Magnetite-driven Bio-Fenton degradation of chloroacetanilide herbicides by a newly isolated hydrogen peroxide producing bacterium Desemzia sp. strain C1.

The efficiency of the Fenton reaction is markedly contingent upon the operational pH related to iron solubility. Therefore, a heterogeneous Fenton reaction has been developed to function at neutral pH. In the present study, the Bio-Fenton reaction was carried out using magnetite (Fe(II)Fe(III)2O4) and H2O2 generated by a newly isolated H2O2-producing bacterium, Desemzia sp. strain C1 at pH 6.8 to degrade chloroacetanilide herbicides. The optimal conditions for an efficient Bio-Fenton reaction were 10 mM of lactate, 0.5% (w/v) of magnetite, and resting-cells (O.D.600 = 1) of strain C1. During the Bio-Fenton reaction, 1.8-2.0 mM of H2O2 was generated by strain C1 and promptly consumed by the Fenton reaction with magnetite, maintaining stable pH conditions. Approximately, 40-50% of the herbicides underwent oxidation through non-specific reactions of •OH, leading to dealkylation, dechlorination, and hydroxylation via hydrogen atom abstraction. These findings will contribute to advancing the Bio-Fenton system for non-specific oxidative degradation of diverse organic pollutants under in-situ environmental conditions with bacteria producing high amount of H2O2 and magnetite under a neutral pH condition.

Fungal biodegradation of poly(butylene adipate-co-terephthalate)-polylactic acid-thermoplastic starch based commercial bio-plastic film at ambient conditions.

Microbial biodegradation of commercially available poly(butylene adipate-co-terephthalate)-polylactic acid-thermoplastic starch based bio-plastic has been pursued at high temperatures exceeding 55 °C. Herein, we first reported three newly isolated fungal strains from farmland soil samples of Republic of Korea namely, Pyrenochaetopsis sp. strain K2, Staphylotrichum sp. S2-1, and Humicola sp. strain S2-3 were capable of degrading a commercial bio-plastic film with degradation rates of 9.5, 8.6, and 12.2%, respectively after 3 months incubation at ambient conditions. Scanning electron microscopy (SEM) analyses showed that bio-plastic film was extensively fragmented with severe cracking on the surface structure after incubation with isolated fungal strains. X-ray diffraction (XRD) analysis also revealed that high crystallinity of the commercial bio-plastic film was significantly decreased after degradation by fungal strains. Liquid chromatography-mass spectrometry (LC-MS) analyses of the fungal culture supernatants containing the bio-plastic film showed the peaks for adipic acid, terephthalic acid (TPA), and terephthalate-butylene (TB) as major metabolites, suggesting cleavage of ester bonds and accumulation of TPA. Furthermore, a consortium of fungal strain K2 with TPA degrading bacterium Pigmentiphaga sp. strain P3-2 isolated from the same sampling site exhibited faster degradation rate of the bio-plastic film within 1 month of incubation with achieving complete biodegradation of accumulated TPA. We assume that the extracellular lipase activity presented in the fungal cultures could hydrolyze the ester bonds of PBAT component of bio-plastic film. Taken together, the fungal and bacterial consortium investigated herein could be beneficial for efficient biodegradation of the commercial bio-plastic film at ambient conditions.

Nodulation Experiment by Cross-Inoculation of Nitrogen-Fixing Bacteria Isolated from Root Nodules of Several Leguminous Plants.

Root-nodule nitrogen-fixing bacteria are known for being specific to particular legumes. This study isolated the endophytic root-nodule bacteria from the nodules of legumes and examined them to determine whether they could be used to promote the formation of nodules in other legumes. Forty-six isolates were collected from five leguminous plants and screened for housekeeping (16S rRNA), nitrogen fixation (nifH), and nodulation (nodC) genes. Based on the 16S rRNA gene sequencing and phylogenetic analysis, the bacterial isolates WC15, WC16, WC24, and GM5 were identified as Rhizobium, Sphingomonas, Methylobacterium, and Bradyrhizobium, respectively. The four isolates were found to have the nifH gene, and the study confirmed that one isolate (GM5) had both the nifH and nodC genes. The Salkowski method was used to measure the isolated bacteria for their capacity to produce phytohormone indole acetic acid (IAA). Additional experiments were performed to examine the effect of the isolated bacteria on root morphology and nodulation. Among the four tested isolates, both WC24 and GM5 induced nodulation in Glycine max. The gene expression studies revealed that GM5 had a higher expression of the nifH gene. The existence and expression of the nitrogen-fixing genes implied that the tested strain had the ability to fix the atmospheric nitrogen. These findings demonstrated that a nitrogen-fixing bacterium, Methylobacterium (WC24), isolated from a Trifolium repens, induced the formation of root nodules in non-host leguminous plants (Glycine max). This suggested the potential application of these rhizobia as biofertilizer. Further studies are required to verify the N2-fixing efficiency of the isolates.

Resistome Study in Aquatic Environments.

Since the first discovery of antibiotics, introduction of new antibiotics has been coupled with the occurrence of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs). Rapid dissemination of ARB and ARGs in the aquatic environments has become a global concern. ARB and ARGs have been already disseminated in the aquatic environments via various routes. Main hosts of most of ARGs were found to belong to Gammaproteobacteria class, including clinically important potential pathogens. Transmission of ARGs also occurs by horizontal gene transfer (HGT) mechanisms between bacterial strains in the aquatic environments, resulting in ubiquity of ARGs. Thus, a few of ARGs and MGEs (e.g., strA, sul1, int1) have been suggested as indicators for global comparability of contamination level in the aquatic environments. With ARB and ARGs contamination, the occurrence of critical pathogens has been globally issued due to their widespread in the aquatic environments. Thus, active surveillance systems have been launched worldwide. In this review, we described advancement of methodologies for ARGs detection, and occurrence of ARB and ARGs and their dissemination in the aquatic environments. Even though numerous studies have been conducted for ARB and ARGs, there is still no clear strategy to tackle antibiotic resistance (AR) in the aquatic environments. At least, for consistent surveillance, a strict framework should be established for further research in the aquatic environments.

Prevalence and Characterization of CRISPR Locus 2.1 Spacers in Escherichia coli Isolates Obtained from Feces of Animals and Humans.

The clustered regularly interspaced short palindromic repeat (CRISPR) has been studied as an immune system in prokaryotes for the survival of bacteriophages. The CRISPR system in prokaryotes records the invasion of bacteriophages or other genetic materials in CRISPR loci. Accordingly, CRISPR loci can reveal a history of infection records of bacteriophages and other genetic materials. Therefore, identification of the CRISPR array may help trace the events that bacteria have undergone. In this study, we characterized and identified the spacers of the CRISPR loci in Escherichia coli isolates obtained from the feces of animals and humans. Most CRISPR spacers were found to stem from phages. Although we did not find any patterns in CRISPR spacers according to sources, our results showed that phage-derived spacers mainly originated from the families Inoviridae, Myoviridae, Podoviridae, and Siphoviridae and the order Caudovirales, whereas plasmid-derived CRISPR spacers were mainly from the Enterobacteriaceae family. In addition, it is worth noting that the isolates from each animal and human source harbored source-specific spacers. Considering that some of these taxa are likely found in the gut of mammalian animals, CRISPR spacers identified in these E. coli isolates were likely derived from the bacteriophageome and microbiome in closed gut environments. Although the bacteriophageome database limits the characterization of CRISPR arrays, the present study showed that some spacers were specifically found in both animal and human sources. Thus, this finding may suggest the possible use of E. coli CRISPR spacers as a microbial source tracking tool. IMPORTANCE We characterized spacers of CRISPR locus 2.1 in E. coli isolates obtained from the feces of various sources. Phage-derived CRISPR spacers are mainly acquired from the order Caudovirales, and plasmid-derived CRISPR spacers are mostly from the Enterobacteriaceae family. This is thought to reflect the microbiome and phageome of the gut environment of the sources. Hence, spacers may help track the encounter of bacterial cells with bacterial cells, viruses, or other genetic materials. Interestingly, source-specific spacers are also observed. The identification of source-specific spacers is thought to help develop the methodology of microbial source tracking and understanding the interactions between viruses and bacteria. However, very few spacers have been uncovered to track where they originate. The accumulation of genome sequences can help identify the hosts of spacers and can be applied for microbial source tracking.

Improvement of thermoalkaliphilic laccase (CtLac) by a directed evolution and application to lignin degradation.

Thermoalkaliphilic laccase (CtLac) from the Caldalkalibacillus thermarum strain TA2.A1 has advantageous properties with potential industrial applications, such as high enzyme activity and stability at 70 °C and pH 8.0. In the present study, a directed evolution approach using a combination of random and site-directed mutagenesis was adopted to enhance the laccase activity of CtLac. Spectrophotometric assay and real-time oxygen measurement techniques were employed to compare and evaluate the enzyme activity among mutants. V243 was targeted for site-directed mutagenesis based on library screening. V243D showed a 25-35% higher laccase activity than wild-type CtLac in the spectrophotometric assay and oxygen consumption measurement results. V243D also showed higher catalytic efficiency than wild-type CtLac with decreased Km and increased kcat values. In addition, V243D enhanced oxidative degradation of the lignin model compound, guaiacylglycerol-ß-guaiacyl ether (GGGE), by 10% and produced a 5-30% increase in high-value aldehydes than wild-type CtLac under optimal enzymatic conditions (i.e., 70 °C and pH 8.0). Considering the lack of protein structural information, we used the directed evolution approach to predict Val at the 243 position of CtLac as one of the critical amino acids contributing to the catalytic efficiency of the enzyme. Moreover, it found that the real-time oxygen measurement technique could overcome the limitations of the spectrophotometric assay, and apply to evaluate oxidase activity in mutagenesis research. KEY POINTS: • CtLac was engineered for enhanced laccase activity through directed evolution approach • V243D showed higher catalytic efficiency (kcat/Km) than wild-type CtLac • V243D produced higher amounts of high-value aldehydes from rice straw than wild-type CtLac.

Highly effective biosorption capacity of Cladosporium sp. strain F1 to lead phosphate mineral and perovskite solar cell PbI2.

Fungus Cladosporium sp. strain F1 showed highly effective biosorption capacity to lead phosphate mineral and perovskite solar cells lead iodide compared to other fungi Aspergillus niger VKMF-1119 and Mucor ramannianus R-56. Scanning electron microscopy and transmission electron microscopy analyses shows that Cladosporium sp. strain F1, which previously showed high biosorption capacity to uranium phosphate nanorods and nanoplates, can accumulate lead phosphate mineral and lead iodide on the fungal hyphae surface in large amounts under a wide range of pH conditions, while A. niger VKMF-1119 and M. ramannianus R-56 adsorbed small amounts of minerals. After biosorption of lead iodide minerals on Cladosporium sp. strain F1, aqueous dimethyl sulfoxide (50%) at pH 2 (70 °C) released the mineral more than 99%. Based on the fungal surface analyses, hydrophobic properties on the surfaces of Cladosporium sp. strain F1 could affect the higher biosorption capacity of strain F1 to lead phosphate mineral and lead iodide as compared to other tested fungi. Cladosporium sp. strain F1 may be the novel biosorbents to remediate the phosphate rich environment and to recover lead from perovskite solar cells lead iodide.

Dynamics of Genotypic and Phenotypic Antibiotic Resistance in a Conventional Wastewater Treatment Plant in 2 Years.

Wastewater treatment plants (WWTPs) are considered a sink and a source of antibiotic resistance. In this study, we applied both culture-dependent and SmartChip-based culture-independent approaches for the investigation of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) at Jungnang (JN), located in the Han River, Seoul, South Korea, for 2 years, i.e., 2017 and 2018. The JN WWTP reduced the diversity and abundance of ARB and ARGs but was not sufficient for removing them all. Interestingly, through the treatment process in the JN WWTP, the composition of diverse multidrug-resistant (MDR) bacteria was concentrated mainly into some genera of the Gammaproteobacteria class (Citrobacter, Escherichia-Shigella, and Stenotrophomonas), which could be key carriages to spread ARGs into the environments. In addition, SmartChip analyses showed that the relative abundance and the number of ARGs were significantly decreased from the influents to the effluents in both 2017 and 2018. SmartChip analyses for 2 years also allowed to notify the core ARGs in the influents and the effluents with the presence of clinically relevant core ARGs, such as vanC, bla OXA , and bla NDM , which persisted in the treatment process. Considering diverse bacterial mechanisms for exchanging and transferring ARGs, the occurrence of MDR bacteria and core ARGs could be a source for the blooming of the antibiotic resistome in the WWTP and nearby environments.

Analysis of phylogenetic markers for classification of a hydrogen peroxide producing Streptococcus oralis isolated from saliva by a newly devised differential medium.

Hydrogen peroxide (H2O2) is produced by alpha-hemolytic streptococci in aerobic conditions. However, the suitable method for detection of H2O2-producing streptococci in oral microbiota has not been setup. Here we show that o-dianisidine dye and horseradish peroxidase were useful in tryptic soy agar medium to detect and isolate H2O2-producing bacteria with the detection limit of one target colony in > 106 colony-forming units. As a proof, we isolated the strain HP01 (KCTC 21190) from a saliva sample using the medium and analyzed its characteristics. Further tests showed that the strain HP01 belongs to Streptococcus oralis in the Mitis group and characteristically forms short-chain streptococcal cells with a high capacity of acid tolerance and biofilm formation. The genome analysis revealed divergence of the strain HP01 from the type strains of S. oralis. They showed distinctive phylogenetic distances in their ROS-scavenging proteins, including superoxide dismutase SodA, thioredoxin TrxA, thioredoxin reductase TrxB, thioredoxin-like protein YtpP, and glutaredoxin-like protein NrdH, as well as a large number of antimicrobial resistance genes and horizontally transferred genes. The concatenated ROS-scavenging protein sequence can be used to identify and evaluate Streptococcus species and subspecies based on phylogenetic analysis.

Draft Genome Sequence of Desemzia sp. Strain C1, Producing Hydrogen Peroxide, Isolated from Oil-Contaminated Soil.

Here, we report the draft genome sequence of Desemzia sp. strain C1, which was isolated from oil-contaminated soil in South Korea and produces hydrogen peroxide (H2O2). The genome of Desemzia sp. strain C1 contains genes encoding various oxidases involved in H2O2 production and resistance to oxidative stress.

Antibacterial effect of copper nanoparticles produced in a Shewanella-supported non-external circuit bioelectrical system on bacterial plant pathogens.

The use of copper nanoparticles for the inhibition of plant pathogens Ralstonia solanacearum, which causes wilt disease, and Xanthomonas axonopodis, which causes citrus canker, was investigated in this study. To avoid the inhibiting effect of Cu2+ ions on the bacterial cells, the copper nanoparticles were synthesized in the cathode chamber of a non-external circuit bioelectrochemical system (nec_BES) inoculated with Shewanella sp. HN-41 at the anode. The electrons produced by the oxidation of lactate by Shewanella sp. HN-41 were directly transferred to the anolyte in the cathode via a graphite electrode connecting the anode and cathode chambers. SEM images of the produced particles revealed that the copper nanoparticles were aggregated into spherical shapes with an average size of 2.9 µm from smaller particles with a size range from 30 nm to approximately 190 nm. X-ray diffraction demonstrated that the copper nanoparticles were mainly in the form of a single-phase crystal mixture of atacamite (Cu2Cl(OH)3) and paracatamite (Cu2Cl(OH)3). Finally, for the application of synthesized nanoparticles, an agar diffusion test was applied to assess the antibacterial activity of the formed copper nanoparticles in propylene glycol solvent against R. solanacearum and X. axonopodis. The results showed that the nanoparticles damaged the cells of R. solanacearum, with a half maximum inhibition (IC50) value of 42 ppm, but did not damage X. axonopodis cells.

Phylogenetic diversity and spatiotemporal dynamics of bacterial and microeukaryotic plankton communities in Gwangyang Bay of the Korean Peninsula.

Nutrient dynamics function globally, flowing from rivers to the ocean (estuarine-coastal zone), and are vulnerable to climate change. Microbial habitats can be affected by marine nutrient dynamics and may provide a clue to predict microbial responses to environmental heterogeneity in estuarine-coastal zones. We surveyed surface seawater in Gwangyang Bay, a semi-enclosed estuary in Korea, from 2016 to 2018 using a metabarcoding approach with prokaryotic 16S and eukaryotic 18S rRNA genes. Bacterial and microeukaryotic communities in these waters showed distinct local communities in response to environmental heterogeneity and community transition at spatiotemporal scales in the estuarine-coastal zone. The relative abundance of prokaryotic and eukaryotic operational taxonomic units suggested a microbial trophic interaction in the Gwangyang Bay waters. We found that the community assembly process in prokaryotic communities was primarily influenced by biological interaction (immigration-emigration), whereas that in eukaryotic communities was more affected by environmental stress (habitat specificity) rather than by biotic factors. Our findings in the Gwangyang Bay waters may provide information on underlying (biotic or abiotic) factors of the assembly process in microbial communities in the estuarine-coastal zone.

Non-specific degradation of chloroacetanilide herbicides by glucose oxidase supported Bio-Fenton reaction.

Bio-Fenton reaction supported by glucose oxidase (GOx) for producing H2O2 was applied to degrade persistent chloroacetanilide herbicides in the presence of Fe (Ⅲ)-citrate at pH 5.5. There were pH decrease to 4.3, the production of 8 mM H2O2 and simultaneous consumption to produce •OH radicals which non-specifically degraded the herbicides. The degradation rates followed the order acetochlor ≈ alachlor ≈ metolachlor > propachlor ≈ butachlor with the degradation percent of 72.8%, 73.4%, 74.0%, 47.4%, and 43.8%, respectively. During the Bio-Fenton degradation, alachlor was dechlorinated and filtered into catechol via the production of intermediates formed through a series of hydrogen atom abstraction and hydrogen oxide radical addition reactions. The current Bio-Fenton reaction leading to the production of •OH radicals could be applied for non-specific oxidative degradation to various persistent organic pollutants under in-situ environmental conditions, considering diverse microbial metabolic systems able to continuously supply H2O2 with ubiquitous Fe(II) and Fe(III) and citrate.

Higher abundance of core antimicrobial resistant genes in effluent from wastewater treatment plants.

Wastewater treatment plants (WWTPs) receive sewage water from a variety of sources, including livestock farms, hospitals, industries, and households, that contain antimicrobial resistant bacteria (ARB) and antimicrobial resistant genes (ARGs). Current treatment technologies are unable to completely remove ARB and ARGs, which are eventually released into the aquatic environment. This study focused on the core resistome of urban WWTPs that are persistent through wastewater treatment processes. We adopted the Hiseq-based metagenomic sequencing approach to identify the core resistome, their genetic context, and pathogenic potential of core ARGs in the influent (IN) and effluent (EF) samples of 12 urban WWTPs in South Korea. In this study, the abundance of ARGs ranged from 0.32 to 3.5 copies of ARGs per copy of the 16S rRNA gene, where the IN samples were relatively higher than the EF samples, especially for the macrolide-lincosamide-streptogramin (MLS)- and tetracycline- resistant genes. On the other hand, there were 43 core ARGs sharing up to 90% of the total, among which the relative abundance of sul1, APH(3'')-lb, and RbpA was higher in EF than in IN (p < 0.05). Moreover, tetracycline and sulfonamide-related core ARGs in both EF and IN were significantly more abundant on plasmids than on chromosomes (p < 0.05). We also found that the majority of core ARGs were carried by opportunistic pathogens such as Acinetobacter baumannii, Enterobacter cloacae, and Pseudomonas aeruginosa in both IN and EF. In addition, phages were the only mobile elements whose abundance correlated with that of core ARGs in EF, suggesting that transduction may play a major role in disseminating ARGs in the receiving water environment of the urban WWTP. The persistent release of core ARGs with pathogenic potential into environmental water is of immediate concern. The mobility of ARGs and ARBs in the environment is a major public health concern. These results should be taken into consideration when developing policy to mitigate environmental dissemination of ARG by WWTPs.

Degradation of sulfonated polyethylene by a bio-photo-fenton approach using glucose oxidase immobilized on titanium dioxide.

Polyethylene (PE) plastics are highly recalcitrant and resistant to photo-oxidative degradation due to its chemically inert backbone structure. We applied two novel reactions such as, Bio-Fenton reaction using glucose oxidase (GOx) enzyme alone and Bio-Photo-Fenton reaction using GOx immobilized on TiO2 nanoparticles (TiO2-GOx) under UV radiation, for (bio)degradation of pre-activated PE with sulfonation (SPE). From both the reactions, GC-MS analyses identified small organic acids such as, acetic acid and butanoic acid as a major metabolites released from SPE. In the presence of UV radiation, 21 fold and 17 fold higher amounts of acetic acid (4.78 mM) and butanoic acid (0.17 mM) were released from SPE after 6 h of reaction using TiO2-GOx than free GOx, which released 0.22 mM and 0.01 mM of acetic acid and butanoic acid, respectively. Our results suggest that (bio)degradation and valorization of naturally weathered and oxidized PE using combined reactions of biochemistry, photochemistry and Fenton chemistry could be possible.

Layers of Uranium Phosphate Nanorods and Nanoplates Encrusted on Fungus Cladosporium sp. Strain F1 Hyphae.

The hyphae of Cladosporium sp. strain F1 (CFGR 2020-301-00084) were heavily encrusted with pre-synthesized uranium phosphate minerals under a wide range of pH conditions. SEM and TEM images showed that nanorods and nanoplates of uranium phosphate minerals at pH 4 and 5 and at pH 6, 7, and 8, respectively, were tightly adsorbed along the hyphae of Cladosporium sp. strain F1, while only a few uranium phosphate minerals were observed on the hyphae of Aspergillus niger VKMF 1119. Based on the physical mobility and chemical stability of uranium phosphate minerals under in situ oxidizing environmental conditions, the application of Cladosporium sp. strain F1 has potential as a novel strategy for the remediation of uranium contamination in sediments and aquifers under a wide range of pH conditions where larger amounts of phosphate are present in the environment.

Structure and substrate specificity determinants of NfnB, a dinitroaniline herbicide-catabolizing nitroreductase from Sphingopyxis sp. strain HMH.

Nitroreductases are emerging as attractive bioremediation enzymes, with substrate promiscuity toward both natural and synthetic compounds. Recently, the nitroreductase NfnB from Sphingopyxis sp. strain HMH exhibited metabolic activity for dinitroaniline herbicides including butralin and pendimethalin, triggering the initial steps of their degradation and detoxification. However, the determinants of the specificity of NfnB for these herbicides are unknown. In this study, we performed structural and biochemical analyses of NfnB to decipher its substrate specificity. The homodimer NfnB is a member of the PnbA subgroup of the nitroreductase family. Each monomer displays a central α + ß fold for the core domain, with a protruding middle region and an extended C-terminal region. The protruding middle region of Val75-Tyr129 represents a structural extension that is a common feature to members of the PnbA subgroup and functions as an opening wall connecting the coenzyme FMN-binding site to the surface, therefore serving as a substrate binding site. We performed mutational, kinetic, and structural analyses of mutant enzymes and found that Tyr88 in the middle region plays a pivotal role in substrate specificity by determining the dimensions of the wall opening. The mutation of Tyr88 to phenylalanine or alanine caused significant changes in substrate selectivity toward bulkier dinitroaniline herbicides such as oryzalin and isopropalin without compromising its activity. These results provide a framework to modify the substrate specificity of nitroreductase in the PnbA subgroup, which has been a challenging issue for its biotechnological and bioremediation applications.

Direct and Indirect Reduction of Cr(VI) by Fermentative Fe(III)-Reducing Cellulomonas sp. Strain Cellu-2a.

Hexavalent chromium (Cr(VI)) is recognized to be carcinogenic and toxic and registered as a contaminant in many drinking water regulations. It occurs naturally and is also produced by industrial processes. The reduction of Cr(VI) to Cr(III) has been a central topic for chromium remediation since Cr(III) is less toxic and less mobile. In this study, fermentative Fe(III)-reducing bacterial strains (Cellu-2a, Cellu-5a, and Cellu-5b) were isolated from a groundwater sample and were phylogenetically related to species of Cellulomonas by 16S rRNA gene analysis. One selected strain, Cellu-2a showed its capacity of reduction of both soluble iron (ferric citrate) and solid iron (hydrous ferric oxide, HFO), as well as aqueous Cr(VI). The strain Cellu-2a was able to reduce 15 µM Cr(VI) directly with glucose or sucrose as a sole carbon source under the anaerobic condition and indirectly with one of the substrates and HFO in the same incubations. The heterogeneous reduction of Cr(VI) by the surface-associated reduced iron from HFO by Cellu-2a likely assisted the Cr(VI) reduction. Fermentative features such as large-scale cell growth may impose advantages on the application of bacterial Cr(VI) reduction over anaerobic respiratory reduction.

Emergence of High Level Carbapenem and Extensively Drug Resistant Escherichia coli ST746 Producing NDM-5 in Influent of Wastewater Treatment Plant, Seoul, South Korea.

High level carbapenem and extensively drug resistant (XDR) Escherichia coli strain N7, which produces a variant of New Delhi metallo-ß-lactamase (NDM-5), was isolated from the influent of the Jungnang wastewater treatment plant located on Han River, Seoul, South Korea. Phenotypic and genotypic resistances to carbapenem were tested using agar and broth dilution methods, and polymerase chain reaction. Whole-genome sequencing was performed to characterize the genetic structure of strain N7. E. coli strain N7, which harbors the bla NDM-5 gene, showed high level of carbapenem resistance at concentrations of doripenem (512 mg/L) and meropenem (256 mg/L), and XDR to 15 antibiotics. Based on the genomic sequence analysis, two plasmids, a hybrid IncHI2/N-type and an IncX3 type, were present. The former contains a cluster (bla NDM-5-ble MBL -trpF-dsbD) bracketed by multi-insertional sequences, IS3000, ISAba125, IS5, and IS26. The latter carries the following resistance genes: bla CTX-14, aac(3)-IV, aadA1, aadA2, aph(3')-Ia, aph(4)-Ia, sul1, sul2, sul3, dfrA12, fosA3, oqxA, oqxB, mph(A), and floR, and cmlA1. The chromosome, contig3, and contig5 also carry bla CTX-64 and mdf(A), tet(A), and erm(B), tet(M) and aadA22, respectively. Strain N7 also harbors virulence factors such as fimH, flu, ecpABCDE, sfmA, hlyE, and gadA. This study demonstrates the emergence of high level carbapenem resistant XDR E. coli strain N7 containing bla NDM-5 in aquatic environment, Seoul, South Korea. Due to the presence of mobile genetic elements, this strain could horizontally transfer resistance genes, including bla NDM-5 to environmental bacteria. Thus, it is necessary to conduct continuous surveillance for carbapenem resistance in various aquatic environments.

Metagenomic exploration of antibiotic resistome in treated wastewater effluents and their receiving water.

Environmental dissemination of antimicrobial resistance is a global health problem. Antimicrobial-resistant bacteria and antibiotic-resistant genes (ARGs) are constantly released into the environment through effluents (EFs) from wastewater treatment plants (WWTPs). Thus, requiring a better understanding of the selection and fate of ARGs in wastewater treatment processes. Therefore, we investigated the impacts of urban WWTP EFs on receiving water in the context of their resistomes and mobilomes. We used a HiSeq-based short read metagenomic approach to address the dynamics and diversity of ARGs in WWTP EF as well as the upstream (UP) and downstream (DN) river waters, followed by an investigation of plasmid-mediated ARGs. The abundance of ARGs at each site varied from 7.2 × 10-2 to 7.4 × 10-1 ARG copies per 16S rRNA gene copy, and EF samples showed the highest abundance, followed by DN and UP water samples. ARG diversity ranged from 121 to 686 types per site, and EF had the most diverse ARGs. Commonly identified ARGs in the EF and DN samples were clinically important and were absent in UP samples. The abundance of ARGs, mobile genetic elements (MGEs), and plasmid contigs found only in EF and DN were positively correlated with each other, indicating the importance of mobilomes in the dissemination of ARGs in the environment. Moreover, the proportions of plasmid-mediated ARGs was highest in the EF samples, followed by the DN and UP samples. These findings suggest that WWTP EF may act as a driving factor shaping the resistomes and mobilomes of receiving waters. In particular, a higher abundance of plasmid-mediated ARGs in WWTP EF suggests higher transmissibility in the DN environment.