Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants and could produce oxidative toxicity to plants. Our previous study has shown that miR398 is involved in response to phenanthrene treatment by targeting CSD1 and CSD2. However, it is not clear which is essential for CSD1 and CSD2 and how miR398 changes. In this study, we performed discontinuous PAGE to separate superoxide dismutase (SOD) isozymes and found that two bands of the cytosolic Cu/Zn-SOD are induced by phenanthrene at day 5 and 7. Low expression of pri-miR398 and high expression of pre-miR398 indicate that the conversion process from pri-miR398 to pre-miR398 is impeded, which causes decrease in mature miR398. The relative expression of CSD1 is entirely up-regulated, further confirming the important role of CSD1 in response to phenanthrene exposure. Besides, the overexpression of WRKY implies its potential function in answering the call from phenanthrene stress. Therefore, it is concluded that the gene silencing of CSD1 recedes due to the biosynthesis inhibition of miR398, causing the increase of SOD activity in response to phenanthrene exposure in wheat roots. Our results are useful not only for better understanding miRNAs regulation in detoxication of reactive oxygen species, but also for alleviating the toxicity to crops caused by PAHs.

To investigate the differential contribution of extracellular polymeric substances (EPS) to polycyclic aromatic hydrocarbon (PAH) degradation by fungi and bacteria, the PAH-degrading ability and characteristics of EPS from the bacterium Mycobacterium gilvum SN12 and the fungus Mucor mucedo were compared. The fungus degraded 11% more pyrene and 21% more benzo[a]pyrene (B[a]P) than the bacterium. The biodegradation of pyrene and B[a]P increased after EPS were introduced into the PAH degradation solution, and 5.0% more pyrene and 4.5% more B[a]P were achieved for the added EPS from the fungal compared with the bacterial isolate. The comparison of two EPS indicated that the amount of protein and carbohydrate in EPS from the fungus was greater than that from the bacterium and was especially enriched for tryptophan, which is positively related to the increase of PAH biodegradation by fungal EPS. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) further revealed that higher molecular weight (HMW) of proteins over 200 kDa only existed in EPS from the fungus, and the polymorphism of proteins in EPS from the fungus was more abundant than that from the bacterium. The HMW proteins with stronger hydrophobicity in the fungal EPS also enabled the fungus to absorb more PAH than the bacterium. The results demonstrated that the pronounced differences in the characteristics of EPS from the fungal and the bacterial sources are responsible for the differential effects on PAH biodegradation.

Collagen is a promising candidate for food and pharmaceutical applications due to its excellent biocompatibility, low antigenicity, and controlled biodegradability; however, its heavy price restricts its utilization. Fish scales generated during the processing are generally regarded as waste material and an environmental pollutant, though they are a promising source of collagen. In the present study, Cirrhinus mrigala scales were demineralized and extracted for acid-soluble collagen (ASC) using acetic acid, with a collagen yield of 2.7%. UV–Vis spectra, SDS-PAGE, FTIR analyses, and amino acid composition confirmed the type I nature of the collagen extracted. The denaturation temperature of the collagen was found to be 30.09 °C using differential scanning calorimetry (DSC). The collagen was highly soluble at acidic pH and lower NaCl concentrations while its solubility was lowered in alkaline conditions and NaCl concentrations above 0.5 M. The collagen exhibited good emulsifying potential with an emulsion activity index (EAI) and emulsion stability index (ESI) of 21.49 ± 0.22 m² g⁻¹ and 15.67 ± 0.13 min, respectively. Owing to the good physicochemical characteristics of the extracted collagen, collagen-chitosan-neem extract (CCN) films were prepared subsequently which showed good antimicrobial activity against Bacillus subtilis NCIM 2635, Staphylococcus aureus NCIM 2654, Escherichia coli NCIM 2832, and Pseudomonas aeruginosa NCIM 5032, suggesting the potential of collagen in the development of antimicrobial films. These results demonstrate that the collagen from fish waste could be valorized and used effectively along with chitosan and neem extract for the synthesis of novel biodegradable films with antimicrobial efficacy.

The epidermal mucus of fish performs diverse functions from prevention of mechanical abrasion to limit pathogen invasions. The current experiment was designed to extract skin mucus proteins of three freshwater fish, i.e. common carp (Cyprinus carpio), mrigal (Cirrhinus mrigala) and rohu (Labeo rohita) with organic solvent (methanol) and dissolve in different pH of Tris-HCl buffers to examine the significance of pH in the solubilisation of skin mucus proteins. The protein profiles of different pH solubilised methanol fish skin mucus extracts were determined by SDS-PAGE. The non-specific immune enzymes, alkaline phosphatase, lysozyme and protease of fish skin mucus were compared and this present study demonstrated that these enzymes differed in their activity depending on pH buffers. The higher lysozyme and protease activity were observed at the pH of 8.0 and higher alkaline phosphatase activity in the pH 9.0 of C. mrigala fish skin mucus methanol extract. In addition, the bactericidal activity was evaluated against the pathogens Proteus vulgaris and Pseudomonas aeruginosa. The pH 8.0 of C. mrigala skin mucus extract revealed better bactericidal activity than other fish species mucus pH buffers against both P. vulgaris and P. aeruginosa. In the case of protein profile from SDS-PAGE, based on pH buffers and the solubilisation of proteins, differences in the resolution of bands were observed. The higher alkaline pH of 9.0 showed smeared gel bands in all the three fish skin mucus methanol extract. The present study suggests that methanol extracted C. mrigala fish skin mucus at pH 8.0 showed better innate immune enzymes and bactericidal activity. The additional examinations of C. mrigala skin mucus methanol extract in this pH aids in identifying novel bioactive molecules. This is the study of proteome of three fish species skin mucus in the effect of pH. Further analyses are required to evaluate proteins present in fish skin mucus extracted with methanol and the influence of pH on protein solubility. These findings could be helpful in exploring natural alternatives to antibiotics in aquaculture industry against infectious pathogens.

In this study, in-house isolated laccase isoforms, i.e., Lac-I and Lac-II of the basidiomycete Pycnoporus sanguineus (CS43), were evaluated in relation to their Remazol Brilliant Blue R (RBBR) dye degradation capacity. A modified Dhouib medium additionally supplemented with 3% ethanol as a secondary inducer was used to propagate P. sanguineus CS43 for enhanced production of laccase under liquid state fermentation. The crude laccase extract was purified by passing through ion exchange diethylaminoethanol (DEAE)-Sepharose and gel filtration-based Sephadex G-200 column chromatography. The purified laccase fractions were subjected to the electrophoresis, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed two laccase isoforms Lac-I and Lac-II with 66 and 68 kDa, respectively. To explore the industrial applicability, for RBBR dye, degradation efficiencies ranged from 82 to 88% after 3 h of incubation for both; Lac-I and Lac-II at both concentrations were recorded. However, with 8 U/mL, the degradation ranged between 70 to 80% during the first 5 min of incubation. Enhanced degradation of RBBR dye was obtained in the presence of violuric acid and N-hydroxypthalamide as laccase mediators. Finally, using RBBR as a substrate kinetic characterization of both Lac-I and Lac-II isoforms was performed that revealed K ₘ (0.243 and 0.117 mM for Lac-I and Lac-II) and V ₘₐₓ (1.233 and 1.012 mM/Sec for Lac-I and Lac-II) values, respectively.

The main objective of this study was to characterize the Giardia duodenalis isolates from Iranian patients in Fars Province, south of Iran by biochemical and molecular methods. Fifteen mass cultivated of G. duodenalis isolates in modified TYI-S-33 medium were analyzed using isoenzyme electrophoresis and PCR genotyping. Polyacrylamide gel electrophoresis (PAGE) of five different enzyme systems was used to characterize isolates: (i) glucose-6-phosphate dehydrogenase, (ii) glucose phosphate isomerase, (iii) malate dehydrogenase, (iv) malic enzyme, and (v) phosphoglucomutase. As well, a fragment of the SSU-rDNA (292 bp) gene was amplified by PCR using the primers RH11 and RH4. The sequencing of the PCR products and phylogenetic tree were performed. The isoenzyme electrophoretic profiles divided fifteen G. duodenalis isolates into four zymodemes. G6PD, GPI, MDH, ME, and PGM enzyme systems showed 1, 2, 2, 3, and 3 enzyme pattern, respectively. G6PD isoenzyme pattern had the most homogeneity, while isoenzyme patterns of ME and PGM had the most heterogeneity in our study. Genotyping results indicated that the zymodemes 1–4 were categorized in assemblage A based on the SSU-rDNA gene. Phylogenetic analysis showed that all four zymodemes were distributed within the cluster of assemblage A. Our results indicated that both isoenzyme and DNA analyses were useful to characterize the isolates of Giardia and distinguishing various zymodemes and assemblages. It could be suggested that the genetic diversity among isoenzymes profiles of G. duodenalis may explain the variable clinical manifestations, pathogenicity, host response, drug susceptibility, and treatment efficacy of human giardiasis.

Cadmium is a toxic element for living organisms. This metal causes different damages to the cell, generating oxidative stress. In order to elucidate cadmium tolerance mechanism and increase tomato plant tolerance by inoculating a Cd-tolerant Burkholderia strain, we analyzed malondialdehyde, hydrogen peroxide content, and the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and glutathione S-transferase of two strains, one isolated from a soil contaminated with Cd (strain SCMS54) and the other from a soil without Cd (strain SNMS32). Strains SNMS32 and SCMS54 exhibited different SOD, CAT, and GR isoenzyme profiles in non-denaturing polyacrylamide gel electrophoresis analysis, with strain SCMS54 exhibiting an extra isoenzyme for all enzymes (Mn-SOD, CAT I, and GR IV, respectively). Despite accumulating more Cd, strain SCMS54 did not increase peroxide hydrogen and presented a fast antioxidant response (increasing SOD and CAT after 5 h of Cd exposure). In this way, strain SCMS54 exhibited a higher metabolic diversity and plasticity when compared to strain SNMS32, so it was selected for Cd–Burkholderia–tomato interaction studies. Inoculated tomato plants in the presence of Cd grew more than non-inoculated plants with Cd indicating that the SCMS54 increased tomato Cd tolerance. It appears that the strain isolated from Cd-contaminated soil (SCMS54) triggers a global stress response in tomato increasing plant tolerance, which may enable plants to be cultivated in Cd-contaminated soils.

In the present work, polyphenol oxidase (PPO) enzyme was purified from potato peel using three-phase partitioning (TPP). In this method, ammonium sulfate and t-butanol were added to precipitate the protein/enzyme from the crude aqueous extract. The PPO enzyme precipitated as an interfacial layer between the upper organic solvent phase and lower aqueous phase. Different purification parameters such as crude extract to t-butanol ratio, ammonium sulfate concentration, temperature, and pH were optimized for TPP. About 69% PPO enzyme activity was recovered in a single step of TPP with 9.2-fold purification. The sodium dodecyl sulfate–polyacrylamide gel electrophoresis profile of partially purified PPO enzyme showed molecular weight in the range of about 30–40 kDa. The PPO enzyme was then investigated for the fabrication of a portable, cost-effective, and disposable colorimetric paper biosensor or colorimetric “test strips” for detection of phenolic contaminants. PPO and a chromophore reagent (3-methyl-2-benzothiazolinone hydrazine) generated a range of color in the presence of phenolic compounds (catechol, phenol, p-cresol, 4-methyl catechol) within 15 min, and limit of detection was found to be 0.5 μM. The biosensor worked in a broad range of pH from 3 to 11 and showed good storage stability at 25 °C and 4 °C for 30 days with no significant loss of activity. The biosensor was also applied on environmental water and urine sample to show reliability of biosensor.

The genetic basis and biochemical aspects of heavy metal endurance abilities have been precisely studied in planktonic bacteria; however, in nature, bacteria mostly grows as surface-attached communities called biofilms. A hallmark trait of biofilm is increased resistance to heavy metals compared with the resistance of planktonic bacteria. A proposed mechanism that contributes to this increased resistance is the enhanced expression of metal-resistant genes. bmtA gene coding for metallothionein protein is one such metal-resistant gene found in many bacterial spp. In the present study, lead (Pb) remediation potential of a biofilm-forming marine bacterium Pseudomonas aeruginosa N6P6 was explored. Biofilm-forming marine bacterium P. aeruginosa N6P6 possess bmtA gene and shows resistance towards many heavy metals, i.e., Pb, Cd, Hg, Cr, and Zn. The expression of metallothionein encoding gene bmtA is significantly high in 48-h-old biofilm culture (11. 4 fold) followed by 24-h-old biofilm culture of P. aeruginosa N6P6 (4.7 fold) (P < 0.05). However, in the case of planktonically grown culture of P. aeruginosa N6P6, the highest expression of bmtA gene was observed in 24-h-old culture. The expression of bmtA also increased significantly with increase in Pb concentration up to 800 ppm. CSLM analysis indicated significant reduction in the raw integrated density of biofilm-associated lipids and polysaccharides (PS) of P. aeruginosa N6P6 biofilm grown in Pb (sub-lethal concentration)-amended medium (P < 0.05), whereas no significant reduction was observed in the raw integrated density of EPS-associated protein. The role of bmtA gene as Pb(II)-resistant determinant was characterized by overexpressing the bmtA gene derived from P. aeruginosa N6P6 in Escherichia coli BL21(DE3). ESI-MS and SDS-PAGE analyses validated the presence of 11.5-kDa MT protein isolated from Pb(II)-induced recombinant E. coli BL21(DE3) harboring bmtA gene.