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September 22, 2019  |  

Characteristics of ARG-carrying plasmidome in the cultivable microbial community from wastewater treatment system under high oxytetracycline concentration.

Studies on antibiotic production wastewater have shown that even a single antibiotic can select for multidrug resistant bacteria in aquatic environments. It is speculated that plasmids are an important mechanism of multidrug resistance (MDR) under high concentrations of antibiotics. Herein, two metagenomic libraries were constructed with plasmid DNA extracted from cultivable microbial communities in a biological wastewater treatment reactor supplemented with 0 (CONTROL) or 25 mg/L of oxytetracycline (OTC-25). The OTC-25 plasmidome reads were assigned to 72 antibiotic resistance genes (ARGs) conferring resistance to 13 types of antibiotics. Dominant ARGs, encoding resistance to tetracycline, aminoglycoside, sulfonamide, and multidrug resistance genes, were enriched in the plasmidome under 25 mg/L of oxytetracycline. Furthermore, 17 contiguous multiple-ARG carrying contigs (carrying =?2 ARGs) were discovered in the OTC-25 plasmidome, whereas only nine were found in the CONTROL. Mapping of the OTC-25 plasmidome reads to completely sequenced plasmids revealed that the conjugative IncU resistance plasmid pFBAOT6 of Aeromonas caviae, carrying multidrug resistance transporter (pecM), tetracycline resistance genes (tetA, tetR), and transposase genes, might be a potential prevalent resistant plasmid in the OTC-25 plasmidome. Additionally, two novel resistant plasmids (containing contig C301682 carrying multidrug resistant operon mexCD-oprJ and contig C301632 carrying the tet36 and transposases genes) might also be potential prevalent resistant plasmids in the OTC-25 plasmidome. This study will be helpful to better understand the role of plasmids in the development of MDR in water environments under high antibiotic concentrations.

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September 22, 2019  |  

Genome analysis of Taraxacum kok-saghyz Rodin provides new insights into rubber biosynthesis

The Russian dandelion Taraxacum kok-saghyz Rodin (TKS), a member of the Composite family and a potential alternative source of natural rubber (NR) and inulin, is an ideal model system for studying rubber biosynthesis. Here we present the draft genome of TKS, the first assembled NR-producing weed plant. The draft TKS genome assembly has a length of 1.29 Gb, containing 46,731 predicted protein-coding genes and 68.56% repeats, in which the LTR-RT elements predominantly contribute to the genome enlargement. We analyzed the heterozygous regions/genes, suggesting its possible involvement in inbreeding depression. Through comparative studies between rubber-producing and non-rubber-producing plants, we found that enzymes of the mevalonate (MVA) pathway and rubber elongation might be critical for rubber biosynthesis, and several key isoforms have been isolated showing predominantly expressed in the latex, indicating their crucial functions in rubber biosynthesis. Moreover, for two important families in rubber elongation, the CPT/CPTL and REF/SRPP families, diverse evolutionary tracks have been revealed. These results provide valuable resources and new insights into the mechanism of NR biosynthesis, and facilitate the development of alternative NR producing crops.

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September 22, 2019  |  

The non-specific adenine DNA methyltransferase M.EcoGII.

We describe the cloning, expression and characterization of the first truly non-specific adenine DNA methyltransferase, M.EcoGII. It is encoded in the genome of the pathogenic strain Escherichia coli O104:H4 C227-11, where it appears to reside on a cryptic prophage, but is not expressed. However, when the gene encoding M.EcoGII is expressed in vivo – using a high copy pRRS plasmid vector and a methylation-deficient E. coli host-extensive in vivo adenine methylation activity is revealed. M.EcoGII methylates adenine residues in any DNA sequence context and this activity extends to dA and rA bases in either strand of a DNA:RNA-hybrid oligonucleotide duplex and to rA bases in RNAs prepared by in vitro transcription. Using oligonucleotide and bacteriophage M13mp18 virion DNA substrates, we find that M.EcoGII also methylates single-stranded DNA in vitro and that this activity is only slightly less robust than that observed using equivalent double-stranded DNAs. In vitro assays, using purified recombinant M.EcoGII enzyme, demonstrate that up to 99% of dA bases in duplex DNA substrates can be methylated thereby rendering them insensitive to cleavage by multiple restriction endonucleases. These properties suggest that the enzyme could also be used for high resolution mapping of protein binding sites in DNA and RNA substrates.© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

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September 22, 2019  |  

Genomic diversity in the endosymbiotic bacterium Rhizobium leguminosarum.

Rhizobium leguminosarum bv. viciae is a soil a-proteobacterium that establishes a diazotrophic symbiosis with different legumes of the Fabeae tribe. The number of genome sequences from rhizobial strains available in public databases is constantly increasing, although complete, fully annotated genome structures from rhizobial genomes are scarce. In this work, we report and analyse the complete genome of R. leguminosarum bv. viciae UPM791. Whole genome sequencing can provide new insights into the genetic features contributing to symbiotically relevant processes such as bacterial adaptation to the rhizosphere, mechanisms for efficient competition with other bacteria, and the ability to establish a complex signalling dialogue with legumes, to enter the root without triggering plant defenses, and, ultimately, to fix nitrogen within the host. Comparison of the complete genome sequences of two strains of R. leguminosarum bv. viciae, 3841 and UPM791, highlights the existence of different symbiotic plasmids and a common core chromosome. Specific genomic traits, such as plasmid content or a distinctive regulation, define differential physiological capabilities of these endosymbionts. Among them, strain UPM791 presents unique adaptations for recycling the hydrogen generated in the nitrogen fixation process.

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September 22, 2019  |  

Translating genomics into practice for real-time surveillance and response to carbapenemase-producing Enterobacteriaceae: evidence from a complex multi-institutional KPC outbreak.

Until recently, Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae were rarely identified in Australia. Following an increase in the number of incident cases across the state of Victoria, we undertook a real-time combined genomic and epidemiological investigation. The scope of this study included identifying risk factors and routes of transmission, and investigating the utility of genomics to enhance traditional field epidemiology for informing management of established widespread outbreaks.All KPC-producing Enterobacteriaceae isolates referred to the state reference laboratory from 2012 onwards were included. Whole-genome sequencing was performed in parallel with a detailed descriptive epidemiological investigation of each case, using Illumina sequencing on each isolate. This was complemented with PacBio long-read sequencing on selected isolates to establish high-quality reference sequences and interrogate characteristics of KPC-encoding plasmids.Initial investigations indicated that the outbreak was widespread, with 86 KPC-producing Enterobacteriaceae isolates (K. pneumoniae 92%) identified from 35 different locations across metropolitan and rural Victoria between 2012 and 2015. Initial combined analyses of the epidemiological and genomic data resolved the outbreak into distinct nosocomial transmission networks, and identified healthcare facilities at the epicentre of KPC transmission. New cases were assigned to transmission networks in real-time, allowing focussed infection control efforts. PacBio sequencing confirmed a secondary transmission network arising from inter-species plasmid transmission. Insights from Bayesian transmission inference and analyses of within-host diversity informed the development of state-wide public health and infection control guidelines, including interventions such as an intensive approach to screening contacts following new case detection to minimise unrecognised colonisation.A real-time combined epidemiological and genomic investigation proved critical to identifying and defining multiple transmission networks of KPC Enterobacteriaceae, while data from either investigation alone were inconclusive. The investigation was fundamental to informing infection control measures in real-time and the development of state-wide public health guidelines on carbapenemase-producing Enterobacteriaceae surveillance and management.

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September 22, 2019  |  

Complete genome sequence of Lactobacillus pentosus SLC13, isolated from mustard pickles, a potential probiotic strain with antimicrobial activity against foodborne pathogenic microorganisms.

Lactobacillus pentosus SLC13 is a high exopolysaccharide (EPS)-producing strain with broad-spectrum antimicrobial activity and the ability to grow in simulated gastrointestinal conditions. SLC13 was isolated from mustard pickles in Taiwan for potential probiotic applications. To better understand the molecular base for its antimicrobial activity and high EPS production, entire genome of SLC13 was determined by PacBio SMRT sequencing.L. pentosus SLC13 contains a genome with a 3,520,510-bp chromosome and a 62,498-bp plasmid. GC content of the complete genome was 46.5% and that of plasmid pSLC13 was 41.3%. Sequences were annotated at the RAST prokaryotic genome annotation server, and the results showed that the genome contained 3172 coding sequences and 82 RNA genes. Seventy-six protein-coding sequences were identified on the plasmid pSLC13. A plantaricin gene cluster, which is responsible for bacteriosins biosynthesis and could be associated with its broad-spectrum antimicrobial activity, was identified based on comparative genomic analysis. Two gene clusters involved in EPS production were also identified.This genomic sequence might contribute to a future application of this strain as probiotic in productive livestock potentially inhibiting competing and pathogenic organisms.

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September 22, 2019  |  

Convergent evolution driven by rifampin exacerbates the global burden of drug-resistant Staphylococcus aureus.

Mutations in the beta-subunit of bacterial RNA polymerase (RpoB) cause resistance to rifampin (Rifr), a critical antibiotic for treatment of multidrug-resistantStaphylococcus aureus.In vitrostudies have shown that RpoB mutations confer decreased susceptibility to other antibiotics, but the clinical relevance is unknown. Here, by analyzing 7,099S. aureusgenomes, we demonstrate that the most prevalent RpoB mutations promote clinically relevant phenotypic plasticity resulting in the emergence of stableS. aureuslineages, associated with increased risk of therapeutic failure through generation of small-colony variants (SCVs) and coresistance to last-line antimicrobial agents. We found eight RpoB mutations that accounted for 93% (469/505) of the total number of Rifrmutations. The most frequently selected amino acid substitutions affecting residue 481 (H481N/Y) were associated with worldwide expansions of Rifrclones spanning decades. Recreating the H481N/Y mutations confirmed no impact onS. aureusgrowth, but the H481N mutation promoted the emergence of a subpopulation of stable RifrSCVs with reduced susceptibility to vancomycin and daptomycin. Recreating the other frequent RpoB mutations showed similar impacts on resistance to these last-line agents. We found that 86% of all Rifrisolates in our global sample carried the mutations promoting cross-resistance to vancomycin and 52% to both vancomycin and daptomycin. As four of the most frequent RpoB mutations confer only low-level Rifr, equal to or below some international breakpoints, we recommend decreasing these breakpoints and reconsidering the appropriate use of rifampin to reduce the fixation and spread of these clinically deleterious mutations. IMPORTANCE Increasing antibiotic resistance in the major human pathogenStaphylococcus aureusis threatening the ability to treat patients with these infections. Recent laboratory studies suggest that mutations in the gene commonly associated with rifampin resistance may also impact susceptibility to other last-line antibiotics inS. aureus; however, the overall frequency and clinical impact of these mutations are unknown. By mining a global collection of clinicalS. aureusgenomes and by mutagenesis experiments, this work reveals that common rifampin-inducedrpoBmutations promote phenotypic plasticity that has led to the global emergence of stable, multidrug-resistantS. aureuslineages that are associated with increased risk of therapeutic failure through coresistance to other last-line antimicrobials. We recommend decreasing susceptibility breakpoints for rifampin to allow phenotypic detection of criticalrpoBmutations conferring low resistance to rifampin and reconsidering the appropriate use of rifampin to reduce the fixation and spread of these deleterious mutations globally.

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September 22, 2019  |  

The repeat structure of two paralogous genes, Yersinia ruckeri invasin (yrInv) and a “Y. ruckeri invasin-like molecule”, (yrIlm) sheds light on the evolution of adhesive capacities of a fish pathogen.

Inverse autotransporters comprise the recently identified type Ve secretion system and are exemplified by intimin from enterohaemorrhagic Escherichia coli and invasin from enteropathogenic Yersiniae. These proteins share a common domain architecture and promote bacterial adhesion to host cells. Here, we identified and characterized two putative inverse autotransporter genes in the fish pathogen Yersinia ruckeri NVH_3758, namely yrInv (for Y. ruckeri invasin) and yrIlm (for Y. ruckeri invasin-like molecule). When trying to clone the highly repetitive genes for structural and functional studies, we experienced problems in obtaining PCR products. PCR failures and the highly repetitive nature of inverse autotransporters prompted us to sequence the genome of Y. ruckeri NVH_3758 using PacBio sequencing, which produces some of the longest average read lengths available in the industry at this moment. According to our sequencing data, YrIlm is composed of 2603 amino acids (7812bp) and has a molecular mass of 256.4kDa. Based on the new genome information, we performed PCR analysis on four non-sequenced Y. ruckeri strains as well as the sequenced. Y. ruckeri type strain. We found that the genes are variably present in the strains, and that the length of yrIlm, when present, also varies. In addition, the length of the gene product for all strains, including the type strain, was much longer than expected based on deposited sequences. The internal repeats of the yrInv gene product are highly diverged, but represent the same bacterial immunoglobulin-like domains as in yrIlm. Using qRT-PCR, we found that yrIlm and yrInv are differentially expressed under conditions relevant for pathogenesis. In addition, we compared the genomic context of both genes in the newly sequenced Y. ruckeri strain to all available PacBio-sequenced Y. ruckeri genomes, and found indications of recent events of horizontal gene transfer. Taken together, this study demonstrates and highlights the power of Single Molecule Real-Time technology for sequencing highly repetitive proteins, and sheds light on the genetic events that gave rise to these highly repetitive genes in a commercially important fish pathogen. Copyright © 2017 Elsevier Inc. All rights reserved.

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September 22, 2019  |  

Characterization and heterologous expression of the neoabyssomicin/abyssomicin biosynthetic gene cluster from Streptomyces koyangensis SCSIO 5802.

The deep-sea-derived microbe Streptomyces koyangensis SCSIO 5802 produces neoabyssomicins A-B (1-2) and abyssomicins 2 (3) and 4 (4). Neoabyssomicin A (1) augments human immunodeficiency virus-1 (HIV-1) replication whereas abyssomicin 2 (3) selectively reactivates latent HIV and is also active against Gram-positive pathogens including methicillin-resistant Staphylococcus aureus (MRSA). Structurally, neoabyssomicins A-B constitute a new subtype within the abyssomicin family and feature unique structural traits characteristic of extremely interesting biosynthetic transformations.In this work, the biosynthetic gene cluster (BGC) for the neoabyssomicins and abyssomicins, composed of 28 opening reading frames, was identified in S. koyangensis SCSIO 5802, and its role in neoabyssomicin/abyssomicin biosynthesis was confirmed via gene inactivation and heterologous expression experiments. Bioinformatics and genomics analyses enabled us to propose a biosynthetic pathway for neoabyssomicin/abyssomicin biosynthesis. Similarly, a protective export system by which both types of compounds are secreted from the S. koyangensis producer was identified, as was a four-component ABC transporter-based import system central to neoabyssomicin/abyssomicin biosynthesis. Furthermore, two regulatory genes, abmI and abmH, were unambiguously shown to be positive regulators of neoabyssomicin/abyssomicin biosynthesis. Consistent with their roles as positive regulatory genes, the overexpression of abmI and abmH (independent of each other) was shown to improve neoabyssomicin/abyssomicin titers.These studies provide new insight into the biosynthesis of the abyssomicin class of natural products, and highlight important exploitable features of its BGC for future efforts. Elucidation of the neoabyssomicin/abyssomicin BGC now enables combinatorial biosynthetic initiatives aimed at improving both the titers and pharmaceutical properties of these important natural products-based drug leads.

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September 22, 2019  |  

Emergence of an extensively drug-resistant Salmonella enterica serovar Typhi clone harboring a promiscuous plasmid encoding resistance to fluoroquinolones and third-generation cephalosporins.

Antibiotic resistance is a major problem in Salmonella enterica serovar Typhi, the causative agent of typhoid. Multidrug-resistant (MDR) isolates are prevalent in parts of Asia and Africa and are often associated with the dominant H58 haplotype. Reduced susceptibility to fluoroquinolones is also widespread, and sporadic cases of resistance to third-generation cephalosporins or azithromycin have also been reported. Here, we report the first large-scale emergence and spread of a novel S. Typhi clone harboring resistance to three first-line drugs (chloramphenicol, ampicillin, and trimethoprim-sulfamethoxazole) as well as fluoroquinolones and third-generation cephalosporins in Sindh, Pakistan, which we classify as extensively drug resistant (XDR). Over 300 XDR typhoid cases have emerged in Sindh, Pakistan, since November 2016. Additionally, a single case of travel-associated XDR typhoid has recently been identified in the United Kingdom. Whole-genome sequencing of over 80 of the XDR isolates revealed remarkable genetic clonality and sequence conservation, identified a large number of resistance determinants, and showed that these isolates were of haplotype H58. The XDR S. Typhi clone encodes a chromosomally located resistance region and harbors a plasmid encoding additional resistance elements, including the blaCTX-M-15 extended-spectrum ß-lactamase, and carrying the qnrS fluoroquinolone resistance gene. This antibiotic resistance-associated IncY plasmid exhibited high sequence identity to plasmids found in other enteric bacteria isolated from widely distributed geographic locations. This study highlights three concerning problems: the receding antibiotic arsenal for typhoid treatment, the ability of S. Typhi to transform from MDR to XDR in a single step by acquisition of a plasmid, and the ability of XDR clones to spread globally. IMPORTANCE Typhoid fever is a severe disease caused by the Gram-negative bacterium Salmonella enterica serovar Typhi. Antibiotic-resistant S. Typhi strains have become increasingly common. Here, we report the first large-scale emergence and spread of a novel extensively drug-resistant (XDR) S. Typhi clone in Sindh, Pakistan. The XDR S. Typhi is resistant to the majority of drugs available for the treatment of typhoid fever. This study highlights the evolving threat of antibiotic resistance in S. Typhi and the value of antibiotic susceptibility testing and whole-genome sequencing in understanding emerging infectious diseases. We genetically characterized the XDR S. Typhi to investigate the phylogenetic relationship between these isolates and a global collection of S. Typhi isolates and to identify multiple genes linked to antibiotic resistance. This S. Typhi clone harbored a promiscuous antibiotic resistance plasmid previously identified in other enteric bacteria. The increasing antibiotic resistance in S. Typhi observed here adds urgency to the need for typhoid prevention measures.

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September 22, 2019  |  

The genome sequence of a new strain of Mycobacterium ulcerans ecovar Liflandii, emerging as a sturgeon pathogen

Mycobacterium ulcerans ecovar Liflandii (MuLiflandii) is emerging as a non-mycobacterial pathogen in amphibians. Here, we make the first report on the prevalence of a new strain of MuLiflandii infection in Chinese sturgeon. All the diseased fish showed the classic clinical symptoms of ascites and/or muscle ulceration. A new slow-growing and acid-fast bacillus ASM001 strain was obtained from the ascites of infected fish; this strain demonstrated pathogenicity when tested in hybrid sturgeon. The complete genome sequence of MuLiflandii ASM001 is a circular chromosome of 6,167,296?bp, with a G?+?C content of 65.57%, containing 4518 predicted coding DNA sequences and 999 pseudo-genes, 3 rRNA operons, and 47 transfer RNA sequences. In addition, we found 245 copies of IS2404, 34 microsatellites, and 36 CRISPR sequences in the whole MuLiflandii ASM001 genome. Among the predicted genes of MuLiflandii ASM001, we found orthologs of 203 virulence factors of clinical MuLiflandii 128FXT operating in host cell invasion, modulation of phagocyte function, and survival inside the macrophages. These virulence factor candidates provide a key basis for understanding their pathogenic mechanisms at the molecular level. A comparative analysis that used complete, existing genomes showed that MuLiflandii ASM001 has high synteny with MuLiflandii 128FXT. We anticipate the availability of the complete MuLiflandii ASM001 genome sequence will provide a valuable resource for comparative genomic studies of MuLiflandii isolates, as well as provide new insights into the host, ecological, and functional diversity of the genus Mycobacterium.

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September 22, 2019  |  

Genomic diversity of Taylorella equigenitalis introduced into the United States from 1978 to 2012.

Contagious equine metritis is a disease of worldwide concern in equids. The United States is considered to be free of the disease although sporadic outbreaks have occurred over the last few decades that were thought to be associated with the importation of horses. The objective of this study was to create finished, reference quality genomes that characterize the diversity of Taylorella equigenitalis isolates introduced into the USA, and identify their differences. Five isolates of T. equigenitalis associated with introductions into the USA from unique sources were sequenced using both short and long read chemistries allowing for complete assembly and annotation. These sequences were compared to previously published genomes as well as the short read sequences of the 200 isolates in the National Veterinary Services Laboratories’ diagnostic repository to identify unique regions and genes, potential virulence factors, and characterize diversity. The 5 genomes varied in size by up to 100,000 base pairs, but averaged 1.68 megabases. The majority of that diversity in size can be explained by repeat regions and 4 main regions of difference, which ranged in size from 15,000 to 45,000 base pairs. The first region of difference contained mostly hypothetical proteins, the second contained the CRISPR, the third contained primarily hemagglutinin proteins, and the fourth contained primarily segments of a type IV secretion system. As expected and previously reported, little evidence of recombination was found within these genomes. Several additional areas of interest were also observed including a mechanism for streptomycin resistance and other virulence factors. A SNP distance comparison of the T. equigenitalis isolates and Mycobacterium tuberculosis complex (MTBC) showed that relatively, T. equigenitalis was a more diverse species than the entirety of MTBC.

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September 22, 2019  |  

Conserved genomic and amino acid traits of cold adaptation in subzero-growing Arctic permafrost bacteria.

Permafrost accounts for 27% of all soil ecosystems and harbors diverse microbial communities. Our understanding of microorganisms in permafrost, their activities and adaptations, remains limited. Using five subzero-growing (cryophilic) permafrost bacteria, we examined features of cold adaptation through comparative genomic analyses with mesophilic relatives. The cryophiles possess genes associated with cold adaptation, including cold shock proteins, RNA helicases, and oxidative stress and carotenoid synthesis enzymes. Higher abundances of genes associated with compatible solutes were observed, important for osmoregulation in permafrost brine veins. Most cryophiles in our study have higher transposase copy numbers than mesophiles. We investigated amino acid (AA) modifications in the cryophiles favoring increased protein flexibility at cold temperatures. Although overall there were few differences with the mesophiles, we found evidence of cold adaptation, with significant differences in proline, serine, glycine and aromaticity, in several cryophiles. The use of cold/hot AA ratios of >1, used in previous studies to indicate cold adaptation, was found to be inadequate on its own. Comparing the average of all cryophiles to all mesophiles, we found that overall cryophiles had a higher ratio of cold adapted proteins for serine (more serine), and to a lesser extent, proline and acidic residues (fewer prolines/acidic residues).

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September 22, 2019  |  

Multi-omics approach identifies novel pathogen-derived prognostic biomarkers in patients with Pseudomonas aeruginosa bloodstream infection

Pseudomonas aeruginosa is a human pathogen that causes health-care associated blood stream infections (BSI). Although P. aeruginosa BSI are associated with high mortality rates, the clinical relevance of pathogen-derived prognostic biomarker to identify patients at risk for unfavorable outcome remains largely unexplored. We found novel pathogen-derived prognostic biomarker candidates by applying a multi-omics approach on a multicenter sepsis patient cohort. Multi-level Cox regression was used to investigate the relation between patient characteristics and pathogen features (2298 accessory genes, 1078 core protein levels, 107 parsimony-informative variations in reported virulence factors) with 30-day mortality. Our analysis revealed that presence of the helP gene encoding a putative DEAD-box helicase was independently associated with a fatal outcome (hazard ratio 2.01, p = 0.05). helP is located within a region related to the pathogenicity island PAPI-1 in close proximity to a pil gene cluster, which has been associated with horizontal gene transfer. Besides helP, elevated protein levels of the bacterial flagellum protein FliL (hazard ratio 3.44, p < 0.001) and of a bacterioferritin-like protein (hazard ratio 1.74, p = 0.003) increased the risk of death, while high protein levels of a putative aminotransferase were associated with an improved outcome (hazard ratio 0.12, p < 0.001). The prognostic potential of biomarker candidates and clinical factors was confirmed with different machine learning approaches using training and hold-out datasets. The helP genotype appeared the most attractive biomarker for clinical risk stratification due to its relevant predictive power and ease of detection.

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September 22, 2019  |  

PhdA catalyzes the first step of phenazine-1-carboxylic acid degradation in Mycobacterium fortuitum.

Phenazines are a class of bacterially produced redox-active metabolites that are found in natural, industrial, and clinical environments. In Pseudomonas spp., phenazine-1-carboxylic acid (PCA)-the precursor of all phenazine metabolites-facilitates nutrient acquisition, biofilm formation, and competition with other organisms. While the removal of phenazines negatively impacts these activities, little is known about the genes or enzymes responsible for phenazine degradation by other organisms. Here, we report that the first step of PCA degradation by Mycobacterium fortuitum is catalyzed by a phenazine-degrading decarboxylase (PhdA). PhdA is related to members of the UbiD protein family that rely on a prenylated flavin mononucleotide cofactor for activity. The gene for PhdB, the enzyme responsible for cofactor synthesis, is present in a putative operon with the gene encoding PhdA in a region of the M. fortuitum genome that is essential for PCA degradation. PhdA and PhdB are present in all known PCA-degrading organisms from the ActinobacteriaM. fortuitum can also catabolize other Pseudomonas-derived phenazines such as phenazine-1-carboxamide, 1-hydroxyphenazine, and pyocyanin. On the basis of our previous work and the current characterization of PhdA, we propose that degradation converges on a common intermediate: dihydroxyphenazine. An understanding of the genes responsible for degradation will enable targeted studies of phenazine degraders in diverse environments.IMPORTANCE Bacteria from phylogenetically diverse groups secrete redox-active metabolites that provide a fitness advantage for their producers. For example, phenazines from Pseudomonas spp. benefit the producers by facilitating anoxic survival and biofilm formation and additionally inhibit competitors by serving as antimicrobials. Phenazine-producing pseudomonads act as biocontrol agents by leveraging these antibiotic properties to inhibit plant pests. Despite this importance, the fate of phenazines in the environment is poorly understood. Here, we characterize an enzyme from Mycobacterium fortuitum that catalyzes the first step of phenazine-1-carboxylic acid degradation. Knowledge of the genetic basis of phenazine degradation will facilitate the identification of environments where this activity influences the microbial community structure. Copyright © 2018 American Society for Microbiology.

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