Biodegradation of polystyrene nanoplastics by Achromobacter xylosoxidans M9 offers a mealworm gut-derived solution for plastic pollution.

Nanoplastics pose significant environmental problems due to their high mobility and increased toxicity. These particles can cause infertility and inflammation in aquatic organisms, disrupt microbial signaling and act as pollutants carrier. Despite extensive studies on their harmful impact on living organisms, the microbial degradation of nanoplastics is still under research. This study investigated the degradation of nanoplastics by isolating bacteria from the gut microbiome of Tenebrio molitor larvae fed various plastic diets. Five bacterial strains capable of degrading polystyrene were identified, with Achromobacter xylosoxidans M9 showing significant nanoplastic degradation abilities. Within 6 days, this strain reduced nanoplastic particle size by 92.3%, as confirmed by SEM and TEM analyses, and altered the chemical composition of the nanoplastics, indicating a potential for enhanced bioremediation strategies. The strain also caused a 7% weight loss in polystyrene film over 30 days, demonstrating its efficiency in degrading nanoplastics faster than polystyrene film. These findings might enhance plastic bioremediation strategies.

New report of endophytic bacterium Achromobacter xylosoxidans from root tissue of Musa spp.

BACKGROUND: Cavendish (AAA) banana plant (Musa spp.) worldwide cultivated crop harbors many endophytic bacteria. Endophytic bacteria are those that live inside plant tissues without producing any visible symptoms of infection. RESULTS: Endophytic bacterium (MRH 11), isolated from root tissue of Musa spp.was identified as Achromobacter xylosoxidans (ON955872) which showed positive effects in IAA production, phosphate solubilization, catalase production. A. xylosoxidans also showed in vitro antagonism against Curvularia lunata causing leaf spot disease of Cavendish (AAA) banana (G-9 variety). The GC-MS analysis of culture filtrate of A. xylosoxidans (ON955872) confirmed this finding. GC-MS analysis was carried by using two solvent etheyl acetate and chloroform and it showed several antifungal compounds. The identification of these bioactive secondary metabolites compounds was based on the peak area, retention time, molecular weight, molecular formula and antimicrobial actions. GC-MS analysis result revealed the presence of major components including Cyclododecane, 1-Octanol, Cetene, Diethyl phthalate. In vivo test to banana plants was carried out in separate field as well as in potted conditions. Appearance of leaf spots after foliar spray of spore of pathogen and reduction in leaf spots after application of bacterial suspension was found. CONCLUSION: The present study has highlighted the role of endophytic bacterium as antagonist to the pathogen Curvularia lunata.

Biochar-assisted degradation of oxytetracycline by Achromobacter denitrificans and underlying mechanisms.

Contamination of the environment with large amounts of residual oxytetracycline (OTC) and the corresponding resistance genes poses a potential threat to natural ecosystems and human health. In this study, an effective OTC-degrading strain, identified as Achromobacter denitrificans OTC-F, was isolated from activated sludge. In the degradation experiment, the degradation rates of OTC in the degradation systems with and without biochar addition were 95.01-100% and 73.72-99.66%, respectively. Biochar promotes the biodegradation of OTC, particularly under extreme environmental conditions. Toxicity evaluation experiments showed that biochar reduced biotoxicity and increased the proportion of living cells by 17.36%. Additionally, biochar increased the activity of antioxidant enzymes by 34.1-91.0%. Metabolomic analysis revealed that biochar promoted the secretion of antioxidant substances such as glutathione and tetrahydrofolate, which effectively reduced oxidative stress induced by OTC. This study revealed the mechanism at the molecular level and provided new strategies for the bioremediation of OTC in the environment.

Effects of bioaugmentation by isolated Achromobacter xylosoxidans BP1 on PAHs degradation and microbial community in contaminated soil.

Polycyclic aromatic hydrocarbons (PAHs) are a group of organic pollutants ubiquitous and persistent in soil. In order to provide a viable solution for bioremediation of PAHs-contaminated soil, a strain of Achromobacter xylosoxidans BP1 with superior PAHs degradation ability was isolated from contaminated soil at a coal chemical site in northern China. The degradation of phenanthrene (PHE) and benzo[a]pyrene (BaP) by strain BP1 was investigated in three different liquid phase cultures, and the removal rates of PHE and BaP by strain BP1 were 98.47% and 29.86% after 7 days under the conditions of PHE and BaP as the only carbon source, respectively. In the medium with the coexistence of PHE and BaP, the removal rates of BP1 were 89.44% and 9.42% after 7 days, respectively. Then, strain BP1 was investigated for its feasibility in remediating PAH-contaminated soil. Among the four PAHs-contaminated soils treated differently, the treatment inoculated with BP1 exhibited higher removal rates of PHE and BaP (p < 0.05), especially the CS-BP1 treatment (inoculation of BP1 into unsterilized PAHs-contaminated soil) showed 67.72%, 13.48% removal of PHE and BaP, respectively, over 49 days of incubation. Bioaugmentation also significantly increased the activity of dehydrogenase and catalase in the soil (p＜0.05). Furthermore, the effect of bioaugmentation on the removal of PAHs was investigated by measuring the activity of dehydrogenase (DH) and catalase (CAT) during incubation. Among them, the DH and CAT activities of CS-BP1 and SCS-BP1 (inoculation of BP1 into sterilized PAHs-contaminated soil) treatments inoculated with strain BP1 were significantly higher than those of treatments without BP1 addition during incubation (p < 0.01). The structure of the microbial community varied among treatments, but the Proteobacteria phylum showed the highest relative abundance in all treatments of the bioremediation process, and most of the bacteria with higher relative abundance at the genus level also belonged to the Proteobacteria phylum. Prediction of microbial functions in soil by FAPROTAX analysis showed that bioaugmentation enhanced microbial functions associated with the degradation of PAHs. These results demonstrate the effectiveness of Achromobacter xylosoxidans BP1 as a PAH-contaminated soil degrader for the risk control of PAHs contamination.

Insights into the Binding Interaction of Catechol 1,2-Dioxygenase with Catechol in Achromobacter xylosoxidans DN002.

Microbial remediation has become one of the promising ways to eliminate polycyclic aromatic hydrocarbons (PAHs) pollution due to its efficient enzyme metabolism system. Catechol 1,2-dioxygenase (C12O) is a crucial rate-limiting enzyme in the degradation pathway of PAHs in Achromobacter xylosoxidans DN002 that opens the benzene ring through the ortho-cleavage pathway. However, little attention has been given to explore the interaction mechanism of relevant enzyme-substrate. This study aims to investigate the binding interaction between C12O of strain DN002 and catechol by means of a molecular biological approach combined with homology modeling, molecular docking, and multiple spectroscopies. The removal rate of catechol in the mutant strain of cat A deletion was only 12.03%, compared to the wild-type strain (54.21%). A Ramachandran plot of active site regions of the primary amino acid sequences in the native enzyme showed that 93.5% sequences were in the most favored regions on account of the results of homology modeling, while an additional 6.2% amino acid sequences were found in conditionally allowed regions, and 0.4% in generously allowed regions. The binding pocket of C12O with catechol was analyzed to obtain that the catalytic trimeric group of Tyr164-His224-His226 was proven to be great vital for the ring-opening reaction of catechol by molecular docking. In the native enzyme, binding complexes were spontaneously formed by hydrophobic interactions. Binding constants and thermodynamic potentials from fluorescence spectra indicated that catechol effectively quenched the intrinsic fluorescence of C12O in the C12O/catechol complex via conventional static and dynamic quenching mechanisms of C12O. The results of ultraviolet and visible (UV) spectra, synchronous fluorescence, and circular dichroism (CD) spectra revealed conspicuous changes in the local conformation, and site-directed mutagenesis confirmed the role of predicted key residues during catalysis, wherein His226 had a significant effect on catechol utilization by C12O. This is the first report to reveal interactions of C12O with substrate from the molecular docking results, providing the mechanistic understanding of representative dioxygenases involved in aromatic compound degradation, and a solid foundation for further site modifications as well as strategies for the directed evolution of this enzyme.

The In Vitro Replication Cycle of Achromobacter xylosoxidans and Identification of Virulence Genes Associated with Cytotoxicity in Macrophages.

Achromobacter xylosoxidans is an opportunistic pathogen implicated in a wide variety of human infections including the ability to colonize the lungs of cystic fibrosis (CF) patients. The role of A. xylosoxidans in human pathology remains controversial due to the lack of optimized in vitro and in vivo model systems to identify and test bacterial gene products that promote a pathological response. We have previously identified macrophages as a target host cell for A. xylosoxidans-induced cytotoxicity. By optimizing our macrophage infection model, we determined that A. xylosoxidans enters macrophages and can reside within a membrane bound vacuole for extended periods of time. Intracellular replication appears limited with cellular lysis preceding an enhanced, mainly extracellular replication cycle. Using our optimized in vitro model system along with transposon mutagenesis, we identified 163 genes that contribute to macrophage cytotoxicity. From this list, we characterized a giant RTX adhesin encoded downstream of a type one secretion system (T1SS) that mediates bacterial binding and entry into host macrophages, an important first step toward cellular toxicity and inflammation. The RTX adhesin is encoded by other human isolates and is recognized by antibodies present in serum isolated from CF patients colonized by A. xylosoxidans, indicating this virulence factor is produced and deployed in vivo. This study represents the first characterization of A. xylosoxidans replication during infection and identifies a variety of genes that may be linked to virulence and human pathology. IMPORTANCE Patients affected by CF develop chronic bacterial infections characterized by inflammatory exacerbations and tissue damage. Advancements in sequencing technologies have broadened the list of opportunistic pathogens colonizing the CF lung. A. xylosoxidans is increasingly recognized as an opportunistic pathogen in CF, yet our understanding of the bacterium as a contributor to human disease is limited. Genomic studies have identified potential virulence determinants in A. xylosoxidans isolates, but few have been mechanistically studied. Using our optimized in vitro cell model, we identified and characterized a bacterial adhesin that mediates binding and uptake by host macrophages leading to cytotoxicity. A subset of serum samples from CF patients contains antibodies that recognize the RTX adhesion, suggesting, for the first time, that this virulence determinant is produced in vivo. This work furthers our understanding of A. xylosoxidans virulence factors at a mechanistic level.

Biodegradation of thermo-oxidative pretreated low-density polyethylene (LDPE) and polyvinyl chloride (PVC) microplastics by Achromobacter denitrificans Ebl13.

Microplastics pretreatment of prior to biodegradation is an efficient approach for their bioremediation. We isolated Achromobacter denitrificans from compost and used it for biodegradation of thermo-oxidative pretreated polyvinyl chloride (PVC) and low-density polyethylene (LDPE). About 12.3 % and 6.5 % weight loss, and 326.4 and 112.32 mg L-1 extracellular protein were observed in bacterial flasks with PVC and LDPE, respectively. The pH in treated PVC reached to 5.12 and the thermal stability increased by 29 °C. The chemical modification in LDPE was demonstrated through oxidation of antioxidants (Phenol group), formation of new groups (Aldehyde group), and chain fracture in the main backbone by Fourier transform infrared spectroscopy. Formation of peaks at the range of 1700-1850 cm-1 in LDPE attributed to formation of carbonyl groups as the degradation result. Scanning electron microscopy confirmed LDPE and PVC degradation by surface alterations. Consequently, thermo-oxidative pretreatment can be considered as a suitable strategy for improving microplastics biodegradation.

A D-2-hydroxyglutarate biosensor based on specific transcriptional regulator DhdR.

D-2-Hydroxyglutarate (D-2-HG) is a metabolite involved in many physiological metabolic processes. When D-2-HG is aberrantly accumulated due to mutations in isocitrate dehydrogenase or D-2-HG dehydrogenase, it functions in a pro-oncogenic manner and is thus considered a therapeutic target and biomarker in many cancers. In this study, DhdR from Achromobacter denitrificans NBRC 15125 is identified as an allosteric transcriptional factor that negatively regulates D-2-HG dehydrogenase expression and responds to the presence of D-2-HG. Based on the allosteric effect of DhdR, a D-2-HG biosensor is developed by combining DhdR with amplified luminescent proximity homogeneous assay (AlphaScreen) technology. The biosensor is able to detect D-2-HG in serum, urine, and cell culture medium with high specificity and sensitivity. Additionally, this biosensor is used to identify the role of D-2-HG metabolism in lipopolysaccharide biosynthesis of Pseudomonas aeruginosa, demonstrating its broad usages.

Bioaccumulation and detoxification of trivalent arsenic by Achromobacter xylosoxidans BHW-15 and electrochemical detection of its transformation efficiency.

Arsenotrophic bacteria play an essential role in lowering arsenic contamination by converting toxic arsenite [As (III)] to less toxic and less bio-accumulative arsenate [As (V)]. The current study focused on the qualitative and electrocatalytic detection of the arsenite oxidation potential of an arsenite-oxidizing bacteria A. xylosoxidans BHW-15 (retrieved from As-contaminated tube well water), which could significantly contribute to arsenic detoxification, accumulation, and immobilization while also providing a scientific foundation for future electrochemical sensor development. The minimum inhibitory concentration (MIC) value for the bacteria was 15 mM As (III). Scanning Electron Microscopy (SEM) investigation validated its intracellular As uptake capacity and demonstrated a substantial association with the MIC value. During the stationary phase, the strain's As (III) transformation efficiency was 0.0224 mM/h. Molecular analysis by real-time qPCR showed arsenite oxidase (aioA) gene expression increased 1.6-fold in the presence of As (III) compared to the untreated cells. The immobilized whole-cell also showed As (III) conversion up to 18 days. To analyze the electrochemical oxidation in water, we developed a modified GCE/P-Arg/ErGO-AuNPs electrode, which successfully sensed and quantified conversion of As (III) into As (V) by accepting electrons; implying a functional As oxidase enzyme activity in the cells. To the best of our knowledge, this is the first report on the electrochemical observation of the As-transformation mechanism with Achromobacter sp. Furthermore, the current work highlighted that our isolate might be employed as a promising candidate for arsenic bioremediation, and information acquired from this study may be helpful to open a new window for the development of a cost-effective, eco-friendly biosensor for arsenic species detection in the future.

Nitrofurazone Removal from Water Enhanced by Coupling Photocatalysis and Biodegradation.

(1) Background: Environmental contamination with antibiotics is particularly serious because the usual methods used in wastewater treatment plants turn out to be insufficient or ineffective. An interesting idea is to support natural biodegradation processes with physicochemical methods as well as with bioaugmentation with efficient microbial degraders. Hence, the aim of our study is evaluation of the effectiveness of different methods of nitrofurazone (NFZ) degradation: photolysis and photodegradation in the presence of two photocatalysts, the commercial TiO2-P25 and a self-obtained Fe3O4@SiO2/TiO2 magnetic photocatalyst. (2) Methods: The chemical nature of the photocatalysis products was investigated using a spectrometric method, and then, they were subjected to biodegradation using the strain Achromobacter xylosoxidans NFZ2. Additionally, the effects of the photodegradation products on bacterial cell surface properties and membranes were studied. (3) Results: Photocatalysis with TiO2-P25 allowed reduction of NFZ by over 90%, demonstrating that this method is twice as effective as photolysis alone. Moreover, the bacterial strain used proved to be effective in the removal of NFZ, as well as its intermediates. (4) Conclusions: The results indicated that photocatalysis alone or coupled with biodegradation with the strain A. xylosoxidans NFZ2 leads to efficient degradation and almost complete mineralization of NFZ.

Inflammation in children with cystic fibrosis: contribution of bacterial production of long-chain fatty acids.

BACKGROUND: Cystic fibrosis (CF) affects >70,000 people worldwide, yet the microbiologic trigger for pulmonary exacerbations (PExs) remains unknown. The objective of this study was to identify changes in bacterial metabolic pathways associated with clinical status. METHODS: Respiratory samples were collected at hospital admission for PEx, end of intravenous (IV) antibiotic treatment, and follow-up from 27 hospitalized children with CF. Bacterial DNA was extracted and shotgun DNA sequencing was performed. MetaPhlAn2 and HUMAnN2 were used to evaluate bacterial taxonomic and pathway relative abundance, while DESeq2 was used to evaluate differential abundance based on clinical status. RESULTS: The mean age of study participants was 10 years; 85% received combination IV antibiotic therapy (beta-lactam plus a second agent). Long-chain fatty acid (LCFA) biosynthesis pathways were upregulated in follow-up samples compared to end of treatment: gondoate (p = 0.012), oleate (p = 0.048), palmitoleate (p = 0.043), and pathways of fatty acid elongation (p = 0.012). Achromobacter xylosoxidans and Escherichia sp. were also more prevalent in follow-up compared to PEx (p < 0.001). CONCLUSIONS: LCFAs may be associated with persistent infection of opportunistic pathogens. Future studies should more closely investigate the role of LCFA production by lung bacteria in the transition from baseline wellness to PEx in persons with CF. IMPACT: Increased levels of LCFAs are found after IV antibiotic treatment in persons with CF. LCFAs have previously been associated with increased lung inflammation in asthma. This is the first report of LCFAs in the airway of persons with CF. This research provides support that bacterial production of LCFAs may be a contributor to inflammation in persons with CF. Future studies should evaluate LCFAs as predictors of future PExs.

Combing multiple site-directed mutagenesis of penicillin G acylase from Achromobacter xylosoxidans PX02 with improved catalytic properties for cefamandole synthesis.

Penicillin G acylase (PGA) was an important biocatalyst for enzymatic production of second-generation cephalosporin. PGA from Achromobacter xylosoxidans PX02 (AxPGA) showed relatively lower identity to EcPGA (54.9% in α subunit and 51.7% in ß subunit), which could synthesize cefamandole in the kinetically controlled N-acylation (kcNa). Semi-rational design of AxPGA and "small and smart" mutant libraries were developed with minimal screening to improve cefamandole production. A triple mutant αR141A/αF142I/ßF24G by combining the mutational sites (ßF24, αR141, and αF142) from different subunits of AxPGA showed better performance in cefamandole production, with 4.2-fold of improvement in the (kcat/Km)AD value for activated acyl donor (R)-Methyl mandelate. Meanwhile, the (kcat/Km)Ps value for cefamandole by mutant αR141A/αF142I/ßF24G was sharply dropped by 25.5 times, indicating its highly synthetic activity and extremely low hydrolysis of cefamandole. Strikingly, the triple mutant αR141A/αF142I/ßF24G could form cefamandole with a yield of 85% at an economical substrate ratio (acyl donor/nucleophile) of 1.3:1 (82% at 1.1:1), which advanced the greener and more sustainable process of cefamandole production than the wild type. Furtherly, the improved synthetic ability and lower hydrolysis of cefamandole by mutant were rationalized using molecular docking.

Anaerobic biodegradation of phenanthrene by a newly isolated nitrate-dependent Achromobacter denitrificans strain PheN1 and exploration of the biotransformation processes by metabolite and genome analyses.

Polycyclic aromatic hydrocarbons (PAHs) are widespread and harmful contaminants and are more persistent under anaerobic conditions. The bioremediation of PAHs in anaerobic zones has been enhanced by treating the contamination with nitrate, which is thermodynamically favourable, cost-effective, and highly soluble. However, anaerobic PAHs biotransformation processes that employ nitrate as an electron acceptor have not been fully explored. In this study, we investigated the anaerobic biotransformation of PAHs by strain PheN1, a newly isolated phenanthrene-degrading denitrifier, using phenanthrene as a model compound. PheN1 is phylogenetically closely related to Achromobacter denitrificans and reduces nitrate to nitrite (not N2 ) during the anaerobic phenanthrene degradation process. Phenanthrene biotransformation processes were detected using gas chromatography-mass spectrometry and were further examined by reverse transcription-quantitative PCR and genome analyses. Carboxylation and methylation were both found to be the initial steps in the phenanthrene degradation process. Downstream biotransformation processed benzene compounds and cyclohexane derivatives. This study describes the isolation of an anaerobic phenanthrene-degrading bacterium along with the pure-culture evidence of phenanthrene biotransformation processes with nitrate as an electron acceptor. The findings in this study can improve our understanding of anaerobic PAHs biodegradation processes and guide PAHs bioremediation by adding nitrate to anaerobic environments.

Improved grain yield and lowered arsenic accumulation in rice plants by inoculation with arsenite-oxidizing Achromobacter xylosoxidans GD03.

Arsenite is the predominant arsenic species in flooded paddy soil, and arsenite bioaccumulation in rice grains has been identified as a major problem in many Asian countries. Lowering arsenite level in rice plants and grain via accelerating arsenite oxidation is a potential strategy to help populations, who depended on rice consumption, to reduce the internal exposure level of arsenic. We herein isolated a strain, Achromobacter xylosoxidans GD03, with the high arsenite-oxidizing ability and plant growth-promoting traits. We observed that arsenite exposure could promote A. xylosoxidans GD03 to excrete indole-3-acetic acid and thus promoted rice growth. The pot culture experiments of Indica rice cultivar Guang You Ming 118 (GYM118) demonstrated that A. xylosoxidans GD03 inoculation of paddy soil (4.5-180 × 108 CFU GD03/kg soil) significantly accelerated arsenite oxidation in flooded soil. The daily arsenic oxidation rate with GD03 inoculation was 1.5-3.3 times as that without strain GD03 inoculation within the whole growth period of Indica GYM118 in the presence of the native microflora. It thus led to a 34-69%, 43-74%, 24-76% and 35-57% decrease in arsenite concentration of the stems, leaves, bran and grain of Indica GYM118 respectively and a 59-96% increase in rice grain yield. The paddy soil inoculated with 40.0 mL/kg of A. xylosoxidans GD03 resulted in a lowest As(III) concentrations in all rice organs of Indica GYM118, which equivalent to only 24-50% of the As(III) concentrations in the group without GD03 inoculation. The results highlight that a highly arsenite-oxidizing bacterium could accelerate arsenite oxidation of paddy soil when facing competition with the native microflora, thus decrease arsenic toxicity and bioavailable soil arsenic.

Complete genome sequence of the aromatic-hydrocarbon-degrading bacterium Achromobacter xylosoxidans DN002.

Achromobacter xylosoxidans DN002 is capable of utilizing numerous aromatic hydrocarbons as sole carbon and energy resource. In this study, the whole genome of strain DN002 was sequenced and analyzed, which consisted of one circular chromosome of 5,943,204 bp and a 278,917 bp plasmid with an average GC content of 65.46 mol%, 5694 protein-coding genes, 13 rRNA genes and 57 tRNA genes. Analysis of cluster of orthologous group (COG) demonstrated that strain DN002 had remarkable gene abundance foramino acid transport and metabolism, transcription, inorganic ion transport and metabolism, energy production and conversion, and carbohydrate transport and metabolism. Genes related to biodegradation of aromatic hydrocarbons, chemotaxis and flagella were identified from the genome, which will advance our fundamental understanding the molecular mechanism for degradation and metabolizing of aromatic hydrocarbons.

Quorum sensing mediated response of Achromobacter denitrificans SP1 towards prodigiosin production under phthalate stress.

Quorum sensing is a density-dependent chemical process between bacteria, which may be intergenus or intragenus. N-acyl homoserine lactones (HSLs) are a type of small signaling molecules associated with Gram-negative bacteria for monitoring their own population density. The present study unveils the mechanism of HSLs in Achromobacter denitrificans SP1 while transforming di(2-ethylhexyl) phthalate (DEHP) into prodigiosin in a simple basal salt medium. The primary detection of HSLs was done by the colorimetric method. Fourier-transform infrared spectroscopy and liquid chromatography-mass spectrometry-quadrupole time-of-flight confirmed and identified the HSLs. The maximum production of HSLs was observed between 24 and 72 h of incubation, which is noted to be a peak time of DEHP degradation. A total of 57.2% of DEHP was degraded within 30 h and complete degradation was observed within 72 h of incubation. Regulation in the synthesis of various acyl-HSL molecules, viz. 3OC6-HSL in the initial stage of DEHP stress, 3OC8-HSL, and C10-HSL during the time of degradation and 3OC12-HSL on completion of degradation was noticed. The role of HSLs on the production of prodigiosin was confirmed using vanillin as an HSL inhibitor. Through the selective activation of HSL molecules, A. denitrificans SP1 sustain the changing stressful conditions. Supplementation of acyl-HSL signal molecules may boost up the efficacy of A. denitrificans SP1 in both DEHP degradation and prodigiosin production which offers great potential towards the management of DEHP containing plastic wastes.

Heterologous expression and functional study of nitric oxide reductase catalytic reduction peptide from Achromobacter denitrificans strain TB.

Biological denitrification is a promising and green technology for air pollution control. To investigate the nitric oxide reductase (NOR) that dominates NO reduction efficiency in biological purification, the heterologous prokaryotic expression system of the norB gene, which encodes the core peptide of the catalytic reduction structure in the NOR from Achromobacter denitrificans strain TB, was constructed in Escherichia coli BL21 (DE3). Results showed that the 1218 bp-long norB gene was expressed at the highest level under 1.0 mM IPTG for 5 h at 30 °C, and the relative expression abundance of norB in recombinant E. coli was increased by 16.6 times compared with that of the wild-type TB. However, the NO reduction efficiency and NOR activity of strain TB was 2.7 and 1.83 times higher than those of recombinant E. coli, respectively. On the basis of genomic reassembly and protein structure modeling, the core peptide of the NOR catalytic reduction structure from Achromobacter sp. TB can independently exert NO reduction. The low NO degradation efficiency of recombinant E. coli may be due to the lack of a NorC-like structure that increases the enzyme activity of the NorB protein. The results of this study can be used as basis for further research on the structure and function of NOR.

Response surface optimization of prodigiosin production by phthalate degrading Achromobacter denitrificans SP1 and exploring its antibacterial activity.

The role of various parameters like temperature, pH, blood bag concentration, agitation and incubation that influence the production of prodigiosin by Achromobacter denitrificans SP1 was determined. The Plackett-Burman and Box-Behnken experimental designs were employed to statistically optimize and find out the best combinational effect of parameters for the better yield of prodigiosin using blood bag as sole carbon and energy source for the growth of A. denitrificans SP1. The maximum (1.314 mg/ml) prodigiosin production was attained at a temperature of 24 °C, pH (8.8), and blood bag (1 g) as optimum; while the predicted value was 1.319 mg/ml with a correlation coefficient of 0.987; which signifies the fitness of the model. Antimicrobial activity of the prodigiosin was also evaluated and found to be an effective agent against bacterial pathogens including Staphylococcus aureus and Proteus mirabilis. Utilization of the plasticizer di (2-ethylhexyl)phthalate (DEHP) in blood bag and the production of antibacterial prodigiosin makes A. denitrificans SP1, an effective competitor toward the pathogenic bacterial disinfection and wastewater treatment processes.

Bioaugmentation of membrane bioreactor with Achromobacter denitrificans strain PR1 for enhanced sulfamethoxazole removal in wastewater.

Achromobacter denitrificans strain PR1, previously found to harbour specific degradation pathways with high sulfamethoxazole (SMX) degradation rates, was bioaugmented into laboratory-scale membrane bioreactors (MBRs) operated under aerobic conditions to treat SMX-containing real domestic wastewater. Different hydraulic retention times (HRTs), which is related to reaction time and loading rates, were considered and found to affect the SMX removal efficiency. The availability of primary substrates was important in both bioaugmented and non-bioaugmented activated sludge (AS) for cometabolism of SMX. High HRT (24 h) resulted in low food to microorganism ratio (F/M) and low SMX removal, due to substrate limitation. Decrease in HRT from 24 h to 12 h, 6 h and finally 4 h led to gradual increases in primary substrates availability, e.g. organic compounds and ammonia, resulted in increased SMX removal efficiency and degradation rate, and is more favorable for high-rate wastewater treatment processes. After inoculation into the MBRs, the bioaugmentation strain was sustained in the reactor for a maximum of 31 days even though a significant decrease in abundance was observed. The bioaugmented MBRs showed enhanced SMX removal, especially under SMX shock loads compared to the control MBRs. The results of this study indicate that re-inoculation is required regularly after a period of time to maintain the removal efficiency of the target compound.

Biodegradation of sulfamethoxazole by a bacterial consortium of Achromobacter denitrificans PR1 and Leucobacter sp. GP.

In the last decade, biological degradation and mineralization of antibiotics have been increasingly reported feats of environmental bacteria. The most extensively described example is that of sulfonamides that can be degraded by several members of Actinobacteria and Proteobacteria. Previously, we reported sulfamethoxazole (SMX) degradation and partial mineralization by Achromobacter denitrificans strain PR1, isolated from activated sludge. However, further studies revealed an apparent instability of this metabolic trait in this strain. Here, we investigated this instability and describe the finding of a low-abundance and slow-growing actinobacterium, thriving only in co-culture with strain PR1. This organism, named GP, shared highest 16S rRNA gene sequence similarity (94.6-96.9%) with the type strains of validly described species of the genus Leucobacter. This microbial consortium was found to harbor a homolog to the sulfonamide monooxygenase gene (sadA) also found in other sulfonamide-degrading bacteria. This gene is overexpressed in the presence of the antibiotic, and evidence suggests that it codes for a group D flavin monooxygenase responsible for the ipso-hydroxylation of SMX. Additional side reactions were also detected comprising an NIH shift and a Baeyer-Villiger rearrangement, which indicate an inefficient biological transformation of these antibiotics in the environment. This work contributes to further our knowledge in the degradation of this ubiquitous micropollutant by environmental bacteria.