Bacterial diversity and function of aerobic granules engineered in a sequencing batch reactor for phenol degradation

Aerobic granules are self-immobilized aggregates of microorganisms and represent a relatively new form of cell immobilization developed for biological wastewater treatment. In this study, both culture-based and culture-independent techniques were used to investigate the bacterial diversity and function in aerobic phenol- degrading granules cultivated in a sequencing batch reactor. Denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rRNA genes demonstrated a major shift in the microbial community as the seed sludge developed into granules. Culture isolation and DGGE assays confirmed the dominance of beta-Proteobacteria and high-G+C gram-positive bacteria in the phenol-degrading aerobic granules. Of the 10 phenol-degrading bacterial strains isolated from the granules, strains PG-01, PG-02, and PG-08 possessed 16S rRNA gene sequences that matched the partial sequences of dominant bands in the DGGE fingerprint belonging to the aerobic granules. The numerical dominance of strain PG-01 was confirmed by isolation, DGGE, and in situ hybridization with a strain-specific probe, and key physiological traits possessed by PG-01 that allowed it to outcompete and dominate other microorganisms within the granules were then identified. This strain could be regarded as a functionally dominant strain and may have contributed significantly to phenol degradation in the granules. On the other hand, strain PG-08 had low specific growth rate and low phenol degradation ability but showed a high propensity to autoaggregate. By analyzing the roles played by these two isolates within the aerobic granules, a functional model of the microbial community within the aerobic granules was proposed. This model has important implications for rationalizing the engineering of ecological systems.     

Physiological traits of bacterial strains isolated from phenol-degrading aerobic granules

The physiological characteristics of ten bacterial strains isolated from phenol-degrading aerobic granules were evaluated in order to identify competitive traits for dominant growth in aerobic granules. The ten strains showed a wide diversity in specific growth rates and oxygen utilization kinetics, and could be divided into four catabolic types of phenol degradation. While some strains degraded phenol mainly via the meta pathway or the ortho pathway, other strains degraded phenol via both these pathways. The ten strains also exhibited high levels of autoaggregation and coaggregation activity. Within the collection of ten strains, 36.7% of all possible strain pairings displayed a measurable degree of coaggregation. Strain PG-08 possessed the strongest autoaggregation activity and showed significant coaggregation (coaggregation indices of 67% to 74%) with PG-02. The three strains PG-01, PG-02 and PG-08 belonging to dominant groups in the granules possessed different competitive characteristics. Microcosm experiments showed the three strains could not coexist at the high phenol concentration of 250 mg L(-1), but could coexist at lower phenol concentrations in a spatially heterogeneous environment. This study illustrated that the spatial heterogeneity provided by the aerobic granules led to niche differentiation and increased physiological diversity in the resident microbial community.     

Bioaugmentation and coexistence of two functionally similar bacterial strains in aerobic granules

The survival of the inoculated microbial culture is critical for successful bioaugmentation but impossible to predict precisely. As an alternative strategy, bioaugmentation of a group of microorganisms may improve reliability of bioaugmentation. This study evaluated simultaneous bioaugmentation of two functionally similar bacterial strains in aerobic granules. The two strains, Pandoraea sp. PG-01 and Rhodococcus erythropolis PG-03, showed high phenol degradation and growth rates in phenol medium, but they were characterized as having a poor aggregation activity and weak bioflocculant-producing and biofilm-forming abilities. In the spatially homogeneous batch conditions, strain PG-01 with higher growth rates outcompeted strain PG-03. However, the two strains could stably coexist in the spatially heterogeneous conditions. Then the two strains were mixed and bioaugmented into activated sludge in two sequencing batch reactors, which were operated with the different settling times of 5 and 30 min, respectively. Aerobic granules were developed only in the reactor with a settling time of 5 min. Fluorescence in situ hybridization and denaturing gradient gel electrophoresis showed that the two strains could coexist in aerobic granules but not in activated sludge. These findings suggested that the compact structure of aerobic granules provided spatial isolation for coexistence of competitively superior and inferior strains with similar functions.     

Biodegradation of dimethyl phthalate by Sphingomonas sp. isolated from phthalic-acid-degrading aerobic granules

This study isolated nine strains of aerobic phenol-degrading granules. These isolates (I1–I9) were characterized using 16S rRNA gene sequencing, with γ-Proteobacteria as the dominant strains in the aerobic granules. While most strains demonstrated either high phenol-degrading capabilities or auto-aggregation capabilities, three isolates, I2, I6, and I8 showed both features. These findings contradict the previous view that auto-aggregation and phenol degradation are mutually exclusive in aerobic granules. Strains I2 and I8 independently formed single-culture aerobic granules except for I3. Anti-microbial activity test results indicated that strains I2 and I8 inhibited growth of strain I3. However, co-culturing I3 with I2 or I8 helped to form granules.     

两株高效苯酚降解菌的筛选、鉴定及生物强化-DTRO组合工艺初步验证

【目的】从煤化工废水中分离、筛选苯酚高效降解微生物,初步考察微生物与DTRO技术联用,构建含酚废水生物强化处理工艺的可行性。【方法】采用苯酚浓度梯度培养基对苯酚降解微生物进行分离和筛选;根据菌体形态电子显微镜观察、菌株生理生化特性考察和16S r RNA基因系统发育树构建,对菌株进行初步生物学鉴定;将筛选出的高效苯酚降解菌制备成相应的菌剂与碟管式反渗透(DTRO)技术组合形成"生物强化-DTRO"工艺,并试用于含酚废水的处理。【结果】共获得7株纯化细菌,其中Phe-03和Phe-05为高效苯酚降解菌;该2株菌均可以苯酚为唯一碳源生长。经鉴定Phe-03为壤霉菌属(Agromyces)菌株;Phe-05为棒杆菌属(Corynebacterium)菌株。到目前为止,壤霉菌属(Agromyces)菌株降解苯酚尚未见报道。在初始苯酚浓度达到1 300 mg/L条件下,Phe-03和Phe-05菌株44 h内对苯酚降解率均达到70%以上;76 h后苯酚降解率均超过90%。组合形成的"生物强化-DTRO"工艺不仅可以有效去除废水中的酚类化合物,而且还能减少反渗透膜污染,以及增加膜的通透性。【结论】研究表明微生物技术可与DTRO技术联用,构建含酚废水生物强化处理工艺,可为含酚废水处理技术研究提供一种选择思路。     

Degradation of phenol by aerobic granules and isolated yeast Candida tropicalis

Aerobic granules effectively degrade phenol at high concentrations. This work cultivated aerobic granules that can degrade phenol at a constant rate of 49 mg-phenol/g x VSS/h up to 1,000 mg/L of phenol. Fluorescent staining and confocal laser scanning microscopy (CLSM) tests demonstrated that an active biomass was accumulated at the granule outer layer. A strain with maximum ability to degrade phenol and a high tolerance to phenol toxicity isolated from the granules was identified as Candida tropicalis via 18S rRNA sequencing. This strain degrades phenol at a maximum rate of 390 mg-phenol/g x VSS/h at pH 6 and 30 degrees C, whereas inhibitory effects existed at concentrations >1,000 mg/L. The Haldane kinetic model elucidates the growth and phenol biodegradation kinetics of the C. tropicalis. The fluorescence in situ hybridization (FISH) and CLSM test suggested that the Candida strain was primarily distributed throughout the surface layer of granule; hence, achieving a near constant reaction rate over a wide range of phenol concentration. The mass transfer barrier provided by granule matrix did not determine the reaction rates for the present phenol-degrading granule.     

Comparing activated sludge and aerobic granules as microbial inocula for phenol biodegradation

Activated sludge and acetate-fed granules were used as microbial inocula to start up two sequencing batch reactors (R1, R2) for phenol biodegradation. The reactors were operated in 4-h cycles at a phenol loading of 1.8 kg m^–3 day^–1. The biomass in R1 failed to remove phenol and completely washed out after 4 days. R2 experienced initial difficulty in removing phenol, but the biomass acclimated quickly and effluent phenol concentrations declined to 0.3 mg l^–1 from day 3. The acetate-fed granules were covered with bacterial rods, but filamentous bacteria with sheaths, presumably to shield against toxicity, quickly emerged as the dominant morphotype upon phenol exposure. Bacterial adaptation to phenol also took the form of modifications in enzyme activity and increased production of extracellular polymers. 16S rRNA gene fingerprints revealed a slight decrease in bacterial diversity from day 0 to day 3 in R1, prior to process failure. In R2, a clear shift in community structure was observed as the seed evolved into phenol-degrading granules without losing species-richness. The results highlight the effectiveness of granules over activated sludge as seed for reactors treating toxic wastewaters.     

Development of an efficient method for screening microorganisms by using symbiotic association between <Emphasis Type="Italic">Nasutitermes takasagoensis</Emphasis> and intestinal microorganisms

Screening method of microorganisms that utilized the symbiotic association between insect (Nasutitermes takasagoensis: Nt) and intestinal microorganisms was developed. The existence of desired microorganisms that grew by degrading difficult-to-degrade materials in the gut was detected using survivability of Nt as an indicator. The desired microorganisms were isolated from the survived Nt. It was thought that guts of Nt behave as continuous culture systems whereby microorganisms that cannot degrade diet components are washed out, whereas those that can degrade it are retained and concentrated in the gut. About 60% of Nt fed with phenol artificial diet (PAD) died within 7 days, while 4% of termites survived for 9 days. The structure of intestinal microorganisms of the survived Nt fed with PAD differed from the bacterial communities obtained from enrichment culture (which contained phenol) of wood-feeding Nt. Relatively high colonies (650-times) were detected in the gut of Nt fed on phenol artificial diet compared with those obtained when Nt was fed on wood. Seven denaturing gradient gel electrophoresis (DGGE) bands were detected from gut of wood-feeding Nt, whereas 11 DGGE-bands were detected from that of phenol-feeding Nt. Out of 11 DGGE-bands, 5 of them were sequenced, and bacterial species including phenol-degrading bacteria were identified.     

Properties of phenol-removal aerobic granules during normal operation and shock loading

The physical structure and activity of aerobic granules, and the succession of bacterial community within aerobic granules under constant operational conditions and shock loading were investigated in one sequencing batch reactor over ten months. While the maturation phase of the granulation process began on day 30, the structure of microbial community changed markedly until after three months of reactor operation under constant conditions with a loading rate of 1.5 g phenol L⁻¹ day⁻¹. A shock loading of 6.0 g phenol L⁻¹ day⁻¹ from days 182–192 led to divergence of bacterial community, an inhibition of the biomass activity, and a decrease in phenol removal rate in the reactor. However, phenol was still completely removed under this disturbance. After the shock loading, the mean sizes of aerobic granules increased, and the activity of the microbial population within the granules decreased, although there appeared highly resilient for the dominant bacterial community of aerobic granules which mainly included β-Proteobacteria. Correlation analysis suggested that biomass concentration and biomass loading were significantly related to the community composition of aerobic granules during the whole operational period. The development of a relatively stable bacterial community in aerobic granules implied that those distinct dominant microbes in aerobic granules were favorably selected and proliferated under the operational conditions.     

Biodegradation of Phenol: Mechanisms and Applications

Phenol, or hydroxybenzene, is both a synthetically and naturally produced aromatic compound. Microorganisms capable of degrading phenol are common and include both aerobes and anaerobes. Many aerobic phenol-degrading microorganisms have been isolated and the pathways for the aerobic degradation of phenol are now firmly established. The first steps include oxygenation of phenol by phenol hydroxylase enzymes to form catechol, followed by ring cleavage adjacent to or in between the two hydroxyl groups of catechol. Phenol hydroxylases ranging from simple flavoprotein monooxygenases to multicomponent hydroxylases, as well as the genes coding for these enzymes, have been described for a number of aerobic phenol-degrading microorganisms. Phenol can also be degraded in the absence of oxygen. Our knowledge of this process is less advanced than that of the aerobic process, and only a few anaerobic phenol-degrading bacteria have been isolated to date. Convincing evidence from both pure culture studies with the denitrifying organism Thauera aromatica K172 and with two Clostridium species, as well as from mixed culture studies, indicates that the first step in anaerobic phenol degradation is carboxylation in the para-position to form 4-hydroxybenzoate. Following para-carboxylation, thioesterification of 4-hydroxybenzoate to co-enzyme A allows subsequent ring reduction, hydration, and fission. Para-carboxylation appears to be involved in the anaerobic degradation of a number of aromatic compounds. Numerous practical applications exist for microbial phenol degradation. These include the exploitation of indigenous anaerobic phenol-degrading bacteria in the in situ bioremediation of creosote-contaminated subsurface environments, and the use of phenol as a co-substrate for indigenous aerobic phenol-degrading bacteria to enhance in situ biodegradation of chlorinated solvents.     

Salt-tolerant phenol-degrading microorganisms isolated from Amazonian soil samples

Two phenol-degrading microorganisms were isolated from Amazonian rain forest soil samples after enrichment in the presence of phenol and a high salt concentration. The yeast Candida tropicalis and the bacterium Alcaligenes faecoalis were identified using several techniques, including staining, morphological observation and biochemical tests, fatty acid profiles and 16S/18S rRNA sequencing. Both isolates, A. faecalis and C. tropicalis, were used in phenol degradation assays, with Rhodococcus erythropolis as a reference phenol-degrading bacterium, and compared to microbial populations from wastewater samples collected from phenol-contaminated environments. C. tropicalis tolerated higher concentrations of phenol and salt (16 mM and 15%, respectively) than A. faecalis (12 mM and 5.6%). The yeast also tolerated a wider pH range (3-9) during phenol degradation than A. faecalis (pH 7-9). Phenol degradation was repressed in C. tropicalis by acetate and glucose, but not by lactate. Glucose and acetate had little effect, while lactate stimulated phenol degradation in A. faecalis. To our knowledge, these soils had never been contaminated with man-made phenolic compounds and this is the first report of phenol-degrading microorganisms from Amazonian forest soil samples. The results support the idea that natural uncontaminated environments contain sufficient genetic diversity to make them valid choices for the isolation of microorganisms useful in bioremediation.     

Intraspecific protoplast fusion of Candida tropicalis for enhancing phenol degradation

Auxotrophic mutants and phenol-degrading defective mutants were separately isolated in a phenol-utilizing strain of Candida tropicalis M4, and were hybridized through protoplast fusion. Some protoplast fusants with phenol-degrading ability were obtained and were very stable. Two of the fusants exhibited slightly higher rates of growth than did the wild strain when the cells were grown on phenol medium, and they possessed about 1.9 and 2.2 times respectively higher phenol hydroxylase activity than the wild strain.     

Development of a microtitre plate method for determination of phenol utilization, biofilm formation and respiratory activity by environmental bacterial isolates

Twenty-five aerobic phenol-degrading bacteria, isolated from different environmental samples on phenol agar after several subcultures in phenol broth, utilized phenol (0.2 g l⁻¹) within 24 h, but removal of phenol was more rapid when other carbon sources were also present. A microtitre plate method was developed to determine growth rate, biofilm formation and respiratory activity of the strains isolated. Pseudomonas putida strains C5 and D6 showed maximum growth (as O.D. at 600 nm), P. putida D6 and unidentified bacterial strain M1 were more stable at high concentrations of phenol (0.8 g l⁻¹), and P. putida C5 formed the greatest amount of biofilm in 0.5 g phenol l⁻¹ medium. Measurement of dehydrogenase activity as reduction of triphenyl tetrazolium chloride supported data on growth rate and biofilm formation. The microtitre plate method provided a selective method for detection of the best phenol degrading and biofilm-forming microorganisms, and was also a rapid, convenient means of studying the effect of phenol concentration on growth rate and biofilm formation.     

Isolation and characterization of a new denitrifying spirillum capable of anaerobic degradation of phenol

Two kinds of phenol-degrading denitrifying bacteria, Azoarcus sp. strain CC-11 and spiral bacterial strain CC-26, were isolated from the same enrichment culture after 1 and 3 years of incubation, respectively. Both strains required ferrous ions for growth, but strain CC-26 grew better than strain CC-11 grew under iron-limited conditions, which may have resulted in the observed change in the phenol-degrading bacteria during the enrichment process. Strain CC-26 grew on phenol, benzoate, and other aromatic compounds under denitrifying conditions. Phylogenetic analysis of 16S ribosomal DNA sequences revealed that this strain is most closely related to a Magnetospirillum sp., a member of the alpha subclass of the class Proteobacteria, and is the first strain of a denitrifying aromatic compound-degrading bacterium belonging to this group. Unlike previously described Magnetospirillum strains, however, this strain did not exhibit magnetotaxis. It grew on phenol only under denitrifying conditions. Other substrates, such as acetate, supported aerobic growth, and the strain exhibited microaerophilic features.     

Ruminococcus bromii, identification and isolation as a dominant community member in the rumen of cattle fed a barley diet

AIMS: To identify dominant bacteria in grain (barley)-fed cattle for isolation and future use to increase the efficiency of starch utilization in these cattle. METHODS AND RESULTS: Total DNA was extracted from samples of the rumen contents from eight steers fed a barley diet for 9 and 14 days. Bacterial profiles were obtained using denaturing gradient gel electrophoresis (DGGE) of the PCR-amplified V2/V3 region of the 16S rRNA genes from total bacterial DNA. Apparently dominant bands were excised and cloned, and the clone insert sequence was determined. One of the most common and dominant bacteria present was identified as Ruminococcus bromii. This species was subsequently isolated using traditional culture-based techniques and its dominance in the grain-fed cattle was confirmed using a real-time Taq nuclease assay (TNA) designed for this purpose. In some animals, the population of R. bromii reached densities above 10(10)R. bromii cell equivalents per ml or approximately 10% of the total bacterial population. CONCLUSIONS: Ruminococcus bromii is a dominant bacterial population in the rumen of cattle fed a barley-based diet. SIGNIFICANCE AND IMPACT OF THE STUDY: Ruminococcus bromii YE282 may be useful as a probiotic inoculant to increase the efficiency of starch utilization in barley-fed cattle. The combination of DGGE and real-time TNA has been an effective process for identifying and targeting for isolation, dominant bacteria in a complex ecosystem.     

Biodegradation efficiency of functionally important populations selected for bioaugmentation in phenol- and oil-polluted area

Denaturing gradient gel electrophoresis of amplified fragments of genes coding for 16S rRNA and for the largest subunit of multicomponent phenol hydroxylase (LmPH) was used to monitor the behaviour and relative abundance of mixed phenol-degrading bacterial populations (Pseudomonas mendocina PC1, P. fluorescens strains PC18, PC20 and PC24) during degradation of phenolic compounds in phenolic leachate- and oil-amended microcosms. The analysis indicated that specific bacterial populations were selected in each microcosm. The naphthalene-degrading strain PC20 was the dominant degrader in oil-amended microcosms and strain PC1 in phenolic leachate microcosms. Strain PC20 was not detectable after cultivation in phenolic leachate microcosms. Mixed bacterial populations in oil-amended microcosms aggregated and formed clumps, whereas the same bacteria had a planktonic mode of growth in phenolic leachate microcosms. Colony hybridisation data with catabolic gene specific probes indicated that, in leachate microcosms, the relative proportions of bacteria having meta (PC1) and ortho (PC24) pathways for degradation of phenol and p-cresol changed alternately. The shifts in the composition of mixed population indicated that different pathways of metabolism of aromatic compounds dominated and that this process is an optimised response to the contaminants present in microcosms.     

Polymerase chain reaction-denaturing gradient gel electrophoresis analysis of bacterial community structure in the food,intestines, and feces of earthworms

The bacterial communities in the food, intestines, and feces of earthworms were investigated by PCR-denaturing Gradient gel electrophoresis (DGGE). In this study, PCR-DGGE was optimized by testing 6 universal primer sets for microbial 16S rRNA in 6 pure culture strains of intestinal microbes in earthworms. One primer set effectively amplified 16S rRNA from bacterial populations that were found in the food, intestines, and feces of earthworms. Compared with the reference markers from the pure culture strains, the resulting DGGE profiles contained 28 unique DNA fragments. The dominant microorganisms in the food, intestines, and feces of earthworms included Rhodobacterales bacterium, Fusobacteria, Ferrimonas marina, Aeromonas popoffii, and soil bacteria. Other straisn, such as Acinetobacter, Clostridium, and Veillonella, as well as rumen bacteria and uncultured bacteria also were present. These results demonstrated that PCR-DGGE analysis can be used to elucidate bacterial diversity and identify unculturable microorganisms.     

Aggregation of immobilized activated sludge cells into aerobically grown microbial granules for the aerobic biodegradation of phenol

AIMS: The aim of this study is to evaluate the utility of aerobically grown microbial granules for the biological treatment of phenol-containing wastewater. METHODS AND RESULTS: A column-type sequential aerobic sludge blanket reactor was inoculated with activated sludge and fed with phenol as the sole carbon source, at a rate of 1.5 g phenol l-1 d-1. Aerobically grown microbial granules first appeared on day 9 of reactor operation and quickly grew to displace the seed flocs as the dominant form of biomass in the reactor. These granules were compact and regular in appearance, and consisted of bacterial rods and cocci and fungi embedded in an extracellular polymeric matrix. The granules had a mean size of 0.52 mm, a sludge volume index of 40 ml g-1 and a specific oxygen utilization rate of 110 mg oxygen g VSS-1 h-1 (VSS stands for volatile suspended solids). Specific phenol degradation rates increased with phenol concentration from 0 to 500 mg phenol l-1, peaked at 1.4 g phenol g VSS-1 d-1, and declined with further increases in phenol concentration as substrate inhibition effects became important. CONCLUSIONS: Aerobically grown microbial granules were successfully cultivated in a reactor maintained at a loading rate of 1.5 g phenol l-1 d-1. The granules exhibited a high tolerance towards phenol. Significant rates of phenol degradation were attained at phenol concentrations as high as 2 g l-1. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to demonstrate the ability of aerobically grown microbial granules to degrade phenol. These granules appear to represent an excellent immobilization strategy for microorganisms to biologically remove phenol and other toxic chemicals in high-strength industrial wastewaters.     

采用苯酚羟化酶基因特异引物检测苯酚降解菌

根据苯酚羟化酶基因高度保守序列设计了一对该基因的特异PCR引物。采用该特异引物从苯酚降解菌醋酸钙不动杆菌 (Acinetobactercalcoaceticus)PHEA 2的总DNA中扩增到唯一一条大小为 684bp的片段。该DNA片段与已知的A .calcoaceticusNCIB82 50的苯酚羟化酶基因具有高度的同源性 ,其核苷酸序列的同源性为 84% ,推导的氨基酸序列的同源性为 98%。对苯酚和非苯酚降解菌株的PCR扩增结果表明 :所有苯酚降解菌均能扩增出 684bp的特征片段 ,而非苯酚降解菌则无PCR条带。对炼焦废水中的细菌群落进行PCR扩增和生化特性检测表明 :显示 684bp特征片段的菌株均具有苯酚降解特性。上述结果表明 ,利用苯酚羟化酶基因的特异引物可对环境中的苯酚降解菌株进行准确快速的PCR检测。     

Effects of Soil pH on the Biodegradation of Chlorpyrifos and Isolation of a Chlorpyrifos-Degrading Bacterium

We examined the role of microorganisms in the degradation of the organophosphate insecticide chlorpyrifos in soils from the United Kingdom and Australia. The kinetics of degradation in five United Kingdom soils varying in pH from 4.7 to 8.4 suggested that dissipation of chlorpyrifos was mediated by the cometabolic activities of the soil microorganisms. Repeated application of chlorpyrifos to these soils did not result in the development of a microbial population with an enhanced ability to degrade the pesticide. A robust bacterial population that utilized chlorpyrifos as a source of carbon was detected in an Australian soil. The enhanced ability to degrade chlorpyrifos in the Australian soil was successfully transferred to the five United Kingdom soils. Only soils with a pH of ≥6.7 were able to maintain this degrading ability 90 days after inoculation. Transfer and proliferation of degrading microorganisms from the Australian soil to the United Kingdom soils was monitored by molecular fingerprinting of bacterial 16S rRNA genes by PCR-denaturing gradient gel electrophoresis (DGGE). Two bands were found to be associated with enhanced degradation of chlorpyrifos. Band 1 had sequence similarity to enterics and their relatives, while band 2 had sequence similarity to strains of Pseudomonas. Liquid enrichment culture using the Australian soil as the source of the inoculum led to the isolation of a chlorpyrifos-degrading bacterium. This strain had a 16S rRNA gene with a sequence identical to that of band 1 in the DGGE profile of the Australian soil. DNA probing indicated that genes similar to known organophosphate-degrading (opd) genes were present in the United Kingdom soils. However, no DNA hybridization signal was detected for the Australian soil or the isolated degrader. This indicates that unrelated genes were present in both the Australian soil and the chlorpyrifos-degrading isolate. These results are consistent with our observations that degradation of chlorpyrifos in these systems was unusual, as it was growth linked and involved complete mineralization. As the 16S rRNA gene of the isolate matched a visible DGGE band from the Australian soil, the isolate is likely to be both prominent and involved in the degradation of chlorpyrifos in this soil.