Horizontal transfer of catabolic plasmids in the process of naphthalene biodegradation in model soil systems

The process of naphthalene degradation by indigenous, introduced, and transconjugant strains was studied in laboratory soil microcosms. Conjugation transfer of catabolic plasmids was demonstrated in naphthalene-contaminated soil. Both indigenous microorganisms and an introduced laboratory strain BS394 (pNF142::TnMod-OTc) served as donors of these plasmids. The indigenous bacterial degraders of naphthalene isolated from soil were identified as Pseudomonas putida and Pseudomonas fluorescens. The frequency of plasmid transfer in soil was 10^?5–10^?4 per donor cell. The activity of the key enzymes of naphthalene biodegradation in indigenous and transconjugant strains was studied. Transconjugant strains harboring indigenous catabolic plasmids possessed high salicylate hydroxylase and low catechol-2,3-dioxygenase activities, in contrast to indigenous degraders, which had a high level of catechol-2,3-dioxygenase activity and a low level of salicylate hydroxylase. Naphthalene degradation in batch culture in liquid mineral medium was shown to accelerate due to cooperation of the indigenous naphthalene degrader P. fluorescens AP1 and the transconjugant strain P. putida KT2442 harboring the indigenous catabolic plasmid pAP35. The role of conjugative transfer of naphthalene biodegradation plasmids in acceleration of naphthalene degradation was demonstrated in laboratory soil microcosms.     

Horizontal transfer of catabolic plasmids in the process of naphthalene biodegradation in model soil systems

The process of naphthalene degradation by indigenous, introduced, and transconjugant strains was studied in laboratory soil microcosms. Conjugation transfer of catabolic plasmids was demonstrated in naphthalene-contaminated soil. Both indigenous microorganisms and an introduced laboratory strain BS394 (pNF142::TnMod-OTc) served as donors of these plasmids. The indigenous bacterial degraders of naphthalene isolated from soil were identified as Pseudomonas putida and Pseudomonas fluorescens. The frequency of plasmid transfer in soil was 10(-5)-10(-4) per donor cell. The activity of the key enzymes of naphthalene biodegradation in indigenous and transconjugant strains was studied. Transconjugant strains harboring indigenous catabolic plasmids possessed high salicylate hydroxylase and low catechol-2,3-dioxygenase activities, in contrast to indigenous degraders, which had a high level of catechol-2,3-dioxygenase activity and a low level of salicylate hydroxylase. Naphthalene degradation in batch culture in liquid mineral medium was shown to accelerate due to cooperation of the indigenous naphthalene degrader P. fluorescens AP1 and the transconjugant strain P. putida KT2442 harboring the indigenous catabolic plasmid pAP35. The role of conjugative transfer of naphthalene biodegradation plasmids in acceleration of naphthalene degradation was demonstrated in laboratory soil microcosms.     

Horizontal transfer of catabolic plasmids and naphthalene biodegradation in open soil

The horizontal transfer of naphthalene biodegradation plasmids and the parallel process of its microbial degradation were studied for the first time. The tagged naphthalene-degrading strains bearing labeled biodegradation plasmids were used for the monitoring of horizontal plasmid transfer in open soil. The population kinetics of microorganisms, the survival rate and competitiveness of introduced strains, and the transfer of biodegradation plasmids to indigenous strains were investigated. The transfer of the labeled plasmid pNF142::TnMod-OTc to the introduced plasmid-free recipient P. putida KT2442 and to indigenous soil microorganisms of the genus Pseudomonas was shown both under selection pressure (in the presence of naphthalene) and in its absence. The 16S rRNA gene sequencing showed that the soil strains that had acquired plasmids were close to the species P. lini, P. frederiksbergensis, P. jessenii, P. graminis, P. putida, and P. alcaligenes. Thus, direct evidence of dissemination of the naphthalene biodegradation plasmids in microbial populations in open soil under selective and nonselective conditions has been obtained.     

Bioremediation of soil polluted with fuels by sequential multiple injection of native microorganisms: Field-scale processes in Poland

Research was conducted to estimate impact of the multiple bioaugmentation on the treatment of soil contaminated by fuels - diesel oil and aircraft fuel. The bacteria used to inoculate the remediation plots were isolated from the polluted soil and proliferated in field conditions. The amount of biomass applied to the polluted soil was set to ensure the total number of bacteria in soil 10⁷-10⁸ cfu/g d.w. The multiple inoculation of soil with indigenous bacteria active in diesel oil and engine oil (plot A) degradation increased bioremediation effectiveness by 50% in comparison to the non-inoculated control soil and by 30% in comparison to the soil that was inoculated only once. The multiple inoculation of soil with indigenous microorganisms was then applied in bioremediation of the soil polluted with double high concentration of diesel oil (soil B) and in bioremediation of the soil polluted with aircraft fuel (soil C). The process efficiency was 80% and 98% removal of TPH for soil B and C, respectively.     

Crude Oil Treatment Leads to Shift of Bacterial Communities in Soils from the Deep Active Layer and Upper Permafrost along the China-Russia Crude Oil Pipeline Route

The buried China-Russia Crude Oil Pipeline (CRCOP) across the permafrost-associated cold ecosystem in northeastern China carries a risk of contamination to the deep active layers and upper permafrost in case of accidental rupture of the embedded pipeline or migration of oil spills. As many soil microbes are capable of degrading petroleum, knowledge about the intrinsic degraders and the microbial dynamics in the deep subsurface could extend our understanding of the application of in-situ bioremediation. In this study, an experiment was conducted to investigate the bacterial communities in response to simulated contamination to deep soil samples by using 454 pyrosequencing amplicons. The result showed that bacterial diversity was reduced after 8-weeks contamination. A shift in bacterial community composition was apparent in crude oil-amended soils with Proteobacteria (esp. α-subdivision) being the dominant phylum, together with Actinobacteria and Firmicutes. The contamination led to enrichment of indigenous bacterial taxa like Novosphingobium, Sphingobium, Caulobacter, Phenylobacterium, Alicylobacillus and Arthrobacter, which are generally capable of degrading polycyclic aromatic hydrocarbons (PAHs). The community shift highlighted the resilience of PAH degraders and their potential for in-situ degradation of crude oil under favorable conditions in the deep soils.     

The Potential of Bacterial Isolates for Emulsification with a Range of Hydrocarbons

A study was undertaken to investigate the distribution of biosurfactant producing and crude oil degrading bacteria in the oil contaminated environment. This research revealed that hydrocarbon contaminated sites are the potent sources for oil degraders. Among 32 oil degrading bacteria isolated from ten different oil contaminated sites of gasoline and diesel fuel stations, 80% exhibited biosurfactant production. The quantity and emulsification activity of the biosurfactants varied. Pseudomonas sp. DS10‐129 produced a maximum of 7.5 ± 0.4 g/l of biosurfactant with a corresponding reduction in surface tension from 68 mN/m to 29.4 ± 0.7 mN/m at 84 h incubation. The isolates Micrococcus sp. GS2‐22, Bacillus sp. DS6‐86, Corynebacterium sp. GS5‐66, Flavobacterium sp. DS5‐73, Pseudomonas sp. DS10‐129, Pseudomonas sp. DS9‐119 and Acinetobacter sp. DS5‐74 emulsified xylene, benzene, n‐hexane, Bombay High crude oil, kerosene, gasoline, diesel fuel and olive oil. The first five of the above isolates had the highest emulsification activity and crude oil degradation ability and were selected for the preparation of a mixed bacterial consortium, which was also an efficient biosurfactant producing oil emulsifying and degrading culture. During this study, biosurfactant production and emulsification activity were detected in Moraxella sp., Flavobacterium sp. and in a mixed bacterial consortium, which have not been reported before.     

Microaerophilic alkane degradation in <Emphasis Type="Italic">Pseudomonas extremaustralis</Emphasis>: a transcriptomic and physiological approach

Diesel fuel is one of the most important sources of hydrocarbon contamination worldwide. Its composition consists of a complex mixture of n-alkanes, branched alkanes and aromatic compounds. Hydrocarbon degradation in Pseudomonas species has been mostly studied under aerobic conditions; however, a dynamic spectrum of oxygen availability can be found in the environment. Pseudomonas extremaustralis, an Antarctic bacterium isolated from a pristine environment, is able to degrade diesel fuel and presents a wide microaerophilic metabolism. In this work RNA-deep sequence experiments were analyzed comparing the expression profile in aerobic and microaerophilic cultures. Interestingly, genes involved in alkane degradation, including alkB, were over-expressed in micro-aerobiosis in absence of hydrocarbon compounds. In minimal media supplemented with diesel fuel, n-alkanes degradation (C13–C19) after 7 days was observed under low oxygen conditions but not in aerobiosis. In-silico analysis of the alkB promoter zone showed a putative binding sequence for the anaerobic global regulator, Anr. Our results indicate that some diesel fuel components can be utilized as sole carbon source under microaerophilic conditions for cell maintenance or slow growth in a Pseudomonas species and this metabolism could represent an adaptive advantage in polluted environments.     

Strong Impact on the Polycyclic Aromatic Hydrocarbon (PAH)-Degrading Community of a PAH-Polluted Soil but Marginal Effect on PAH Degradation when Priming with Bioremediated Soil Dominated by Mycobacteria

Bioaugmentation of soil polluted with polycyclic aromatic hydrocarbons (PAHs) is often disappointing because of the low survival rate and low activity of the introduced degrader bacteria. We therefore investigated the possibility of priming PAH degradation in soil by adding 2% of bioremediated soil with a high capacity for PAH degradation. The culturable PAH-degrading community of the bioremediated primer soil was dominated by Mycobacterium spp. A microcosm containing pristine soil artificially polluted with PAHs and primed with bioremediated soil showed a fast, 100- to 1,000-fold increase in numbers of culturable phenanthrene-, pyrene-, and fluoranthene degraders and a 160-fold increase in copy numbers of the mycobacterial PAH dioxygenase gene pdo1. A nonpolluted microcosm primed with bioremediated soil showed a high rate of survival of the introduced degrader community during the 112 days of incubation. A nonprimed control microcosm containing pristine soil artificially polluted with PAHs showed only small increases in the numbers of culturable PAH degraders and no pdo1 genes. Initial PAH degradation rates were highest in the primed microcosm, but later, the degradation rates were comparable in primed and nonprimed soil. Thus, the proliferation and persistence of the introduced, soil-adapted degraders had only a marginal effect on PAH degradation. Given the small effect of priming with bioremediated soil and the likely presence of PAH degraders in almost all PAH-contaminated soils, it seems questionable to prime PAH-contaminated soil with bioremediated soil as a means of large-scale soil bioremediation.     

Succession of Indigenous Pseudomonas spp. and Actinomycetes on Barley Roots Affected by the Antagonistic Strain Pseudomonas fluorescens DR54 and the Fungicide Imazalil

In recent years, the interest in the use of bacteria for biological control of plant-pathogenic fungi has increased. We studied the possible side effects of coating barley seeds with the antagonistic strain Pseudomonas fluorescens DR54 or a commercial fungicide, imazalil. This was done by monitoring the number of indigenous Pseudomonas organisms and actinomycetes on barley roots during growth in soil, harvest after 50 days, and subsequent decomposition. Bacteria were enumerated by traditional plate spreading on Gould's S1 agar (Pseudomonas) and as filamentous colonies on Winogradsky agar (actinomycetes) and by two quantitative competitive PCR assays. For this we developed an assay targeting Streptomyces and closely related genera. DR54 constituted more than 75% of the Pseudomonas population at the root base during the first 21 days but decreased to less than 10% at day 50. DR54 was not successful in colonizing root tips. Initially, DR54 affected the number of indigenous Pseudomonas organisms negatively, whereas imazalil affected Pseudomonas numbers positively, but the effects were transient. Although plate counts were considerably lower than the number of DNA copies, the two methods correlated well for Pseudomonas during plant growth, but after plant harvest Pseudomonas-specific DNA copy numbers decreased while plate counts were in the same magnitude as before. Hence, Pseudomonas was 10-fold more culturable in a decomposition environment than in the rhizosphere. The abundance of actinomycetes was unaffected by DR54 or imazalil amendments, and CFU and quantitative PCR results correlated throughout the experiment. The abundance of actinomycetes increased gradually, mostly in numbers of DNA copies, confirming their role in colonizing old roots.     

Plasmid Transfer from Pseudomonas putida to the Indigenous Bacteria on Alfalfa Sprouts: Characterization, Direct Quantification, and In Situ Location of Transconjugant Cells

The transfer of the plasmids pJKJ5 and TOL (pWWO) from Pseudomonas putida to the indigenous bacterial community on alfalfa sprouts was studied. Tagging with fluorescent protein markers allowed direct quantification of the introduced donor bacteria and of indigenous bacteria that had received the plasmids. The sprouts were observed for 9 days; during this time alfalfa seeds, inoculated with donor bacteria, developed to edible and subsequently decaying sprouts. The first transconjugants were detected on day 6 after donor inoculation and occurred at frequencies of 3.4 × 10⁻⁴ and 2.0 × 10⁻⁶ transconjugant cells per donor cell for pKJK5::gfp and TOL::gfp, respectively. Confocal laser scanning microscopy revealed that the sprouts were heavily colonized with donors and that most transconjugants were located around the hypocotyl and root areas. Randomly selected members of the indigenous bacterial community from both inoculated and uninoculated sprouts, as well as a representative part of the community that had received the plasmids, were characterized by polymorphisms of PCR-amplified ribosomal DNA (rDNA) spacer regions between the 16S and 23S genes, followed by partial 16S rDNA sequencing. This showed that the initially dominating genera Erwinia and Paenibacillus were gradually replaced by Pseudomonas on the fully developed sprouts. Transconjugants carrying either of the investigated plasmids mainly belonged to the genera Pseudomonas and Erwinia. The numbers of transconjugant cells did not reach detectable levels until 6 days after the onset of germination, at which point these species constituted the majority of the indigenous bacteria. In conclusion, the alfalfa sprouts provided an environment that allowed noteworthy frequencies of plasmid transfer from P. putida in the absence of selective pressure that could favor the presence of the investigated plasmids.     

Quantitative Selective PCR of 16S Ribosomal DNA Correlates Well with Selective Agar Plating in Describing Population Dynamics of Indigenous Pseudomonas spp. in Soil Hot Spots

We used a quantitative PCR method targeting 16S ribosomal DNA using competitive PCR for specific detection of indigenous Pseudomonas DNA in soil hot spots. The amount of Pseudomonas DNA corresponded to the number of culturable Pseudomonas bacteria on Gould’s S1 agar. This represents the first use of PCR for quantification of indigenous bacteria in more than one sample of soil.     

Polychlorinated Biphenyl (PCB)-Degrading Bacteria Associated with Trees in a PCB-Contaminated Site

The abundance, identities, and degradation abilities of indigenous polychlorinated biphenyl (PCB)-degrading bacteria associated with five species of mature trees growing naturally in a contaminated site were investigated to identify plants that enhance the microbial PCB degradation potential in soil. Culturable PCB degraders were associated with every plant species examined in both the rhizosphere and root zone, which was defined as the bulk soil in which the plant was rooted. Significantly higher numbers of PCB degraders (2.7- to 56.7-fold-higher means) were detected in the root zones of Austrian pine (Pinus nigra) and goat willow (Salix caprea) than in the root zones of other plants or non-root-containing soil in certain seasons and at certain soil depths. The majority of culturable PCB degraders throughout the site and the majority of culturable PCB degraders associated with plants were identified as members of the genus Rhodococcus by 16S rRNA gene sequence analysis. Other taxa of PCB-degrading bacteria included members of the genera Luteibacter and Williamsia, which have not previously been shown to include PCB degraders. PCB degradation assays revealed that some isolates from the site have broad congener specificities; these isolates included one Rhodococcus strain that exhibited degradation abilities similar to those of Burkholderia xenovorans LB400. Isolates with broad congener specificity were widespread at the site, including in the biostimulated root zone of willow. The apparent association of certain plant species with increased abundance of indigenous PCB degraders, including organisms with outstanding degradation abilities, throughout the root zone supports the notion that biostimulation through rhizoremediation is a promising strategy for enhancing PCB degradation in situ.     

Unveiling metabolic characteristics of an uncultured Gammaproteobacterium responsible for in situ PAH biodegradation in petroleum polluted soil

Exploring the metabolic characteristics of indigenous PAH degraders is critical to understanding the PAH bioremediation mechanism in the natural environment. While stable-isotopic probing (SIP) is a viable method to identify functional microorganisms in complex environments, the metabolic characteristics of uncultured degraders are still elusive. Here, we investigated the naphthalene (NAP) biodegradation of petroleum polluted soils by combining SIP, amplicon sequencing and metagenome binning. Based on the SIP and amplicon sequencing results, an uncultured Gammaproteobacterium sp. was identified as the key NAP degrader. Additionally, the assembled genome of this uncultured degrader was successfully obtained from the ¹³C-DNA metagenomes by matching its 16S rRNA gene with the SIP identified OTU sequence. Meanwhile, a number of NAP degrading genes encoding naphthalene/PAH dioxygenases were identified in this genome, further confirming the direct involvement of this indigenous degrader in the NAP degradation. The degrader contained genes related to the metabolisms of several carbon sources, energy substances and vitamins, illuminating potential reasons for why microorganisms cannot be cultivated and finally realize their cultivation. Our findings provide novel information on the mechanisms of in situ PAH biodegradation and add to our current knowledge on the cultivation of non-culturable microorganisms by combining both SIP and metagenome binning.     

Genome Sequence of Pseudomonas aeruginosa DQ8, an Efficient Degrader of n-Alkanes and Polycyclic Aromatic Hydrocarbons

Pseudomonas aeruginosa DQ8, which was isolated from the crude oil polluted soil in the Daqing oilfield of China, can efficiently degrade diesel, crude oil, n-alkanes, and polycyclic aromatic hydrocarbons (PAHs). Here, we present a 6.8-Mb assembly of its genome sequence. We have annotated 23 coding sequences (CDSs) responsible for catabolism of n-alkanes and PAHs.     

Phenol and 2-naphthol production by toluene 4-monooxygenases using an aqueous/dioctyl phthalate system

A two-phase system is developed here for converting: (1) benzene to phenol and (2) naphthalene to 2-naphthol, using whole cells expressing wild-type toluene 4-monooxygenase (T4MO) and the alpha subunit variant TmoA I100A from Pseudomonas mendocina KR1. Using the T4MO TmoA I100A variant, the solubility of naphthalene was enhanced and the toxicity of the naphthols was prevented by the use of a water/dioctyl phthalate (80:20, vol%) system which yielded 21-fold more 2-naphthol. More than 99% 2-naphthol was extracted to the dioctyl phthalate phase, dihydroxynaphthalene formation was prevented, 92% 2-naphthol was formed, and 12% naphthalene was converted. Similarly, using 50 vol% dioctyl phthalate, an initial concentration of 3.0 g l⁻¹ (39 mM), and wild-type T4MO, a 51±9% conversion of benzene was obtained and phenol was produced at a purity of 97%. Relative to the one-phase system, there was a 12-fold reduction in the formation of the byproduct catechol.     

Bacterial Communities Predominant in the Degradation of 13C-Labeled Pyrene in Red Soil

The bacterial community composition using a consortium (CON) with or without methyl-β-cyclodextrin (MCD), used to bioremediate the pyrene-contaminated soil was evaluated through stable-isotope probing (SIP). Microcosms were artificially contaminated with ¹³C pyrene at 15 ppm, and bacterial community composition was determined over a 14-day period. After 14 days, only 29.1% of pyrene was degraded in control, while 90.6% of pyrene was degraded by the bacterial community with MCD, showing the best bioremediation rate of all treatments. After 14 days, PAH degraders became dominated by Proteobacteria. Sphingomonas and Pseudomonas were the predominant genera in all treatments, and Pseudomonas was the main degrader from the bacterial community. The bacterial community was little affected by MCD. The results indicated that the combination of the bacterial community with MCD can be used to bioremediate PAHs polluted soil.     

Investigating the Diversity of Pseudomonas spp. in Soil Using Culture Dependent and Independent Techniques

Less than 1 % of bacterial populations present in environmental samples are culturable, meaning that cultivation will lead to an underestimation of total cell counts and total diversity. However, it is less clear whether this is also true for specific well-defined groups of bacteria for which selective culture media is available. In this study, we use culture dependent and independent techniques to describe whether isolation of Pseudomonas spp. on selective nutrient-poor NAA 1:100 agar-medium can reflect the full diversity, found by pyrosequencing, of the total soil Pseudomonas community in an urban waste field trial experiment. Approximately 3,600 bacterial colonies were isolated using nutrient-poor NAA 1:100 medium from soils treated with different fertilizers; (i) high N-level sewage sludge (SA), (ii) high N-level cattle manure (CMA), and (iii) unfertilized control soil (U). Based on Pseudomonas specific quantitative-PCR and Pseudomonas CFU counts, less than 4 % of Pseudomonas spp. were culturable using NAA 1:100 medium. The Pseudomonas selectivity and specificity of the culture medium were evaluated by 454 pyrosequencing of 16S rRNA gene amplicons generated using Bacteria- and Pseudomonas-specific primers. Pyrosequencing results showed that most isolates were Pseudomonas and that the culturable fraction of Pseudomonas spp. reflects most clusters of the total Pseudomonas diversity in soil. This indicates that NAA 1:100 medium is highly selective for Pseudomonas species, and reveals the ability of NAA 1:100 medium to culture mostly the dominant Pseudomonas species in soil.     

Microbial degradation of petroleum hydrocarbons in a polluted tropical stream

Summary Crude oil degradation was observed in water samples from three sites along the course of a polluted stream in Lagos, Nigeria. Consistent increase and decrease in the total viable counts (TVCs) of indigenous organisms occurred in the test and control experiments, respectively. Enrichments of the water samples with crude oil resulted in the isolation of nine bacteria belonging to seven genera. A mixed culture was developed from the assemblage of the nine species. The defined microbial consortium utilized a wide range of pure HCs including cycloalkane and aromatic HCs. Utilization of crude oil and petroleum cuts, i.e., kerosene and diesel resulted in an increase in TVC (till day 10) concomitant with decreases in pH and residual oil concentration. Crude oil, diesel and kerosene were degraded by 88, 85 and 78%, respectively, in 14 days. Substrate uptake studies with axenic cultures showed that growth was not sustainable on either cyclohexane or aromatics while degradation of the petroleum fractions fell below 67% in spite of extended incubation period (20 day). From the GC analysis of recovered oil, while reductions in peaks of n-alkane fractions and in biomarkers namely n-C₁₇/pristane and n-C₁₈/phytane ratios were observed in culture fluids of pure strains, complete removal of all the HC components of kerosene, diesel and crude oil including the isoprenoids was obtained with the consortium within 14 days.     

Predominant culturable crude oil-degrading bacteria in the coast of Kuwait

Total of 272 crude oil-degrading bacteria were isolated from seven locations along the coast of Kuwait. The analysis of the 16S rDNA sequences of isolated bacteria revealed the predominance of six bacterial genera: Pseudomonas, Bacillus, Staphylococcus, Acinetobacter, Kocuria and Micrococcus. Investigation of the factors associated with bacterial predominance revealed that, dominant culturable crude oil-degrading bacteria were better crude oil utilizers than the less frequently occurring isolates. Bacterial predominance was also influenced by the ability of bacteria to adapt to the level of organic content available. Predominant culturable bacteria constituted 89.7–54.2% of the total crude oil-degrading bacterial communities. Using 16S-RFLP analyses to assess the diversity of the dominant crude oil-degrading bacterial genera, four phylotypes of Pseudomonas sp. and seven phylotypes of Bacillus sp. were determined. This suggested high degree of diversity of crude oil-degrading bacterial population at the strain level, but low diversity at the genus level.     

Effects of the inoculant strain Pseudomonas putida KT2442 (pNF142) and of naphthalene contamination on the soil bacterial community

The naphthalene-degrading activity of a Pseudomonas sp. strain isolated from a creosote-contaminated soil was shown to be encoded by the IncP9 plasmid pNF142 by transfer to Pseudomonas putida KT2442. The effects of the inoculant strain KT2442 (pNF142) and of naphthalene contamination on the soil bacterial community were studied in microcosms with the following treatments: (I) soil, (II) soil with naphthalene, (III) soil with naphthalene and inoculated with KT2442 (pNF142). The inoculant became the dominant bacterial population in treatment (III) as evidenced by cultivation and denaturing gradient gel electrophoresis (DGGE) analysis. The bacterial DGGE profiles revealed drastically reduced complexity due to the numerical dominance of the inoculant. However, group-specific fingerprints (beta-proteobacteria, actinobacteria) that excluded KT2442 (pNF142) showed less severe changes in the bacterial community patterns. A major effect of naphthalene on the soil bacterial community was observed in treatment (II) after 21 days. Two dominant bands appeared whose sequences showed the highest similarity to those of Burkholderia sp. RP007 and Nocardia vinaceae based on 16S rRNA gene sequencing. These bands were less intense in treatment (III). The increased abundance of RP007-like populations due to naphthalene contamination was also confirmed by PCR amplification of the phnAc gene. The nahAc and nahH genes were detected in DNA and cDNA only in treatment III. Although the inoculant strain KT2442 (pNF142) showed good survival and expression of genes involved in naphthalene degradation, this study suggests that KT2442 (pNF142) suppressed the enrichment of indigenous naphthalene degraders.