1. Oxytetracycline hyper-production through targeted genome reduction of Streptomyces rimosusAlen Pšeničnik, Lucija Slemc, Martina Avbelj, Miha Tome, Martin Šala, Paul R. Herron, Maksym Shmatkov, Marko Petek, Špela Baebler, Peter Mrak, Daslav Hranueli, Antonio Starcevic, Iain S. Hunter, Hrvoje Petković, 2024, izvirni znanstveni članek Povzetek: Most biosynthetic gene clusters (BGC) encoding the synthesis of important microbial secondary metabolites, such as antibiotics, are either silent or poorly expressed; therefore, to ensure a strong pipeline of novel antibiotics, there is a need to develop rapid and efficient strain development approaches. This study uses comparative genome analysis to instruct rational strain improvement, using Streptomyces rimosus, the producer of the important antibiotic oxytetracycline (OTC) as a model system. Sequencing of the genomes of two industrial strains M4018 and R6-500, developed independently from a common ancestor, identified large DNA rearrangements located at the chromosome end. We evaluated the effect of these genome deletions on the parental S. rimosus Type Strain (ATCC 10970) genome where introduction of a 145 kb deletion close to the OTC BGC in the Type Strain resulted in massive OTC overproduction, achieving titers that were equivalent to M4018 and R6-500. Transcriptome data supported the hypothesis that the reason for such an increase in OTC biosynthesis was due to enhanced transcription of the OTC BGC and not due to enhanced substrate supply. We also observed changes in the expression of other cryptic BGCs; some metabolites, undetectable in ATCC 10970, were now produced at high titers. This study demonstrated for the first time that the main force behind BGC overexpression is genome rearrangement. This new approach demonstrates great potential to activate cryptic gene clusters of yet unexplored natural products of medical and industrial value.
Ključne besede: genome reduction, antibiotic biosynthesis, oxytetracycline, cryptic metabolites Objavljeno v DiRROS: 07.08.2024; Ogledov: 83; Prenosov: 111
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2. High-quality genome sequence of the radioresistant bacterium Deinococcus ficus KS 0460Vera Y. Matrosova, Elena K. Gaidamakova, Kira S. Makarova, Olga Grichenko, Polina Klimenkova, Robert P. Volpe, Rok Tkavc, Gözen Ertem, Isabel H. Conze, Evelyne Brambilla, Nina Gunde-Cimerman, Tine Grebenc, Cene Gostinčar, 2017, izvirni znanstveni članek Povzetek: The genetic platforms of Deinococcus species remain the only systems in which massive ionizing radiation (IR)-induced genome damage can be investigated in vivo at exposures commensurate with cellular survival. We report the whole genome sequence of the extremely IR-resistant rod-shaped bacterium Deinococcus ficus KS 0460 and its phenotypic characterization. Deinococcus ficus KS 0460 has been studied since 1987, first under the name Deinobacter grandis, then Deinococcus grandis. The D. ficus KS 0460 genome consists of a 4.019 Mbp sequence (69.7% GC content and 3894 predicted genes) divided into six genome partitions, five of which are confirmed to be circular. Circularity was determined manually by mate pair linkage. Approximately 76% of the predicted proteins contained identifiable Pfam domains and 72% were assigned to COGs. Of all D. ficus KS 0460 proteins, 79% and 70% had homologues in Deinococcus radiodurans ATCC BAA-816 and Deinococcus geothermalis DSM 11300, respectively. The most striking differences between D. ficus KS 0460 and D. radiodurans BAA-816 identified by the comparison of the KEGG pathways were as follows: (i) D. ficus lacks nine enzymes of purine degradation present in D. radiodurans, and (ii) D. ficus contains eight enzymes involved in nitrogen metabolism, including nitrate and nitrite reductases, that D. radiodurans lacks. Moreover, genes previously considered to be important to IR resistance are missing in D. ficus KS 0460, namely, for the Mn-transporter nramp, and proteins DdrF, DdrJ and DdrK, all of which are also missing in Deinococcus deserti. Otherwise, D. ficus KS 0460 exemplifies the Deinococcus lineage. Ključne besede: Deinococcus-Thermus, Deinococcaceae, Deinococcus ficus, radiation-resistant, rod-shaped, phenotype characterization, genome analysis, phylogenetic analysis Objavljeno v DiRROS: 25.07.2024; Ogledov: 125; Prenosov: 108
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3. Newly isolated bacteriophages from the Podoviridae, Siphoviridae, and Myoviridae families have variable effects on putative novel Dickeya spp.Špela Alič, Tina Mikuletič, Magda Tušek-Žnidarič, Maja Ravnikar, Nejc Rački, Matjaž Peterka, Tanja Dreo, 2017, izvirni znanstveni članek Povzetek: Soft rot pathogenic bacteria from the genus Dickeya cause severe economic losses in orchid nurseries worldwide, and there is no effective control currently available. In the last decade, the genus Dickeya has undergone multiple changes as multiple new taxa have been described, and just recently a new putative Dickeya species was reported. This study reports the isolation of three bacteriophages active against putative novel Dickeya spp. isolates from commercially produced infected orchids that show variable host-range profiles. Bacteriophages were isolated through enrichment from Dickeya-infected orchid tissue. Convective interaction media monolith chromatography was used to isolate bacteriophages from wastewaters, demonstrating its suitability for the isolation of infective bacteriophages from natural sources. Based on bacteriophage morphology, all isolated bacteriophages were classified as being in the order Caudovirales, belonging to three different families, Podoviridae, Myoviridae, and Siphoviridae. The presence of three different groups of bacteriophages was confirmed by analyzing the bacteriophage specificity of bacterial hosts, restriction fragment length polymorphism and plaque morphology. Bacteriophage BF25/12, the first reported Podoviridae bacteriophage effective against Dickeya spp., was selected for further characterization. Its genome sequence determined by next-generation sequencing showed limited similarity to other characterized Podoviridae bacteriophages. Interactions among the bacteriophages and Dickeya spp. were examined using transmission electron microscopy, which revealed degradation of electron-dense granules in response to bacteriophage infection in some Dickeya strains. The temperature stability of the chosen Podoviridae bacteriophage monitored over 1 year showed a substantial decrease in the survival of bacteriophages stored at -20∘C over longer periods. It showed susceptibility to low pH and UV radiation but was stable in neutral and alkaline pH. Furthermore, the stability of the tested bacteriophage was also connected to the incubation medium and bacteriophage concentration at certain pH values. Finally, the emergence of bacteriophage-resistant bacterial colonies is highly connected to the concentration of bacteriophages in the bacterial environment. This is the first report on bacteriophages against Dickeya from the Podoviridae family to expand on potential bacteriophages to include in bacteriophage cocktails as biocontrol agents. Some of these bacteriophage isolates also showed activity against Dickeya solani, an aggressive strain that causes the soft rot of potatoes, which indicates their broad potential as biocontrol agents. Ključne besede: bacteriophages, genome sequencing, resistance development, convective interaction media monolith chromatography Objavljeno v DiRROS: 25.07.2024; Ogledov: 118; Prenosov: 109
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4. A model species for agricultural pest genomics : the genome of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)Sean D. Schoville, Yolanda H. Chen, Martin N. Andersson, Joshua B. Benoit, Anita Bhandari, Julia H. Bowsher, Kristian Brevik, Kaat Cappelle, Mei-Ju M. Chen, Anna K. Childers, Kristina Gruden, Marko Petek, 2018, izvirni znanstveni članek Povzetek: The Colorado potato beetle is one of the most challenging agricultural pests to manage. It has shown a spectacular ability to adapt to a variety of solanaceaeous plants and variable climates during its global invasion, and, notably, to rapidly evolve insecticide resistance. To examine evidence of rapid evolutionary change, and to understand the genetic basis of herbivory and insecticide resistance, we tested for structural and functional genomic changes relative to other arthropod species using genome sequencing, transcriptomics, and community annotation. Two factors that might facilitate rapid evolutionary change include transposable elements, which comprise at least 17% of the genome and are rapidly evolving compared to other Coleoptera, and high levels of nucleotide diversity in rapidly growing pest populations. Adaptations to plant feeding are evident in gene expansions and differential expression of digestive enzymes in gut tissues, as well as expansions of gustatory receptors for bitter tasting. Surprisingly, the suite of genes involved in insecticide resistance is similar to other beetles. Finally, duplications in the RNAi pathway might explain why Leptinotarsa decemlineata has high sensitivity to dsRNA. The L. decemlineata genome provides opportunities to investigate a broad range of phenotypes and to develop sustainable methods to control this widely successful pest. Ključne besede: colorado potato beetle, genome Objavljeno v DiRROS: 24.07.2024; Ogledov: 135; Prenosov: 105
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5. Fifty Aureobasidium pullulans genomes reveal a recombining polyextremotolerant generalistCene Gostinčar, Martina Turk, Janja Zajc, Nina Gunde-Cimerman, 2019, izvirni znanstveni članek Povzetek: The black yeast Aureobasidium pullulans is a textbook example of a generalistic and ubiquitous fungus thriving in a wide variety of environments. To investigate whether A. pullulans is a true generalist, or alternatively, whether part of its versatility can be attributed to intraspecific specialization masked by cryptic diversification undetectable by traditional phylogenetic analyses, we sequenced and analysed the genomes of 50 strains of A. pullulans from different habitats and geographic locations. No population structure was observed in the sequenced strains. Decay of linkage disequilibrium over shorter physical distances (<100 bp) than in many sexually reproducing fungi indicates a high level of recombination in the species. A homothallic mating locus was found in all of the sequenced genomes. Aureobasidium pullulans appears to have a homogeneous population genetics structure, which is best explained by good dispersal and high levels of recombination. This means that A. pullulans is a true generalist that can inhabit different habitats without substantial specialization to any of these habitats at the genomic level. Furthermore, in the future, the high level of A. pullulans recombination can be exploited for the identification of genomic loci that are involved in the many biotechnologically useful traits of this black yeast. Ključne besede: black yeast, genome Objavljeno v DiRROS: 24.07.2024; Ogledov: 94; Prenosov: 78
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6. MapMan4 : a refined protein classification and annotation framework applicable to multi-omics data analysisRainer Schwacke, Gabriel Y. Ponce-Soto, Kirsten Krause, Anthony M. Bolger, Borjana Arsova, Asis Hallab, Kristina Gruden, Mark Stitt, Marie E. Bolger, Björn Usadel, 2019, izvirni znanstveni članek Povzetek: Genome sequences from over 200 plant species have already been published, with this number expected to increase rapidly due to advances in sequencing technologies. Once a new genome has been assembled and the genes identified, the functional annotation of their putative translational products, proteins, using ontologies is of key importance as it places the sequencing data in a biological context. Furthermore, to keep pace with rapid production of genome sequences, this functional annotation process must be fully automated. Here we present a redesigned and significantly enhanced MapMan4 framework, together with a revised version of the associated online Mercator annotation tool. Compared with the original MapMan, the new ontology has been expanded almost threefold and enforces stricter assignment rules. This framework was then incorporated into Mercator4, which has been upgraded to reflect current knowledge across the land plant group, providing protein annotations for all embryophytes with a comparably high quality. The annotation process has been optimized to allow a plant genome to be annotated in a matter of minutes. The output results continue to be compatible with the established MapMan desktop application. Ključne besede: genome sequences, MapMan4 Objavljeno v DiRROS: 23.07.2024; Ogledov: 153; Prenosov: 140
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7. Evidence-based unification of potato gene models with the UniTato collaborative genome browserMaja Zagorščak, Jan Zrimec, Carissa Bleker, Nadja Francesca Nolte, Mojca Juteršek, Živa Ramšak, Kristina Gruden, Marko Petek, 2024, izvirni znanstveni članek Povzetek: Potato (Solanum tuberosum) is the most popular tuber crop and a model organism. A variety of gene models for potato exist, and despite frequent updates, they are not unified. This hinders the comparison of gene models across versions, limits the ability to reuse experimental data without significant re-analysis, and leads to missing or wrongly annotated genes. Here, we unify the recent potato double monoploid v4 and v6 gene models by developing an automated merging protocol, resulting in a Unified poTato genome model (UniTato). We subsequently established an Apollo genome browser (unitato.nib.si) that enables public access to UniTato and further community-based curation. We demonstrate how the UniTato resource can help resolve problems with missing or misplaced genes and can be used to update or consolidate a wider set of gene models or genome information. The automated protocol, genome annotation files, and a comprehensive translation table are provided at github.com/NIB-SI/unitato. Ključne besede: bioinformatics analysis, plant genome annotation, gene model annotations, Phureja group, GFF files, poTato genome model, UniTato Objavljeno v DiRROS: 11.06.2024; Ogledov: 171; Prenosov: 201
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8. Recovering high-quality bacterial genomes from cross-contaminated cultures : a case study of marine Vibrio campbelliiNeža Orel, Eduard Fadeev, Gerhard J. Herndl, Valentina Turk, Tinkara Tinta, 2024, izvirni znanstveni članek Povzetek: Background: Environmental monitoring of bacterial pathogens is critical for disease control in coastal marine ecosystems to maintain animal welfare and ecosystem function and to prevent significant economic losses. This requires accurate taxonomic identification of environmental bacterial pathogens, which often cannot be achieved by commonly used genetic markers (e.g., 16S rRNA gene), and an understanding of their pathogenic potential based on the information encoded in their genomes. The decreasing costs of whole genome sequencing (WGS), combined with newly developed bioinformatics tools, now make it possible to unravel the full potential of environmental pathogens, beyond traditional microbiological approaches. However, obtaining a high-quality bacterial genome, requires initial cultivation in an axenic culture, which is a bottleneck in environmental microbiology due to cross-contamination in the laboratory or isolation of non-axenic strains. Results: We applied WGS to determine the pathogenic potential of two Vibrio isolates from coastal seawater. During the analysis, we identified cross-contamination of one of the isolates and decided to use this dataset to evaluate the possibility of bioinformatic contaminant removal and recovery of bacterial genomes from a contaminated culture. Despite the contamination, using an appropriate bioinformatics workflow, we were able to obtain high quality and highly identical genomes (Average Nucleotide Identity value 99.98%) of one of the Vibrio isolates from both the axenic and the contaminated culture. Using the assembled genome, we were able to determine that this isolate belongs to a sub-lineage of Vibrio campbellii associated with several diseases in marine organisms. We also found that the genome of the isolate contains a novel Vibrio plasmid associated with bacterial defense mechanisms and horizontal gene transfer, which may offer a competitive advantage to this putative pathogen. Conclusions: Our study shows that, using state-of-the-art bioinformatics tools and a sufficient sequencing effort, it is possible to obtain high quality genomes of the bacteria of interest and perform in-depth genomic analyses even in the case of a contaminated culture. With the new isolate and its complete genome, we are providing new insights into the genomic characteristics and functional potential of this sub-lineage of V. campbellii. The approach described here also highlights the possibility of recovering complete bacterial genomes in the case of non-axenic cultures or obligatory co-cultures. Ključne besede: whole-genome assembly, non-axenic culture, plasmid, marine bacteria, marine biology Objavljeno v DiRROS: 28.03.2024; Ogledov: 397; Prenosov: 220
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9. The Carniolan honeybee from Slovenia : a complete and annotated mitochondrial genome with comparisons to closely related Apis mellifera subspeciesAjda Moškrič, Andraž Marinč, Polonca Ferk, Branimir Leskošek, Mai-Britt Mosbech, Ignas Bunikis, Olga Vinnere Pettersson, Lucile Soler, Janez Prešern, izvirni znanstveni članek Povzetek: The complete mitochondrial genome of the Carniolan honeybee (Apis mellifera carnica) from Slovenia, a homeland of this subspecies, was acquired in two contigs from WGS data and annotated. The newly obtained mitochondrial genome is a circular closed loop of 16,447 bp. It comprises 37 genes (13 protein coding genes, 22 tRNA genes, and 2 rRNA genes) and an AT-rich control region. The order of the tRNA genes resembles the order characteristic of A. mellifera. The mitogenomic sequence of A. m. carnica from Slovenia contains 44 uniquely coded sites in comparison to the closely related subspecies A. m. ligustica and to A. m. carnica from Austria. Furthermore, 24 differences were recognised in comparison between A. m. carnica and A. m. ligustica subspecies. Among them, there are three SNPs that affect translation in the nd2, nd4, and cox2 genes, respectively. The phylogenetic placement of A. m. carnica from Slovenia within C lineage deviates from the expected position and changes the perspective on relationship between C and O lineages. The results of this study represent a valuable addition to the information available in the phylogenomic studies of A. mellifera—a pollinator species of worldwide importance. Such genomic information is essential for this local subspecies’ conservation and preservation as well as its breeding and selection. Ključne besede: western honeybee, Apis mellifera carnica, mitochondrial genome, phylogeny, C lineage, annotation, whole genome sequencing, protection, conservation Objavljeno v DiRROS: 06.09.2022; Ogledov: 749; Prenosov: 244
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