Aminobacter sp. MSH1 Mineralizes the Groundwater Micropollutant 2,6-Dichlorobenzamide through a Unique Chlorobenzoate Catabolic Pathway.

2,6-Dichlorobenzamide (BAM) is a major groundwater micropollutant posing problems for drinking water treatment plants (DWTPs) that depend on groundwater intake. Aminobacter sp. MSH1 uses BAM as the sole source of carbon, nitrogen, and energy and is considered a prime biocatalyst for groundwater bioremediation in DWTPs. Its use in bioremediation requires knowledge of its BAM-catabolic pathway, which is currently restricted to the amidase BbdA converting BAM into 2,6-dichlorobenzoic acid (2,6-DCBA) and the monooxygenase BbdD transforming 2,6-DCBA into 2,6-dichloro-3-hydroxybenzoic acid. Here, we show that the 2,6-DCBA catabolic pathway is unique and differs substantially from catabolism of other chlorobenzoates. BbdD catalyzes a second hydroxylation, forming 2,6-dichloro-3,5-dihydroxybenzoic acid. Subsequently, glutathione-dependent dehalogenases (BbdI and BbdE) catalyze the thiolytic removal of the first chlorine. The remaining chlorine is then removed hydrolytically by a dehalogenase of the α/ß hydrolase superfamily (BbdC). BbdC is the first enzyme in that superfamily associated with dehalogenation of chlorinated aromatics and appears to represent a new subtype within the α/ß hydrolase dehalogenases. The activity of BbdC yields a unique trihydroxylated aromatic intermediate for ring cleavage that is performed by an extradiol dioxygenase (BbdF) producing 2,4,6-trioxoheptanedioic acid, which is likely converted to Krebs cycle intermediates by BbdG.

Broad-spectrum antimicrobial activity by Burkholderia cenocepacia TAtl-371, a strain isolated from the tomato rhizosphere.

The Burkholderia cepacia complex (Bcc) comprises a group of 24 species, many of which are opportunistic pathogens of immunocompromised patients and also are widely distributed in agricultural soils. Several Bcc strains synthesize strain-specific antagonistic compounds. In this study, the broad killing activity of B. cenocepacia TAtl-371, a Bcc strain isolated from the tomato rhizosphere, was characterized. This strain exhibits a remarkable antagonism against bacteria, yeast and fungi including other Bcc strains, multidrug-resistant human pathogens and plant pathogens. Genome analysis of strain TAtl-371 revealed several genes involved in the production of antagonistic compounds: siderophores, bacteriocins and hydrolytic enzymes. In pursuit of these activities, we observed growth inhibition of Candida glabrata and Paraburkholderia phenazinium that was dependent on the iron concentration in the medium, suggesting the involvement of siderophores. This strain also produces a previously described lectin-like bacteriocin (LlpA88) and here this was shown to inhibit only Bcc strains but no other bacteria. Moreover, a compound with an m/z 391.2845 with antagonistic activity against Tatumella terrea SHS 2008T was isolated from the TAtl-371 culture supernatant. This strain also contains a phage-tail-like bacteriocin (tailocin) and two chitinases, but the activity of these compounds was not detected. Nevertheless, the previous activities are not responsible for the whole antimicrobial spectrum of TAtl-371 seen on agar plates, suggesting the presence of other compounds yet to be found. In summary, we observed a diversified antimicrobial activity for strain TAtl-371 and believe it supports the biotechnological potential of this Bcc strain as a source of new antimicrobials.

A Colicin M-Type Bacteriocin from Pseudomonas aeruginosa Targeting the HxuC Heme Receptor Requires a Novel Immunity Partner.

Pyocins are bacteriocins secreted by Pseudomonas aeruginosa, and they assist in the colonization of different niches. A major subset of these antibacterial proteins adopt a modular organization characteristic of polymorphic toxins. They include a receptor-binding domain, a segment enabling membrane passage, and a toxin module at the carboxy terminus, which eventually kills the target cells. To protect themselves from their own products, bacteriocin-producing strains express an immunity gene concomitantly with the bacteriocin. We show here that a pyocin equipped with a phylogenetically distinct ColM toxin domain, PaeM4, mediates antagonism against a large set of P. aeruginosa isolates. Immunity to PaeM4 is provided by the inner membrane protein PmiC, which is equipped with a transmembrane topology not previously described for the ColM family. Given that strains lacking a pmiC gene are killed by PaeM4, the presence of such an immunity partner likely is a key criterion for escaping cellular death mediated by PaeM4. The presence of a TonB box in PaeM4 and enhanced bacteriocin activity under iron-poor conditions strongly suggested the targeting of a TonB-dependent receptor. Evaluation of PaeM4 activities against TonB-dependent receptor knockout mutants in P. aeruginosa PAO1 revealed that the heme receptor HxuC (PA1302) serves as a PaeM4 target at the cellular surface. Because other ColM-type pyocins may target the ferrichrome receptor FiuA, our results illustrate the versatility in target recognition conferred by the polymorphic nature of ColM-type bacteriocins.IMPORTANCE The antimicrobial armamentarium of a bacterium is a major asset for colonizing competitive environments. Bacteriocins comprise a subset of these compounds. Pyocins are an example of such antibacterial proteins produced by Pseudomonas aeruginosa, killing other P. aeruginosa strains. A large group of these molecules show a modular protein architecture that includes a receptor-binding domain for initial target cell attachment and a killer domain. In this study, we have shown that a novel modular pyocin (PaeM4) that kills target bacteria via interference with peptidoglycan assembly takes advantage of the HxuC heme receptor. Cells can protect themselves from killing by the presence of a dedicated immunity partner, an integral inner membrane protein that adopts a transmembrane topology distinct from that of proteins currently known to provide immunity against such toxin activity. Understanding the receptors with which pyocins interact and how immunity to pyocins is achieved is a pivotal step toward the rational design of bacteriocin cocktails for the treatment of P. aeruginosa infections.

The plant growth-promoting effect of the nitrogen-fixing endophyte Pseudomonas stutzeri A15.

The use of plant growth-promoting rhizobacteria as a sustainable alternative for chemical nitrogen fertilizers has been explored for many economically important crops. For one such strain isolated from rice rhizosphere and endosphere, nitrogen-fixing Pseudomonas stutzeri A15, unequivocal evidence of the plant growth-promoting effect and the potential contribution of biological nitrogen fixation (BNF) is still lacking. In this study, we investigated the effect of P. stutzeri A15 inoculation on the growth of rice seedlings in greenhouse conditions. P. stutzeri A15 induced significant growth promotion compared to uninoculated rice seedlings. Furthermore, inoculation with strain A15 performed significantly better than chemical nitrogen fertilization, clearly pointing to the potential of this bacterium as biofertilizer. To assess the contribution of BNF to the plant growth-promoting effect, rice seedlings were also inoculated with a nitrogen fixation-deficient mutant. Our results suggest that BNF (at best) only partially contributes to the stimulation of plant growth.

Novel Conopeptides of Largely Unexplored Indo Pacific Conus sp.

Cone snails are predatory creatures using venom as a weapon for prey capture and defense. Since this venom is neurotoxic, the venom gland is considered as an enormous collection of pharmacologically interesting compounds having a broad spectrum of targets. As such, cone snail peptides represent an interesting treasure for drug development. Here, we report five novel peptides isolated from the venom of Conus longurionis, Conus asiaticus and Conus australis. Lo6/7a and Lo6/7b were retrieved from C. longurionis and have a cysteine framework VI/VII. Lo6/7b has an exceptional amino acid sequence because no similar conopeptide has been described to date (similarity percentage <50%). A third peptide, Asi3a from C. asiaticus, has a typical framework III Cys arrangement, classifying the peptide in the M-superfamily. Asi14a, another peptide of C. asiaticus, belongs to framework XIV peptides and has a unique amino acid sequence. Finally, AusB is a novel conopeptide from C. australis. The peptide has only one disulfide bond, but is structurally very different as compared to other disulfide-poor peptides. The peptides were screened on nAChRs, NaV and KV channels depending on their cysteine framework and proposed classification. No targets could be attributed to the peptides, pointing to novel functionalities. Moreover, in the quest of identifying novel pharmacological targets, the peptides were tested for antagonistic activity against a broad panel of Gram-negative and Gram-positive bacteria, as well as two yeast strains.

Biosynthetic Origin of the Antibiotic Pseudopyronines†A and B in Pseudomonas putida BW11M1.

Within the framework of our effort to discover new antibiotics from pseudomonads, pseudopyronines A and B were isolated from the plant-derived Pseudomonas putida BW11M1. Pseudopyronines are 3,6-dialkyl-4-hydroxy-2-pyrones and displayed high in vitro activities against several human pathogens, and in our hands also towards the plant pathogen Pseudomonas savastanoi. Here, the biosynthesis of pseudopyronine B was studied by a combination of feeding experiments with isotopically labeled precursors, genomic sequence analysis, and gene deletion experiments. The studies resulted in the deduction of all acetate units and revealed that the biosynthesis of these α-pyrones occurs with a single PpyS-homologous ketosynthase. It fuses, with some substrate flexibility, a 3-oxo-fatty acid and a further unbranched saturated fatty acid, both of medium chain-length and provided by primary metabolism.

Structural determinants for activity and specificity of the bacterial toxin LlpA.

Lectin-like bacteriotoxic proteins, identified in several plant-associated bacteria, are able to selectively kill closely related species, including several phytopathogens, such as Pseudomonas syringae and Xanthomonas species, but so far their mode of action remains unrevealed. The crystal structure of LlpABW, the prototype lectin-like bacteriocin from Pseudomonas putida, reveals an architecture of two monocot mannose-binding lectin (MMBL) domains and a C-terminal ß-hairpin extension. The C-terminal MMBL domain (C-domain) adopts a fold very similar to MMBL domains from plant lectins and contains a binding site for mannose and oligomannosides. Mutational analysis indicates that an intact sugar-binding pocket in this domain is crucial for bactericidal activity. The N-terminal MMBL domain (N-domain) adopts the same fold but is structurally more divergent and lacks a functional mannose-binding site. Differential activity of engineered N/C-domain chimers derived from two LlpA homologues with different killing spectra, disclosed that the N-domain determines target specificity. Apparently this bacteriocin is assembled from two structurally similar domains that evolved separately towards dedicated functions in target recognition and bacteriotoxicity.

Impact of a stereocentre inversion in cyclic lipodepsipeptides from the viscosin group: a comparative study of the viscosinamide and pseudodesmin conformation and self-assembly.

The viscosin group covers a series of cyclic lipodepsipeptides (CLPs) produced by Pseudomonas bacteria, with a range of biological functions and antimicrobial activities. Their oligopeptide moieties are composed of both L- and D-amino acids. Remarkably, the Leu5 amino acid-centrally located in the nonapeptide sequence-is the sole residue found to possess either an L or D configuration, depending on the producing strain. The impact of this D/L switch on the solution conformation was investigated by NMR-restrained molecular modelling of the epimers pseudodesmin A and viscosinamide A. Although the backbone fold remained unaffected, the D/L switch adjusted the segregation between hydrophobic and hydrophilic residues, and thus the amphipathicity. It also influenced the self-assembly capacity in organic solvents. Additionally, several new minor variants of viscosinamide A from Pseudomonas fluorescens DR54 were identified, and an NMR assay is proposed to assess the presence of either an L- or D-Leu5.

MMBL proteins: from lectin to bacteriocin.

Arguably, bacteriocins deployed in warfare among related bacteria are among the most diverse proteinacous compounds with respect to structure and mode of action. Identification of the first prokaryotic member of the so-called MMBLs (monocot mannose-binding lectins) or GNA (Galanthus nivalis agglutinin) lectin family and discovery of its genus-specific killer activity in the Gram-negative bacteria Pseudomonas and Xanthomonas has added yet another kind of toxin to this group of allelopathic molecules. This novel feature is reminiscent of the protective function, on the basis of antifungal, insecticidal, nematicidal or antiviral activity, assigned to or proposed for several of the eukaryotic MMBL proteins that are ubiquitously distributed among monocot plants, but also occur in some other plants, fish, sponges, amoebae and fungi. Direct bactericidal activity can also be effected by a C-type lectin, but this is a mammalian protein that limits mucosal colonization by Gram-positive bacteria. The presence of two divergent MMBL domains in the novel bacteriocins raises questions about task distribution between modules and the possible role of carbohydrate binding in the specificity of target strain recognition and killing. Notably, bacteriocin activity was also demonstrated for a hybrid MMBL protein with an accessory protease-like domain. This association with one or more additional modules, often with predicted peptide-hydrolysing or -binding activity, suggests that additional bacteriotoxic proteins may be found among the diverse chimaeric MMBL proteins encoded in prokaryotic genomes. A phylogenetic survey of the bacterial MMBL modules reveals a mosaic pattern of strongly diverged sequences, mainly occurring in soil-dwelling and rhizosphere bacteria, which may reflect a trans-kingdom acquisition of the ancestral genes.

LlpB represents a second subclass of lectin-like bacteriocins.

Bacteriocins are secreted bacterial proteins that selectively kill related strains. Lectin-like bacteriocins are atypical bacteriocins not requiring a cognate immunity factor and have been primarily studied in Pseudomonas. These so-called LlpAs are composed of a tandem of B-lectin domains. One domain interacts with d-rhamnose residues in the common polysaccharide antigen of Pseudomonas lipopolysaccharide (LPS). The other lectin domain is crucial for interference with the outer membrane protein assembly machinery by interacting with surface-exposed loops of its central component BamA. Via genome mining, we identified a second subclass of Pseudomonas lectin-like proteins, termed LlpB, consisting of a single B-lectin domain. We show that these proteins also display bactericidal activity. Among LlpB-resistant transposon mutants of an LlpB-susceptible Pseudomonas strain, a major subset was hit in an acyltransferase gene, predicted to be involved in LPS core modification, hereby suggesting that LlpBs equally attach to LPS for surface anchoring. This indicates that LPS binding and target strain specificity are condensed in a single B-lectin domain. The identification of this second subclass of lectin-like bacteriocins further expands the toolbox of antibacterial warfare deployed by bacteria and holds potential for their integration in biotechnological applications.

Turning Over a New Leaf: Bacteriocins Going Green.

Bacteriocins are potent antibacterial proteins that selectively kill phylogenetic relatives of the producer. Their polymorphic nature, most prominent in γ-Proteobacteria, offers potential for the design of customized bacteriocin cocktails targeting Gram-negative pathogens. As an alternative to recombinant production in bacteria, they are eligible for large-scale production in plants.

Lectin-Like Bacteriocins.

Bacteria produce a diverse array of antagonistic compounds to restrict growth of microbial rivals. Contributing to this warfare are bacteriocins: secreted antibacterial peptides, proteins and multi-protein complexes. These compounds typically eliminate competitors closely related to the producer. Lectin-like bacteriocins (LlpAs) constitute a distinct class of such proteins, produced by Pseudomonas as well as some other proteobacterial genera. LlpAs share a common architecture consisting of two B-lectin domains, followed by a short carboxy-terminal extension. Two surface-exposed moieties on susceptible Pseudomonas cells are targeted by the respective lectin modules. The carboxy-terminal domain binds D-rhamnose residues present in the lipopolysaccharide layer, whereas the amino-terminal domain interacts with a polymorphic external loop of the outer-membrane protein insertase BamA, hence determining selectivity. The absence of a toxin-immunity module as found in modular bacteriocins and other polymorphic toxin systems, hints toward a novel mode of killing initiated at the cellular surface, not requiring bacteriocin import. Despite significant progress in understanding the function of LlpAs, outstanding questions include the secretion machinery recruited by lectin-like bacteriocins for their release, as well as a better understanding of the environmental signals initiating their expression.

The ColM Family, Polymorphic Toxins Breaching the Bacterial Cell Wall.

Bacteria host an arsenal of antagonism-mediating molecules to combat for ecologic space. Bacteriocins represent a pivotal group of secreted antibacterial peptides and proteins assisting in this fight, mainly eliminating relatives. Colicin M, a model for peptidoglycan-interfering bacteriocins in Gram-negative bacteria, appears to be part of a set of polymorphic toxins equipped with such a catalytic domain (ColM) targeting lipid II. Diversifying recombination has enabled parasitism of different receptors and has also given rise to hybrid bacteriocins in which ColM is associated with another toxin module. Remarkably, ColM toxins have recruited a diverse array of immunity partners, comprising cytoplasmic membrane-associated proteins with different topologies. Together, these findings suggest that different immunity mechanisms have evolved for ColM, in contrast to bacteriocins with nuclease activities.

Hitting with a BAM: Selective Killing by Lectin-Like Bacteriocins.

Lectin-like bacteriocins (LlpAs) are secreted by proteobacteria and selectively kill strains of their own or related species, and they are composed of two B-lectin domains with divergent sequences. In Pseudomonas spp., initial binding of these antibacterial proteins to cells is mediated by the carboxy-terminal domain through d-rhamnose residues present in the common polysaccharide antigen of their lipopolysaccharide, whereas the amino-terminal domain accounts for strain selectivity of killing. Here, we show that spontaneous LlpA-resistant mutants carry mutations in one of three surface-exposed moieties of the essential ß-barrel outer membrane protein insertase BamA, the core component of the BAM complex. Polymorphism of this loop in different Pseudomonas groups is linked to LlpA susceptibility, and targeted cells all share the same signature motif in this loop. Since heterologous expression of such a bamA gene confers LlpA susceptibility upon a resistant strain, BamA represents the primary bacteriocin selectivity determinant in pseudomonads. Contrary to modular bacteriocins that require uptake via the Tol or Ton system, parasitism of BamA as an LlpA receptor advocates a novel bacteriocin killing mechanism initiated by impairment of the BAM machinery.IMPORTANCE Bacteria secrete a variety of molecules to eliminate microbial rivals. Bacteriocins are a pivotal group of peptides and proteins that assist in this fight, specifically killing related bacteria. In Gram-negative bacteria, these antibacterial proteins often comprise distinct domains for initial binding to a target cell's surface and subsequent killing via enzymatic or pore-forming activity. Here, we show that lectin-like bacteriocins, a family of bacteriocins that lack the prototypical modular toxin architecture, also stand out by parasitizing BamA, the core component of the outer membrane protein assembly machinery. A particular surface-exposed loop of BamA, critical for its function, serves as a key discriminant for cellular recognition, and polymorphisms in this loop determine whether a strain is susceptible or immune to a particular bacteriocin. These findings suggest a novel mechanism of contact-dependent killing that does not require cellular uptake. The evolutionary advantage of piracy of an essential cellular compound is highlighted by the observation that contact-dependent growth inhibition, a distinct antagonistic system, can equally take advantage of this receptor.

Community-led comparative genomic and phenotypic analysis of the aquaculture pathogen Pseudomonas baetica a390T sequenced by Ion semiconductor and Nanopore technologies.

Pseudomonas baetica strain a390T is the type strain of this recently described species and here we present its high-contiguity draft genome. To celebrate the 16th International Conference on Pseudomonas, the genome of P. baetica strain a390T was sequenced using a unique combination of Ion Torrent semiconductor and Oxford Nanopore methods as part of a collaborative community-led project. The use of high-quality Ion Torrent sequences with long Nanopore reads gave rapid, high-contiguity and -quality, 16-contig genome sequence. Whole genome phylogenetic analysis places P. baetica within the P. koreensis clade of the P. fluorescens group. Comparison of the main genomic features of P. baetica with a variety of other Pseudomonas spp. suggests that it is a highly adaptable organism, typical of the genus. This strain was originally isolated from the liver of a diseased wedge sole fish, and genotypic and phenotypic analyses show that it is tolerant to osmotic stress and to oxytetracycline.

Novel Immunity Proteins Associated with Colicin M-like Bacteriocins Exhibit Promiscuous Protection in Pseudomonas.

Bacteriocins related to colicin M, acting via cleavage of the cell wall precursor lipid II, have been characterized in γ- and ß-proteobacteria. Depending on the species, immunity is provided by either an inner membrane-anchored periplasmic protein or by an integral membrane protein. In Pseudomonas however, the immunity partner of colicin M-like bacteriocins remains unknown. Based on an in silico analysis in pseudomonad genomes, we here identify a gene encoding a putative immunity partner that represents a novel type of integral membrane protein (PmiA, Pseudomonas colicin M-like immunity type A). By heterologous expression of pmiA genes in susceptible strains, we show that immunity to colicin M-like bacteriocins is indeed provided by the cognate PmiA. Sequence homology among PmiA proteins is essentially absent, except for a short motif with a conserved periplasm-exposed aspartate residue. However, PmiA's protective function is not abolished by changing this acidic residue to the uncharged alanine. Immunity by PmiAs appears promiscuous to the extent that PmiA homologs from a clade sharing <40% pairwise amino acid identity, equally provide protection against the bacteriocin linked to the original PmiA. This study shows that multiple immunity factors have evolved independently to silence lipid II-targeting enzymatic bacteriocins. Their relaxed bacteriocin immunization capacity contrasts to the strict specificity of immunity proteins shielding the enzymatic domain of nuclease bacteriocins. The nature of associated immune functions needs consideration when using such natural protein antibiotics or designing novel variants.

A Natural Chimeric Pseudomonas Bacteriocin with Novel Pore-Forming Activity Parasitizes the Ferrichrome Transporter.

Modular bacteriocins represent a major group of secreted protein toxins with a narrow spectrum of activity, involved in interference competition between Gram-negative bacteria. These antibacterial proteins include a domain for binding to the target cell and a toxin module at the carboxy terminus. Self-inhibition of producers is provided by coexpression of linked immunity genes that transiently inhibit the toxin's activity through formation of bacteriocin-immunity complexes or by insertion in the inner membrane, depending on the type of toxin module. We demonstrate strain-specific inhibitory activity for PmnH, a Pseudomonas bacteriocin with an unprecedented dual-toxin architecture, hosting both a colicin M domain, potentially interfering with peptidoglycan synthesis, and a novel colicin N-type domain, a pore-forming module distinct from the colicin Ia-type domain in Pseudomonas aeruginosa pyocin S5. A downstream-linked gene product confers PmnH immunity upon susceptible strains. This protein, ImnH, has a transmembrane topology similar to that of Pseudomonas colicin M-like and pore-forming immunity proteins, although homology with either of these is essentially absent. The enhanced killing activity of PmnH under iron-limited growth conditions reflects parasitism of the ferrichrome-type transporter for entry into target cells, a strategy shown here to be used as well by monodomain colicin M-like bacteriocins from pseudomonads. The integration of a second type of toxin module in a bacteriocin gene could offer a competitive advantage against bacteria displaying immunity against only one of both toxic activities.IMPORTANCE In their continuous struggle for ecological space, bacteria face a huge load of contenders, including phylogenetically related strains that compete for the same niche. One important group of secreted antibacterial proteins assisting in eliminating these rivals are modular bacteriocins of Gram-negative bacteria, comprising a domain for docking onto the cell envelope of a target cell, a translocation domain enabling subsequent cellular entry, and a toxin module that kills target cells via enzymatic or pore-forming activity. We here demonstrate the antagonistic function of a Pseudomonas bacteriocin with unique architecture that combines a putative enzymatic colicin M-like domain and a novel pore-forming toxin module. For target cell recognition and entry, this bacteriocin hybrid takes advantage of the ferrichrome transporter, also parasitized by enzymatic Pseudomonas bacteriocins devoid of the pore-forming module. Bacteriocins with an expanded toxin potential may represent an inventive bacterial strategy to alleviate immunity in target cells.

Identification and functional analysis of a bacteriocin, pyocin S6, with ribonuclease activity from a Pseudomonas aeruginosa cystic fibrosis clinical isolate.

S-type pyocins are bacteriocins produced by Pseudomonas aeruginosa isolates to antagonize or kill other strains of the same species. They have a modular organization comprising a receptor-binding domain recognizing a surface constituent of the target bacterium, a domain for translocation through the periplasm, and a killing or toxic domain with DNase, tRNase, or pore-forming activity. Pyocins S2, S3, S4, and S5 recognize TonB-dependent ferri-siderophore receptors in the outer membrane. We here describe a new nuclease bacteriocin, pyocin S6, encoded in the genome of a P. aeruginosa cystic fibrosis (CF) clinical isolate, CF_PA39. Similarly to pyocins S1 and S2, the S6 toxin-immunity gene tandem was recruited to the genomic region encoding exotoxin A. The pyocin S6 receptor-binding and translocation domains are identical to those of pyocin S1, whereas the killing domain is similar to the 16S ribonuclease domain of Escherichia coli colicin E3. The cytotoxic activity was abolished in pyocin S6 forms with a mutation in the colicin E3-equivalent catalytic motif. The CF_PA39 S6 immunity gene displays a higher expression level than the gene encoding the killing protein, the latter being only detected when bacteria are grown under iron-limiting conditions. In the S1-pyocinogenic strain P. aeruginosa ATCC 25324 and pyocin S2 producer P. aeruginosa PAO1, a remnant of the pyocin S6 killing domain and an intact S6-type immunity gene are located downstream of their respective pyocin operons. Strain PAO1 is insensitive for pyocin S6, and its S6-type immunity gene provides protection against pyocin S6 activity. Purified pyocin S6 inhibits one-fifth of 110 P. aeruginosa CF clinical isolates tested, showing clearer inhibition zones when the target cells are grown under iron limitation. In this panel, about half of the CF clinical isolates were found to host the S6 genes. The pyocin S6 locus is also present in the genome of some non-CF clinical isolates.

Draft Genome Sequence of Pseudomonas putida BW11M1, a Banana Rhizosphere Isolate with a Diversified Antimicrobial Armamentarium.

In this study, we report the draft genome ofPseudomonas putidaBW11M1, a banana rhizosphere isolate producing various antimicrobial compounds, including a lectin-like bacteriocin, an R-type tailocin, the cyclic lipopeptide xantholysin, and the fatty acid-derived pseudopyronine.

Indexing the Pseudomonas specialized metabolome enabled the discovery of poaeamide B and the bananamides.

Pseudomonads are cosmopolitan microorganisms able to produce a wide array of specialized metabolites. These molecules allow Pseudomonas to scavenge nutrients, sense population density and enhance or inhibit growth of competing microorganisms. However, these valuable metabolites are typically characterized one-molecule-one-microbe at a time, instead of being inventoried in large numbers. To index and map the diversity of molecules detected from these organisms, 260 strains of ecologically diverse origins were subjected to mass-spectrometry-based molecular networking. Molecular networking not only enables dereplication of molecules, but also sheds light on their structural relationships. Moreover, it accelerates the discovery of new molecules. Here, by indexing the Pseudomonas specialized metabolome, we report the molecular-networking-based discovery of four molecules and their evolutionary relationships: a poaeamide analogue and a molecular subfamily of cyclic lipopeptides, bananamides 1, 2 and 3. Analysis of their biosynthetic gene cluster shows that it constitutes a distinct evolutionary branch of the Pseudomonas cyclic lipopeptides. Through analysis of an additional 370 extracts of wheat-associated Pseudomonas, we demonstrate how the detailed knowledge from our reference index can be efficiently propagated to annotate complex metabolomic data from other studies, akin to the way in which newly generated genomic information can be compared to data from public databases.