Effects of pyroligneous acid on acute, chronic, and cyto-genotoxicity to earthworms (Eisenia fetida).

The pyroligneous acid (PA), or wood vinegar, is a byproduct of wood carbonization during the slow pyrolysis process. PA is recognized globally as a safe compound for agriculture due to its various beneficial properties, such as antioxidant, antibacterial, antifungal, and termiticidal properties. However, the impact of different PA concentrations on beneficial soil organisms, such as earthworms has not been investigated. The present study aims to understand the effects of different PA concentrations on earthworm Eisenia fetida. The earthworms were exposed to nine different concentrations of PA in soils, including their control. The acute toxicity assay was performed after 14 days of exposure, and the chronic toxicity assay was performed up to 8 weeks after exposure. The results from the acute toxicity assay demonstrated no significant effect on earthworm mortality. The chronic toxicity assay showed that lower PA concentrations (0.01-0.2% of weight/weight PA in soil) promoted cocoon and juvenile production in soils, whereas higher PA concentrations (0.5 and 1%) had a negative effect. These findings highlight the potential of PA to enhance soil fertility at lower concentrations, up to 0.2%, by stimulating worm activity and subsequent manure production. The outcomes of this study have significant implications for the careful management of PA concentrations within agricultural operations.

Investigation on removal of perfluorooctanoic acid (PFOA), perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHxS) using water treatment sludge and biochar.

This work assessed the adsorption performance of three common PFAS compounds (PFOA, PFOS and PFHxS) on two water treatment sludges (WTS) and two biochars (commercial biomass biochar and semi-pilot scale biosolids biochar). Of the two WTS samples included in this study, one was sourced from poly-aluminium chloride (PAC) and the other from alum (Al2(SO4)3). The results of experiments using a single PFAS for adsorption reinforced established trends in affinity - the shorter-chained PFHxS was less adsorbed than PFOS, and the sulphates (PFOS) were more readily adsorbed than the acid (PFOA). Interestingly, PAC WTS, showed an excellent adsorption affinity for the shorter chained PFHxS (58.8%), than the alum WTS and biosolids biochar at 22.6% and 41.74%, respectively. The results also showed that the alum WTS was less effective at adsorption than the PAC WTS despite having a larger surface area. Taken together, the results suggest that the hydrophobicity of the sorbent and the chemistry of the coagulant were critical factors for understanding PFAS adsorption on WTS, while other factors, such as the concentration of aluminium and iron in the WTS could not explain the trends seen. For the biochar samples, the surface area and hydrophobicity are believed to be the main drivers in the different performances. Adsorption from the solution containing multiple PFAS was also investigated with PAC WTS and biosolids biochar, demonstrating comparable performance on overall adsorption. However, the PAC WTS performed better with the short-chain PFHxS than the biosolids biochar. While both PAC WTS and biosolids biochar are promising candidates for adsorption, the study highlights the need to explore further the mechanisms behind PFAS adsorption, which could be a highly variable source to understand better the potential for WTS to be utilized as a PFAS adsorbent.

Efficient bioremediation of laboratory wastewater co-contaminated with PAHs and dimethylformamide by a methylotrophic enrichment culture.

A methylotrophic enrichment culture, MM34X, has been assessed for its exceptional ability in biodegradation of dimethylformamide (DMF) and bioremediation of laboratory wastewater (LWW) co-contaminated with polycyclic aromatic hydrocarbons (PAHs). The culture MM34X tolerated high concentrations of DMF and efficiently degraded 98% of 20,000 mg L-1 DMF within 120 h. LWW bioremediation was performed in stirred bottle laboratory-scale bioreactor. After 35 days of incubation, 2760.8 ± 21.1 mg L-1 DMF, 131.8 ± 9.7 mg L-1 phenanthrene, 177.3 ± 7.5 mg L-1 pyrene and 39.5 ± 2.7 mg L-1 BaP in LWW were removed. Analysis of post-bioremediation residues indicated the absence of any known toxic intermediates. The efficacy of bioremediation was further evaluated through cyto-genotoxicity assays using Allium cepa. The roots of A. cepa exposed to bioremediated LWW showed improved mitotic index, whereas original LWW completely arrested cell growth. Similarly, the alkaline comet assay indicated alleviation of genotoxicity in bioremediated LWW, as evidenced by significantly lower DNA damage in terms of tail DNA and Olive tail moment. In addition, oxidative stress assays, performed using fluorescent probes 2',7'-dichlorodihydrofluorescein diacetate, C11-BODIPY and dihydrorhodamine 123, revealed significant mitigation of oxidative stress potential in bioremediated LWW. Our findings suggest that the enrichment MM34X may prime the development of inexpensive and efficient large-scale bioremediation of LWW co-contaminated with PAHs and DMF.

Cyto-genotoxicity evaluation of pyroligneous acid using Allium cepa assay.

Pyroligneous acid (PA) is a highly oxygenated organic condensate obtained by cooling the gases generated from the pyrolysis process. PA has been used in agriculture for several years with multiple beneficial effects, including plant health and yields, pest resilience, and seed germination. It is generally applied to agricultural soils in the dilution of 1:1000 to 1:100, corresponding to 0.1-1% PA concentration. In this study, the cyto-genotoxic potential of PA to Allium cepa meristematic root-tips (where all cells undergo repeated division and form primary root tissues) was examined. Exposure to PA concentrations of 0.1% and above showed a reduction in the mitotic index percentage, and at 5%, a complete arrest in the cell division was recorded. However, chromosomal aberrations at 0.5, 1, and 3% PA were reversible types such as bridges, vagrants, laggards, and multipolar anaphase, with a maximum of only 5.8% chromosomal aberration observed at 3% PA. Comet assay (single-cell gel electrophoresis) for genotoxicity assessment determined using PA exposed A. cepa root tips showed that it was not genotoxic. The absence of cyto-genotoxicity in A. cepa, even at concentrations far above what would be typically encountered in agricultural applications, strongly suggests that PA is unlikely to cause adverse effects on crops and ultimately on the biota and human health.

Perfluorooctane sulfonate (PFOS) induces several behavioural defects in Caenorhabditis elegans that can also be transferred to the next generations.

Perfluorooctane sulfonate (PFOS) is a well-known global persistent organic pollutant of grave concern to ecological and human health. Toxicity of PFOS to animals and humans are well studied. Although few studies have reported the behavioral effect of PFOS on nematode Caenorhabditis elegans, it's transgenerational effects were seldom studied. Therefore, we investigated the toxicity of PFOS on several behavioral responses besides bioaccumulation and transgenerational effects in C. elegans. In contrast to the several published studies, we used lower concentrations (0.5-1000 µg/L or 0.001-2.0 µM) that are environmentally relevant and reported to occur close to the contaminated areas. The 48 h median lethal concentration of PFOS was found to be 3.15 µM (1575 µg/L). PFOS (≥0.01 µM) caused severe toxicity to locomotion, and this effect was even transferred to progeny. However, after a few generations, the defect was rectified in the progeny of single-time exposed parent nematodes. Whereas, continuous exposure at 0.001 µM PFOS, no visible defects were observed in the progeny. PFOS (≥0.01 µM) also significantly decreased the brood size in a concentration-dependent manner. Besides, lifespan was affected by the higher concentration of PFOS (≥1.0 µM). These two behavioral endpoints, lifespan and reproduction defects, became less severe in the progeny. Chemotaxis plasticity was also significantly retarded by ≥ 1.0 µM PFOS compared to the control group. Results indicate that PFOS can exert severe neurobehavioral defects that can be transferred from parents to their offspring. The findings of this study have significant implications for the risk assessment of perfluorinated substances in the environment.

Exposure to perfluorooctanesulfonate (PFOS) but not perflurorooctanoic acid (PFOA) at ppb concentration induces chronic toxicity in Daphnia carinata.

Widespread environmental contamination of per- and polyfluoroalkyl substances (PFAS) is well established. Nevertheless, few studies have reported on the aquatic toxicity of PFAS, especially in indicator species such as Daphnia. In this study, the toxicity of two major PFAS, namely perfluorooctanoic acid (PFOA) and perfluorooctanesulfonate (PFOS), was investigated on water flea (Daphnia carinata) using a battery of comprehensive toxicity tests, including a 48 h acute and a 21-day chronic assays. The survival, growth, and reproduction of D. carinata were monitored over a 21-day life cycle. PFOS exhibited higher toxicity than PFOA. The 48 h LC50 values (confidence interval) based on acute toxicity for PFOA and PFOS were 78.2 (54.9-105) mg L-1 and 8.8 (6.4-11.6) mg L-1, respectively. Chronic exposure to PFOS for 21 days displayed mortality and reproductive defects in D. carinata at a concentration as low as 0.001 mg L-1. Genotoxicity assessment using comet assay revealed that exposure for 96 h to PFOS at 1 and 10.0 mg L-1 significantly damaged the organism's genetic makeup. The results of this study have great implications for risk assessment of PFOS and PFOA in aquatic ecosystems, given the potential of PFOS to pose a risk to Daphnia even at lower concentrations (1 µg L-1).

Evaluation of Cyto-genotoxicity of Perfluorooctane Sulfonate (PFOS) to Allium cepa.

Per- and polyfluoroalkyl substances (PFAS) have emerged as contaminants of global concern. Among several PFAS, perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) are persistent and bioaccumulative compounds. We investigated the cyto-genotoxic potential of PFOS to Allium cepa root meristem cells. The A. cepa root tips were exposed to 6 different concentrations (1-100 mg L-1 ) of PFOS for 48 h. Reduction in mitotic index and chromosomal aberrations was measured as genotoxic endpoints in meristematic root cells. Exposure to PFOS significantly affected cell division by reducing the miotic index at higher concentrations (>10 mg L-1 ). The median effect concentration of PFOS to elicit cytotoxicity based on the mitotic index was 43.2 mg L-1 . Exposure to PFOS significantly increased chromosomal aberrations at concentrations >25 mg L-1 . The common aberrations were micronuclei, vagrant cells, and multipolar anaphase. The alkaline comet assay revealed a genotoxic potential of PFOS with increased tail DNA percentage at concentrations >25 mg L-1 . To our knowledge, this is the first study to report the cyto-genotoxic potential of PFOS in higher plants. Environ Toxicol Chem 2021;40:792-798. © 2020 SETAC.

Rhizodegradation of PAHs differentially altered by C3 and C4 plants.

Pyrosequencing of 16S ribosomal RNA (rRNA) was employed to characterize bacterial communities colonizing the rhizosphere of plants with C3 and C4 photosynthetic pathways grown in soil contaminated with polycyclic aromatic hydrocarbons (PAHs) after 60 and 120 days. The results of this study exhibited a clear difference in bacterial diversity between the rhizosphere and non-rhizosphere samples and between the rhizospheres of the C3 and C4 plants after 120 days. In both C3 and C4 rhizospheres, an incremental change in PAHs degrading bacterial genera was observed in the 120th day samples compared to the 60th day ones. Among the PAHs degrading bacterial genera, Pseudomonas showed good resistance to PAHs in the 120th day rhizosphere of both C3 and C4 plants. Conversely, the genus Sphingomonas showed sensitivity to PAHs in the 120th day rhizosphere soils of C3 plants only. Also, a significant increase in the PAHs degrading genera was observed at 120th day in the C4 rhizosphere in comparison to the C3 rhizosphere, which was reflected in a reduced PAHs concentration measured in the soil remediated with C4 plants rather than C3 plants. These results suggest that the rhizoremediation of PAHs was primarily governed by the plant photosystems, which led to differences in root secretions that caused the variation in bacterial diversity seen in the rhizospheres. This study is the first report to demonstrate the greater effectiveness of C4 plants in enhancing the PAHs degrading bacterial community than C3 plants.

Phytoremediation efficacy assessment of polycyclic aromatic hydrocarbons contaminated soils using garden pea (Pisum sativum) and earthworms (Eisenia fetida).

Endpoint assessment using biological systems in combination with the chemical analysis is important for evaluating the residual effect of contaminants following remediation. In this study, the level of residual toxicity of polycyclic aromatic hydrocarbons (PAHs) after 120 days of phytoremediation with five different plant species:- maize (Zea mays), Sudan grass (Sorghum sudanense), vetiver (Vetiveria zizanioides), sunflower (Helianthus annuus) and wallaby grass (Austrodanthonia sp.) has been evaluated by ecotoxicological tests such as root nodulation and leghaemoglobin assay using garden pea (Pisum sativum) and acute, chronic and genotoxicity assays using earthworm (Eisenia fetida). The phytoremediated soil exhibited lesser toxicity supporting improved root nodulation and leghaemoglobin content in P. sativum and reducing DNA damage in E. fetida when compared to contaminated soil before remediation. Also, the results of the ecotoxicological assays with the legume and earthworm performed in this study complemented the results obtained by the chemical analysis of PAHs in phytoremediated soil. Therefore, these findings provide a basis for a framework in which remediation efficacy of PAHs-contaminated sites can be evaluated effectively with simple ecotoxicological bioassays using legumes and earthworms.

Low molecular weight organic acids enhance the high molecular weight polycyclic aromatic hydrocarbons degradation by bacteria.

The biodegradation potential of three bacterial cultures isolated from the rhizosphere of maize (Zea mays) and Sudan grass (Sorghum sudanense) grown in PAHs contaminated soils to degrade benzo[a]pyrene (BaP) and pyrene (PYR) was assessed. Of the three bacterial cultures isolated, two belonged to Gram-positive bacteria of phylum Actinobacteria namely Arthrobacter sp. MAL3 and Microbacterium sp. MAL2. The Gram-negative bacterial culture was Stenotrophomonas sp. MAL1, from the phylum Proteobacteria. The cultures were grown in the presence of BaP and PYR as sole carbon sources and with the addition of low molecular weight organic acids (LMWOAs) mixture. After 10-14 days of exposure, all the bacterial isolates exhibited a complete degradation of PYR with the addition of LMWOAs mixture, whereas only 38.7% of BaP was degraded by Stenotrophomonas sp. MAL1 with the addition of LMWOAs mixture. In addition, enhanced PAHs biodegradation by bacterial culture was observed when the PAHs present as mixture (BaP + PYR) with the addition of LMWOAs. Dioxygenase genes were detected in Stenotrophomonas sp. MAL1 (phnAC), and Arthrobacter sp. MAL3 (nidA and PAH-RHDα). Therefore, this study provides new insights on the influence of LMWOAs in enhancing the degradation of high molecular weight (HMW) PAHs in soil by rhizosphere bacterial cultures.

Metabolomics reveals defensive mechanisms adapted by maize on exposure to high molecular weight polycyclic aromatic hydrocarbons.

Polycyclic aromatic hydrocarbons are an important group of persistent organic pollutants. Using plants to remediate PAHs has been recognized as a cost-effective and environmentally friendly technique. However, the overall impact of PAHs on the regulation of plant metabolism has not yet been explored. In this study, we analyzed the alteration in the maize (Zea mays L.) metabolome on exposure to high molecular weight PAHs such as benzo[a]pyrene (BaP) and pyrene (PYR) in a hydroponic medium, individually and as a mixture (BaP + PYR) using GC-MS. The differences in the metabolites were analyzed using XCMS (an acronym for various forms (X) of chromatography-mass spectrometry), an online-based data analysis tool. A significant variation in metabolites was observed between treatment groups and the unspiked control group. The univariate, multivariate and pathway impact analysis showed there were more significant alterations in metabolic profiles between individual PAHs and the mixture of BaP and PYR. The marked changes in the metabolites of galactose metabolism and aminoacyl tRNA biosynthesis in PAHs treated maize leaves exhibit the adaptive defensive mechanisms for individual and PAHs mixture. Therefore, the metabolomics approach is essential for an understanding of the complex biochemical responses of plants to PAHs contaminants. This knowledge will shed new light in the field of phytoremediation, bio-monitoring, and environmental risk assessment.

Comparison of plants with C3 and C4 carbon fixation pathways for remediation of polycyclic aromatic hydrocarbon contaminated soils.

The phytoremediation technique has been demonstrated to be a viable option for the remediation of polycyclic aromatic hydrocarbons (PAHs) contaminated sites. This study evaluated the potential applicability of plants with C3 and C4 carbon fixation pathways for the phytoremediation of recalcitrant high molecular weight (HMW) PAHs contaminated soil. A 60 and 120-day greenhouse study was conducted which showed higher degradation of HMW PAHs in soil grown with C4 plants when compared to C3 plants. Also, no PAHs were detected in the maize cobs, sunflower, wallaby, and Sudan grass seeds at the end of the experiment. The effect of plants in modifying the microbial community and dynamics in the rhizosphere was also examined by measuring soil biochemical properties such as dehydrogenase activity and water-soluble phenols. The results demonstrate a substantial difference in the microbial populations between planted and unplanted soils, which in turn facilitate the degradation of PAHs. To the best of our knowledge, this study for the first time evaluated the phytoremediation efficacy through the A. cepa cyto- and genotoxicity assay which should be considered as an integral part of all remediation experiments.

Impact of plant photosystems in the remediation of benzo[a]pyrene and pyrene spiked soils.

The phytoremediation potential of 14 different plant species belonging to C3 and C4 carbon fixation pathway for soils spiked with polycyclic aromatic hydrocarbons (PAHs) such as benzo[a]pyrene (B[a]P) and pyrene (PYR) was investigated. A glasshouse experiment was conducted to measure the changes in morphological, physiological, biochemical parameters and the bioaccumulation and biodegradation ability of the plants in soils spiked with 48 and 194 mg kg-1 of B[a]P and PYR, respectively. The per cent removal efficacy of B[a]P and PYR by the tested plant species over a period of 50 days was from 6 to 26% and 14 to 40% respectively. The maximum removal of both B[a]P and PYR was observed in Sudan grass (C4), vetiver (C4), maize (C4), and sunflower (C3). In terms of accumulation in root and shoot, the concentration of PYR was higher in both C3 and C4 plant species when compared to B[a]P. Overall the results indicated that C4 plants were more efficient than their C3 counterparts in terms of morphological, physiological, biochemical and degradation ability of PAHs.