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An abstract of the article "Charting the Clonal Variability Landscape of Pichia pastoris via Genome Sequencing: New Insights into Integration Events and their Effects," by J. P. Schwarzhans and colleagues is presented.

Frankia cluster-2 strains are diazotrophs that engage in root nodule symbiosis with actinorhizal plants of the Cucurbitales and the Rosales. Previous studies have shown that an assimilated nitrogen source, presumably arginine, is exported to the host in nodules of Datisca glomerata (Cucurbitales), while a different metabolite is exported in the nodules of Ceanothus thyrsiflorus (Rosales). To investigate if an assimilated nitrogen form is commonly exported to the host by cluster-2 strains, and which metabolite would be exported in Ceanothus, we analysed gene expression levels, metabolite profiles, and enzyme activities in nodules. We conclude that the export of assimilated nitrogen in symbiosis seems to be a common feature for Frankia cluster-2 strains, but which source is host-dependent. The export of assimilated ammonium to the host suggests that 2-oxoglutarate is drawn from the TCA cycle at a high rate. This specialised metabolism obviates the need for the reductive branch of the TCA cycle. We found several genes encoding enzymes of the central carbon and nitrogen metabolism were lacking in Frankia cluster-2 genomes: the glyoxylate shunt and succinate semialdehyde dehydrogenase. This led to a linearization of the TCA cycle, and we hypothesize this could explain the low saprotrophic potential of Frankia cluster-2., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Nanoscale zero valent iron (nZVI) is used to remediate aquifers polluted by organochlorines or heavy metals and was also suggested to eliminate harmful algal blooms. nZVI can therefore affect microorganisms in the vicinity of the application area, including microalgae. However, studies on early transcriptomic effects of microalgae after exposure to nZVI are rare. Here, we described the early physiological and transcriptomic response of the freshwater ecological indicator green microalga, Raphidocelis subcapitata ATCC 22662, to 100 mg/L of reactive nZVI and non-reactive nano-magnetite (nFe 3 O 4 ). The combined effect of shading and the release of total iron from nZVI posed a short-term inhibition effect leading to 15 % of deformed cells and cytosol leakage, while cells viability increased after 24 h. nZVI triggered a more pronounced transcriptomic response with (7380 differentially expressed genes [DEGs]) compared to nFe 3 O 4 (4601 DEGs) after 1 h. nZVI, but not nFe 3 O 4 increased the expression of genes function in DNA repair and replication, while deactivated carbohydrate-energy metabolisms, mitochondria signaling, and transmembrane ion transport. This study highlights an early fate assessment of algal cells under nZVI and nFe 3 O 4 exposure using next-generation risk assessment methods and will serve as valuable information for safe and sustainable application of nZVI in water remediation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Uptake hydrogenase (Hup) recycles H2 formed by nitrogenase during nitrogen fixation, thereby preserving energy. Among root nodule bacteria, most rhizobial strains examined are Hup-, while only one Hup-  Frankia inoculum had been identified. Previous analyses had led to the identification of two different [NiFe] hydrogenase syntons. We analysed the distribution of different types of [NiFe] hydrogenase in the genomes of different Frankia species. Our results show that Frankia strains can contain four different [NiFe] hydrogenase syntons representing groups 1f, 1h, 2a and 3b according to Søndergaard et al. (2016); no more than three types were found in any individual genome. The phylogeny of the structural proteins of groups 1f, 1h and 2a follows Frankia phylogeny; the phylogeny of the accessory proteins does not consistently. An analysis of different [NiFe] hydrogenase types in Actinomycetia shows that under the most parsimonious assumption, all four types were present in the ancestral Frankia strain. Based on Hup activities analysed and the losses of syntons in different lineages of genome reduction, we can conclude that groups 1f and 2a are involved in recycling H2 formed by nitrogenase while group 1h and group 3b are not., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Research data management (RDM) is central to the implementation of the FAIR (Findable Accessible, Interoperable, Reusable) and Open Science principles. Recognising the importance of RDM, ELIXIR Platforms and Nodes have invested in RDM and launched various projects and initiatives to ensure good data management practices for scientific excellence. These projects have resulted in a rich set of tools and resources highly valuable for FAIR data management. However, these resources remain scattered across projects and ELIXIR structures, making their dissemination and application challenging. Therefore, it becomes imminent to coordinate these efforts for sustainable and harmonised RDM practices with dedicated forums for RDM professionals to exchange knowledge and share resources. The proposed ELIXIR RDM Community will bring together RDM experts to develop ELIXIR's vision and coordinate its activities, taking advantage of the available assets. It aims to coordinate RDM best practices and illustrate how to use the existing ELIXIR RDM services. The Community will be built around three integral pillars, namely, a network of RDM professionals, RDM knowledge management and RDM training expertise and resources. It will also engage with external stakeholders to leverage benefits and provide a forum to RDM professionals for regular knowledge exchange, capacity building and development of harmonised RDM practices, keeping in line with the overall scope of the RDM Community. In the short term, the Community aims to build upon the existing resources and ensure that the content of these remain up to date and fit for purpose. In the long run, the Community will aim to strengthen the skills and knowledge of its RDM professionals to support the emerging needs of the scientific community. The Community will also devise an effective strategy to engage with other ELIXIR structures and international stakeholders to influence and align with developments and solutions in the RDM field., Competing Interests: No competing interests were disclosed., (Copyright: © 2024 D'Anna F et al.)

Tritrichomonas foetus is a parasitic protist responsible for bovine trichomonosis, a reproductive disease associated with significant economic burden to the livestock industry throughout the world. Here, we present a chromosome-level reference genome of T. foetus -KV-1 (ATCC 30924) using short-read (Illumina Miseq), long-read (Oxford Nanopore) and chromatin-linked (Hi-C) sequencing. This is the first chromosome-level genome of a parasitic protist of the order Tritrichomonadida and the second within the Parabasalia lineage, after Trichomonas vaginalis, the human-associated causative agent of the sexually transmitted infection in humans. Our constructed genome is 148 Mb in size, with a N50 length of the scaffolds of 22.9 Mb. The contigs are anchored in five super-scaffolds, corresponding to the expected five chromosomes of the species and covering 78% of the genome assembly. We predict 41,341 protein-coding genes, of which 95.10% have been functionally annotated. This high-quality genome assembly serves as a valuable reference genome for T. foetus to support future studies in functional genomics, genetic conservation and taxonomy., (© 2024. The Author(s).)

Objectives: The ascomycotic yeast-like fungus Aureobasidium exhibits the natural ability to synthesize several secondary metabolites, like polymalic acid, pullulan, or polyol lipids, with potential biotechnological applications. Combined with its polyextremotolerance, these properties make Aureobasidium a promising production host candidate. Hence, plenty of genomes of Aureobasidia have been sequenced recently. Here, we provide the annotated draft genome sequence of the polyol lipid-producing strain A. pullulans NRRL 62042., Data Description: The genome of A. pullulans NRRL 62042 was sequenced using Illumina NovaSeq 6000. Genome assembly revealed a genome size of 24.2 Mb divided into 39 scaffolds with a GC content of 50.1%. Genome annotation using Genemark v4.68 and GenDBE yielded 9,596 genes., (© 2024. The Author(s).)

Coriariaceae are a small plant family of 14-17 species and subspecies that currently have a global but disjunct distribution. All species can form root nodules in symbiosis with diazotrophic Frankia cluster-2 strains, which form the earliest divergent symbiotic clade within this bacterial genus. Studies on Frankia cluster-2 mostly have focused on strains occurring in the northern hemisphere. Except for one strain from Papua New Guinea, namely Candidatus Frankia meridionalis Cppng1, no complete genome of Frankia associated with Coriaria occurring in the southern hemisphere has been published thus far, yet the majority of the Coriariaceae species occur here. We present field sampling data of novel Frankia cluster-2 strains, representing two novel species, which are associated with Coriaria arborea and Coriaria sarmentosa in New Zealand, and with Coriaria ruscifolia in Patagonia (Argentina), in addition to identifying Ca. F. meridionalis present in New Zealand. The novel Frankia species were found to be closely related to both Ca. F. meridionalis, and a Frankia species occurring in the Philippines, Taiwan, and Japan. Our data suggest that the different Frankia cluster-2 species diverged early after becoming symbiotic circa 100 million years ago., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

By investigating wet and dry age-related ripening of beef, Pseudomonas strains V3/3/4/13 T and V3/K/3/5 T were isolated. Strain V3/3/4/13 T exhibited more than 99 % 16S rRNA gene-based similarity to Pseudomonas fragi and other members of this group, while isolate V3/K/3/5 T was very close to Pseudomonas veronii and a number of relatives within the Pseudomonas fluorescens group. Additional comparisons of complete rpoB sequences and draft genomes allowed us to place isolate V3/3/4/13 T close to Pseudomonas deceptionensis DSM 26521 T . In the case of V3/K/3/5 T the closest relative was P. veronii DSM 11331 T . Average nucleotide identity (ANIb) and digital DNA-DNA hybridization (dDDH) values calculated from the draft genomes of V3/3/4/13 T and P. deceptionensis DSM 26521 T were 88.5 and 39.8 %, respectively. For V3/K/3/5 T and its closest relative P. veronii DSM 11331 T , the ANIb value was 95.1 % and the dDDH value was 60.7 %. The DNA G+C contents of V3/3/4/13 T and V3/K/3/5 T were 57.4 and 60.8 mol%, respectively. Predominant fatty acids were C 16 : 0 , C 18 : 1  ω7 c , C 17 : 0 cyclo and summed feature C 16 : 1 ω7 ct /C 15 : 0 iso 2OH. The main respiratory quinones were Q9, with minor proportions of Q8 and, in the case of V3/K/3/5 T , additional Q10. The main polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and, in the case of V3/K/3/5 T , additional phosphatidylcholine. Based on the combined data, isolates V3/3/4/13 T and V3/K/3/5 T should be considered as representatives of two novel Pseudomonas species. The type strain of the newly proposed Pseudomonas kulmbachensis sp. nov. is V3/3/4/13 T (=DSM 113654 T =LMG 32520 T ), a second strain belonging to the same species is FLM 004-28 (=DSM 113604=LMG 32521); the type strain for the newly proposed Pseudomonas paraveronii sp. nov. is V3/K/3/5 T (=DSM 113573 T =LMG 32518 T ) with a second isolate FLM 11 (=DSM 113572=LMG 32519).

Background: Industrial by-products accrue in most agricultural or food-related production processes, but additional value chains have already been established for many of them. Crude glycerol has a 60% lower market value than commercial glucose, as large quantities are produced in the biodiesel industry, but its valorisation is still underutilized. Due to its high carbon content and the natural ability of many microorganisms to metabolise it, microbial upcycling is a suitable option for this waste product., Results: In this work, the use of crude glycerol for the production of the value-added compound itaconate is demonstrated using the smut fungus Ustilago maydis. Starting with a highly engineered strain, itaconate production from an industrial glycerol waste stream was quickly established on a small scale, and the resulting yields were already competitive with processes using commercial sugars. Adaptive laboratory evolution resulted in an evolved strain with a 72% increased growth rate on glycerol. In the subsequent development and optimisation of a fed-batch process on a 1.5-2 L scale, the use of molasses, a side stream of sugar beet processing, eliminated the need for other expensive media components such as nitrogen or vitamins for biomass growth. The optimised process was scaled up to 150 L, achieving an overall titre of 72 g L - 1 , a yield of 0.34 g g - 1 , and a productivity of 0.54 g L - 1 h - 1 ., Conclusions: Pilot-scale itaconate production from the complementary waste streams molasses and glycerol has been successfully established. In addition to achieving competitive performance indicators, the proposed dual feedstock strategy offers lower process costs and carbon footprint for the production of bio-based itaconate., (© 2024. The Author(s).)

Cnidarians thriving in biofouling communities on aquaculture net pens represent a significant health risk for farmed finfish due to their stinging cells. The toxins coming into contact with the fish, during net cleaning, can adversely affect their behavior, welfare, and survival, with a particularly serious health risk for the gills, causing direct tissue damage such as formation of thrombi and increasing risks of secondary infections. The hydroid Ectopleura larynx is one of the most common fouling organisms in Northern Europe. However, despite its significant economic, environmental, and operational impact on finfish aquaculture, biological information on this species is scarce and its venom composition has never been investigated. In this study, we generated a whole transcriptome of E. larynx, and identified its putative expressed venom toxin proteins (predicted toxin proteins, not functionally characterized) based on in silico transcriptome annotation mining and protein sequence analysis. The results uncovered a broad and diverse repertoire of putative toxin proteins for this hydroid species. Its toxic arsenal appears to include a wide and complex selection of toxin proteins, covering a large panel of potential biological functions that play important roles in envenomation. The putative toxins identified in this species, such as neurotoxins, GTPase toxins, metalloprotease toxins, ion channel impairing toxins, hemorrhagic toxins, serine protease toxins, phospholipase toxins, pore-forming toxins, and multifunction toxins may cause various major deleterious effects in prey, predators, and competitors. These results provide valuable new insights into the venom composition of cnidarians, and venomous marine organisms in general, and offer new opportunities for further research into novel and valuable bioactive molecules for medicine, agronomics and biotechnology., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Twenty oxygenated aristolochene congeners were rapidly synthesised by combining genes from four different fungal pathways in the fungal host organism Aspergillus oryzae . Compounds produced in a single step include the natural product hypoxylan A and an epimer of guignaderemophilane C. A new fungal aromatase was discovered that produces phenols by oxidative demethylation., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Strain TÜ4103 T was originally sampled from Java, Indonesia and deposited in the Tübingen strain collection under the name ' Streptomyces sp.'. The strain was found to be an antibiotic producer as strain TÜ4103 T showed bioactivity against Gram-positive bacteria, such as Bacillus subtilis and Kocuria rhizophila in bioassays. Strain TÜ4103 T showed 16S rRNA gene sequence similarity of 99.65 % to Kitasatospora cheerisanensis DSM 101999 T and 98.82 % to Kitasatospora niigatensis DSM 44781 T and Kitasatospora cineracea DSM 44780 T . Genome-based phylogenetic analysis revealed that strain TÜ4103 T is closely related to K. cineracea DSM 44780 T and K. niigatensis DSM 44781 T . The digital DNA-DNA hybridization values between the genome sequences of strain TÜ4103 T and its closest phylogenomic relatives, strains DSM 44780 T and DSM 44781 T , were 43.0 and 42.9 %, respectively. Average nucleotide identity (ANI) values support this claim, with the highest ANI score of 91.14 % between TÜ4103 T and K. niigatensis being closely followed by an ANI value of 91.10 % between K. cineracea and TÜ4103 T . The genome of TÜ4103 T has a size of 7.91 Mb with a G+C content of 74.05 mol%. Whole-cell hydrolysates of strain TÜ4103 T are rich in meso -diaminopimelic acid, and rhamnose, galactose and mannose are characteristic as whole-cell sugars. The phospholipid profile contains phosphatidylethanolamine, diphosphatidylglycerol and glycophospholipid. The predominant menaquinones (>93.5 %) are MK-9(H 8 ) and MK-9(H 6 ). Based on the phenotypic, genotypic and genomic characteristics, strain TÜ4103 T (=DSM 114396 T =CECT 30712 T ) merits recognition as the type strain of a novel species of the genus Kitasatospora , for which the name Kitasatospora fiedleri sp. nov. is proposed.

Male sex represents one of the major risk factors for severe COVID-19 outcome. However, underlying mechanisms that mediate sex-dependent disease outcome are as yet unknown. Here, we identify the CYP19A1 gene encoding for the testosterone-to-estradiol metabolizing enzyme CYP19A1 (also known as aromatase) as a host factor that contributes to worsened disease outcome in SARS-CoV-2-infected males. We analyzed exome sequencing data obtained from a human COVID-19 cohort (n = 2,866) using a machine-learning approach and identify a CYP19A1-activity-increasing mutation to be associated with the development of severe disease in men but not women. We further analyzed human autopsy-derived lungs (n = 86) and detect increased pulmonary CYP19A1 expression at the time point of death in men compared with women. In the golden hamster model, we show that SARS-CoV-2 infection causes increased CYP19A1 expression in the lung that is associated with dysregulated plasma sex hormone levels and reduced long-term pulmonary function in males but not females. Treatment of SARS-CoV-2-infected hamsters with a clinically approved CYP19A1 inhibitor (letrozole) improves impaired lung function and supports recovery of imbalanced sex hormones specifically in males. Our study identifies CYP19A1 as a contributor to sex-specific SARS-CoV-2 disease outcome in males. Furthermore, inhibition of CYP19A1 by the clinically approved drug letrozole may furnish a new therapeutic strategy for individualized patient management and treatment., Competing Interests: Declaration of interests Method for predicting the course of a viral disease. Inventors: G.G. and S.S.-B. Filing date: 04.30.2021. Pending patent applications: Europe (EP21722231.4), USA (US17995728), Japan (JP2022-566073), China (CN202180031796.5)., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Systems biology tools offer new prospects for industrial strain selection. For bacteria that are significant for industrial applications, whole-genome sequencing coupled to flux balance analysis (FBA) can help unpack the complex relationships between genome mutations and carbon trafficking. This work investigates the l-tyrosine (l-Tyr) overproducing model system Corynebacterium glutamicum ATCC 21573 with an eye to more rational and precision strain development. Using genome-wide mutational analysis of C. glutamicum , we identified 27,611 single nucleotide polymorphisms and 479 insertion/deletion mutations. Mutations in the carbon uptake machinery have led to phosphotransferase system-independent routes as corroborated with FBA. Mutations within the central carbon metabolism of C. glutamicum impaired the carbon flux, as evidenced by the lower growth rate. The entry to and flow through the tricarboxylic acid cycle was affected by mutations in pyruvate and α-ketoglutarate dehydrogenase complexes, citrate synthase, and isocitrate dehydrogenase. FBA indicated that the estimated flux through the shikimate pathway became larger as the l-Tyr production rate increased. In addition, protocatechuate export was probabilistically impossible, which could have contributed to the l-Tyr accumulation. Interestingly, aroG and cg0975 , which have received previous attention for aromatic amino acid overproduction, were not mutated. From the branch point molecule, prephenate, the change in the promoter region of pheA could be an influential contributor. In summary, we suggest that genome sequencing coupled with FBA is well poised to offer rational guidance for industrial strain development, as evidenced by these findings on carbon trafficking in C. glutamicum ATCC 21573.

An anaerobic bacterial strain, designated strain M3/9 T , was isolated from a laboratory-scale biogas fermenter fed with maize silage supplemented with 5 % wheat straw. Cells were straight, non-motile rods, which stained Gram-negative. Optimal growth occurred between 30 and 40°C, at pH 7.5-8.5, and up to 3.9 % (w/v) NaCl was tolerated. When grown on peptone from casein and soymeal, strain M3/9 T produced mainly acetic acid, ethanol, and isobutyric acid. The major cellular fatty acids of the novel strain were C 16 : 0 and C 16 : 0 DMA. The genome of strain M3/9 T is 3757  330 bp in size with a G+C content of 38.45 mol%. Phylogenetic analysis allocated strain M3/9 T within the family Lachnospiraceae with Clostridium colinum DSM 6011 T and Anaerotignum lactatifermentans DSM 14214 T being the most closely related species sharing 57.86 and 56.99% average amino acid identity and 16S rRNA gene sequence similarities of 91.58 and 91.26 %, respectively. Based on physiological, chemotaxonomic and genetic data, we propose the description of a novel species and genus Anaeropeptidivorans aminofermentans gen. nov., sp. nov., represented by the type strain M3/9 T (=DSM 100058 T =LMG 29527 T ). In addition, an emended description of Clostridium colinum is provided.

An actinobacterial strain, CMB-FB, was isolated from surface-sterilized root nodules of a Coriaria intermedia plant growing along Halsema Highway in the province of Benguet (Luzon, Philippines). The 16S rRNA gene sequence of CMB-FB showed high sequence similarity to those of the type strains of Streptomyces rishiriensis (99.4 %), Streptomyces humidus (99.1 %), Streptomyces cacaoi subsp. asoensis (99.0 %), and Streptomyces phaeofaciens (98.6 %). The major menaquinones of CMB-FB were composed of MK-9(H 4 ), MK-9(H 6 ) and MK-9(H 8 ), and there was a minor contribution of MK-9(H 10 ). The polar lipid profile consisted of phosphatidylethanolamine, unidentified aminolipids and phospholipids, a glycophospholipid and four unidentified lipids. The diagnostic diamino acid of the peptidoglycan was meso -diaminopimelic acid. The major fatty acids were iso-C 16 : 0 , anteiso-C 15 : 0 and anteiso-C 17 : 0 . The results of physiological analysis indicated that CMB-FB was mesophilic. The results of phylogenetic, genome-genome distance calculation and average nucleotide identity analysis indicated that the isolated strain represents the type strain of a novel species. On the basis of these results, strain CMB-FB (=DSM 112754 T =LMG 32457 T ) is proposed as the type strain of the novel species Streptomyces coriariae sp. nov.

Research on biogas-producing microbial communities aims at elucidation of correlations and dependencies between the anaerobic digestion (AD) process and the corresponding microbiome composition in order to optimize the performance of the process and the biogas output. Previously, Lachnospiraceae species were frequently detected in mesophilic to moderately thermophilic biogas reactors. To analyze adaptive genome features of a representative Lachnospiraceae strain, Anaeropeptidivorans aminofermentans M3/9 T was isolated from a mesophilic laboratory-scale biogas plant and its genome was sequenced and analyzed in detail. Strain M3/9 T possesses a number of genes encoding enzymes for degradation of proteins, oligo- and dipeptides. Moreover, genes encoding enzymes participating in fermentation of amino acids released from peptide hydrolysis were also identified. Based on further findings obtained from metabolic pathway reconstruction, M3/9 T was predicted to participate in acidogenesis within the AD process. To understand the genomic diversity between the biogas isolate M3/9 T and closely related Anaerotignum type strains, genome sequence comparisons were performed. M3/9 T harbors 1,693 strain-specific genes among others encoding different peptidases, a phosphotransferase system (PTS) for sugar uptake, but also proteins involved in extracellular solute binding and import, sporulation and flagellar biosynthesis. In order to determine the occurrence of M3/9 T in other environments, large-scale fragment recruitments with the M3/9 T genome as a template and publicly available metagenomes representing different environments was performed. The strain was detected in the intestine of mammals, being most abundant in goat feces, occasionally used as a substrate for biogas production., Competing Interests: IM, DW, PB and AS were employed by company Forschungszentrum Jülich GmbH. CS and BB were employed by company Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Maus, Wibberg, Belmann, Hahnke, Huang, Spröer, Bunk, Blom, Sczyrba, Pühler, Klocke and Schlüter.)

It is currently assumed that around 100 million years ago, the common ancestor to the Fabales, Fagales, Rosales and Cucurbitales in Gondwana, developed a root nodule symbiosis with a nitrogen-fixing bacterium. The symbiotic trait evolved first in Frankia cluster-2; thus, strains belonging to this cluster are the best extant representatives of this original symbiont. Most cluster-2 strains could not be cultured to date, except for Frankia coriariae, and therefore many aspects of the symbiosis are still elusive. Based on phylogenetics of cluster-2 metagenome-assembled genomes (MAGs), it has been shown that the genomes of strains originating in Eurasia are highly conserved. These MAGs are more closely related to Frankia cluster-2 in North America than to the single genome available thus far from the southern hemisphere, i.e., from Papua New Guinea.To unravel more biodiversity within Frankia cluster-2 and predict routes of dispersal from Gondwana, we sequenced and analysed the MAGs of Frankia cluster-2 from Coriaria japonica and Coriaria intermedia growing in Japan, Taiwan and the Philippines. Phylogenetic analyses indicate there is a clear split within Frankia cluster-2, separating a continental from an island lineage. Presumably, these lineages already diverged in Gondwana.Based on fossil data on the host plants, we propose that these two lineages dispersed via at least two routes. While the continental lineage reached Eurasia together with their host plants via the Indian subcontinent, the island lineage spread towards Japan with an unknown host plant., (© 2022. The Author(s).)

The gilled mushroom Clitopilus passeckerianus (Entolomataceae, Agaricales, Basidiomycota) is well known to produce the terpenoid pleuromutilin, which is the biotechnological basis for medically important antibiotics such as lefamulin and retapamulin. Their unique mode of action and good tolerance entails an increasing demand of pleuromutilin-derived antibiotics in veterinary and human health care. Surprisingly, despite their pharmaceutical importance, no genome sequence is available of any pleuromutilin-producing fungus. Here, we present the high-quality draft genome sequence of the pleuromutilin-producer C. passeckerianus DSM1602 including functional genome annotation. More precisely, we employed a hybrid assembly strategy combining Illumina sequencing and Nanopore sequencing to assemble the mitochondrial genome as well as the nuclear genome. In accordance with the dikaryotic state of the fungus, the nuclear genome has a diploid character. Interestingly, the mitochondrial genome appears duplicated. Bioinformatic analysis revealed a versatile secondary metabolism with an emphasis on terpenoid biosynthetic enzymes in C. passeckerianus and also in related strains. Two alleles of biosynthetic gene clusters for pleuromutilin were found in the genome of C. passeckerianus . The pleuromutilin genes were reassembled with yeast-specific elements for heterologous expression in Saccharomyces cerevisiae . Our work lays the foundation for metabolic strain engineering towards higher yields of the valuable compound pleuromutilin.

Pseudomonas sp. strain 1008 was isolated from the rhizosphere of field grown wheat plants at the tillering stage in an agricultural plot near Pergamino city, Argentina. Based on its in vitro phosphate solubilizing capacity and the production of IAA, strain 1008 was formulated as an inoculant for bacterization of wheat seeds and subjected to multiple field assays within the period 2010-2017. Pseudomonas sp. strain 1008 showed a robust positive impact on the grain yield (+8% on average) across a number of campaigns, soil properties, seed genotypes, and with no significant influence of the simultaneous seed treatment with a fungicide, strongly supporting the use of this biostimulant bacterium as an agricultural input for promoting the yield of wheat. Full genome sequencing revealed that strain 1008 has the capacity to access a number of sources of inorganic and organic phosphorus, to compete for iron scavenging, to produce auxin, 2,3-butanediol and acetoin, and to metabolize GABA. Additionally, the genome of strain 1008 harbors several loci related to rhizosphere competitiveness, but it is devoid of biosynthetic gene clusters for production of typical secondary metabolites of biocontrol representatives of the Pseudomonas genus. Finally, the phylogenomic, phenotypic, and chemotaxonomic comparative analysis of strain 1008 with related taxa strongly suggests that this wheat rhizospheric biostimulant isolate is a representative of a novel species within the genus Pseudomonas , for which the name Pseudomonas pergaminensis sp. nov. (type strain 1008 T = DSM 113453 T = ATCC TSD-287 T ) is proposed., Competing Interests: MD, TB, FN, and WC, were employed by Rizobacter Argentina S.A. GGA was employed by Indrasa Biotecnología S.A. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declare that this study received funding from Rizobacter Argentina S.A. The funder had the following involvement in the study: isolation and characterization of the bacterial strain; development of the commercial formulation tested in field assays; design, implementation, and monitoring of the agronomical evaluation of the formulation in field trials., (Copyright © 2022 Díaz, Bach, González Anta, Agaras, Wibberg, Noguera, Canciani and Valverde.)

Today, one of the biggest challenges in antibiotic research is a targeted prioritization of natural compound producer strains and an efficient dereplication process to avoid undesired rediscovery of already known substances. Thereby, genome sequence-driven mining strategies are often superior to wet-lab experiments because they are generally faster and less resource-intensive. In the current study, we report on the development of a novel in silico screening approach to evaluate the genetic potential of bacterial strains to produce protein synthesis inhibitors (PSI), which was termed the protein synthesis inhibitor ('psi') target gene footprinting approach = Ψ-footprinting. The strategy is based on the occurrence of protein synthesis associated self-resistance genes in genome sequences of natural compound producers. The screening approach was applied to 406 genome sequences of actinomycetes strains from the DSMZ strain collection, resulting in the prioritization of 15 potential PSI producer strains. For twelve of them, extract samples showed protein synthesis inhibitory properties in in vitro transcription/translation assays. For four strains, namely Saccharopolyspora flava DSM 44771, Micromonospora aurantiaca DSM 43813, Nocardioides albertanoniae DSM 25218, and Geodermatophilus nigrescens DSM 45408, the protein synthesis inhibitory substance amicoumacin was identified by HPLC-MS analysis, which proved the functionality of the in silico screening approach., (© The Author(s) 2022. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Ustilago maydis is an important plant pathogen that causes corn smut disease and serves as an effective biotechnological production host. The lack of a comprehensive metabolic overview hinders a full understanding of the organism's environmental adaptation and a full use of its metabolic potential. Here, we report the first genome-scale metabolic model (GSMM) of Ustilago maydis (iUma22) for the simulation of metabolic activities. iUma22 was reconstructed from sequencing and annotation using PathwayTools, and the biomass equation was derived from literature values and from the codon composition. The final model contains over 25% annotated genes (6909) in the sequenced genome. Substrate utilization was corrected by BIOLOG phenotype arrays, and exponential batch cultivations were used to test growth predictions. The growth data revealed a decrease in glucose uptake rate with rising glucose concentration. A pangenome of four different U. maydis strains highlighted missing metabolic pathways in iUma22. The new model allows for studies of metabolic adaptations to different environmental niches as well as for biotechnological applications.

Endophytic fungi have been recognized as prolific producers of chemically diverse secondary metabolites. In this work, we describe a new representative of the order Helotiales isolated from the medicinal plant Bergenia pacumbis. Several bioactive secondary metabolites were produced by this Helotiales sp. BL 73 isolate grown on rice medium, including cochlioquinones and isofusidienols. Sequencing and analysis of the approximately 59-Mb genome revealed at least 77 secondary metabolite biosynthesis gene clusters, of which several could be associated with detected compounds or linked to previously reported molecules. Four terpene synthase genes identified in the BL73 genome were codon optimized and expressed, together with farnesyl-, geranyl-, and geranylgeranyl-pyrophosphate synthases, in Streptomyces spp. An analysis of recombinant strains revealed the production of linalool and its oxidized form, terpenoids typically associated with plants, as well as a yet unidentified terpenoid. This study demonstrates the importance of a complex approach to the investigation of the biosynthetic potential of endophytic fungi using both conventional methods and genome mining. IMPORTANCE Endophytic fungi represent an as yet underexplored source of secondary metabolites, of which some may have industrial and medical applications. We isolated a slow-growing fungus belonging to the order Helotiales from the traditional medicinal plant Bergenia pacumbis and characterized its potential to biosynthesize secondary metabolites. We used cultivation of the isolate with a subsequent analysis of compounds produced, bioinformatics-based mining of the genome, and heterologous expression of several terpene synthase genes. Our study revealed that this Helotiales isolate has enormous potential to produce structurally diverse natural products, including polyketides, nonribosomally synthesized peptides, terpenoids, and ribosomally synthesized and posttranslationally modified peptides (RiPPs). Identification of meroterpenoids and xanthones, along with establishing a link between these molecules and their putative biosynthetic genes, sets the stage for investigation of the respective biosynthetic pathways. The heterologous production of terpenoids suggests that this approach can be used for the discovery of new compounds belonging to this chemical class using Streptomyces bacteria as hosts.

The family of Ustilaginaceae belongs to the order of Basidiomycetes. Despite their plant pathogenicity causing, e.g., corn smut disease, they are also known as natural producers of value-added chemicals such as extracellular glycolipids, organic acids, and polyols. Here, we present 17 high-quality draft genome sequences (N50 > 1 Mb) combining third-generation nanopore and second-generation Illumina sequencing. The data were analyzed with taxonomical genome-based bioinformatics methods such as Percentage of Conserved Proteins (POCP), Average Nucleotide Identity (ANI), and Average Amino Acid Identity (AAI) analyses indicating that a reclassification of the Ustilaginaceae family might be required. Further, conserved core genes were determined to calculate a phylogenomic core genome tree of the Ustilaginaceae that also supported the results of the other phylogenomic analysis. In addition, to genomic comparisons, secondary metabolite clusters (e.g., itaconic acid, mannosylerythritol lipids, and ustilagic acid) of biotechnological interest were analyzed, whereas the sheer number of clusters did not differ much between species.

The continuing reports of plastic pollution in various ecosystems highlight the threat posed by the ever-increasing consumption of synthetic polymers. Therefore, Pseudomonas capeferrum TDA1, a strain recently isolated from a plastic dump site, was examined further regarding its ability to degrade polyurethane (PU) compounds. The previously reported degradation pathway for 2,4-toluene diamine, a precursor and degradation intermediate of PU, could be confirmed by RNA-seq in this organism. In addition, different cell fractions of cells grown on a PU oligomer were tested for extracellular hydrolytic activity using a standard assay. Strikingly, purified outer membrane vesicles (OMV) of P. capeferrum TDA1 grown on a PU oligomer showed higher esterase activity than cell pellets. Hydrolases in the OMV fraction possibly involved in extracellular PU degradation were identified by mass spectrometry. On this basis, we propose a model for extracellular degradation of polyester-based PUs by P. capeferrum TDA1 involving the role of OMVs in synthetic polymer degradation., (© 2022. The Author(s).)

The microbial biogas network is complex and intertwined, and therefore relatively stable in its overall functionality. However, if key functional groups of microorganisms are affected by biotic or abiotic factors, the entire efficacy may be impaired. Bacteriophages are hypothesized to alter the steering process of the microbial network. In this study, an enriched fraction of virus-like particles was extracted from a mesophilic biogas reactor and sequenced on the Illumina MiSeq and Nanopore GridION sequencing platforms. Metagenome data analysis resulted in identifying 375 metagenome-assembled viral genomes (MAVGs). Two-thirds of the classified sequences were only assigned to the superkingdom Viruses and the remaining third to the family Siphoviridae , followed by Myoviridae , Podoviridae , Tectiviridae , and Inoviridae . The metavirome showed a close relationship to the phage genomes that infect members of the classes Clostridia and Bacilli . Using publicly available biogas metagenomic data, a fragment recruitment approach showed the widespread distribution of the MAVGs studied in other biogas microbiomes. In particular, phage sequences from mesophilic microbiomes were highly similar to the phage sequences of this study. Accordingly, the virus particle enrichment approach and metavirome sequencing provided additional genome sequence information for novel virome members, thus expanding the current knowledge of viral genetic diversity in biogas reactors.

Activity-based protein profiling (ABPP) has so far scarcely been applied in Archaea in general and, especially, in extremophilic organisms. We herein isolated a novel Thermococcus strain designated sp. strain 2319x1E derived from the same enrichment culture as the recently reported Thermococcus sp. strain 2319x1. Both strains are able to grow with xylan as the sole carbon and energy source, and for Thermococcus sp. strain 2319x1E (optimal growth at 85°C, pH 6-7), the induction of xylanolytic activity in the presence of xylan was demonstrated. Since the solely sequence-based identification of xylanolytic enzymes is hardly possible, we established a complementary approach by conducting comparative full proteome analysis in combination with ABPP using α- or β-glycosidase selective probes and subsequent mass spectrometry (MS)-based analysis. This complementary proteomics approach in combination with recombinant protein expression and classical enzyme characterization enabled the identification of a novel bifunctional maltose-forming α-amylase and deacetylase (EGDIFPOO_00674) belonging to the GH57 family and a promiscuous β-glycosidase (EGIDFPOO_00532) with β-xylosidase activity. We thereby further substantiated the general applicability of ABPP in archaea and expanded the ABPP repertoire for the identification of glycoside hydrolases in hyperthermophiles., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Klaus, Ninck, Albersmeier, Busche, Wibberg, Jiang, Elcheninov, Zayulina, Kaschani, Bräsen, Overkleeft, Kalinowski, Kublanov, Kaiser and Siebers.)

Yarrowia lipolytica is an oleaginous yeast that is particularly suitable for the sustainable production of secondary metabolites. The genome of this yeast is characterized by its relatively large size and its high number of different rDNA clusters located in its telomeric regions. However, due to the presence of long repetitive elements in the sub-telomeric regions, rDNA clusters and telomeres are missing in current genome assemblies of Y. lipolytica . Here, we present the near-contiguous genome sequence of the biotechnologically relevant strain DSM 3286. We employed a hybrid assembly strategy combining Illumina and nanopore sequencing reads to integrate all six rDNA clusters as well as telomeric repeats into the genome sequence. By fine-tuning of DNA isolation and library preparation protocols, we were able to create ultra-long reads that not only contained multiples of mitochondrial genomes but also shed light on the inter- and intra-chromosomal diversity of rDNA cluster types. We show that there are ten different rDNA units present in this strain that additionally appear in a predefined order in a cluster. Based on single reads, we also demonstrate that the number of rDNA repeats in a specific cluster varies from cell to cell within a population., (© The Author(s) 2021. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Rhizoctonia solani AG1-IB of the phylum Basidiomycota is known as phytopathogenic fungus affecting various economically important crops, such as bean, rice, soybean, figs, cabbage and lettuce. The isolates 1/2/21 and O8/2 of the anastomosis group AG1-IB originating from lettuce plants with bottom rot symptoms represent two less aggressive R. solani isolates, as confirmed in a pathogenicity test on lettuce. They were deeply sequenced on the Illumina MiSeq system applying the mate-pair and paired-end mode to establish their genome sequences. Assemblies of obtained sequences resulted in 2092 and 1492 scaffolds, respectively, for isolate 1/2/21 and O8/2, amounting to a size of approximately 43 Mb for each isolate. Gene prediction by applying AUGUSTUS (v. 3.2.1.) yielded 12,827 and 12,973 identified genes, respectively. Based on automatic functional annotation, genes potentially encoding cellulases and enzymes involved in secondary metabolite synthesis were identified in the AG1-IB genomes. The annotated genome sequences of the less aggressive AG1-IB isolates were compared with the isolate 7/3/14, which is highly aggressive on lettuce and other vegetable crops such as bean, cabbage and carrot. This analysis revealed the first insights into core genes of AG1-IB isolates and unique determinants of each genome that may explain the different aggressiveness levels of the strains.

This article describes some use case studies and self-assessments of FAIR status of de.NBI services to illustrate the challenges and requirements for the definition of the needs of adhering to the FAIR (findable, accessible, interoperable and reusable) data principles in a large distributed bioinformatics infrastructure. We address the challenge of heterogeneity of wet lab technologies, data, metadata, software, computational workflows and the levels of implementation and monitoring of FAIR principles within the different bioinformatics sub-disciplines joint in de.NBI. On the one hand, this broad service landscape and the excellent network of experts are a strong basis for the development of useful research data management plans. On the other hand, the large number of tools and techniques maintained by distributed teams renders FAIR compliance challenging., (© The Author(s) 2021. Published by Oxford University Press.)

By culturing microorganisms under standard laboratory conditions, most biosynthetic gene clusters (BGCs) are not expressed, and thus, the products are not produced. To explore this biosynthetic potential, we developed a novel "semi-targeted" approach focusing on activating "silent" BGCs by concurrently introducing a group of regulator genes into streptomycetes of the Tübingen strain collection. We constructed integrative plasmids containing two classes of regulatory genes under the control of the constitutive promoter ermE*p (cluster situated regulators (CSR) and Streptomyces antibiotic regulatory proteins (SARPs)). These plasmids were introduced into Streptomyces sp. TÜ17, Streptomyces sp . TÜ10 and Streptomyces sp . TÜ102. Introduction of the CSRs-plasmid into strain S. sp . TÜ17 activated the production of mayamycin A. By using the individual regulator genes, we proved that Aur1P, was responsible for the activation. In strain S. sp . TÜ102, the introduction of the SARP-plasmid triggered the production of a chartreusin-like compound. Insertion of the CSRs-plasmid into strain S. sp . TÜ10 resulted in activating the warkmycin-BGC. In both recombinants, activation of the BGCs was only possible through the simultaneous expression of aur1PR3 and griR in S. sp . TÜ102 and aur1P and pntR in of S. sp . TÜ10.

Microplastics in marine ecosystems are colonized by diverse prokaryotic and eukaryotic communities. How these communities and their functional profiles are shaped by the artificial surfaces remains broadly unknown. In order to close this knowledge gap, we set up an in situ experiment with pellets of the polyolefin polymer polyethylene (PE), the aromatic hydrocarbon polymer polystyrene (PS), and wooden beads along a coastal to estuarine gradient in the Baltic Sea, Germany. We used an integrated metagenomics/metaproteomics approach to evaluate the genomic potential as well as protein expression levels of aquatic plastic biofilms. Our results suggest that material properties had a minor influence on the plastic-associated assemblages, as genomic and proteomic profiles of communities associated with the structurally different polymers PE and PS were highly similar, hence polymer-unspecific. Instead, it seemed that these communities were shaped by biogeographic factors. Wood, on the other hand, induced the formation of substrate-specific biofilms and served as nutrient source itself. Our study indicates that, while PE and PS microplastics may be relevant in the photic zone as opportunistic colonization grounds for phototrophic microorganisms, they appear not to be subject to biodegradation or serve as vectors for pathogenic microorganisms in marine habitats., (© 2021 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

A polyphasic approach was applied to investigate the diversity of microbiota that evolved during cold storage beef ripening. Isolate V4/DAB/S4/2a T with a unique BOX-rep-PCR fingerprint profile revealed more than 99 % nucleotide identities upon pairwise comparisons of 16S rDNA sequences from the type strains Pseudomonas versuta DSM 101070 T , Pseudomonas saxonica DSM 108989 T , Pseudomonas deceptionensis DSM 26521 T and Pseudomonas weihenstephanensis DSM 29166 T , placing it within the Pseudomonas fragi / lundensis branch of the genus Pseudomonas . Additional rpoB based comparison revealed P. versuta DSM 101070 T as the nearest relative, with 98.5 % nucleotide identity. Calculation of ANIb values of the V4/DAB/S4/2a T draft genome identified P. versuta DSM 101070 T with 90.1 %, P. deceptionensis DSM 26521 T with 85.1 %, P. fragi DSM 3456 T with 84.4 %, Pseudomonas psychrophila DSM 17535 T and Pseudomonas bubulae DSM 107389 T with 84.2 % similarities each. Pairwise genome-to-genome distance calculations [digital DNA-DNA hybridization (dDDH)] resulted in values of 47.1, 35.1, 34.8, 34.2 and 34.1 %, respectively. A second isolate was detected years later in ground beef and showed ANIb values of 99.3 % and dDDH of 96.1 % relatedness to V4/DAB/S4/2a T . The DNA G+C content was 58.6 mol% for both isolates. The predominant cellular fatty acids of V4/DAB/S4/2a T were C 16 : 0 , C 18 : 1 ω7 c , C 17 : 0 cyclo and a summed feature containing C 16 : 1 ω7 c and/or C 15 : 0 iso 2-OH. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol, the major respiratory quinone was Q9, with a small portion of Q8. The combined data on genotypic and phenotypic features support the proposal of a novel species, for which the name Pseudomonas paraversuta sp. nov. is proposed. The type strain is V4/DAB/S4/2a T (=DSM 111361 T =LMG 31844 T ) and a second isolate is UBT376 (=DSM 111360=LMG 31845).

The methylotrophic thermophile Bacillus methanolicus can utilize the non-food substrate methanol as its sole carbon and energy source. Metabolism of L-lysine, in particular its biosynthesis, has been studied to some detail, and methanol-based L-lysine production has been achieved. However, little is known about L-lysine degradation, which may proceed via 5-aminovalerate (5AVA), a non-proteinogenic ω-amino acid with applications in bioplastics. The physiological role of 5AVA and related compounds in the native methylotroph was unknown. Here, we showed that B. methanolicus exhibits low tolerance to 5AVA, but not to related short-chain (C4-C6) amino acids, diamines, and dicarboxylic acids. In order to gain insight into the physiological response of B. methanolicus to 5AVA, transcriptomic analyses by differential RNA-Seq in the presence and absence of 5AVA were performed. Besides genes of the general stress response, RNA levels of genes of histidine biosynthesis, and iron acquisition were increased in the presence of 5AVA, while an Rrf2 family transcriptional regulator gene showed reduced RNA levels. In order to test if mutations can overcome growth inhibition by 5AVA, adaptive laboratory evolution (ALE) was performed and two mutants-AVA6 and AVA10-with higher tolerance to 5AVA were selected. Genome sequencing revealed mutations in genes related to iron homeostasis, including the gene for an iron siderophore-binding protein. Overexpression of this mutant gene in the wild-type (WT) strain MGA3 improved 5AVA tolerance significantly at high Fe 2+ supplementation. The combined ALE, omics, and genetics approach helped elucidate the physiological response of thermophilic B. methanolicus to 5AVA and will guide future strain development for 5AVA production from methanol., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Haupka, Brito, Busche, Wibberg and Wendisch.)

During a project focusing on the diversity of meat microbiota associated with beef ripening, a Pseudomonas strain was isolated exhibiting high 16S rRNA gene sequence similarities (>99 %) to Pseudomonas carnis DSM 107652 T , P. lactis DSM 29167 T , P. paralactis DSM 29164 T and P. azotoformans DSM 18862 T . Phylogenetic analysis of the complete rpoB gene sequences of the isolate V5/DAB/2/5 T indicated a separate branch with about 99.0 % nucleotide identities to the closest relatives P. carnis DSM 107652 T , P. lactis DSM 29167 T and P. paralactis DSM 29164 T , while average nucleotide identities (ANIb) calculated from the draft genomes were 94.8, 94.2 and 90.2 %, respectively. Pairwise genome-to-genome distance calculations (GGDC) resulted in values of 67.7, 63.5 and 45.7 %, respectively, lying below the actual species demarcation line as well. A second isolate, UBT403, was detected some years later by using matrix-assisted laser desorption ionization-time of flight MS of the microbiota of minced beef. The fatty acid profile of V5/DAB/2/5 T consisted of C 16 : 0 , summed feature C 16 : 1 ω 7 c /iso-C 15 : 0 2-OH, C 18 : 1 ω7 c, C 17 : 0 cyclo, C 12 : 0 , C 12 : 0 3-OH, C 10 : 0 3-OH and C 12 : 0 2-OH. The major cellular lipids were aminopholipids, phospholipids, phosphatidylethanolamine and phosphatidylglycerol; the major quinone was Q9 with a minor proportion of Q8. Based on phenotypic and chemotaxonomic characterizations, the isolates can be considered as representing a novel species, for which the name Pseudomonas paracarnis sp. nov. is proposed. The type strain is V5/DAB/2/5 T (=DSM 111363 T =LMG 31846 T ); a second strain is UBT403 (=DSM 111362=LMG 31847).

Bacteria occupy all major ecosystems and maintain an intensive relationship to the eukaryotes, developing together into complex biomes (i.e., phycosphere and rhizosphere). Interactions between eukaryotes and bacteria range from cooperative to competitive, with the associated microorganisms affecting their host`s development, growth and health. Since the advent of non-culture dependent analytical techniques such as metagenome sequencing, consortia have been described at the phylogenetic level but rarely functionally. Multifaceted analysis of the microbial consortium of the ancient phytoplankton Botryococcus as an attractive model food web revealed that its all abundant bacterial members belong to a niche of biotin auxotrophs, essentially depending on the microalga. In addition, hydrocarbonoclastic bacteria without vitamin auxotrophies seem adversely to affect the algal cell morphology. Synthetic rearrangement of a minimal community consisting of an alga, a mutualistic and a parasitic bacteria underpins the model of a eukaryote that maintains its own mutualistic microbial community to control its surrounding biosphere. This model of coexistence, potentially useful for defense against invaders by a eukaryotic host could represent ecologically relevant interactions that cross species boundaries. Metabolic and system reconstruction is an opportunity to unravel the relationships within the consortia and provide a blueprint for the construction of mutually beneficial synthetic ecosystems.

Acidic environments naturally occur worldwide and inappropriate agricultural management may also cause acidification of soils. Low soil pH values are an important barrier in the plant-rhizobia interaction. Acidic conditions disturb the establishment of the efficient rhizobia usually used as biofertilizer. This negative effect on the rhizobia-legume symbiosis is mainly due to the low acid tolerance of the bacteria. Here, we describe the identification of relevant factors in the acid tolerance of Rhizobium favelukesii using transcriptome sequencing. A total of 1924 genes were differentially expressed under acidic conditions, with ∼60% underexpressed. Rhizobium favelukesii acid response mainly includes changes in the energy metabolism and protein turnover, as well as a combination of mechanisms that may contribute to this phenotype, including GABA and histidine metabolism, cell envelope modifications and reverse proton efflux. We confirmed the acid-sensitive phenotype of a mutant in the braD gene, which showed higher expression under acid stress. Remarkably, 60% of the coding sequences encoded in the symbiotic plasmid were underexpressed and we evidenced that a strain cured for this plasmid featured an improved performance under acidic conditions. Hence, this work provides relevant information in the characterization of genes associated with tolerance or adaptation to acidic stress of R. favelukesii., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Mycobacterium avium ( M. a. ) subsp. paratuberculosis (MAP) is a worldwide-distributed obligate pathogen in ruminants causing Johne's disease. Due to a lack of complete subtype III genome sequences, there is not yet conclusive information about genetic differences between strains of cattle (MAP-C, type II) and sheep (MAP-S) type, and especially between MAP-S subtypes I, and III. Here we present the complete, circular genome of MAP-S/type III strain JIII-386 (DE) closed by Nanopore-technology and its comparison with MAP-S/type I closed genome of strain Telford (AUS), MAP-S/type III draft genome of strain S397 (U.S.), twelve closed MAP-C strains, and eight closed M.-a. -complex-strains. Structural comparative alignments revealed clearly the mosaic nature of MAP, emphasized differences between the subtypes and the higher diversity of MAP-S genomes. The comparison of various genomic elements including transposases and genomic islands provide new insights in MAP genomics. MAP type specific phenotypic features may be attributed to genes of known large sequence polymorphisms (LSP S s) regions I-IV and deletions #1 and #2, confirmed here, but could also result from identified frameshifts or interruptions of various virulence-associated genes (e.g., mbtC in MAP-S). Comprehensive core and pan genome analysis uncovered unique genes (e.g., cytochromes) and genes probably acquired by horizontal gene transfer in different MAP-types and subtypes, but also emphasized the highly conserved and close relationship, and the complex evolution of M.-a. -strains.

Tropolone sesquiterpenoids (TS) are an intriguing family of biologically active fungal meroterpenoids that arise through a unique intermolecular hetero Diels-Alder (hDA) reaction between humulene and tropolones. Here, we report on the combinatorial biosynthesis of a series of unprecedented analogs of the TS pycnidione 1 and xenovulene A 2. In a systematic synthetic biology driven approach, we recombined genes from three TS biosynthetic gene clusters (pycnidione 1, xenovulene A 2 and eupenifeldin 3) in the fungal host Aspergillus oryzae NSAR1. Rational design of the reconstituted pathways granted control over the number of hDA reactions taking place, the chemical nature of the fused polyketide moiety (tropolono- vs. monobenzo-pyranyl) and the degree of hydroxylation. Formation of unexpected monobenzopyranyl sesquiterpenoids was investigated using isotope-feeding studies to reveal a new and highly unusual oxidative ring contraction rearrangement., (© 2020 Wiley-VCH GmbH.)

Members of the genera Proteiniphilum and Petrimonas were speculated to represent indicators reflecting process instability within anaerobic digestion (AD) microbiomes. Therefore, Petrimonas mucosa ING2-E5A T was isolated from a biogas reactor sample and sequenced on the PacBio RSII and Illumina MiSeq sequencers. Phylogenetic classification positioned the strain ING2-E5A T in close proximity to Fermentimonas and Proteiniphilum species (family Dysgonomonadaceae). ING2-E5A T encodes a number of genes for glycosyl-hydrolyses (GH) which are organized in Polysaccharide Utilization Loci (PUL) comprising tandem sus CD-like genes for a TonB-dependent outer-membrane transporter and a cell surface glycan-binding protein. Different GHs encoded in PUL are involved in pectin degradation, reflecting a pronounced specialization of the ING2-E5A T PUL systems regarding the decomposition of this polysaccharide. Genes encoding enzymes participating in amino acids fermentation were also identified. Fragment recruitments with the ING2-E5A T genome as a template and publicly available metagenomes of AD microbiomes revealed that Petrimonas species are present in 146 out of 257 datasets supporting their importance in AD microbiomes. Metatranscriptome analyses of AD microbiomes uncovered active sugar and amino acid fermentation pathways for Petrimonas species. Likewise, screening of metaproteome datasets demonstrated expression of the Petrimonas PUL-specific component SusC providing further evidence that PUL play a central role for the lifestyle of Petrimonas species.