed the significant in vitro cytotoxicity activity against MCF-7 cell lines at 500µg/ml when compared to the crude extract. Among the various concentrations (25, 50, 100, 250, and 500µg/ml), at higher concentration the cell viability was pronounced and also compared with that of the control. It was first time to report that the isolated flavonoid showed photocatalytic against azo dye-methyl orange. The dye degradation was monitored by UV-Vis spectrophotometry. The isolated compound showed dye degradation of 91.66% with the crude extract 82.47% at 160min. Hence in the present findings, the photocatalytic degradation of MO dye under UV irradiation was investigated over isolated compound of L. speciosa. Hence we expect that this can be used to treat the waste water in near future based on the photocatalytic technique.

Isolation, development and identification of salt-tolerant bacterial consortium from crude-oil-contaminated soil for degradation of di-azo dye Reactive Blue 220.

The objective of this study was development and characterization of a halophilic bacterial consortium for rapid decolorization and degradation of a wide range of dyes and their mixtures. The 16S rRNA gene analysis of developed halophilic consortium VN.1 showed that the bacterial consortium contained six bacterial strains, which were identified as Pseudomonas fluorescens HM480360, Enterobacter aerogenes HM480361, Shewanella sp. HM589853, Arthrobacter nicotianae HM480363, Bacillus beijingensis HM480362 and Pseudomonas aeruginosa JQ659549. Halophilic consortium VN.1 was able to decolorize up to 2,500 mg/L RB220 with >85% chemical oxygen demand (COD) reduction under static condition at 30 °C and pH 8.0 in the presence of 7% NaCl. VN.1 also exhibited more than 85% COD reduction with >25 mg/(L h) rate of decolorization in the case of different reactive dye mixtures. We propose the symmetric cleavage of RB220 using Fourier transform infrared, high-performance liquid chromatography (HPLC), nuclear magnetic resonance and gas chromatography-mass spectrometry analysis, and confirmed the formation of sodium-4-aminobenzenesulfonate, sodium-6-aminonepthalenesulfonate, and sodiumbenzene/nepthalenesulfonate. Toxicity studies confirm that the biodegraded products of RB220 effluent stimulate the growth of plants as well as the bacterial community responsible for soil fertility.

Sunlight mediated synthesis of silver nanoparticles using redox phytoprotein and their application in catalysis and colorimetric mercury sensing.

Owing to the benign nature, plant extracts mediated green synthesis of metal nanoparticles (NPs) is rapidly expanding. In this study, we demonstrated the successful green synthesis of silver nanoparticles (AgNPs) by utilizing natural sunlight and redox protein complex composed of ferredoxin-NADP(+) reductase (FNR) and ferredoxin (FD). The capping and stabilization of the AgNPs by the redox protein was confirmed by Fourier transform infrared spectroscopy. Light and redox protein is the prerequisite factor for the formation of AgNPs. The obtained result shows that the photo generated free radicals by the redox protein is responsible for the reduction of Ag(+) to Ag(0). Transmission electron microscopy revealed the formation of spherical AgNPs with size ranging from 10 to 15 nm. As-prepared AgNPs exhibit excellent catalytic activity toward the degradation of hazardous organic dyes, such as methylene blue, methyl orange and methyl red. These bio-inspired AgNPs is highly sensitive and selective in sensing hazardous mercury ions in the water at micromolar concentration. In addition, FNR/FD extract stabilized AgNPs showed good antimicrobial activity against gram positive and gram negative bacteria.

Isolation and characterization of chicken bile matrix metalloproteinase.

Avian bile is rich in matrix metalloproteinases (MMP), the enzymes that cleave extracellular matrix proteins such as collagens and proteoglycans. Changes in bile MMP expression have been correlated with hepatic and gall bladder pathologies, but the significance of their expression in normal, healthy bile is not understood. We hypothesized that the MMP in bile may aid the digestion of native collagens that are resistant to conventional gastric proteases. Hence, the objective of this study was to characterize the bile MMP and check its regulation in association with dietary factors. We used substrate zymography, azocoll protease assay, and gelatin affinity chromatography to identify and purify the MMP from chicken bile. Using zymography and SDS PAGE, 5 bands at 70, 64, 58, 50, and 42 kDa were detected. The bands corresponding to 64, 50, and 42 kDa were identified as MMP2 using trypsin in-gel digestion and matrix-assisted laser desorption time-of-flight mass spectrometry and peptide mass fingerprinting. Chickens fed diets containing gelatin supplements showed higher levels of MMP expression in the bile by both azocoll assay and zymography. We conclude that the bile MMP may be associated with the digestion of collagens and other extracellular matrix proteins in avian diets.

Oxidative stress response in dye degrading bacterium Lysinibacillus sp. RGS exposed to Reactive Orange 16, degradation of RO16 and evaluation of toxicity.

Lysinibacillus sp. RGS degrades sulfonated azo dye Reactive Orange 16 (RO16) efficiently. Superoxide dismutase and catalase activity were tested to study the response of Lysinibacillus sp. RGS to the oxidative stress generated by RO16. The results demonstrated that oxidative stress enzymes not only protect the cell from oxidative stress but also has a probable role in decolorization along with an involvement of oxidoreductive enzymes. Formation of three different metabolites after degradation of RO16 has been confirmed by GC-MS analysis. FTIR analysis verified the degradation of functional groups of RO16, and HPTLC confirmed the removal of auxochrome group from the RO16 after degradation. Toxicity studies confirmed the genotoxic, cytotoxic, and phytotoxic nature of RO16 and the formation of less toxic products after the treatment of Lysinibacillus sp. RGS. Therefore, Lysinibacillus sp. RGS has a better perspective of bioremediation for textile wastewater treatment.

Effects of Alisma Decoction on lipid metabolism and inflammatory response are mediated through the activation of the LXRα pathway in macrophage-derived foam cells.

The liver X receptor α (LXRα)/ATP-binding cassette transporter A1 (ABCA1) pathway and LXR-modulated cytokines play an important role in macrophages which mediate lipid engulfment and the inflammatory response, and participate in the process of atherosclerosis. Therefore, lipid-lowering and anti-inflammatory therapy through the activation of the LXRα/ABCA1 pathway and LXRα-modulated cytokines may prove to be one of the main treatment strategies for atherosclerosis. Alisma Decoction (AD) has long been used in China to clinically treat cardiovascular and cerebral diseases; however, the precise mechanisms involved remain to be elucidated. In the present study, we evaluated the regulation of lipids and the anti-inflammatory effects exerted by AD and investigated the underlying molecular mechanisms using oxidized low-density lipoprotein (ox-LDL)-stimulated foam cells derived from rat peritoneal macrophages. We first found that AD markedly relieved lipid deposition in foam cells as it increased LXRα and ABCA1 expression and decreased the ox-LDL-induced expression of inflammatory cytokines, such as matrix metalloproteinase-9 and interleukin-1ß. Collectively, our findings suggest that blocking lipid deposition and inhibiting inflammatory response through the activation of the LXRα pathway may be one of the main mechanisms through which AD exerts its anti-atherosclerotic effects.

Induction of laccases in Trametes versicolor by aqueous wood extracts.

The induction of laccase isoforms in Trametes versicolor HEMIM-9 by aqueous extracts (AE) from softwood and hardwood was studied. Samples of sawdust of Pinus sp., Cedrela sp., and Quercus sp. were boiled in water to obtain AE. Different volumes of each AE were added to fungal cultures to determine the amount of AE needed for the induction experiments. Laccase activity was assayed every 24 h for 15 days. The addition of each AE (50 to 150 µl) to the fungal cultures increased laccase production compared to the control (0.42 ± 0.01 U ml(-1)). The highest laccase activities detected were 1.92 ± 0.15 U ml(-1) (pine), 1.87 ± 0.26 U ml(-1) (cedar), and 1.56 ± 0.34 U ml(-1) (oak); laccase productivities were also significantly increased. Larger volumes of any AE inhibited mycelial growth. Electrophoretic analysis revealed two laccase bands (lcc1 and lcc2) for all the treatments. However, when lcc2 was analyzed by isoelectric focusing, inducer-dependent isoform patterns composed of three (pine AE), four (oak AE), and six laccase bands (cedar AE) were observed. Thus, AE from softwood and hardwood had induction effects in T. versicolor HEMIM-9, as indicated by the increase in laccase activity and different isoform patterns. All of the enzymatic extracts were able to decolorize the dye Orange II. Dye decolorization was mainly influenced by pH. The optimum pH for decolorization was pH 5 (85%), followed by pH 7 (50%) and pH 3 (15%). No significant differences in the dye decolorizing capacity were detected between the control and the differentially induced laccase extracts (oak, pine and cedar). This could be due to the catalytic activities of isoforms with pI 5.4 and 5.8, which were detected under all induction conditions.

Biodegradation of the textile dye Mordant Black 17 (Calcon) by Moraxella osloensis isolated from textile effluent-contaminated site.

The bacterium with dye degrading ability was isolated from effluent disposal sites of textile industries, Tirupur and was identified as Moraxella osloensis based on the biochemical and morphological characterization as well as 16S rRNA sequencing. This organism was found to decolorize 87 % of Mordant Black 17 at 100 mg l⁻¹ under shake culture condition compared to 92 % under stationary culture condition. Maximum degradation of the dye by M. osloensis was achieved when the mineral salt medium was supplemented with 0.5 % glucose and 0.1 % ammonium nitrate at 35 °C. Degradation of dye was found to follow first order kinetics with the k value of 0.06282 h⁻¹ and a R² value of 0.955. Analyses for the identification of intermediate compounds confirmed the presence of naphthalene, naphthol, naphthoquinone, salicylic acid and catechol. Based on this finding a probable pathway for the degradation of Mordant Black 17 by M. osloensis has been proposed.

Super-resolution imaging with Pontamine Fast Scarlet 4BS enables direct visualization of cellulose orientation and cell connection architecture in onion epidermis cells.

BACKGROUND: In plants, a complex cell wall protects cells and defines their shape. Cellulose fibrils form a multilayered network inside the cell-wall matrix that plays a direct role in controlling cell expansion. Resolving the structure of this network will allow us to comprehend the relationship of cellulose fibril orientation and growth.The fluorescent dye Pontamine Fast Scarlet 4BS (PFS) was shown to stain cellulose with high specificity and could be used to visualize cellulose bundles in cell walls of Arabidopsis root epidermal cells with confocal microscopy. The resolution limit of confocal microscopy of some 200 nm in xy and 550 nm in z for green light, restricts the direct visualization of cellulose to relatively large bundles, whereas the structure of cellulose microfibrils with their diameter below 10 nm remains unresolved. Over the last decade, several so-called super-resolution microscopy approaches have been developed; in this paper we explore the potential of such approaches for the direct visualization of cellulose. RESULTS: To ensure optimal imaging we determined the spectral properties of PFS-stained tissue. PFS was found not to affect cell viability in the onion bulb scale epidermis. We present the first super-resolution images of cellulose bundles in the plant cell wall produced by direct stochastic optical reconstruction microscopy (dSTORM) in combination with total internal reflection fluorescence (TIRF) microscopy. Since TIRF limits observation to the cell surface, we tested as alternatives 3D-structured illumination microscopy (3D-SIM) and confocal microscopy, combined with image deconvolution. Both methods offer lower resolution than STORM, but enable 3D imaging. While 3D-SIM produced strong artifacts, deconvolution gave good results. The resolution was improved over conventional confocal microscopy and the approach could be used to demonstrate differences in fibril orientation in different layers of the cell wall as well as particular cellulose fortifications around plasmodesmata. CONCLUSIONS: Super-resolution light microscopy of PFS-stained cellulose fibrils is possible and the increased resolution over conventional approaches makes it a valuable tool for the investigation of the cell-wall structure. This is one step in method developments that will close the gap to more invasive techniques, such as atomic force and electron microscopy.