Synergistic impact of two autochthonous saprobic fungi (A. niger and T. pseudokoningii) on the growth, ionic contents, and metals uptake in Brassica juncea L. and Vigna radiata L. under tannery solid waste contaminated soil.

Unrestricted disposal of tannery solid waste (TSW) into agricultural soils has resulted in the contamination of heavy metals (HMs) such as chromium (Cr) cadmium (Cd), Copper (Cu), and Zinc (Zn) along with the severe potential to degrade the environmental quality around the world. In the present study, a combined phyto- and myco-remediation strategy was evaluated to enhance the growth, ionic contents, and phytoextraction potential of Brassica juncea and Vigna radiata for HMs from TSW-contaminated soil. A pot experiment was conducted in the greenhouse using single or combined inoculation of Trichoderma pseudokoningii (Tp) and Aspergillus niger (An) in B. juncea and V. radiata under TSW-contaminated soil at different doses (0, 50, and 100%). The results showed that the growth parameters of both B. juncea and V. radiata were severely affected under 50 and 100% TSW treatment. The combined inoculation of both the fungal species ameliorated the positive impacts of 50 and 100% TSW application on growth and ionic contents accumulation in B. juncea and V. radiata. The combined application of An + Tp at 100% TSW enhanced the shoot length (87.8, 157.2%), root length (123.9, 120.6%), number of leaves (184.2, 175.0%), number of roots (104.7, 438.9%), and dry weight (179.4, 144.8%) of B. juncea and V. radiata, respectively as compared to control with any fungal treatment at 100% TSW. A single application of An at different doses of TSW enhanced the metal concentration in B. juncea, whereas Tp increased the concentration of the metals in V. radiata. The concentration of Cr in roots (196.2, 263.8%), shoots (342.4, 182.2%), Cu in roots (187.6, 137.0%), shoots (26.6, 76.0%), Cd in roots (245.2, 184.6%), shoots (142.1, 73.4%), Zn in roots (73.4, 57.5%), shoots (62.9, 57.6%), in B. juncea were increased by the application of An at 50 and 100% treatment levels of TSW, respectively compared to control (C). Moreover, the HMs (Cr, Cu, Cd, and Zn) uptake was also improved under 50 and 100% TSW with the combined inoculation of Tp + An in both B. juncea and V. radiata. In conclusion, the combined inoculation of Tp + An was more effective in metal removal from TSW-treated soil.NOVELTY STATEMENTLimited studies have been conducted on filamentous fungi systematically under metal-contaminated sites for their diversity, metal tolerance, and their potential in enhancing the phytoremediation potential of different crop plants.In the present study, single and/or combined inoculation of fungal strains was found effective in alleviating different metals stress in tannery solid waste contaminated soil by improving defense mechanisms and plant growth due to the association between fungal strains and plants.The combined application of both fungal strains had an additive effect in enhancing the bioaccumulation capacity of B. juncea and V. radiata compared to their single inoculation.

Metabolic profile and molecular characterization of endophytic bacteria isolated from Pinus sylvestris L. with growth-promoting effect on sunflower.

Endophytic bacteria inhabit plant tissues such as roots, stems, leaves, fruits, and seeds and can multiply inside plant tissue without damaging them. This study involves the isolation, characterization, metabolic profiling, and effect of endophytic bacteria isolated from the roots of Scots pine (Pinus sylvestris), on the growth of sunflower. In the current study, fifteen isolates of endophytic bacteria were obtained from the roots of Scots pine, and their molecular characterization was performed using 16 s rRNA ribotyping. The molecular characterization revealed that the strains belonged to Bacillus spp., Pseudomonas spp., Micrococcus sp., Serratia sp., Enterobacter sp., Pantoea sp., Staphylococcus sp., and Microbacterium sp. Among the isolated strains, 9 strains showed positive results for ammonium production, 12 strains for calcium solubilization, 11 strains for magnesium solubilization, 5 strains for zinc solubilization, 12 strains for phosphate solubilization, 8 strains for potassium solubilization, 10 strains for indole acetic acid (IAA) production, 9 strains for siderophore, and 6 strains for hydrogen cyanide (HCN) production. The greenhouse experiment results demonstrated that all isolated endophytic bacteria improved the shoot length, dry weight, and chlorophyll content of sunflower, whereas a significant increase was observed by PS-3 (Bacillus cereus), PS-6 (Serratia marcescens), and PS-8 (Pseudomonas putida). Besides, the concentration of nitrogen, phosphorus, and potassium were also measured in sunflower shoots, and results asserted that bacterial inoculation increased the bioavailability of these essential nutrients to plants compared to uninoculated control. Thus, these endophytic bacteria could be used as an encouraging option to improve plant growth and performance.

Autochthonous strains of Trichoderma isolated from tannery solid waste improve phytoextraction potential of heavy metals by sunflower.

This research work was aimed at isolating and demonstrating the significant potential of autochthonous fungi for phytoextraction of hazardous metals in metal polluted soil using Helianthus annuus. Four multi-metal resistant strains of Trichoderma were selected from a total of 21 strains isolated from tannery polluted soil and tannery solid waste. Autochthonous Trichoderma strains were used singly and in the form of consortium (TC). Sunflower was grown in pots for 90 days having eight different amendments of tannery polluted soil with and without Trichoderma inoculation. Growth and biochemical attributes of the plants were observed along with metal content extract by different plant parts. The results revealed that TC enhanced shoot length, shoot dry weight, and metal uptake as compared to single specie inoculation. Similarly, BCF (72.8-118.23%) and TF were significantly pronounced in shoots of H. annuus grown with TC at 40% amended soil. The biochemical analysis of the plants showed that Trichoderma strains boosted the enzymatic (catalase, peroxidase, and superoxide dismutase) antioxidants in the plants. The use of indigenous fungi with metal accumulating plants like sunflower can help to alleviate metal contamination from industrial sites and can make the soil cultivable for energy crops.

The genus Trichoderma is among the most common cosmopolitan soil fungi that enhance phytoextraction capability of plants. Hence, the isolation and identification of diversified and potent Trichoderma strains from contaminated environments is the need of the hour for broad spectrum applications in bioremediation. In the present study, contaminated soil mycoflora was explored and multi-metal resistant strains of Trichoderma were isolated. Their application in myco-assisted phytoextraction with Helianthus annuus was assessed to analyze their impact on the metal removal efficacy and enhancing growth in highly contaminated soil.

Role of composted tannery solid waste and its autochthonous microbes in enhancing phytoextraction of toxic metals and stress abatement in sunflower.

The excessive concentration of multiple heavy metals in the tannery solid waste (TSW) needs integrated process solutions for its decontamination. This study is aimed at deriving TSW compost and autochthonous microbe synergies for improving phytoextraction potential of sunflower. In-vessel composting of TSW was carried out by using fruit waste as an inoculum to achieve the optimized conditions. Autochthonous strains of Trichoderma viride and Bacilllus sp. isolated from TSW were utilized individually as well as in combination with TSWC amendments of 2.5, 5 and 10% (w/w) prepared in our pilot scale experiment. Analyses of TSW compost based on FTIR and SEM illustrated the wide range of functionality and porosity along the mesh of fungal hyphae and inorganic moieties present on the compost surface. Plant biomass and TMs uptake (Cr 540 mg kg-1 > Cd 330 mg kg-1 > Pb 285 mg kg-1) were significantly pronounced in shoots of sunflower under combined treatments at 10% TSWC amended soils. However, in seeds, TMs were found below detection limit (BDL) through atomic absorption spectrophotometry. Biochemical assays of sunflower including total chlorophyll content (18%), total soluble protein (45%), superoxide dismutase (80%) and catalase (75%) activities were also increased significantly at higher level of amendment in combination with microbes than in the control. Despite being high in TMs, high biomass in sunflower and associated elevation in biochemical products demonstrate the potential of TSW for valorization.Novelty statement: This study identifies the cost-effective management of multi metal contaminated tannery solid waste through deriving its compost along with autochthonous microbes as phytoextraction assistants by yielding higher plant biomass. This study suggests the use of composted TSW inoculated with selected autochthonous fungi and bacteria for enhancing sunflower's biomass and enhancing the bioavailable fractions of toxic metals for phytoextraction.

Equilibrium kinetics and thermodynamic studies on biosorption of heavy metals by metal-resistant strains of Trichoderma isolated from tannery solid waste.

This study was aimed at finding the metal sorption potential of six indigenous Trichoderma strains by using batch experiments for Cd (II), Cr (VI), Cu (II), and Pb (II). Trichoderma atrobrunneum showed maximum metal biosorption potential at 800 mg L-1 of initial concentration. Two adsorption isotherm models, (1) Langmuir (2) Freundlich models, were employed on the biosorption data obtained at various initial metal concentrations (10 mg L-1-200 mg L-1) and pseudo-first (PSI) and pseudo-second (PSII) order equilibrium kinetic models were subjected to data of agitation time (3-7 days). A maximum correlation coefficient value (R2) of ≤ 1 was observed for the Langmuir and PSII model. Results revealed that pH 6-7 was the best for metal sorption, while metal removal efficiency was increased by increasing temperature (298 K, 303 K, 308 K, 313 K). The results of thermodynamic study parameters (∆G°, ∆H°, ∆S°) indicated that heavy metal biosorption by Trichoderma strains was an endothermic, spontaneous, and feasible process. Moreover, surface characterization analysis through SEM, BET, FTIR, and XRD showed that T. atrobrunneum and Trichoderma sp. could adsorb more metal ions when grown in high metal concentrations. The results indicate that living biomass of T. atrobrunneum and Trichoderma sp. is an effective multi-metal biosorbent that can be used for efficacious bioremediation of bio-treatment of heavy metal polluted wastewater.

Biosorption potential and molecular characterization of metal-resistant autochthonous microbes from tannery solid waste.

This study encompasses isolation and screening of heavy metal-resistant fungal and bacterial strains from tannery solid waste (TSW). Twelve fungal strains and 25 bacterial strains were isolated from TSW. The growth of fungal strains was observed against different heavy metals ranging from 10 to 1050 mg L-1 and the growth of bacteria was observed in metal concentrations ranging from 10 to 1200 mg L-1. Five multi-metal-resistant fungal isolates belonging to the genus Trichoderma and ten bacterial isolates belonging to the genus Bacillus showed good metal resistance and biosorption potential. They were identified through molecular techniques, fungi based on ITS region ribotyping, and bacteria based on 16S rRNA ribotyping. The fungal strains were characterized as T. hamatum (TSWF-06), T. harzianum (TSWF-11), T. lixii (TSWF-02), and T. pseudokoningii (TSWF-03, TSWF-10). The bacterial strains were characterized as Bacillus xiamenensis (TSW-02), B. velezensis (TSW-05), B. piscis (TSW-06), B. safensis (TSW-10), B. subtilis (TSW-14, TSW-15, TSW-17) B. licheniformis (TSW-19), B. cereus (TSW-20), and B. thuringiensis (TSW-22). The fungal strains, namely, T. pseudokoningii (TSWF-03) and T. harzianum, proved to be two multi-metal-resistant strains with good biosorption efficiency. Unlike fungi, bacterial strains showed metal-specific resistance. The strains Bacillus xiamenensis, B. subtilis (TSW-14), and B. subtilis (TSW-15) showed good biosorption efficiency against Cr, B. safensis against Cu, B. piscis, and B. subtilis (TSW-17) against Pb and B. licheniformis and B. thuringiensis against Zn. The autochthonous fungal and bacterial strains can therefore be employed to clean metal-contaminated environments.

Application of microbe-impregnated tannery solid waste biochar in soil enhances growth performance of sunflower.

Synergistic effect of biochar and microbes in soil enhances performance of plants. Hazardous tannery solid waste can be reduced by one-third in volume by conversion to biochar. A greenhouse trial was set up with soil having different doses of metal resistant microbe-impregnated biochar (MIBC) prepared from tannery solid waste. Consortia of autochthonous strains of Trichoderma and Bacillus were inoculated on BC and the behavior and fate of metals were evaluated for their bioavailability to sunflower. Sunflower was grown in pots for 80 days having six different amendments of tannery solid waste biochar (0-10% w/w) with and without Trichoderma and Bacillus consortia and its morphological and biochemical attributes as well as metal uptake were observed. The results illustrated that application of BC at 2% rate without inoculation increased the shoot length and dry biomass by 19.8% and 77.4%, respectively, while plant growth and performance were reduced at higher amendments of BC. However, application of MIBC with Trichoderma or/and Bacillus consortium significantly improved the plant attributes at all levels of amendment. The results indicated that MIBC having Trichoderma and Bacillus consortia at 10% rate increased shoot length and dry biomass by 65.3% and 516% compared to control without BC. Application of BC without inoculation reduced the uptake of Cu, Fe, and Ni and increased the mobilization of all other metals for uptake in sunflower. Mobilization and uptake of Cd, Cr, Cu, Ni, Pb, and Zn decreased with MIBC having Trichoderma and Bacillus consortia whereas that of Fe and Mg were noted. A considerable decrease in proline and total phenolic content was demonstrated by MIBC-grown sunflower. The data of metal fractionation in BC also supported the above findings. Therefore, MIBC can be used as a promising option for enhancing growth performance and ensuring the physiological safety of sunflower as an energy crop.

Fungal biostimulant-driven phytoextraction of heavy metals from tannery solid waste contaminated soils.

Two of the multiple limitations of phytoextraction efficiency (PE %) of TSW polluted soils are: (i) low growth of plant performance, (ii) poor bioavailability of excessive essential and heavy metals (ascribed as Category-I and II metals respectively) The current study reports biostimulant role of allochthonous Trichoderma harzianum (F1) and autochthonous Trichoderma pseudokoningii (F2) in growth of Tagetes patula L. and uptake of Category-I & II metals from TSW-soil (0, 5 & 10%). Significantly higher growth (27.5-47.8% dry wt. than Control) and highly significantly higher uptake of Category-I & II metals (72-80% Ca, 32-69% K, 72-76% Na & 73-86% Cd, 63-100% Cr, 72-77% Cu, 73-78% Fe, 43-77% Mg, 22-33% Ni, 70-73% Zn) was observed in T. patula applied with F1 + F2 treatment. The PE (%) parameters viz.specific extraction yield, tolerance and translocation index of Category-I & II metals were higher in plants cultivated on fungal inoculated TSW:soil. The Trichoderma spp. acted as strong biostimulants for enhancing plant growth and conc. of catalase (CAT, 44-52% than control), superoxide dismutase (SOD, 37-43%), soluble proteins (37-68%) and total chlorophyll (10-26%) in T. patula during metal phytoextraction of TSW:soil. Novelty statement Due to multiple socio-economic constraints for effective management of tannery solid waste (TSW), the heavy metal phytoextraction seems to be one of the promising approaches. However, due to complex composition of TSW, that is, with more than 37 components, high pH, multiple types and high conc. of metals; there lies huge challenge of enhancing phytoextraction efficiency (PE %). This can be done by enhancing growth of hyperaccumulator plants and increasing bioavailable fraction of metals. The current study suggests application of selected fungal biostimulants for increasing growth of T. patula while improving bioavailable fraction of the total metal contents of the TSW: soil.

Zinc-lysine prevents chromium-induced morphological, photosynthetic, and oxidative alterations in spinach irrigated with tannery wastewater.

Anthropogenic activities have resulted in severe environmental degradation. Untreated wastewater from tanneries is hazardous to all kinds of life on earth. Effluent from tanning industries, containing large amount of Cr, is used to irrigate the crops in Pakistan. The current experiment was carried out to study the effects of tannery wastewater on spinach and the role of lysine-Zn in mitigating the severity of stress. The plants were grown in soil and the following treatments were used: irrigation with 0%, 33%, 66%, and 100% wastewater (ww) along with two doses (0 mM, 10 mM) of Zn-lysine. Foliar application of zinc-lysine enhanced the plant growth, biomass, Zn contents, photosynthesis, and enzyme activities in different tissues of plant. Zinc-lysine (10 mM) considerably decreased the Cr content in roots and shoots, along with ameliorating the oxidative stress by enhancing the activities of antioxidant enzymes in plants. Addition of Zn-lys (10 mM) improved the plant height by 19%, root length by 57%, leaf dry weight by 19%, and root dry weight by33% under 100% Cr treatment. Zn-lys significantly reduces the oxidative stress and concentration of Cr as compared with the Cr treatments alone. Application of Zn-lys (10 mM) reduced the Cr contents in roots by 27 and 22 under 33 and 66% Cr treatment, respectively. Taken together, Zn-lys chelates efficiently ameliorated the toxic effects of chromium. Zn-lysine has the extravagant potential of mitigating the heavy metal toxicity without harming the normal growth and development of the plants.

Modeling of Cr contamination in the agricultural lands of three villages near the leather industry in Kasur, Pakistan, using statistical and GIS techniques.

Kasur is one of the hubs of leather industry in the Punjab, Pakistan, where chrome tanning method of leather processing is extensively being used. Chromium (Cr) accumulation levels in the irrigation water, soil, and seasonal vegetables were studied in three villages located in the vicinity of wastewater treatment plant and solid waste dumping site operated by the Kasur Tanneries Waste Management Agency (KTWMA). The data was interpreted using analysis of variance (ANOVA), clustering analysis (CA), and principal component analysis (PCA). Interpolated surface maps for Cr were generated using the actual data obtained for the 30 sampling sites in each of the three villages for irrigation water, soil, and seasonal vegetables. The level of contamination in the three villages was directly proportional to their distance from KTWMA wastewater treatment plant and the direction of water runoff. The highest level of Cr contamination in soil (mg kg-1) was observed at Faqeeria Wala (37.67), intermediate at Dollay Wala (30.33), and the least in Maan (25.16). A gradational variation in Cr accumulation was observed in the three villages from contaminated wastewater having the least contamination level (2.02-4.40 mg L-1), to soil (25.16-37.67 mg kg-1), and ultimately in the seasonal vegetable crops (156.67-248.33 mg kg-1) cultivated in the region, having the highest level of Cr contamination above the permissible limit. The model used not only predicted the current situation of Cr contamination in the three villages but also indicated the trend of magnification of Cr contamination from irrigation water to soil and to the base of the food chain. Among the multiple causes of Cr contamination of vegetables, soil irrigation with contaminated groundwater was observed to be the dominant one.

Differential mobilization and metal uptake versus leaching in multimetal soil columns using EDTA and three metal bioaccumulators.

Tannery waste is a major environmental concern that needs proper management. Tannery solid waste (TSW) can be added to the soil as an organic amendment but needs metal removal. Chelant-assisted phytoremediation hastens the process of metal removal but also poses risk of leaching at the same time. This research evaluates Ethylenediaminetetraacetic acid (EDTA)-assisted phytoextraction and associated leaching hazard using metal-tolerant plants. Greenhouse trials were carried out with sunflower, spinach, and marigold using columns of uniform diameter packed with field soil and multimetal contamination of TSW (5% and 10%) with four EDTA doses. The amounts of metal absorbed or leached conformed to amounts in the soil amendment and the dose of EDTA. The mobilization of metals by EDTA was however metal-specific. The metals that were extracted in greater amounts by the plants were leached less compared to Cr and Cu. A significant amount of other metals was leached down and thus less amount was phytoextracted by the plants e.g. Cd and Ni. A high correlation was observed between the amount of metal absorbed by the plant and the amount in leachate except for Cr in all the plants. Antioxidant activities like SOD and catalase were also found to be high in sunflower and spinach.