The desert woodrat (Neotoma lepida) induces a diversity of biotransformation genes in response to creosote bush resin.

Liver biotransformation enzymes have long been thought to enable animals to feed on diets rich in xenobiotic compounds. However, despite decades of pharmacological research in humans and rodents, little is known about hepatic gene expression in specialized mammalian herbivores feeding on toxic diets. Leveraging a recently identified population of the desert woodrat (Neotoma lepida) found to be highly tolerant to toxic creosote bush (Larrea tridentata), we explored the expression changes of suites of biotransformation genes in response to diets enriched with varying amounts of creosote resin. Analysis of hepatic RNA-seq data indicated a dose-dependent response to these compounds, including the upregulation of several genes encoding transcription factors and numerous phase I, II, and III biotransformation families. Notably, elevated expression of five biotransformation families - carboxylesterases, cytochromes P450, aldo-keto reductases, epoxide hydrolases, and UDP-glucuronosyltransferases - corresponded to species-specific duplication events in the genome, suggesting that these genes play a prominent role in N. lepida's adaptation to creosote bush. Building on pharmaceutical studies in model rodents, we propose a hypothesis for how the differentially expressed genes are involved in the biotransformation of creosote xenobiotics. Our results provide some of the first details about how these processes likely operate in the liver of a specialized mammalian herbivore.

Indoor air levels of polycyclic aromatic compounds (PAC) in public buildings with creosote impregnated constructions - A pilot case study using passive samplers.

Creosote has been used in Sweden as a wood preservative in buildings since the 19th century. These buildings can function as workplaces, homes, and cultural buildings to which the public has access. Creosote contains polycyclic aromatic hydrocarbons (PAH) which are well known carcinogens. To understand exposure and risks in an indoor environment, it is important to determine air levels of parent PAHs as well as the more toxic nitrated and oxygenated PAH derivatives (NPAH, OPAH). This study aims to investigate indoor air levels of polycyclic aromatic compounds (PACs) e.g., PAH, NPAH, OPAH and dibenzothiophenes in buildings containing creosote sources and whether these levels pose a health risk. Four cultural buildings were studied, all located within a radius of 130 m. Two were known to have creosote sources, and two had not. Polyurethane foam passive air samplers (PUF-PAS) were used to indicate possible point sources. PUF-PAS measurements were performed for one month in each building winter and summer. Simultaneously, PAC outdoor level measurements were performed. Buildings with creosote impregnated constructions had notably higher indoor air levels of PAC (31-1200 ng m-3) compared to the two buildings without creosote sources (14-45 ng m-3). The PAH cancer potency (sum of benzo[a]pyrene equivalents (BaPeq)) was more than one order of magnitude higher in the buildings containing creosote impregnated wood compared to reference buildings. The highest value was 5.1 BaPeq ng m-3 which was significantly higher than the outdoor winter measurement (1.3 BaPeq ng m-3). Fluoranthene and phenanthrene, with significant distribution in gas phase, but also several particulate NPAHs contributed significantly to the total cancer risk. Thus, creosote containing buildings can still contaminate the indoor air with PACs despite being over a hundred years old. The PUF-PAS was shown to be a good tool providing quantitative/semiquantitative measures of PACs exposure in indoor microenvironments.

Study of creosote transport properties in sandy and clay soils.

Creosote is an organic pollutant formed by a complex mixture of highly toxic and carcinogenic compounds and classified as a dense non-aqueous phase liquid (DNAPL). Its migration depends on media and fluid properties that control the multiphase flow in the subsurface. Residual saturation and hydraulic conductivity are essential parameters to accurately simulate fluid displacement in porous media. This work shows the behavior of creosote in porous medium for sandy and clay soils, collected in a contaminated area in the state of São Paulo, Brazil. Creosote retention was evaluated and compared to water. The retention curve parameters were obtained based on van Genuchten and Brooks and Corey models. The hydraulic conductivities of creosote and water are presented for both soils. The results show that, in the clay soil, water was more retained than creosote, while in the sandy soil, creosote retention was higher. The hydraulic conductivity values obtained in the clay soil show a difference of two orders of magnitude between creosote and water. Although creosote is a viscous fluid, it presents considerable mobility in the clay soil, which is relevant in remediation processes. This study advances our knowledge about DNAPL behavior in clay and sand, and no other study of creosote parameters in these porous media was found. A more accurate estimate of the time required for a liquid spill to reach groundwater can then be predicted, so that appropriate actions can be taken and risk management can be carried out.

Development of a safer and improved analytical method for polycyclic aromatic hydrocarbons in creosote products.

Polycyclic aromatic hydrocarbons (PAHs) in creosote products used for wood preservation are regulated in Japan. Although the analytical method for this regulation has been stipulated by law, two main problems have been highlighted, namely the use of dichloromethane, a potential carcinogen, as a solvent and inadequate purification. Therefore, an analytical method to solve these problems was developed in this study. Actual creosote-treated wood samples were examined, and it was found that acetone could be used as an alternative solvent. Purification methods using centrifugation, silica gel cartridges, and strong anion exchange (SAX) cartridges were also developed. It was found that the SAX cartridges strongly retained PAHs, and an effective purification method was developed using this phenomenon, in which contaminants were removed by washing with diethyl ether/hexane (1/9 v/v), which could not be achieved with a silica gel cartridge. This strong retention was attributed to cation-π interactions. The analytical method developed in this study yielded good recoveries (81.4-113.0%) with low relative standard deviations (<6.8%), and the limit of quantification (0.02-0.29 µg/g) was significantly lower than the current creosote product regulation. Therefore, this method can safely and effectively extract and purify PAHs from creosote products.

Application of Ozonation-Biodegradation Hybrid System for Polycyclic Aromatic Hydrocarbons Degradation.

Creosote, a mixture of polycyclic aromatic hydrocarbons (PAHs), was and is a wood impregnate of widespread use. Over the years the accumulation of creosote PAHs in soils and freshwaters has increased, causing a threat to ecosystems. The combined ozonation-biodegradation process is proposed to improve the slow and inefficient biodegradation of creosote hydrocarbons. The impact of different ozonation methods on the biodegradation of model wastewater was evaluated. The biodegradation rate, the changes in chemical oxygen demand, and the total organic carbon concentration were measured in order to provide insight into the process. Moreover, the bacteria consortium activity was monitored during the biodegradation step of the process. The collected data confirmed the research hypothesis, which was that the hybrid method can improve biodegradation. The pre-ozonation followed by inoculation with a bacteria consortium resulted in a significant increase in the biodegradation rate. It allows for the shortening of the time required for the consortium to reach maximum degradation effectiveness and cell activity. Hence, the study gives an important and useful perspective for the decontamination of creosote-polluted ecosystems.

Increased expression of glucagon-like peptide-1 and cystic fibrosis transmembrane conductance regulator in the ileum and colon in mouse treated with metformin.

Metformin, an oral medication, is prescribed to patients with type 2 diabetes mellitus. Although the efficacy, safety, and low economic burden of metformin on patients have long been recognized, approximately 5% of the patients treated with this drug develop severe diarrhea and discontinue the treatment. We previously reported that 1,000 mg·kg-1·day-1 of metformin induced diarrhea in diabetic obese (db/db) mice and wood creosote (traditional medication for diarrhea) ameliorated the symptoms. In this study, we attempted to elucidate the molecular mechanisms by which metformin induces diarrhea. Cystic fibrosis transmembrane conductance regulator (CFTR) is a key ion (chloride) channel in cyclic adenosine monophosphate (cAMP)-induced diarrhea. Metformin treatment increased bile flow (bile acids and bilirubin) in the ileum of mice. In addition, the treatment was accompanied by an increase in mRNA and protein levels of CFTR in the mucosa of the ileum and colon in both wild-type (C57BL/6J) and db/db mice. Glucagon-like peptide-1 (GLP-1), as well as cholic acid, induces CFTR mRNA expression in human colon carcinoma Caco-2 cells through cAMP signaling. Although wood creosote (10 mg/kg) ameliorated diarrhea symptoms, it did not alter the mRNA levels of Glp-1 or Cftr. Similar to overeating, metformin upregulated GLP-1 and CFTR expression, which may have contributed to diarrhea symptoms in mice. Although we could not identify db/db mouse-specific factors associated with metformin-induced diarrhea, these factors may modulate colon function. Wood creosote may not interact with these factors but ameliorates diarrhea symptoms.

Magnetic signatures of a creosote oil contaminated site: case study in São Paulo, Brazil.

Soils and groundwater contamination modifies the physical-chemical conditions of the environment, altering natural biogeochemical processes of the ground. As a result, several mineral transformations occur, in which iron plays a decisive role. The presence of iron enables the study of magnetic properties, improving the understanding of the geophysical signatures of highly dynamic environments (e.g., biogeochemical hotspots and contamination plumes). In this work, we seek to identify creosote biodegradation related to the precipitation of magnetic minerals on sediments at a contaminated site in São Paulo, Brazil. Several rock magnetism analyses were carried out to provide the magnetic mineralogy of the samples in terms of their composition, size, and abundance. We conducted high-temperature thermomagnetic curves, frequency-dependent magnetic susceptibility, anesthetic remanent magnetization (ARM) and isothermal remanent magnetization (IRM) data, superparamagnetic concentration and dipole moment (SPCDM), and scanning electron microscopy (SEM) analyses. The magnetic signatures of the contaminated samples suggest an increase of superparamagnetic grains in the water table fluctuation zone if compared to the magnetic signatures of the uncontaminated samples. Thermomagnetic curves of contaminated samples showed a lower heterogeneity of the magnetic mineral phases than the uncontaminated ones. This work contributes to the advancement of the understanding of how natural biogeochemical processes are impacted by human actions, such as soil contamination, and even by climate change, which should affect soil redox conditions in periods of drought and flooding.

PUF-Immobilized Bjerkandera adusta DSM 3375 as a Tool for Bioremediation of Creosote Oil Contaminated Soil.

Creosote oil, a byproduct of coal distillation, is primarily composed of aromatic compounds that are difficult to degrade, such as polycyclic aromatic hydrocarbons, phenolic compounds, and N-, S-, and O-heterocyclic compounds. Despite its toxicity and carcinogenicity, it is still often used to impregnate wood, which has a particularly negative impact on the condition of the soil in plants that impregnate wooden materials. Therefore, a rapid, effective, and eco-friendly technique for eliminating the creosote in this soil must be developed. The research focused on obtaining a preparation of Bjerkandera adusta DSM 3375 mycelium immobilized in polyurethane foam (PUF). It contained mold cells in the amount of 1.10 ± 0.09 g (DW)/g of the carrier. The obtained enzyme preparation was used in the bioremediation of soil contaminated with creosote (2% w/w). The results showed that applying the PUF-immobilized mycelium of B. adusta DSM 3375 over 5, 10, and 15 weeks of bioremediation, respectively, removed 19, 30, and 35% of creosote from the soil. After 15 weeks, a 73, 79, and 72% level of degradation of fluoranthene, pyrene, and fluorene, respectively, had occurred. The immobilized cells have the potential for large-scale study, since they can degrade creosote oil in soil.

Embryonic exposure to benzo[a]pyrene causes age-dependent behavioral alterations and long-term metabolic dysfunction in zebrafish.

Polycyclic aromatic hydrocarbons (PAH) are products of incomplete combustion which are ubiquitous pollutants and constituents of harmful mixtures such as tobacco smoke, petroleum and creosote. Animal studies have shown that these compounds exert developmental toxicity in multiple organ systems, including the nervous system. The relative persistence of or recovery from these effects across the lifespan remain poorly characterized. These studies tested for persistence of neurobehavioral effects in AB* zebrafish exposed 5-120 h post-fertilization to a typical PAH, benzo[a]pyrene (BAP). Study 1 evaluated the neurobehavioral effects of a wide concentration range of BAP (0.02-10 µM) exposures from 5 to 120 hpf during larval (6 days) and adult (6 months) stages of development, while study 2 evaluated neurobehavioral effects of BAP (0.3-3 µM) from 5 to 120 hpf across four stages of development: larval (6 days), adolescence (2.5 months), adulthood (8 months) and late adulthood (14 months). Embryonic BAP exposure caused minimal effects on larval motility, but did cause neurobehavioral changes at later points in life. Embryonic BAP exposure led to nonmonotonic effects on adolescent activity (0.3 µM hyperactive, Study 2), which attenuated with age, as well as startle responses (0.2 µM enhanced, Study 1) at 6 months of age. Similar startle changes were also detected in Study 2 (1.0 µM), though it was observed that the phenotype shifted from reduced pretap activity to enhanced posttap activity from 8 to 14 months of age. Changes in the avoidance (0.02-10 µM, Study 1) and approach (reduced, 0.3 µM, Study 2) of aversive/social cues were also detected, with the latter attenuating from 8 to 14 months of age. Fish from study 2 were maintained into aging (18 months) and evaluated for overall and tissue-specific oxygen consumption to determine whether metabolic processes in the brain and other target organs show altered function in late life based on embryonic PAH toxicity. BAP reduced whole animal oxygen consumption, and overall reductions in total basal, mitochondrial basal, and mitochondrial maximum respiration in target organs, including the brain, liver and heart. The present data show that embryonic BAP exposure can lead to neurobehavioral impairment across the life-span, but that these long-term risks differentially emerge or attenuate as development progresses.

Extensive chemical and bioassay analysis of polycyclic aromatic compounds in a creosote-contaminated superfund soil following steam enhanced extraction.

Polycyclic aromatic compounds (PACs) are organic compounds commonly found in contaminated soil. Previous studies have shown the removal of polycyclic aromatic hydrocarbons (PAHs) in creosote-contaminated soils during steam enhanced extraction (SEE). However, less is known about the removal of alkyl-PAHs and heterocyclic compounds, such as azaarenes, and oxygen- and sulfur-heterocyclic PACs (OPACs and PASHs, respectively). Further, the impact of SEE on the freely dissolved concentration of PACs in soil as well as the soil bioactivity pre- and post-SEE have yet to be addressed. To fulfil these research gaps, chemical and bioanalytical analysis of a creosote-contaminated soil, collected from a U.S. Superfund site, pre- and post-SEE were performed. The decrease of 64 PACs (5-100%) and increase in the concentrations of nine oxygenated-PAHs (OPAHs) (150%) during SEE, some of which are known to be toxic and can potentially contaminate ground water, were observed. The freely dissolved concentrations of PACs in soil were assessed using polyoxymethylene (POM) strips and the concentrations of 66 PACs decreased post-SEE (1-100%). Three in vitro reporter gene bioassays (DR-CALUX®, ERα-CALUX® and anti-AR CALUX®) were used to measure soil bioactivities pre- and post-SEE and all reporter gene bioassays measured soil bioactivity decreases post-SEE. Mass defect suspect screening tentatively identified 27 unique isomers of azaarenes and OPAC in the soil. As a remediation technique, SEE was found to remove alkyl-PAHs and heterocyclic PACs, reduce the concentrations of freely dissolved PACs, and decrease soil bioactivities.

Ebullition-facilitated mobilization of trapped dense non-aqueous phase liquid at residual saturation from sandy sediments.

Gas ebullition can mobilize dense non-aqueous phase liquids (DNAPLs) from sediments to the overlying water column, increasing the DNAPL-impacted area and posing serious challenges to the remediation and management of contaminated sediments. Despite this, there have been few laboratory studies focused on gas ebullition-facilitated transport of DNAPL. In this study, bubble-facilitated transport was investigated by injecting gas (air or nitrogen) at 1 mL/min through a creosote source zone (∼25% saturation) capped with sand layers of different thicknesses. Three short-term experiments (8.3-8.7 h) were capped with 11.4, 7.0 or 4.5 cm of sand to estimate DNAPL flux. One long-term experiment (30 days) was capped with 8 cm of sand to investigate DNAPL removal. Heptane placed on a layer of water above the sand was used as a solvent trap and analyzed for petroleum hydrocarbons (PHCs). Results showed that creosote travelled as thin coatings and films surrounding gas bubbles migrating out of the source zone. Gas invasion was dominated by capillarity in the 11.4 cm-thick sand layer and by fracturing in the 7.0 and 4.5 cm-thick sand layers. Migration through these fractures often led to the formation of creosote tails on mobilized bubbles that drained towards the rear end of the bubble. The mass released decreased exponentially with sand cap thickness. In the long-term experiment, images showed significant depletion of the source zone in 30 days. Linear regression analysis showed that relationships with high predictive capabilities for ebullition-facilitated fluxes of hydrophobic organic contaminants can be obtained by incorporating gas ebullition flux and source strength, based on results from this study along with others from the field and laboratory. To our knowledge, this is the first study to compile and integrate data collected from laboratory and field studies to develop an assessment tool to facilitate the management of contaminated sediments affected by gas ebullition.

Successes and limitations of quantitative diet metabarcoding in a small, herbivorous mammal.

DNA metabarcoding is widely used to determine wild animal diets, but whether this technique provides accurate, quantitative measurements is still under debate. To test our ability to accurately estimate the abundance of dietary items using metabarcoding, we fed wild-caught desert woodrats (Neotoma lepida) diets consisting of constant amounts of juniper (Juniperus osteosperma, 15%) and varying amounts of creosote (Larrea tridentata, 1%-60%), cactus (Opuntia sp., 0%-100%) and commercial chow (0%-85%). Using metabarcoding, we compared the representation of items in the original diet samples to that in the faecal samples to test the sensitivity and accuracy of diet metabarcoding, the performance of different bioinformatic pipelines and our ability to correct sequence counts. Metabarcoding, using standard trnL primers, detected creosote, juniper and chow. Different pipelines for assigning taxonomy performed similarly. While creosote was detectable at dietary proportions as low as 1%, we failed to detect cactus in most samples, probably due to a primer mismatch. Creosote read counts increased as its proportion in the diet increased, and we could differentiate when creosote was a minor and major component of the diet. However, we found that estimates of juniper and creosote varied. Using previously suggested methods to correct these errors did not improve accuracy estimates of creosote, but did reduce error for juniper and chow. Our results indicate that metabarcoding can provide quantitative information on dietary composition, but may be limited. We suggest that researchers use caution when quantitatively interpreting diet metabarcoding results unless they first experimentally determine the extent of possible biases.

Using solid-phase microextraction during ultrasound reveals higher aqueous PAHs release from contaminated sediment.

Ultrasound (US) releases polycyclic aromatic hydrocarbons (PAHs) from contaminated creosote sediments and degrades PAHs in aqueous solution. However, it is unclear how much PAHs release occurs during active US compared to after US is stopped. In this study, solid-phase microextraction (SPME) was used to determine aqueous PAH concentrations desorbed from Little Scioto River creosote contaminated sediment during exposure to 20 kHz ultrasound (US) at a power of 430 W L-1. First, SPME fiber-water partition coefficients,KSPME, were experimentally determined and shown to be comparable with previous studies. Next, PAH concentrations released into aqueous solution were determined by sequentially exposing fresh, conditioned SPME fibers in a reactor containing the contaminated sediment and DI for 10 min periods. Three consecutive 10 min periods each were measured during US and after US. Compared to mixing only, PAHs desorbed during ultrasound was significantly higher. In addition, for phenanthrene, anthracene, and fluoranthene, US showed significantly higher aqueous concentrations during US compared with after US. For these less soluble PAHs, desorption into the aqueous phase reaches and surpasses expected equilibrium aqueous concentrations. However, when US is ceased, PAHs appear to resorb onto sediment resulting in the lower concentrations measured in these PAH compounds after US that are similar to expected equilibrium aqueous concentrations. Typical analytical extraction methods for determining the effects of US require stopping treatment and may underestimate the potential US has for release of contaminants from sediment during US. Using SPME during US treatment reveals that the concentration of PAHs during US may surpass the expected equilibrium aqueous concentration of some PAHs congeners. To our knowledge, this higher concentration observed during US compared to after US has not been shown previously.

The Inhibitory Effect of Wood Creosote on the Movement of Anisakis Larvae: An Implication for the Treatment of Acute Anisakiasis.

INTRODUCTION: Anisakiasis is a common disease in countries such as Japan, where raw or undercooked marine fish are frequently consumed. The disease is caused by accidental ingestion of a live larva of Anisakis in raw or undercooked marine fish. In typical cases, it causes abrupt gastrointestinal symptoms, such as epigastric pain, nausea, and vomiting. According to a published report, the disease was alleviated by oral ingestion of an over-the-counter drug containing wood creosote. METHODS: We performed an in vitro experiment to elucidate whether wood creosote can inhibit the motor activity of Anisakis larvae, using infrared locomotion tracking and agarose gel penetration techniques. RESULTS: Our results clearly demonstrate that wood creosote inhibits the motor activity of Anisakis larvae. The concentration of wood creosote used in our experiment is similar to that found in stomach juice when a usual oral dose is taken of the medicine containing wood creosote. DISCUSSION/CONCLUSION: Our results suggest the potential usefulness of the medicine containing wood creosote in the treatment of acute Anisakis infection of the gastrointestinal tract.

Initial Observation of Factors Interfering with the Treatment of Alveolar Osteitis Using Hyaluronic Acid with Octenidine-A Series of Case Reports.

Alveolar osteitis (AO) is a common complication following the extraction of the teeth, particularly the lower third molars. It starts within a few days after the extraction and manifests mainly as pain in the extraction site. Several strategies of treatment are available in order to relieve pain and heal the extraction wound. Recently, a novel medical device combining hyaluronic acid (HA) and octenidine (OCT) was introduced for the treatment of AO. This series of case reports aims to summarize the initial clinical experiences with this new device and to highlight factors possibly interfering with this treatment. The medical documentation of five patients with similar initial situations treated for AO with HA + OCT device was analyzed in detail. Smoking and previous treatment with Alveogyl (Septodont, Saint-Maur-des-Fossés, France) were identified as factors interfering with the AO treatment with the HA + OCT device. In three patients without these risk factors, the treatment led to recovery within two or three days. The patient pretreated with Alveogyl and the smoker required six and seven applications of the HA + OCT device, respectively. According to these initial observations, it seems smoking and previous treatment with Alveogyl prolong the treatment of AO using the HA + OCT device that, in turn, shows a rapid effect if these risk factors are not present.

Wood­feeding termites as an obscure yet promising source of bacteria for biodegradation and detoxification of creosote-treated wood along with methane production enhancement.

This study aims to explore distinct bacterial strains from wood-feeding termites and to construct novel bacterial consortium for improving the methane yield during anaerobic digestion by degrading birchwood sawdust (BSD) and removing creosote (CRO) compounds simultaneously. A novel bacterial consortium CTB-4 which stands for the molecularly identified species Burkholderia sp., Xanthomonas sp., Shewanella sp., and Pseudomonas mosselii was successfully developed. The CTB-4 consortium showed high efficiency in the removal of naphthalene and phenol. It also revealed reduction in lignin, hemicellulose, and cellulose by 19.4, 52.5, and 76.8%, respectively. The main metabolites after the CRO degradation were acetic acid, succinate, pyruvate, and acetaldehyde. Pretreatment of treated BSD mixed with CRO enhanced the total methane yield (162 L/kg VS) by 82.7% and biomass reduction by 54.7% compared to the untreated substrate. CRO showed a toxicity decrease of >90%, suggesting the efficiency of constructed bacterial consortia in bioremediation and biofuel production.

Construction of a novel microbial consortium valued for the effective degradation and detoxification of creosote-treated sawdust along with enhanced methane production.

Lignocellulosic biomass represents an unlimited and ubiquitous energy source, which can effectively address current global challenges, including climate change, greenhouse gas emissions, and increased energy demand. However, lignocellulose recalcitrance hinders microbial degradation, especially in case of contaminated materials such as creosote (CRO)-treated wood, which necessitates appropriate processing in order to eliminate pollution. This study might be the first to explore a novel bacterial consortium SST-4, for decomposing birchwood sawdust, capable of concurrently degrading lignocellulose and CRO compounds. Afterwards, SST-4 which stands for molecularly identified bacterial strains Acinetobacter calcoaceticus BSW-11, Shewanella putrefaciens BSW-18, Bacillus cereus BSW-23, and Novosphingobium taihuense BSW-25 was evaluated in terms of biological sawdust pre-treatment, resulting in effective lignocellulose degradation and 100% removal of phenol and naphthalene. Subsequently, the maximum biogas production observed was 18.7 L/kg VS, while cumulative methane production was 162.8 L/kg VS, compared to 88.5 without microbial pre-treatment. The cumulative energy production from AD-I and AD-II through biomethanation was calculated as 3177.1 and 5843.6 KJ/kg, respectively. The pretreatment process exhibited a significant increase in the energy yield by 83.9%. Lastly, effective CRO detoxification was achieved with EC50 values exceeding 90%, showing the potential for an integrated process of effective contaminated wood management and bioenergy production.

The In Vitro Proinflammatory Properties of Water Accommodated Sediment Extracts from a Creosote-Contaminated US Environmental Protection Agency Superfund Site.

The southern branch of the Elizabeth River near Portsmouth, Virginia, USA, is one of the most creosote-polluted subestuaries in North America and the former location of the Atlantic Wood US Environmental Protection Agency Superfund site. We previously demonstrated that adult Atlantic Wood killifish collected in situ had severe hepatic lesions, including hepatoblastoma and hepatocellular carcinoma, as well as suppressed circulating antibody responses compared to a historical reference site. Moreover, several innate immune functions were higher in Atlantic Wood fish, including elevated expression of hepatic cyclooxygenase-2 (COX-2), suggesting a proinflammatory environment. To further examine the potential of Atlantic Wood contaminants to modulate innate immune function(s), the present study used RAW264.7 mouse macrophages as an in vitro model to develop new approach methodologies for rapid screening. Lipopolysaccharide (LPS)-stimulated nitric oxide secretion by macrophages is a rapid, sensitive, and predictive in vitro system for screening potentially immunotoxic contaminants as single compounds or as complex mixtures. Compared to the reference site, filter-sterilized Atlantic Wood sediment extracts (water accommodated fractions) induced nitric oxide and IL-6 secretion as well as inducible nitric oxide synthase and COX-2 proteins at levels comparable to or higher than those induced by LPS treatments alone. Extracts also increased phagocytic activity by macrophages. Using a limulus lysate assay, we show that bacterial endotoxin levels in Atlantic Wood extracts are higher than in reference extracts and that polymyxin-B chelation ameliorates proinflammatory effects. These findings illuminate the reality of sediment constituents other than toxic compounds previously associated with developmental abnormalities and carcinogenesis in killifish from the Atlantic Wood site. Perhaps these data also suggest the presence of contaminant-adapted consortia of sediment microbes at many heavily polluted sites worldwide compared to less contaminated sites. Environ Toxicol Chem 2021;40:1576-1585. © 2021 SETAC.

Biodegradation of creosote-treated wood by two novel constructed microbial consortia for the enhancement of methane production.

Lignocellulose biodegradation is limited because of its recalcitrant structure particularly when polluted by toxic and carcinogenic compounds such as creosote oil (CRO). As far as we know, this might be the first report that explores the biodegradation of creosote treated wood (CTW) to serve biomethane production. Two novel CTW-degrading microbial consortia, designated as CTW-1 and CTW-2, were screened and constructed to enhance methane production from CRO-treated pine sawdust. After 12 days of biological pretreatment by CTW-1 and CTW-2, a significant reduction in lignocellulosic content of CTW was recorded; estimated as 49 and 43%, respectively. More than 64 and 91% of cumulative biogas and methane yields were obtained from biodegraded CTW over control. Ecotoxicity of treated and untreated CTW was compared by Microtox test. The biodegraded CTW hydrolysates showed a toxicity decrease of more than 80%, suggesting the promising role of constructed microbial consortia for biofuel production and bioremediation.

Inhibition of Acetylcholinesterase by Wood Creosote and Simple Phenolic Compounds.

Anisakiasis is common in countries where raw or incompletely cooked marine fish are consumed. Currently, effective therapeutic methods to treat anisakiasis are unavailable. A recent study found that wood creosote inactivates the movement of Anisakis species. Essential oil of Origanum compactum containing carvacrol and thymol, which are similar to the constituents of wood creosote, was reported to inactivate Anisakis by inhibiting its acetylcholinesterase. We examined whether wood creosote can also inhibit acetylcholinesterase. We examined the effect of components of wood creosote using the same experimental method. A computer simulation experiment (molecular docking) was also performed. Here, we demonstrate that wood creosote inactivated acetylcholinesterase in a dose-dependent manner with an IC50 of 0.25 mg/mL. Components of wood creosote were also tested individually: 5-methylguaiacol, p-cresol, guaiacol, o-cresol, 2,4-dimethylphenol, m-cresol, phenol and 4-methylguaiacol inactivated the enzyme with an IC50 of 14.0, 5.6, 17.0, 6.3, 3.9, 10.0, 15.2 and 27.2 mM, respectively. The mechanism of acetylcholinesterase inactivation was analyzed using a computer-based molecular docking simulation, which employed a three-dimensional structure of acetylcholinesterase and above phenolic compounds as docking ligands. The simulation indicated that the phenolic compounds bind to the active site of the enzyme, thereby competitively blocking entry of the substrate acetylcholine. These findings suggest that the mechanism for the inactivation of Anisakis movement by wood creosote is due to inhibition of acetylcholinesterase needed for motor neuron activity.