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Pseudomonas lipopeptides (LPs) are involved in diverse ecological functions and have biotechnological application potential associated with their antimicrobial and/or antiproliferative activities. They are synthesized by multimodular nonribosomal peptide synthetases which, together with transport and regulatory proteins, are encoded by large biosynthetic gene clusters (BGCs). These secondary metabolites are classified in distinct families based on the sequence and length of the oligopeptide and size of the macrocycle, if present. The phylogeny of PleB, the MacB-like transporter that is part of a dedicated ATP-dependent tripartite efflux system driving export of Pseudomonas LPs, revealed a strong correlation with LP chemical diversity. As each LP BGC carries its cognate pleB , PleB is suitable as a diagnostic sequence for genome mining, allowing assignment of the putative metabolite to a particular LP family. In addition, pleB proved to be a suitable target gene for an alternative PCR method for detecting LP-producing Pseudomonas sp. and did not rely on amplification of catalytic domains of the biosynthetic enzymes. Combined with amplicon sequencing, this approach enabled typing of Pseudomonas strains as potential producers of a LP belonging to one of the known LP families, underscoring its value for strain prioritization. This finding was validated by chemical characterization of known LPs from three different families secreted by novel producers isolated from the rice or maize rhizosphere, namely, the type strains of Pseudomonas fulva (putisolvin), Pseudomonas zeae (tensin), and Pseudomonas xantholysinigenes (xantholysin). In addition, a new member of the Bananamide family, prosekin, was discovered in the type strain of Pseudomonas prosekii, which is an Antarctic isolate. IMPORTANCE Pseudomonas spp. are ubiquitous bacteria able to thrive in a wide range of ecological niches, and lipopeptides often support their lifestyle but also their interaction with other micro- and macro-organisms. Therefore, the production of lipopeptides is widespread among Pseudomonas strains. Consequently, Pseudomonas lipopeptide research not only affects chemists and microbiologists but also touches a much broader audience, including biochemists, ecologists, and plant biologists. In this study, we present a reliable transporter gene-guided approach for the detection and/or typing of Pseudomonas lipopeptide producers. Indeed, it allows us to readily assess the lipopeptide diversity among sets of Pseudomonas isolates and differentiate strains likely to produce known lipopeptides from producers of potentially novel lipopeptides. This work provides a valuable tool that can also be integrated in a genome mining strategy and adapted for the typing of other specialized metabolites.

Background and Aims: Lichens represent a symbiotic relationship between at least one fungal and one photosynthetic partner. The association between the lichen-forming fungus Mastodia tessellata (Verrucariaceae) and different species of Prasiola (Trebouxiophyceae) has an amphipolar distribution and represents a unique case study for the understanding of lichen symbiosis because of the macroalgal nature of the photobiont, the flexibility of the symbiotic interaction and the co-existence of free-living and lichenized forms in the same microenvironment. In this context, we aimed to (1) characterize the photosynthetic performance of co-occurring populations of free-living and lichenized Prasiola and (2) assess the effect of the symbiosis on water relations in Prasiola, including its tolerance of desiccation and its survival and performance under sub-zero temperatures., Methods: Photochemical responses to irradiance, desiccation and freezing temperature and pressure-volume curves of co-existing free-living and lichenized Prasiola thalli were measured in situ in Livingston Island (Maritime Antarctica). Analyses of photosynthetic pigment, glass transition and ice nucleation temperatures, surface hydrophobicity extent and molecular analyses were conducted in the laboratory., Key Results: Free-living and lichenized forms of Prasiola were identified as two different species: P. crispa and Prasiola sp., respectively. While lichenization appears to have no effect on the photochemical performance of the alga or its tolerance of desiccation (in the short term), the symbiotic lifestyle involves (1) changes in water relations, (2) a considerable decrease in the net carbon balance and (3) enhanced freezing tolerance., Conclusions: Our results support improved tolerance of sub-zero temperature as the main benefit of lichenization for the photobiont, but highlight that lichenization represents a delicate equilibrium between a mutualistic and a less reciprocal relationship. In a warmer climate scenario, the spread of the free-living Prasiola to the detriment of the lichen form would be likely, with unknown consequences for Maritime Antarctic ecosystems., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Bacterial adhesins are modular cell-surface proteins that mediate adherence to other cells, surfaces, and ligands. The Antarctic bacterium Marinomonas primoryensis uses a 1.5-MDa adhesin comprising over 130 domains to position it on ice at the top of the water column for better access to oxygen and nutrients. We have reconstructed this 0.6-μm-long adhesin using a "dissect and build" structural biology approach and have established complementary roles for its five distinct regions. Domains in region I (RI) tether the adhesin to the type I secretion machinery in the periplasm of the bacterium and pass it through the outer membrane. RII comprises ~120 identical immunoglobulin-like β-sandwich domains that rigidify on binding Ca 2+ to project the adhesion regions RIII and RIV into the medium. RIII contains ligand-binding domains that join diatoms and bacteria together in a mixed-species community on the underside of sea ice where incident light is maximal. RIV is the ice-binding domain, and the terminal RV domain contains several "repeats-in-toxin" motifs and a noncleavable signal sequence that target proteins for export via the type I secretion system. Similar structural architecture is present in the adhesins of many pathogenic bacteria and provides a guide to finding and blocking binding domains to weaken infectivity.

The large size of a 1.5-MDa ice-binding adhesin [MpAFP (Marinomonas primoryensis antifreeze protein)] from an Antarctic Gram-negative bacterium, M. primoryensis, is mainly due to its highly repetitive RII (Region II). MpAFP_RII contains roughly 120 tandem copies of an identical 104-residue repeat. We have previously determined that a single RII repeat folds as a Ca2+-dependent immunoglobulin-like domain. Here, we solved the crystal structure of RII tetra-tandemer (four tandem RII repeats) to a resolution of 1.8 Å. The RII tetra-tandemer reveals an extended (~190-Å × ~25-Å), rod-like structure with four RII-repeats aligned in series with each other. The inter-repeat regions of the RII tetra-tandemer are strengthened by Ca2+ bound to acidic residues. SAXS (small-angle X-ray scattering) profiles indicate the RII tetra-tandemer is significantly rigidified upon Ca2+ binding, and that the protein's solution structure is in excellent agreement with its crystal structure. We hypothesize that >600 Ca2+ help rigidify the chain of ~120 104-residue repeats to form a ~0.6 μm rod-like structure in order to project the ice-binding domain of MpAFP away from the bacterial cell surface. The proposed extender role of RII can help the strictly aerobic, motile bacterium bind ice in the upper reaches of the Antarctic lake where oxygen and nutrients are most abundant. Ca2+-induced rigidity of tandem Ig-like repeats in large adhesins might be a general mechanism used by bacteria to bind to their substrates and help colonize specific niches.