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Downy mildew of hops represents a serious disease affecting hops production in all growing regions. Disease management is primarily based on the application of fungicides at regular intervals based on a short-term forecasting methodology that is essential for evaluating the occurrence of theoretical infections. To enable a more reliable assessment of the pathogen’s presence in a given area, spore traps capturing airborne Pseudoperonospora humuli sporangia can be utilized. The use of quantitative real-time PCR (qRT-PCR) for the detection of sporangia collected by these traps allows for the elimination of laborious and time-consuming microscopic counting. Among four tested P. humuli-specific nuclear DNA sequences, an effective qRT-PCR detection method was developed based on the c127233.5e3 sequence. This detection approach was used for the quantification of sporangia from volumetric spore trap samples collected in situ under field conditions at three selected localities in Bohemia and Moravia during the 2021–2022 period. The obtained results were compared with the short-term forecasting method of the downy mildew (HDM) weather index (I) based on meteorological data. The overall course of the HDM weather index (I) closely correlated with the occurrence of sporangia: after reaching the maximum HDM weather index (I) value, the highest sporangium detection was observed with a time delay of 1–2 weeks at all the monitored sites. The results corresponded well with data obtained from volumetric spore traps in Germany, and the qRT-PCR method proved to be fully comparable to light microscopy. The combination of volumetric spore traps and qRT-PCR can significantly improve the precision of short-term forecasting systems for P. humuli infection, thereby enabling more efficient fungicide application programs in hops protection and contributing to a better understanding of the pathogen’s dispersal dynamics.

Background Hop ( Humulus lupulus L.) bitter acids are valuable metabolites for the brewing industry. They are biosynthesized and accumulate in glandular trichomes of the female inflorescence (hop cone). The content of alpha bitter acids, such as humulones, in hop cones can differentiate aromatic from bitter hop cultivars. These contents are subject to genetic and environmental control but significantly correlate with the number and size of glandular trichomes (lupulin glands). Results We evaluated the expression levels of 37 genes involved in bitter acid biosynthesis and morphological and developmental differentiation of glandular trichomes to identify key regulatory factors involved in bitter acid content differences. For bitter acid biosynthesis genes, upregulation of humulone synthase genes, which are important for the biosynthesis of alpha bitter acids in lupulin glands, could explain the higher accumulation of alpha bitter acids in bitter hops. Several transcription factors, including HlETC1, HlMYB61 and HlMYB5 from the MYB family, as well as HlGLABRA2, HlCYCB2–4, HlZFP8 and HlYABBY1, were also more highly expressed in the bitter hop cultivars; therefore, these factors may be important for the higher density of lupulin glands also seen in the bitter hop cultivars. Conclusions Gene expression analyses enabled us to investigate the differences between aromatic and bitter hops. This study confirmed that the bitter acid content in glandular trichomes (lupulin glands) is dependent on the last step of alpha bitter acid biosynthesis and glandular trichome density.

This study quantifies the effects of weather conditions, irrigation, and plant age on yield and alpha-acids content of Czech hop cultivars Saaz, Sládek, Premiant and Agnus in a 25-year long period, i.e., from 1993 to 2018. The yields of Czech hop cultivars were increasing for the first three years of plant age until they stabilized and then started to decline slightly for 15 to 20 years until they reached the optimal time for replanting. The highest alpha-acids content in hop cones was achieved in the first year of cultivation, followed by a logarithmic decline in the upcoming years. Rainfall was the most significant factor that positively influenced the yield of Saaz hops with correlations of r = 0.59 and 0.61 (P < 0.01) for total seasonal rainfall (April–August), 0.65 (P < 0.001) and 0.60 (P < 0.01) for daily rainfall above 3 mm, 0.37 (P < 0.05) and 0.58 (P < 0.01) for rainfall in May and 0.50 (P < 0.01) and 0.32 (P < 0.05) in July in the Saaz region and the Stekník farm, respectively. The yield of cultivars Sládek, Premiant and Agnus was not statistically influenced by the amount of precipitation, but there was a positive effect of irrigation level on yield increase with correlations 0.58 (P < 0.01), 0.55 and 0.49 (P < 0.05), respectively. High air temperatures during summer were the most significant factor that negatively influenced the alpha-acids content with the correlations ranging from –0.56 to –0.83. However, cv. Agnus showed a stable weather-independent alpha-acids content.

Satellite DNA (satDNA) is a rapidly evolving component of plant genomes, typically found in (peri)centromeric, (sub)telomeric, and other heterochromatic regions. Due to their variability and species- or population-specific distribution, satDNA serves as valuable cytogenetic markers for studying chromosomal rearrangements and karyotype evolution among closely related species. Previous studies have identified species-specific subtelomeric repeats CS-1 in , HSR1 in , and HJSR in . These satellites have been used to differentiate sex chromosomes from autosomes, however, their evolutionary origins, sequence variation and conservation pattern across related species remain largely unexplored. In this study, we analyze sequence similarity among these satellites and assess their interspecific chromosomal localization using fluorescence in situ hybridization (FISH). Our results reveal that the HSR1 and HJSR satellites are shared across all studied species, suggesting their common origin from a shared pool of satDNA in their common ancestor. In contrast, the CS-1 satellite exhibits higher sequence divergence. Although all three satellites are predominantly localized in subtelomeric regions, we identified species-specific exceptions. These findings provide new insight into the evolutionary dynamics of satDNA within the Cannabaceae family and offer further support for the divergence of species.

Verification of clonal identity of hop ( Humulus lupulus L.) cultivars within breeding programs and germplasm collections is vital to conserving genetic resources. Accurate and economic DNA-based tools are needed in dioecious hop to confirm identity and parentage, neither of which can be reliably determined from morphological observations. In this study, we developed two fingerprinting sets for hop: a 9-SSR fingerprinting set containing high-core repeats that can be run in a single PCR reaction and a kompetitive allele specific PCR (KASP) assay of 25 single nucleotide polymorphisms (SNPs). The SSR set contains a sex-linked primer pair, HI-AGA7, that was used to genotype 629 hop accessions from the US Department of Agriculture (USDA) National Clonal Germplasm Repository (NCGR), the USDA Forage Seed and Cereal Research (FSCR), and the University of Nebraska-Lincoln (UNL) collections. The SSR set identified unique genotypes except for 89 sets of synonymous samples. These synonyms included: cultivars with different designations, the same cultivars from different sources, heat-treated clones, and clonal variants. Population structure analysis clustered accessions into wild North American (WNA) and cultivated groups. Diversity was slightly higher in the cultivated samples due to larger sample size. Parentage and sib-ship analyses were used to identify true-to-type cultivars. The HI-AGA7 marker generated two male- and nine female-specific alleles among the cultivated and WNA samples. The SSR and KASP fingerprinting sets were compared in 190 samples consisting of cultivated and WNA accession for their ability to confirm identity and assess diversity and population structure. The SSR fingerprinting set distinguished cultivars, selections and WNA accessions while the KASP assays were unable to distinguish the WNA samples and had lower diversity estimates than the SSR set. Both fingerprinting sets are valuable tools for identity confirmation and parentage analysis in hop for different purposes. The 9-SSR assay is cost efficient when genotyping a small number of wild and cultivated hop samples (<96) while the KASP assay is easy to interpret and cost efficient for genotyping a large number of cultivated samples (multiples of 96).

Traditional hop (Humulus lupulus L.) cultivars have been used in the brewing industry for a long time. Globally, about ten new breeding lines were released to the market in each decade from ~1970 to 1999. Since 2006, the rate of release of new cultivars has increased tenfold. It is, therefore, important to identify their genotype and origin. Molecular genetic methods based on DNA are the most appropriate technology for this purpose. Recently, we developed an efficient marker system for the authenticity control of hop genotypes based on expressed sequence tag-simple sequence repeats (EST-SSR). In the present study, we enlarged the previously established EST-SSR set with 27 new polymorphic markers and evaluated molecular genetic variability within 135 traditional and new world hop cultivars. Two sets of 10 markers effectively differentiated all used cultivars, with the exception of cultivars derived from the same original genotype such as Saaz, Spalt, Tettnang and Nadwislawsky. Results of molecular genetic variability analyses corresponded with the genealogical and geographical origin of the key cultivars.

Viroids are small infectious pathogens, composed of a short single-stranded circular RNA. Hop (Humulus lupulus L.) plants are hosts to four viroids from the family Pospiviroidae. Hop latent viroid (HLVd) is spread worldwide in all hop-growing regions without any visible symptoms on infected hop plants. In this study, we evaluated the influence of HLVd infection on the content and the composition of secondary metabolites in maturated hop cones, together with gene expression analyses of involved biosynthesis and regulation genes for Saaz, Sládek, Premiant and Agnus cultivars. We confirmed that the contents of alpha bitter acids were significantly reduced in the range from 8.8% to 34% by viroid infection. New, we found that viroid infection significantly reduced the contents of xanthohumol in the range from 3.9% to 23.5%. In essential oils of Saaz cultivar, the contents of monoterpenes, terpene epoxides and terpene alcohols were increased, but the contents of sesquiterpenes and terpene ketones were decreased. Secondary metabolites changes were supported by gene expression analyses, except essential oils. Last-step biosynthesis enzyme genes, namely humulone synthase 1 (HS1) and 2 (HS2) for alpha bitter acids and O-methytransferase 1 (OMT1) for xanthohumol, were down-regulated by viroid infection. We found that the expression of ribosomal protein L5 (RPL5) RPL5 and the splicing of transcription factor IIIA-7ZF were affected by viroid infection and a disbalance in proteosynthesis can influence transcriptions of biosynthesis and regulatory genes involved in of secondary metabolites biosynthesis. We suppose that RPL5/TFIIIA-7ZF regulatory cascade can be involved in HLVd replication as for other viroids of the family Pospiviroidae.

The hop variety Saaz is well known over the world and is usually used for brewing of lager beers. Recently, the new related varieties Saaz Late, Brilliant, Comfort, and Shine were registered. Information about these varieties is splintered and often available only in the Czech language in regional journals. This review (i) summarizes previously published data (breeding history, genetic data, basic parameters such as yield, sensory profile, concentrations of key technologically important hop compounds), (ii) presents long-term data (2004–2021), and (iii) shows similarities/differences among these varieties. All Saaz varieties are typically fine aroma hops with a relatively low content of alpha bitter acids ranging from 5 to 7 wt%, cohumulone amounts lower than 30% rel., and hop oil content of about 1.0 wt%. Even though the new varieties have no identical chemical parameters to the original Saaz, they can substitute this established standard as well. Furthermore, the varieties Saaz Comfort and Saaz Shine show high resistance to Pseudoperonospora humuli as well as very good tolerance to drought.

Background The hop plant ( Humulus lupulus L.) is a valuable source of several secondary metabolites, such as flavonoids, bitter acids, and essential oils. These compounds are widely implicated in the beer brewing industry and are having potential biomedical applications. Several independent breeding programs around the world have been initiated to develop new cultivars with enriched lupulin and secondary metabolite contents but met with limited success due to several constraints. In the present work, a pioneering attempt has been made to overexpress master regulator binary transcription factor complex formed by Hl WRKY1 and Hl WDR1 using a plant expression vector to enhance the level of prenylflavonoid and bitter acid content in the hop. Subsequently, we performed transcriptional profiling using high-throughput RNA-Seq technology in leaves of resultant transformants and wild-type hop to gain in-depth information about the genome-wide functional changes induced by Hl WRKY1 and Hl WDR1 overexpression. Results The transgenic WW-lines exhibited an elevated expression of structural and regulatory genes involved in prenylflavonoid and bitter acid biosynthesis pathways. In addition, the comparative transcriptome analysis revealed a total of 522 transcripts involved in 30 pathways, including lipids and amino acids biosynthesis, primary carbon metabolism, phytohormone signaling and stress responses were differentially expressed in WW-transformants. It was apparent from the whole transcriptome sequencing that modulation of primary carbon metabolism and other pathways by Hl WRKY1 and Hl WDR1 overexpression resulted in enhanced substrate flux towards secondary metabolites pathway. The detailed analyses suggested that none of the pathways or genes, which have a detrimental effect on physiology, growth and development processes, were induced on a genome-wide scale in WW-transgenic lines. Conclusions Taken together, our results suggest that Hl WRKY1 and Hl WDR1 simultaneous overexpression positively regulates the prenylflavonoid and bitter acid biosynthesis pathways in the hop and thus these transgenes are presented as prospective candidates for achieving enhanced secondary metabolite content in the hop.

In vitro meristem cultures have been used for the production of hop (Humulus lupulus L.) virus-free rootstocks worldwide, because multipropagation is considered to preserve the genetic stability of the produced plantlet. Nevertheless, in vitro tissue cultures can cause genetic and epigenetic changes. Therefore, we studied the genetic and epigenetic variability of Saaz Osvald's clones, Sládek and Premiant cultivars on the DNA methylation level by methylation-sensitive amplification polymorphism (MSAP). In vitro propagated plants, acclimatised glasshouse rootstocks as well as derived mericlones and control plants under field conditions were used for the analyses. A total of 346 clearly and highly reproducible amplified products were detected in the MSAP analyses within the studied hop plants. We found 16 polymorphic products (4.6% of products) and 64 products with methylation changes (18.5% of products) in the analyses. The demethylation events were comparable to the de novo methylation events. Most demethylation changes were found in the in vitro plants, but only a few of them were found in the derived mericlones under field conditions. In contrast, the de novo methylation changes persisted in the acclimatised plants under glasshouse or field conditions. A hierarchical cluster analysis was used for the evaluation of the molecular genetic variability within the individual samples. The dendrogram showed that the individual samples of the same variety, more or less, clustered together. Because the methylation status varied during the virus-free rootstock production process, we suppose that de/methylation process is a natural tool of epigenetics and evolution in vegetatively propagated plants.