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Pentachlorophenol (PCP) is a toxic compound, which is widely used as a wood preservative product and general biocide. It is persistent in the environment and has been classified as a persistent organic pollutant to be reclaimed in many countries. Bioremediation is an emerging approach to rehabilitating areas polluted by recalcitrant xenobiotics. In the present study, we evaluated the potential of three strains of Pseudomonas (P. putida S121, P. rhizophila S211, and P. fuscovagiceae S115) as bioremediation agents in depletion and detoxification of PCP in soil microcosms. PCP removal was effectively optimized using a central-composite experimental design and response surface methodology (RSM). The optimum conditions for maximum PCP removal yield (85 ± 5%) were: 500 mg/kg PCP concentration, 10 8 UFC/g soil inoculum size of each strain and 55 days incubation period. The bacterial strains, P. putida, P. rhizophila, and P. fuscovagiceae, showed good capability to tolerate and degrade PCP so that they could be successfully used in synergistic effect to treat PCP polluted soils., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Soil microorganisms play a crucial role in the bioremediation of pentachlorophenol (PCP)-contaminated soils. However, whether and how soil bacterial networks with keystone taxa affect PCP dechlorination is not well understood. The present study investigated the effects of citrate on soil bacterial networks mediating PCP dechlorination by direct and indirect transformation in iron-rich upland and paddy soils. The rates of PCP dechlorination and Fe(II) generation were accelerated by citrate addition, particularly in the paddy soils. Network analysis revealed that the topological properties of bacterial networks were changed by citrate addition; more modules and keystone taxa were significantly correlated with PCP dechlorination and Fe(II) generation in the networks. Random forest modeling indicated that Clostridiales was the most important bacterial order; it was significantly involved in both the direct and indirect pathways of PCP dechlorination. Citrate addition had less influence on the balance between the direct and indirect pathways of PCP dechlorination in the upland soils, whereas it enhanced biological PCP dechlorination more directly and efficiently in the paddy soils. Our results suggested that land-use type and citrate addition play a critical role in controlling the biogeochemical mechanisms of PCP dechlorination., (Copyright © 2019 Elsevier B.V. All rights reserved.)

In this study, experiments were performed using network analysis to investigate the effects of different carbon sources, including blank, citrate, glucose and lactate, on indigenous bacterial communities and on the pentachlorophenol (PCP) dechlorination in two soils. Kinetics results demonstrate that PCP dechlorination is significantly enhanced by adding citrate/lactate, but to a lesser extent by adding glucose. High-throughput sequencing results revealed that Firmicutes and Proteobacteria were the dominant groups in these four different treatments during the PCP dechlorination, whereas random forest analysis indicated that the orders Clostridiales, Haloplasmatales, Bacillales, Pseudomonadales and Gaiellales were the critical bacterial orders in modules that were significantly correlated with PCP dechlorination. Among them, the relative abundance of Clostridiales dramatically increased in both citrate and lactate treatment, further accelerating the PCP dechlorination. Addition of citrate/lactate as the carbon source increased the bacterial co-occurrence network density, average clustering coefficient and modularity. Moreover, more modules significantly correlated with PCP dechlorination in the citrate/lactate networks compared with the glucose/blank networks. Random forest modeling suggested that Clostridiales played a critical role in these functional modules. Taken together, our results provide insight into the biological mechanism of the impact of exogenous carbon sources on PCP dechlorination pathways by modifying soil bacterial networks., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Atmospheric release of persistent organic pollutants (POPs) constitutes a silent threat through chronic contamination of soils at global scale; yet fundamental understanding of their occurrence, sources and fate is still largely lacking. Similar to a three act play, this review comprises Setup, Confrontation and Resolution. The first emphasises the eighty years of the history of pentachlorophenol (PCP) usage, only recently classified as POP. The second focus on active sources of PCP pollution, including inside cork oak forests in N.W. Tunisia; a threat partially neutralised by the soil microbial diversity, especially fungi. As Resolution, the need for improved knowledge on the global distribution and impacts of PCP in soil microbial diversity as means to preserve the multi-functionality of terrestrial ecosystem is emphasised., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Pentachlorophenol (PCP) is a common persistent pesticide in soil that has generated a significant environmental problem worldwide. Therefore, anaerobic degradation of PCP by the soil indigenous microbial community has gained increasing attention. However, little information is available concerning the functional microorganisms and the potential shifts in the microbial community associated with PCP degradation. In this study, we conducted a set of experiments to determine which components of the indigenous microbial community were capable of degrading PCP in soils of two land use types (upland and paddy soils) in southern China. Our results showed that the PCP degradation rate was significantly higher in paddy soils than that in upland soils. 16S ribosomal RNA (rRNA) high-throughput sequencing revealed significant differences in microbial taxonomic composition between the soil with PCP and blank (soil without PCP) with Acinetobacter, Clostridium, Coprococcus, Oxobacter, and Sedimentibacter dominating the PCP-affected communities. Acinetobacter was also apparently enriched in the paddy soils with PCP (up to 52.2 %) indicated this genus is likely to play an important role in PCP degradation. Additionally, the Fe(III)-reducing bacteria Clostridium may also be involved in PCP degradation. Our data further revealed hitherto unknown metabolisms of potential PCP degradation by microorganisms including Coprococcus, Oxobacter, and Ruminiclostridium. Overall, these findings indicated that land use types may affect the PCP anaerobic degradation rate via the activities of indigenous bacterial populations and extend our knowledge of the bacterial populations responsible for PCP degradation.

Soil humic substances can be used as redox mediators in accelerating the biotransformation of organic pollutants, and humus-respiring bacteria are widely distributed in soils. However, the impact of humic substances on the soil microbial community during the biotransformation of organic pollutants is expected to be crucial while remains to be unclear. In this study, the biostimulation of indigenous microbial communities and the consequent effects on anaerobic transformation of pentachlorophenol (PCP) by a model humic substance, anthraquinone-2,6-disulfonate (AQDS), were systematically investigated in a paddy soil. The addition of AQDS was observed to increase the production of HCl-extractable Fe(II) and enhance the PCP transformation rates consequently. The pseudo-first-order rate constants of the PCP transformation showed a positive exponential relationship with the AQDS dosage. The terminal restriction fragment length polymorphism (T-RFLP) results indicated the substantial effect of added AQDS on soil microbial community. The enhanced abundance of Geobacter sp. was disclosed to be most critical for accelerated PCP transformation when with AQDS, in which Geobacter sp. functioned for promoting the generation of active Fe(II) and consequently enhancing the PCP transformation rates. The transformation rates of PCP were exponentially correlated with the abundance of Geobacter sp. positively. The findings are expected to improve the understanding of diversity and ubiquity of microorganisms in humic substances-rich soils for accelerating the transformations of soil chlorinated pollutants., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The ecological effect of earthworms on the fate of soil pentachlorophenol (PCP) differs with species. This study addressed the roles and mechanisms by which two earthworm species (epigeic Eisenia fetida and endogeic Amynthas robustus E. Perrier) affect the soil microbial community and enzyme activity during the bioremediation of PCP-contaminated soils. A. robustus removed more soil PCP than did E. foetida. A. robustus improved nitrogen utilisation efficiency and soil oxidation more than did E. foetida, whereas the latter promoted the organic matter cycle in the soil. Both earthworm species significantly increased the amount of cultivable bacteria and actinomyces in soils, enhancing the utilisation rate of the carbon source (i.e. carbohydrates, carboxyl acids, and amino acids) and improving the richness and evenness of the soil microbial community. Additionally, earthworm treatment optimized the soil microbial community and increased the amount of the PCP-4-monooxygenase gene. Phylogenic classification revealed stimulation of indigenous PCP bacterial degraders, as assigned to the families Flavobacteriaceae, Pseudomonadaceae and Sphingobacteriacea, by both earthworms. A. robustus and E. foetida specifically promoted Comamonadaceae and Moraxellaceae PCP degraders, respectively., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Pentachlorophenol (PCP) is globally dispersed and contamination of soil with this biocide adversely affects its functional biodiversity, particularly of fungi - key colonizers. Their functional role as a community is poorly understood, although a few pathways have been already elucidated in pure cultures. This constitutes here our main challenge - elucidate how fungi influence the pollutant mitigation processes in forest soils. Circumstantial evidence exists that cork oak forests in N. W. Tunisia - economically critical managed forests are likely to be contaminated with PCP, but the scientific evidence has previously been lacking. Our data illustrate significant forest contamination through the detection of undefined active sources of PCP. By solving the taxonomic diversity and the PCP-derived metabolomes of both the cultivable fungi and the fungal community, we demonstrate here that most strains (predominantly penicillia) participate in the pollutant biotic degradation. They form an array of degradation intermediates and by-products, including several hydroquinone, resorcinol and catechol derivatives, either chlorinated or not. The degradation pathway of the fungal community includes uncharacterized derivatives, e.g. tetrachloroguaiacol isomers. Our study highlights fungi key role in the mineralization and short lifetime of PCP in forest soils and provide novel tools to monitor its degradation in other fungi dominated food webs., (© 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.)

As the most prevalent preservative worldwide for many years, pentachlorophenol (PCP) has attracted much interest in the study of biodegradation in soil and aquatic ecosystems. However, the key microorganisms involved in anaerobic degradation are less well understood. Hence, we used DNA-based stable isotope probing (SIP) to identify the PCP-degrading microorganisms in iron-rich paddy soil under anaerobic conditions. (12)C- and (13)C-labeled PCP were almost completely degraded in 30 days under iron-reducing conditions. The results of terminal restriction fragment length polymorphism (T-RFLP) of 16S rRNA genes showed that 197 and 217 bp (HaeIII digests) restriction fragments (T-RFs) were enriched in heavy DNA fractions of (13)C-labeled samples, and the information from 16S rRNA gene clone libraries suggested that the microorganisms corresponding to these T-RF fragments, which increased in relative abundance during incubation, belonged to the order of Burkholderiales, in which 197 and 217 bp were classified as unclassified Burkholderiales and the genus Achromobacter, respectively. The results of the present study indicated that Burkholderiales-affiliated microorganisms were responsible for PCP degradation in anaerobic paddy soil and shed new light on in situ bioremediation in anaerobic PCP contaminated soil.

Carbon isotopic analysis and molecular-based methods were used in conjunction with geochemical data sets to assess the dechlorination of pentachlorophenol (PCP) when coupled to biogeochemical processes in a mangrove soil having no prior history of anthropogenic contamination. The PCP underwent 96% dechlorination in soil amended with acetate, compared to 21% dehalogenation in control soil. Carbon isotope analysis of residual PCP demonstrated an obvious enrichment of 13C (εC, -3.01±0.1%). Molecular and statistical analyses demonstrated that PCP dechlorination and Fe(III) reduction were synergistically combined electron-accepting processes. Microbial community analysis further suggested that enhanced dechlorination of PCP during Fe(III) reduction was mediated by members of the multifunctional family of Geobacteraceae. In contrast, PCP significantly suppressed the growth of SO4(2-) reducers, which, in turn, facilitated the production of CH4 by diversion of electrons from SO4(2-) reduction to methanogenesis. The integrated data regarding stoichiometric alterations in this study gives direct evidence showing PCP, Fe(III), and SO4(2-) reduction, and CH4 production are coupled microbial processes during changes in soil redox.

Vertical variations of pentachlorophenol (PCP) dissipation and microbial community were investigated in a paddy soil with the addition of electron acceptors (NO3(-), SO4(2-)) and donors (crop residues). Crop residues enhanced PCP dissipation by supplying dissolved organic carbon (DOC) as an electron donor, whereas NO3(-) and SO4(2-) inhibited it. The dissipation of PCP in electron donor treatments resulted in the accumulation of 3,4,5-trichlorophenol (3,4,5-TCP) except for wheat residues. The abundance and diversity of phospholipid fatty acids (PLFAs) decreased with increasing soil depth. The succession of predominant PLFAs shifted from aerobic bacteria to anaerobic bacteria when electron acceptors were changed to electron donors. The saturated/monounsaturated fatty acids (S/M) ratio increased with soil depth, which probably implied that nutrient turnover rate declined after the accumulation of 3,4,5-TCP. The results showed that the addition of electron donors and acceptors modified the microbial communities, which then further influenced the degradation pathway of PCP.

Pesticides are a class of xenobiotics intentionally released into the environment. Hexachlorobenzene (HCB) was used as a fungicide from 1945, leaving behind many contaminated sites. Very few studies have examined the biodegradation of HCB or the fate of HCB-derived carbon. Here we report that certain bacterial populations are capable of deriving carbon from HCB in contaminated soil under aerobic conditions. These populations are primarily Proteobacteria, including Methylobacterium and Pseudomonas, which predominated as detected by stable isotope probing (SIP) and 16S rRNA gene amplicon pyrosequencing. Due to the nature of SIP, which can be used as a functional method solely for assimilatory processes, it is not possible to elucidate whether these populations metabolized directly HCB or intermediates of its metabolism produced by different populations. The possibility exists that HCB is degraded via the formation of pentachlorophenol (PCP), which is further mineralized. With this in mind, we designed primers to amplify PCP 4-monooxygenase-coding sequences based on the available pcpB gene sequence from Methylobacterium radiotolerans JCM 2831. Based on 16S rRNA gene analysis, organisms closely related to this strain were detected in (13)C-labeled DNA. Using the designed primers, we were able to amplify pcpB genes in both total community DNA and (13)C-DNA. This indicates that HCB might be transformed into PCP before it gets assimilated. In summary, this study is the first report on which bacterial populations benefit from carbon originating in the pesticide HCB in a contaminated soil., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Reductive dechlorination is a crucial pathway for anaerobic biodegradation of highly chlorinated organic contaminants. Under an anoxic environment, reductive dechlorination of organic contaminants can be affected by many redox processes such as nitrate reduction and iron reduction. In the present study, batch incubation experiments were conducted to investigate the effect of nitrate addition on reductive dechlorination of PCP in paddy soil with consideration of iron transformation. Study results demonstrate that low concentrations (0, 0.5 and 1 mM) of nitrate addition can enhance the reductive dechlorination of PCP and Fe(III) reduction, while high concentrations (5, 10, 20 and 30 mM) of nitrate addition caused the contrary. Significant positive correlations between PCP degradation rates and the formation rates of dissolved Fe(II) (pearson correlation coefficients r = 0.965) and HCl-extractable Fe(II) (r = 0.921) suggested that Fe(III) reduction may enhance PCP dechlorination. Furthermore, consistent variation trends of PCP degradation and the abundances of the genus Comamonas, capable of Fe(III) reduction coupled to reductive dechlorination, and of the genus Dehalobacter indicated the occurrence of microbial community variation induced by nitrate addition as a response to PCP dechlorination., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

This study explored biostimulation mechanisms with an electron donor and a shuttle for accelerating pentachlorophenol (PCP) transformation in iron-rich soils. The results indicated that indigenous microbial communities are important for PCP transformation in soils. Biostimulation of indigenous microbial communities by the addition of lactate and anthraquinone-2,6-disulfonate (AQDS) led to the enhanced rates of PCP dechlorination by the dechlorinating- and iron-reducing bacteria in soils. The electrochemical studies using cyclic voltammograms and microbial current measurements confirmed the high reduction potential and the large amount of electrons generated under biostimulation conditions, which were responsible for the higher rates of PCP transformation. After biostimulation treatments by the additions of lactate and/or AQDS during PCP dechlorination processes, microbial community analysis by the terminal restriction fragment length polymorphism (T-RFLP) method showed the abundance terminal restricted fragments (T-RFs), an indicator of bacterial abundance, which represents the dechlorinating- and iron-reducing bacteria, suggesting their critical roles in PCP dechlorination in soils.

A polyphasic taxonomic approach was applied to the study of a Gram-negative bacterium (B2(T)) isolated from soil by selective enrichment with pentachlorophenol. 16S rRNA gene sequence analysis of strain B2(T) showed that the strain belongs to the genus Achromobacter within the Betaproteobacteria. The 16S rRNA gene sequence displayed more than 99 % similarity to the sequences of the type strains of all species of Achromobacter, with the highest sequence similarity to those of Achromobacter spanius CCM 7183(T) and A. piechaudii CCM 2986(T) (99.8 %). On the basis of phylogenetic analysis, genomic DNA-DNA relatedness and phenotypic characteristics, including chemotaxonomic (cellular fatty acid profile) analysis, a novel species is proposed, Achromobacter marplatensis sp. nov., with the type strain B2(T) ( = CCM 7608(T)  = CCUG 56371(T)  = CECT 7342(T)).

Two composting piles were prepared by adding to a mixture of rice straw, vegetables and bran: (i) raw soil free from pentachlorophenol (PCP) contamination (pile A) and (ii) PCP-contaminated soil (pile B). It was shown by the results that compost maturity characterized by water soluble carbon (WSC), TOC/TN ratio, germination index (GI) and dehydrogenase activity (DA) was significantly affected by PCP exposure, which resulted in an inferior degree of maturity for pile B. DGGE analysis revealed an inhibited effect of PCP on compost microbial abundance. The bacteria community shifts were mainly consistent with composting factors such as temperature, pH, moisture content and substrates. By contrast, the fungal communities were more sensitive to PCP contamination due to the significant correlation between fungal community shifts and PCP removal. Therefore, the different microbial community compositions for properly evaluating the degree of maturity and PCP contamination were suggested., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Bioaugmentation is a promising technology to clean up sites contaminated with recalcitrant chemicals. White-rot fungi have proven to be effective in the degradation of pentachlorophenol. Here, we report the bioremediation of soil contaminated with pentachlorophenol (PCP) by Anthracophyllum discolor and its impact on the soil microbial community. In this study three types of microcosms were established: fresh soil (C(0)), fresh soil plus wheat straw (WS(0)) and, fresh soil plus wheat straw inoculated with A. discolor (WSAD(0)). Additionally, similar treatments and a control of sterile soil spiked with PCP (C(250), WS(250) and WSAD(250)) were used to evaluate the remediation and adsorption of PCP. The PCP removal, total microbial activity, and enzymatic activities were evaluated. This study also investigated the structure of soil microbial community by denaturing gradient gel electrophoresis (DGGE), identifying some of the dominant bacterial and fungal species. The results showed that PCP was effectively degraded in soils by A. discolor and by indigenous soil microorganisms. The addition of wheat straw increased the PCP degradation and enzymatic activities. Only laccase activity was negatively affected by PCP contamination. The PCP degradation was associated with changes in microbial communities, mainly stimulation of members of bacterial phylum Proteobacteria (Xanthomonadaceae, Burkholderiaceae and Enterobacteriaceae), and fungal phylum Ascomycota and Basidiomycota. This study shows the ability of A. discolor to degrade PCP from contaminated soil, and demonstrates that agricultural residues, such as wheat straw, can be used as growth substrate by microorganisms in PCP-contaminated soil, demonstrating a great potential of autochthonous microorganisms for soil remediation., (Copyright 2010 Elsevier B.V. All rights reserved.)

This study investigated the use of electrokinetics in unsaturated soil to promote biodegradation of pentachlorophenol through increased contact between bacteria and contaminant. Soil microcosms, contaminated with approximately 100 mg kg(-1) pentachlorophenol (containing [(14)C]-PCP as a tracer), and inoculated with a specific pentachlorophenol-degrading bacterium (Sphingobium sp. UG30-1 x 10(8) cfu g(-1)) were subjected to constant and regularly reversed electric currents (10 mA). The former caused large pH and moisture content changes due to water electrolysis and electroosmotic effects, with subsequent negative impacts on biodegradation parameters including enzyme activity and contaminant mineralisation (as measured by (14)CO(2) evolution rate). The reversed field caused little change in pH and moisture content and led to more rapid contaminant mineralisation, lower soil contaminant concentration in the majority of the microcosms and increased soil enzyme activity (with the exception of soil immediately adjacent to the anode). The presence of an electric field, if suitably applied, may therefore enhance contaminant biodegradation in unsaturated soil.

Dechlorination of PCP has been observed previously under anaerobic condition in paddy soil. However, there is poor information about the dechlorination pathway of PCP and the microbial community associated with the PCP dechlorination in paddy soil. In this study, an anaerobic microbial community dechlorinating PCP was enriched by serial transfers from a paddy soil using a medium containing PCP, lactate and the steam-sterilized paddy soil. The enriched microbial community dechlorinated PCP completely to phenol under the anaerobic condition by a dechlorinating pathway as follows; PCP-->2,3,4,5-tetrachlorophenol-->3,4,5-trichlorophenol-->3,5-dichlorophenol-->3-chlorophenol-->phenol. Intermediate products such as 3-chlorophenol were not accumulated, which were immediately dechlorinated to phenol. The enriched microbial community was characterized physiologically by testing the effects of electron donors and electron acceptors on the dechlorinating activity. The dechlorinating activity was promoted with lactate, pyruvate, and hydrogen as electron donors but not with acetate. Electron acceptors, nitrate and sulphate, inhibited the dechlorinating activity competitively but not iron (III). The microbial group associated with the anaerobic dechlorination was characterized by the effect of specific inhibitors on the PCP dechlorination. Effects of specific metabolic inhibitors and antibiotics indicated the involvement of Gram-positive spore-forming bacteria with the PCP dechlorinating activity, which was represented as bacteria of phylum Firmicutes. The structure of the microbial community was characterized by fluorescence in situ hybridization, quinone profiling, and PCR-DGGE (denaturing gel gradient electrophoresis). The combined results indicated the predominance of Clostridium species of phylum Firmicutes in the microbial community. Desulfitobacterium spp. known as anaerobic Gram-positive spore-forming bacteria dechlorinating PCP were not detected by PCR using a specific primer set. These indicated a probable presence of novel anaerobic Gram-positive spore-forming bacteria dechlorinating PCP in the microbial community.

Goal, Scope and Background: Pentachlorophenol (PCP) is the second highest volume pesticide used in the United States. It is a mutagenic compound whose exposure poses significant health effects, One of the most desirable, environmentally friendly treatment methods is bioremediation. For soil-based contamination, the effectiveness of bioremediation will also be affected by the presence of an active indigenous population, sorption of the contaminant onto the soil, and environmental parameters., Methods: Two pure strains and their mixed culture were used to evaluate PCP biodegradation in two different field soils, Columbia (CO) and New Mexico (NM). Biostimulation of the indigenous microbes was evaluated by adding nutrients. The efficiency of adding bacteria strains (bioaugmentation) for degrading PCP was determined with Arthrobacter sp., Flavobacterium sp. and a 50:50 mixture of the two bacteria strains., Results: In CO soil, only 24%, 12% and 25% of the initial PCP concentration were degraded by Flavobacterium sp., Arthrobacter sp. and mixed culture, respectively. Arthrobacter sp. was used in NM soil with two initial concentrations and achieved degradation efficiencies of 57% and 61% for 361 and 95 mg kg- concentrations, respectively. Discussion. Analysis via statistical methods showed that the bacteria had different efficiencies on PCP degradation in each soil. 2, Conclusions: All bacteria catalyzed a higher PCP degradation when present in NM soil. Second, Flavobacterium sp. degraded more PCP than Arthrobacter sp. in CO soil. The mixed culture achieved the highest degradation efficiency regardless of the initial concentration or soil origin., Recommendations and Perspectives: The effect of the soil properties, such as the soil organic matter (SOM) on PCP biodegradation should be investigated. Future work can also investigate the effect of aging time on biodegradation.

Though phytoremediation is widely studied in remediation of metal contaminated soils or organic contaminated soils, little information is available regarding the effectiveness and processes of phytoremediation of sites co-contaminated with organic and metal pollutants. Sites co-contaminated with organic and metal pollutants are common and considered to be a more complex problem as the two components often cause a synergistic effect on cytotoxicity as measured both by growth inhibition and colony-forming ability. In this paper, the dissipation mechanisms for pentachlorophenol (PCP) in copper co-contaminated soil by Lolium perenne L, and Raphanus sativus was investigated in a greenhouse experiment by monitoring the growth response of plants, evaluating the removal efficiency of extractable PCP, differentiating PCP residuals in strongly and loosely adhering rhizosphere soils, and analyzing the microbial activity in the rhizosphere. In copper co-contaminated soil with the initial PCP concentration of 50 mg/kg, plants grew better with the increment of soil Cu level (0, 150, 300 mg/kg), which implied that combinations of inorganic and organic pollutants sometimes exerted antagonistic effects on plant cytotoxicity. The observed higher PCP dissipation in soil spiked with 50 mg/kg PCP in the presence of Cu and the less difference of PCP residual between strongly and loosely adhering soils further suggests the occurrence of Cu-PCP interaction and the enhanced degradation and mass flow are two possible explanations. In copper co-contaminated soil with the initial PCP concentration of 100 mg/kg, however, both plant growth and microbial activity were inhibited with the increment of soil Cu level. The lowered degrading activity of microorganisms and the reduced mass flow were probably responsible for the significantly lower levels of PCP dissipation in copper co-contaminated soil. These results showed that remediation of sites co-contaminated with organic and metal pollutants is a complex problem and a more thorough understanding of the extent and mechanisms by which metals inhibit organic degradation is needed to develop phytoremediation of co-contaminated sites.

Specific physiological groups can be used to evaluate the recovery of soil microbial communities following disturbance. In this study, soil was contaminated with pentachlorophenol (PCP) to assess the resiliency of microorganisms responsible for carbon and nitrogen cycling. Methane fluxes were monitored in soil microcosms to evaluate the effects of contamination and augmentation on microbial populations involved in carbon cycling. The addition of a PCP-mineralizing bacterium, Sphingomonas chlorophenolica strain RA2, enhanced the recovery of methane-oxidizing capacity relative to uninoculated treatments when soil was contaminated with low concentrations of PCP (10 and 50 microg/g soil, ppm). At the highest level of PCP contamination tested (300 ppm), there was no recovery of methane-oxidizing capacity whether or not the PCP-mineralizing bacterium was added to soil. The nitrogen-cycling capacity of contaminated and augmented soil was tested by measuring nitrification potentials. The addition of PCP resulted in a concentration-dependent reduction in nitrification rates. After three months of incubation, all augmented soils had nitrification rates equivalent to the uncontaminated control, and the uninoculated soils contaminated with 100 and 300 ppm PCP still were impaired in their nitrification potentials. These results indicate that biological removal of a contaminant has the potential to restore microbially mediated processes to levels observed prior to contamination.

A study was conducted using two pilot-scale land-treatment units (LTUs) to evaluate the efficacy of different cultivation and maintenance schedules during bioremediation of contaminated soil from a wood treatment facility using landfarming technology. The soil contained high concentrations of polycyclic aromatic hydrocarbons (PAHs, approximately 13000 ppm) as well as of pentachlorophenol (PCP, approximately 1500 ppm). An initial 6-month intensive-treatment phase was followed by 24 months of less-intensive treatment. During the first phase, traditional landfarming practice of regular cultivation was compared with a gas-phase composition based cultivation strategy, and both the landfarming units were intensively monitored and maintained with respect to moisture control and delivery of nutrients. The two strategies resulted in similar contaminant concentration profiles with time during this phase, although different microbial populations developed in the two-landfarming units. The second (less-intensive) treatment phase involved no moisture control and nutrient delivery beyond the initial adjustments, and compared natural attenuation (no cultivation) with quarterly cultivation of soil. Both the strategies showed similar behavior again. GC/MS analysis of the soil samples showed PAH removal including four-ring homologues. Leachability tests at zero time and after 6 and 22 months of operation showed significant reductions in leaching of PCP and low molecular weight PAHs. Extended treatment resulted in some leaching of high molecular weight PAHs. Significant biological activity was demonstrated, even at the high contaminant concentrations. Phospholipid ester-linked fatty acid (PLFA) analysis showed an increase in biomass and a divergence in community composition in soils depending on the treatment conducted.

To develop a dependable approach to introduce laboratory selected, pesticide-degrading microorganisms into soil environments for the purpose of in situ bioremediation, we tested the possibility of utilizing plant rhizospheres as the vehicle. We first established the rhizosphere competency of a strain of the soil fungus Trichoderma harzianum, a biocontrol species well studied by plant pathologists to colonize plant rhizosphere in many parts of the world. The strain we chose, T.h.2023 is resistant to many fungicides, and it has been shown to metabolize several pesticides. Second, we found that it readily metabolized pentachlorophenol (PCP), which is quickly and stochiometrically converted to pentachloroanisole (PCA) in liquid culture. Taking advantage of this specific feature, we have developed a sensitive metabolic marker approach that allowed us to monitor for the continuous presence and activity of this fungal strain in the corn rhizosphere soil in situ over time.

Anaerobic biodegradation of pentachlorophenol (PCP) was studied in rotative bioreactors containing 200 g of PCP-contaminated soil and 250 ml of liquid medium. Reactors were bioaugmented with cells of Desulfitobacterium frappieri strain PCP-1, a bacterium able to dehalogenate PCP to 3-chlorophenol. Cells of strain PCP-1 were detected by quantitative PCR for at least 21 days in reactors containing 500 mg of PCP per kg of soil but disappeared after 21 days in reactors with 750 mg of PCP per kg of soil. Generally, PCP was completely removed in less than 9 days in soils contaminated with 189 mg of PCP per kg of soil. Sorption of PCP to soil organic matter reduced its toxicity and enhanced the survival of strain PCP-1. In some non-inoculated reactors, the indigenous microorganisms of some soils were also able to degrade PCP. These results suggest that anaerobic dechlorination of PCP in soils by indigenous PCP-degrading bacteria, or after augmentation with D. frappieri PCP-1, should be possible in situ and ex situ when the conditions are favourable for the survival of the degrading microorganisms.

A new most probable number (MPN) method for the determination of pentachlorophenol (PCP) degraders in soil using the change in pH due to PCP degradation is compared with a well documented MPN method using radiolabeled PCP. The results of all MPN counts were similar within a 95% confidence limit. The results obtained in MPN per gram of dry soil using pH measurements were 1.8 (+3.1, -1.03) x10 (4) compared to 0.64 (+1.34, -0.42) x 10(4) when using production of [(14)C]CO(2).

We examined the mineralization of pentachlorophenol (PCP) in sterile and non-sterile soil with or without added bacteria (Mycobacterium chlorophenolicum PCP-1). The soil used had no history of PCP contamination. Microcosms (30 g dry weight of soil) were incubated with labelled PCP (6.76% 13C, a non-radioactive stable isotope, 22 mg kg-1 dry weight) for 60 days. M. chlorophenolicum PCP-1 (7.8 x 10(6) cells g-1 dry weight) was added to some samples. 50% of the PCP was mineralized in non-sterile soil with or without the exogenous bacteria. Only 5% of the PCP was mineralized in sterile soil with or without bacteria. These data suggest that the PCP was not accessible to M. chlorophenolicum and that the indigenous soil microflora can mineralize PCP.

Anaerobic biodegradation of pentachlorophenol (PCP) in a contaminated soil from a wood-treating industrial site was studied in soil slurry microcosms inoculated with a PCP-degrading methanogenic consortium. When the microcosms containing 10%-40% (w/v) soil were inoculated with the consortium, more than 90% of the PCP was removed in less than 30 days at 29 degrees C. Less-chlorinated phenols, mainly 3-chlorophenol were slowly degraded and accumulated in the cultures. Addition of glucose and sodium formate to the microcosms was not necessary, suggesting that the organic compounds in the soil can sustain the dechlorinating activity. Inoculation of Desulfitobacterium frappieri strain PCP-1 along with a 3-chlorophenol-degrading consortium in the microcosms also resulted in the rapid dechlorination of PCP and the slow degradation of 3-chlorophenol. Competitive polymerase chain reaction experiments showed that PCP-1 was present at the same level throughout the 21-day biotreatment. D. frappieri, strain PCP-1, inoculated into the soil microcosms, was able to remove PCP from soil containing up to 200 mg PCP/kg soil. However, reinoculation of the strain was necessary to achieve more than 95% PCP removal with a concentration of 300 mg and 500 mg PCP/kg soil. These results demonstrate that D. frappieri strain PCP-1 can be used effectively to dechlorinate PCP to 3-chlorophenol in contaminated soils.

A pentachlorophenol (PCP)-degrading bacterium was isolated from possible PCP-contaminated soil from Pusan, Korea and identified as a member of the genus Pseudomonas. It used PCP as its sole source of carbon and energy. This micro-organism was capable of degrading PCP more effectively, certified by the increase in cell density and the decrease in PCP substrate. Pseudomonas sp. Bu34 was able to degrade a much higher concentration of PCP (4000 mg l-1) than any previously reported PCP-degrading bacteria and fungi and to grow in mineral salts solution containing one of a variety of chlorophenols. In non-acclimated strain Bu34, the cell number decreased from 87 to 99.9% in 75-4000 mg l-1 PCP at 24 h. In the acclimated strain the PCP toxic effect did not appear with 75 mg l-1 PCP treatment, but 1000-4000 mg l-1 PCP decreased the cell number of strain Bu34 by 25% to 24 h and then the cell number slightly increased at 48 h. Therefore, it suggested that the maximum resistance of acclimated strain Bu34 to PCP was 4000 mg l-1 PCP. We suggest that strain Bu34 could be used as a micro-organism for the bioremediation of highly PCP-contaminated soils, water or wood products.

Bacteria isolated from a pentachlorophenol (PCP) contaminated site grew in the presence of 50 micrograms PCP/mL but were not able to degrade it in either liquid medium or the presence of 1% sterile potting soil as a solid support. Probes developed using the gene sequence of PCP-4-monooxygenase (pcpB) from Sphingomonas chlorophenolica sp.nov hybridized to two separate isolates. Identification based on fatty acid methyl ester profiles (Sherlock), substrate utilization (BIOLOG), and 16S rRNA showed that the two strains were different from each other and from Sphingomonas chlorophenolica. Sequences from these isolates, amplified by polymerase chain reaction, confirmed the homology with pcpB. The presence of pcpB sequences in these nondegraders indicated that growth and hybridization data alone were insufficient for predicting degradation capability.

Columns of an Alfisol under permanent pasture were polluted by repeated additions of pentachlorophenol (PCP) (7 mg l-1) to levels of 102 and 510 mg Kg-1, to simulate a dynamic diffuse pollution. PCP was rapidly sorbed to the soil organic matter, and was only slightly degraded. Measurements of soil microbial biomass-C revealed a 25% decrease in total biomass-C caused by both leaching and PCP toxicity. Microbial biomass-C measurements performed on soil fractions showed that only microorganisms located in the outer compartment of the aggregates were affected. Microorganisms protected by soil micro-aggregates were not affected, suggesting that they were not in contact with PCP, which was thus unavailable for biodegradation. Three gram negative bacterial strains (Si, C3 and C2), able to use PCP as a sole carbon and energy source, were isolated after 0, 1 and 3 months of PCP enrichment respectively, and were identified as Pseudomonas (Si) and Acinetobacter (C3 and C2). In liquid degradation tests, the strains C2 and C3 degraded 60% of PCP within 26 days whereas the Pseudomonas degraded only 25%. A specific immuno-labeling of the three strains permitted to show that repeated PCP additions to soil had a positive, negative or absence of effect on the populations C2, C3 and Si respectively.

Sterilised and non-sterilised soils contaminated with pentachlorophenol (PCP) were inoculated with solid substrate cultures of Lentinula edodes LE2 ("shiitake" mushroom) to simulate monoculture bioremediation treatments and treatments in which the fungus competes with natural microflora. With monocultures of L. edodes, rates of PCP depletion were rapid for the initial 4 weeks and, although thereafter the rate decreased, 99% biotransformation was obtained in 10 weeks. In mixed culture, PCP biotransformation by L. edodes was markedly slower and only 42% of the PCP was depleted after 10 weeks. Maximal rates of PCP transformation, biomass (ergosterol) accumulation and oxidative enzymes (phenol oxidase and manganese-peroxidase) production were observed after 2 weeks of incubation. In monocultures, phenol oxidase activity was 195.5 U g-1 and Mn-peroxidase 138.4 U g-1. In mixed cultures, fungal enzyme activities were markedly lower: 70.33 U g-1 for phenol oxidase and 85.0 g-1 for Mn-peroxidase. Analyses of soil metabolites after 10 weeks revealed that monocultures of L. edodes had eliminated both PCP and pentachloroanisole. Pentachloroanisole, however, was detected in soils with the mixed microflora. Both dechlorination and mineralisation of the xenobiotic compound were effected by L. edodes LE2.

Recovery of 97.5% of the pentachlorophenol (PCP) in contaminated wood powder was obtained after extraction with 0.1% KOH solution at 60 degrees C for 75 min. Extraction with NaOH and Na2CO3 was less effective than KOH. The neutralized extract was treated using a methanogenic consortium in an upflow anaerobic fixed-film reactor. The reactor was operated at 29 degrees C for over 600 d. The best performance of the reactor was observed when the PCP liquor was supplemented with glucose and formate. Complete dechlorination of PCP and phenol removal was obtained for a PCP loading rate of 13.3-18.0 mg l(-1) of reactor volume d(-1) with recirculation of the effluent and a hydraulic retention time (HRT) of 0.5-0.6 d.

The fate of pentachlorophenol (PCP) in soil under natural conditions was investigated. It was revealed that the total amount of PCP significantly decreased when soil was inoculated by Streptomyces rochei 303, a strain-destructor of chlorophenols. The products of PCP transformation, such as tetra- and trichlorophenols, pentachlorobenzene, chlorinated dioxins, were identified after the first month of the experiment. Their quantity was less in the variant with the introduced strain compared to control.

The nutristat, a substrate concentration-controlled continuous culture, was used to grow pentachlorophenol (PCP)-degrading microorganisms. The PCP concentration control system consisted of on-line measurement of the PCP concentration in the culture vessel with a tangential filter and a flowthrough spectrophotometer. With PCP concentrations between 45 and 77 microM, a stable situation was established in the nutristat, with an average dilution rate of 0.035 +/- 0.003 h-1. Compared with those of fed-batch cultures and chemostat cultures, the growth rates of microorganisms in the PCP nutristat were significantly higher, leading to considerable time savings in the enrichment procedure. In addition, PCP accumulation to severe inhibitory levels in the culture is prevented because the set point determines the (maximum) PCP concentration in the culture. The use of the nutristat as a tool for the growth of bacteria that degrade toxic compounds is discussed.

A Gram-negative bacterium, Pseudomonas sp. strain SR3, was isolated from soil at a former wood treatment plant in north central Florida. The ability of this bacterium to degrade pentachlorophenol (PCP) was confirmed by growing cells in a basal salts medium in which PCP was the only source of carbon and energy. Degradation from a measured concentration of 39-40 micrograms PCP/ml to 0.0006 micrograms PCP/ml was observed within 120 h of incubation in the presence of PCP-induced cells of Pseudomonas sp. strain SR3. The initial cell density in these cultures was 6 x 10(6) cfu/ml. Microtox 5 min EC50 toxicity tests revealed that aqueous solutions of PCP, measured concentrations 39-40 micrograms/ml were toxic but that final biodegraded samples, 0.0006 micrograms PCP/ml were nontoxic. However, bioassays with embryonic inland silversides, Menidia beryllina, showed that the biodegraded samples were embryotoxic or teratogenic. Water containing added PCP at concentrations up to 30 times higher than measured in the final biodegraded samples was less toxic/teratogenic. These results indicate that while biodegradation of PCP was nearly complete, intermediate metabolites of the degradation process or undegraded impurities in PCP were toxic or teratogenic. Thus, the M. beryllina bioassay allows extremely sensitive assessment of toxicity associated with biodegraded environmental pollutants and may be a useful criterion for determining whether bioremediated water or soil is safe for discharge back into the environment.

A collection of 1056 strains in our laboratory were incubated with various xenobiotics among which were two potent fungicides: pentachlorophenol (PCP) and pentachloronitrobenzene (PCNB). The production of extracellular phenoloxidases were examined, using a series of ten different reagents. On the whole, PCNB is less accessible to fungal degradation than PCP. Although no correlation was found between the biodegradative capability of individual fungal strains for PCP or for PCNB, when taxonomic groups were considered as a whole, the same activity profiles were found. Zygomycetes were the most efficient; yeasts the least efficient towards both substrates. A more detailed study of the metabolism of both substrates on selected strains is in progress.

The reductive dechlorination of pentachlorophenol (PCP) was investigated in anaerobic sediments that contained nonadapted or 2,4- or 3,4-dichlorophenol (DCP)-adapted microbial communities. Adaptation of sediment communities increased the rate of conversion of 2,4- or 3,4-DCP to monochlorophenols (CPs) and eliminated the lag phase before dechlorination was observed. Both 2,4- and 3,4-DCP-adapted sediment communities dechlorinated the six DCP isomers to CPs. The specificity of chlorine removal from the DCP isomers indicated a preference for ortho-chlorine removal by 2,4-DCP-adapted sediment communities and for para-chlorine removal by 3,4-DCP-adapted sediment communities. Sediment slurries containing nonadapted microbial communities either did not dechlorinate PCP or did so following a lag phase of at least 40 days. Sediment communities adapted to dechlorinate 2,4- or 3,4-DCP dechlorinated PCP without an initial lag phase. The 2,4-DCP-adapted communities initially removed the ortho-chlorine from PCP, whereas the 3,4-DCP-adapted communities initially removed the para-chlorine from PCP. A 1:1 mixture of the adapted sediment communities also dechlorinated PCP without a lag phase. Dechlorination by the mixture was regiospecific, following a para greater than ortho greater than meta order of chlorine removal. Intermediate products of degradation, 2,3,5,6-tetrachlorophenol, 2,3,5-trichlorophenol, 3,5-DCP, 3-CP, and phenol, were identified by a combination of cochromatography (high-pressure liquid chromatography) with standards and gas chromatography-mass spectrometry.

The effects of physical, chemical, and biological treatments on biodegradation of pentachlorophenol (PCP) were studied in a silt-loam soil contaminated with 175 mg PCP/kg and uniformly 14C-labelled PCP. Biodegradation of 14C-labelled PCP and technical-grade PCP were monitored over 210 days incubation. Mineralization of labelled PCP was significantly (p=0.05) influenced by soil treatments. Negligible biodegradation occurred in either the sterile control soil or the uninoculated control soil, with less than 1% of added 14C recovered as 14 CO2. Inoculation of unamended soil with a strain of Flavobacterium (ATCC 39723) known to degrade PCP increased biodegradation of PCP; approximately 60% of the [14C]PCP was recovered as 14CO2. Increased soil water content (60% versus 30% w/w) enhanced biodegradation (67% recovery of 14C as CO2), while increased chloride ion concentration and anoxic conditions were inhibitory (20 and 1% recoveries, respectively). Residual soil PCP concentrations were also influenced by various treatments. In the sterile control soil and noninoculated control, after 210 days incubation, concentrations of PCP were 143 and 1223 mg/kg, respectively, while the PCP concentration in the inoculated soil was 21 mg/kg. When soil organic matter was increased by adding finely ground red clover leaf and stem material, the residual PCP concentration was reduced to 6 mg/kg after 210 days. Increased soil water content resulted in a residual PCP concentration of 5 mg/kg. High-pressure liquid chromatography of soil extracts revealed no accumulation of partial PCP degradation products. The results indicated that biodegradation of PCP in soil was significantly influenced by various soil amendments.

Shake flask studies examined the rate and extent of biodegradation of pentachlorophenol (PCP) and 42 components of coal-tar creosote present in contaminated groundwater recovered from the American Creosote Works Superfund site, Pensacola, Fla. The ability of indigenous soil microorganisms to remove these contaminants from aqueous solutions was determined by gas chromatographic analysis of organic extracts of biotreated groundwater. Changes in potential environmental and human health hazards associated with the biodegradation of this material were determined at intervals by Microtox assays and fish toxicity and teratogenicity tests. After 14 days of incubation at 30 degrees C, indigenous microorganisms effectively removed 100, 99, 94, 88, and 87% of measured phenolic and lower-molecular-weight polycyclic aromatic hydrocarbons (PAHs) and S-heterocyclic, N-heterocyclic, and O-heterocyclic constituents of creosote, respectively. However, only 53% of the higher-molecular-weight PAHs were degraded; PCP was not removed. Despite the removal of a majority of the organic contaminants through biotreatment, only a slight decrease in the toxicity and teratogenicity of biotreated groundwater was observed. Data suggest that toxicity and teratogenicity are associated with compounds difficult to treat biologically and that one may not necessarily rely on indigenous microorganisms to effectively remove these compounds in a reasonable time span; to this end, alternative or supplemental approaches may be necessary. Similar measures of the toxicity and teratogenicity of treated material may offer a simple, yet important, guide to bioremediation effectiveness.

With a new enrichment protocol, pentachlorophenol (PCP)-degrading bacteria were isolated from soil, water, and sewage. When characterized, all isolates were related and shared characteristics of the genus Arthrobacter. Growth rates for strain NC were determined for a number of substrates, including PCP and 2,4,6-trichlorophenol. Changes in PCP concentration affected growth rate and length of the lag phase but not cell yield. Increasing the pH from 6.8 to 7.8 decreased the length of the lag phase for growth on PCP. Cessation of growth, upon incremental addition of PCP, was found to be related to a decrease in pH rather than to a buildup of a toxic metabolite. Degradation of PCP by strain NC was shown to be complete.

Metabolism of pentachlorophenol (PCP)-ring-14C(U) by a microorganism (Pseudomonas sp.) isolated from soil was examined. The microorganism degraded PCP-14C rapidly, and released 14CO2 equivalent to approximately 50% of the PCP-14c added to the bacterial cell suspension in 1 hour of incubation. The results of amino acid analysis of the bacterial cells incubated with PCP-14C showed that radioactive carbon derived from PCP-14C was incorporated rapidly into the cell constituents, and that the pattern of 14C-amino acids in the cell constituents was not much different between the 15 minute and 24 hour incubation periods. Intermediate metabolities of PCP isolated from the incubation medium were identified as tetrachlorocatechol and tetrachlorohydroquinone by spectral analyses.