BACKGROUND: This paper brings new information about the genome and phenotypic characteristics of Pantoea agglomerans strain DBM 3797, isolated from fresh Czech hop (Humulus lupulus) in the Saaz hop-growing region. Although P. agglomerans strains are frequently isolated from different materials, there are not usually thoroughly characterized even if they have versatile metabolism and those isolated from plants may have a considerable potential for application in agriculture as a support culture for plant growth. METHODS: P. agglomerans DBM 3797 was cultured under aerobic and anaerobic conditions, its metabolites were analyzed by HPLC and it was tested for plant growth promotion abilities, such as phosphate solubilization, siderophore and indol-3-acetic acid productions. In addition, genomic DNA was extracted, sequenced and de novo assembly was performed. Further, genome annotation, pan-genome analysis and selected genome analyses, such as CRISPR arrays detection, antibiotic resistance and secondary metabolite genes identification were carried out. RESULTS AND DISCUSSION: The typical appearance characteristics of the strain include the formation of symplasmata in submerged liquid culture and the formation of pale yellow colonies on agar. The genetic information of the strain (in total 4.8 Mb) is divided between a chromosome and two plasmids. The strain lacks any CRISPR-Cas system but is equipped with four restriction-modification systems. The phenotypic analysis focused on growth under both aerobic and anaerobic conditions, as well as traits associated with plant growth promotion. At both levels (genomic and phenotypic), the production of siderophores, indoleacetic acid-derived growth promoters, gluconic acid, and enzyme activities related to the degradation of complex organic compounds were found. Extracellular gluconic acid production under aerobic conditions (up to 8 g/l) is probably the result of glucose oxidation by the membrane-bound pyrroloquinoline quinone-dependent enzyme glucose dehydrogenase. The strain has a number of properties potentially beneficial to the hop plant and its closest relatives include the strains also isolated from the aerial parts of plants, yet its safety profile needs to be addressed in follow-up research.
- Publikační typ
- časopisecké články MeSH
Staphylococcus (S.) aureus is an important causative agent of wound infections with increasing incidence in the past decades. Specifically, the emergence of methicillin-resistant S. aureus (MRSA) causes serious problems, especially in nosocomial infections. Therefore, there is an urgent need to develop of alternative or supportive antimicrobial therapeutic modalities to meet these challenges. Purified compounds from hops have previously shown promising antimicrobial effects against MRSA isolates in vitro. In this study, purified beta-acids from hops were tested for their potential antimicrobial and healing properties using a porcine model of wounds infected by MRSA. The results show highly significant antimicrobial effects of the active substance in both the powder and Ambiderman-based application forms compared to both no-treatment control and treatment with Framycoin. Moreover, the macroscopic evaluation of the wounds during the treatment using the standardized Wound Healing Continuum indicated positive effects of the beta-acids on the overall wound healing. This is further supported by the microscopic data, which showed a clear improvement of the inflammatory parameters in the wounds treated by beta-acids. Thus, using the porcine model, we demonstrate significant therapeutic effects of hops compounds in the management of wounds infected by MRSA. Beta-acids from hops, therefore, represent a suitable candidate for the treatment of non-responsive nosocomial tissue infections by MRSA.
- Publikační typ
- časopisecké články MeSH
The hop plant (Humulus lupulus L.) produces several valuable secondary metabolites, such as prenylflavonoid, bitter acids, and essential oils. These compounds are biosynthesized in glandular trichomes (lupulin glands) endowed with pharmacological properties and widely implicated in the beer brewing industry. The present study is an attempt to generate exhaustive information of transcriptome dynamics and gene regulatory mechanisms involved in biosynthesis and regulation of these compounds, developmental changes including trichome development at three development stages, namely leaf, bract, and mature lupulin glands. Using high-throughput RNA-Seq technology, a total of 61.13, 50.01, and 20.18 Mb clean reads in the leaf, bract, and lupulin gland libraries, respectively, were obtained and assembled into 43,550 unigenes. The putative functions were assigned to 30,996 transcripts (71.17%) based on basic local alignment search tool similarity searches against public sequence databases, including GO, KEGG, NR, and COG families, which indicated that genes are principally involved in fundamental cellular and molecular functions, and biosynthesis of secondary metabolites. The expression levels of all unigenes were analyzed in leaf, bract, and lupulin glands tissues of hop. The expression profile of transcript encoding enzymes of BCAA metabolism, MEP, and shikimate pathway was most up-regulated in lupulin glands compared with leaves and bracts. Similarly, the expression levels of the transcription factors and structural genes that directly encode enzymes involved in xanthohumol, bitter acids, and terpenoids biosynthesis pathway were found to be significantly enhanced in lupulin glands, suggesting that production of these metabolites increases after the leaf development. In addition, numerous genes involved in primary metabolism, lipid metabolism, photosynthesis, generation of precursor metabolites/energy, protein modification, transporter activity, and cell wall component biogenesis were differentially regulated in three developmental stages, suggesting their involvement in the dynamics of the lupulin gland development. The identification of differentially regulated trichome-related genes provided a new foundation for molecular research on trichome development and differentiation in hop. In conclusion, the reported results provide directions for future functional genomics studies for genetic engineering or molecular breeding for augmentation of secondary metabolite content in hop.
- MeSH
- flavonoidy biosyntéza chemie metabolismus MeSH
- genová ontologie MeSH
- Humulus chemie metabolismus MeSH
- listy rostlin genetika metabolismus MeSH
- propiofenony chemie metabolismus MeSH
- regulace genové exprese u rostlin MeSH
- rostlinné proteiny genetika metabolismus MeSH
- sekvenování transkriptomu MeSH
- terpeny chemie metabolismus MeSH
- transkripční faktory metabolismus MeSH
- transkriptom genetika MeSH
- trichomy genetika metabolismus ultrastruktura MeSH
- Publikační typ
- časopisecké články MeSH
Lupulin glands localized in female hop (Humulus lupulus L.) cones are valuable source of bitter acids, essential oils and polyphenols. These compounds are used in brewing industry and are important for biomedical applications. In this study we describe the potential effect of transcription factors from WRKY family in the activation of the final steps of lupulin biosynthesis. In particular, lupulin gland-specific transcription factor HlWRKY1 that shows significant similarity to AtWRKY75, has ability to activate the set of promoters driving key genes of xanthohumol and bitter acids biosynthesis such as chalcone synthase H1, valerophenone synthase, prenyltransferase 1, 1L and 2 and O-methyltransferase-1. When combined with co-factor HlWDR1 and silencing suppressor p19, HlWRKY1 is able to enhance transient expression of gus gene driven by Omt1 and Chs_H1 promoters to significant level as compared to 35S promoter of CaMV in Nicotiana. benthamiana. Transformation of hop with dual Agrobacterium vector bearing HlWRKY1/HlWDR1 led to ectopic overexpression of these transgenes and further activation of lupulin-specific genes expression in hop leaves. It was further showed that (1) HlWRKY1 is endowed with promoter autoactivation; (2) It is regulated by post-transcriptional gene silencing (PTGS) mechanism; (3) It is stimulated by kinase co-expression. Since HlWRKY1 promotes expression of lupulin-specific HlMyb3 gene therefore it can constitute a significant component in hop lupulin regulation network. Putative involvement of HlWRKY1 in the regulation of lupulin biosynthesis may suggest the original physiological function of lupulin components in hop as flower and seed protective compounds.
- MeSH
- Humulus enzymologie genetika metabolismus MeSH
- listy rostlin enzymologie genetika MeSH
- promotorové oblasti (genetika) genetika MeSH
- regulace genové exprese u rostlin * MeSH
- rostlinné proteiny genetika metabolismus MeSH
- terpeny * MeSH
- transkripční faktory genetika metabolismus MeSH
- umlčování genů fyziologie MeSH
- Publikační typ
- časopisecké články MeSH
The female flower of hop (Humulus lupulus var. lupulus) is an essential ingredient that gives characteristic aroma, bitterness and durability/stability to beer. However, the molecular genetic basis for identifying DNA markers in hop for breeding and to study its domestication has been poorly established. Here, we provide draft genomes for two hop cultivars [cv. Saazer (SZ) and cv. Shinshu Wase (SW)] and a Japanese wild hop [H. lupulus var. cordifolius; also known as Karahanasou (KR)]. Sequencing and de novo assembly of genomic DNA from heterozygous SW plants generated scaffolds with a total size of 2.05 Gb, corresponding to approximately 80% of the estimated genome size of hop (2.57 Gb). The scaffolds contained 41,228 putative protein-encoding genes. The genome sequences for SZ and KR were constructed by aligning their short sequence reads to the SW reference genome and then replacing the nucleotides at single nucleotide polymorphism (SNP) sites. De novo RNA sequencing (RNA-Seq) analysis of SW revealed the developmental regulation of genes involved in specialized metabolic processes that impact taste and flavor in beer. Application of a novel bioinformatics tool, phylogenetic comparative RNA-Seq (PCP-Seq), which is based on read depth of genomic DNAs and RNAs, enabled the identification of genes related to the biosynthesis of aromas and flavors that are enriched in SW compared to KR. Our results not only suggest the significance of historical human selection process for enhancing aroma and bitterness biosyntheses in hop cultivars, but also serve as crucial information for breeding varieties with high quality and yield.
- MeSH
- délka genomu MeSH
- dieta MeSH
- fylogeneze MeSH
- genom rostlinný * MeSH
- Humulus genetika metabolismus MeSH
- kvantitativní znak dědičný MeSH
- květy genetika MeSH
- organely genetika MeSH
- pivo * MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- regulace genové exprese u rostlin MeSH
- repetitivní sekvence nukleových kyselin genetika MeSH
- rostlinné geny MeSH
- sekvenční analýza DNA MeSH
- sekvenční analýza RNA MeSH
- stanovení celkové genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Hops represent an important source of β-acids with antimicrobial and sensory properties. Transformation products of β-acids formed during their oxidation, mainly hulupones, have been shown to have an interesting kind of bitterness. Their structures were recently elucidated using LC-TOFMS and 1D/2D NMR in solution after thermal treatment of the hop β-acids. This study demonstrates the advantages of MS detection with high resolution and accurate mass measurements. The structure of transformation products in an experimental solution of oxidized β-acids was elucidated using a newly developed method by hybrid quadrupole-Orbitrap MS. In addition to already known structures, two new ones were identified and named epoxycohulupone and epoxyhulupone. The method was verified on real samples; the profiles of these products in Sládek hops harvested in 2008 and 2012 and in corresponding beers were compared. For this purpose, a new QuEChERS assay was used for the preparation of beer samples.
- MeSH
- chuť MeSH
- Humulus chemie MeSH
- magnetická rezonanční spektroskopie metody MeSH
- molekulární struktura MeSH
- oxidace-redukce MeSH
- pivo analýza MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- terpeny analýza chemie MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Wild hops (Humulus lupulus L.) are potential new germplasms to expand the variability of genetic resources for hop breeding. We evaluated Canadian (62 plants) and Caucasian (58 plants) wild hops by their chemical characteristics and with molecular genetic analyses using sequence-tagged site and simple sequence repeat markers, in comparison with European (104 plants) and North American (27 plants) wild hops. The contents of alpha and beta acids varied from 0.36% to 5.11% and from 0.43% to 6.66% in Canadian wild hops, and from 0.85% to 3.65% and from 1.22% to 4.81% in Caucasian wild hops, respectively. The contents of cohumulone and colupulone distinctly differed between European and North American wild hops: the cohumulone level in alpha acids was in the range 46.1%-68.4% among North American wild hops and in the range 13.6%-30.6% among European wild hops. The high content of myrcene and the low contents of humulene, farnesene, and selinenes were typical for wild hops from Canada, in contrast to wild hops from the Caucasus region. We compared the chemical characteristics with molecular genetic data. Chemical characteristics differentiated wild hops into North American and Eurasian groups. Molecular genetic analysis was able to separate Caucasian wild hops from European wild hops. We proved a hop phylogeny by means of wide molecular analysis.
- MeSH
- cyklohexanony metabolismus MeSH
- fylogeneze MeSH
- Humulus klasifikace genetika metabolismus MeSH
- minisatelitní repetice genetika MeSH
- místa se sekvenční adresou MeSH
- polymorfismus genetický genetika MeSH
- systém (enzymů) cytochromů P-450 metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
- Geografické názvy
- Kanada MeSH
- Rusko MeSH
