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Hop Latent Viroid (HLVd)-Caused Pathogenesis: Effects of HLVd Infection on Lupulin Composition of Meristem Culture-Derived Humulus lupulus
Abstract
Season-dependent fluctuation of hop latent viroid in meristem tips enabled selection of viroid-free cultures from Osvald's clone 31, 72, 114, and cv. Premiant. These mericlones were used to evaluate effect of viroid infection on the composition of hop resins and essential oils in the first production year. Healthy plants were compared with naturally re-infected individuals under field conditions. On average, viroid infection decreased content of -bitter acids by 40 %. The content of -bitter acids, as well as the ratios of humulone/cohumulone and lupulone/colupulone was not influenced by viroid infection. The content of all monoterpenes was for 29, 37.4 and 41.6 % higher for myrcene, - and -pinene, respectively, in infected plants compared with the healthy controls. The contents of sesquiterpenes like -caryophyllen -humulene, -copaene, -muurolene, -bisabolene, -cadinene, and -cadinene decreased by 13.7, 13, 14, 18.5, 29, 21.7 and 18.5 %, respectively, due to viroid infection. The possible influence of some oxidative-reduction processes activated by viroid-caused pathogenesis was assumed to be involved in the accumulation of terpenes alcohols like geraniol and methylgeranate, and in the reduction of the contents of the majority of ketones detected in the spectra of essential oils.
... HLVd is also known to affect both the oil and the terpene profiles of hops. For instance, HLVd infection increased the levels of monoterpenes such as myrcene and both αand β-pinene by 29% to 41.6% as compared to healthy plants [20]. On the contrary, HLVd infection decreased the levels of sesquiterpenes such as β-caryophyllene, α-humulene, α-copaene, γ-muurolene, β-bisabolene, γ-cadinene and δ-cadinene by 13 to 29%. ...
... On the contrary, HLVd infection decreased the levels of sesquiterpenes such as β-caryophyllene, α-humulene, α-copaene, γ-muurolene, β-bisabolene, γ-cadinene and δ-cadinene by 13 to 29%. The possible influence of some oxidative-reduction processes that are activated by the viroid-caused pathogenesis was assumed to be involved in the accumulation of terpenes alcohols such as geraniol and methylgeranate, as well as in the reduction in the levels of the majority of the ketones detected in the spectra of the essential oils [20]. However, these changes in the composition of the essential oils present in the hop cones are both genotype-dependent and cultivar-specific [21]. ...
... For instance, the linalool content in the cones of infected plants was found to be higher for cultivars Sybillla, Lubelski and Pulawski, but lower for cultivars Marynka and Magnat. The methylgeranate content was found to be lower in the infected plants from all cultivars [20]. In terms of the importance of the HLVd infection of hops with economic importance, even the slightest differences in either the content or composition in α-bitter acid, βbitter acid, oil content and in the terpene profile can change the resulting beer's aroma [10] ...
Hop latent viroid (HLVd) is the biggest concern for cannabis and hop growers worldwide. Although most HLVd-infected plants remain asymptomatic, research on hops has demonstrated a decrease in both the α-bitter acid and terpene content of hop cones, which affects their economic value. The HLVd-associated "dudding" or "duds" disease of cannabis was first reported in 2019 in California. Since then, the disease has become widespread in cannabis-growing facilities across North America. Although severe yield loss associated with duds disease has been recorded, little scientific information is available to growers in order to contain HLVd. Consequently, this review aims to summarise all of the scientific information available on HLVd so as to be able to understand the effect of HLVd on yield loss, canna-binoid content, terpene profile, disease management and inform crop protection strategies.
... Krofta and Nesvadba (2003) reported a reduction up to 25% of hop's alpha acid content due to powdery mildew infection. Patzak et al. (2001) demonstrated by biochemical analysis a significant influence on the lupulin chemical composition of cone when Hop latent viroid (HLVd) infects the plant. ...
... HLVd is generally latent on most cultivars however, in some susceptible varieties HLVd can induce chlorosis, slow growth, reduction of cones size and yield and a decrease in alpha-acids level (Barbara et al., 1990;Adams et al., 1991;Patzak et al., 2001). For this viroid an high or ubiquitous incidence is reported in the majority of the areas where it is known to occur (Pethybridge et al., 2008). ...
Over the last few decades, Italy has seen the rise
and the establishment of a new type of business in the
brewing industry: the craft beer industry, which became
one of the most significant phenomena of the
Italian agri-food sector. In spite of this, the national
production of hop is still not sufficient to satisfy the
request of Italian craft breweries. As a consequence,
Italian craft-brewers are forced to buy and use imported
hops and cannot rely on local products for their
craft beers. To face this challenge, the Italian Ministry
of Agricultural, Food and Forestry Policies funded the
Project “Luppolo.it” on hop cultivation in Italy. As 15
for all new crops, the information on Italian hop is
very incomplete and in some cases non-existent; one
of the main issues to be tackled is the lack of hopregistered
plant protection products in Italy. In order
to fill this lack of knowledge, this work reported the
first survey on the main pests and diseases inhabiting/
infesting Italian hop crops. The results of this survey
was then integrated with a bibliographic review
on the main hop pests and infections reported in the
world hop growing areas with similar pedoclimatic
conditions to those of Italy. Moreover, to compare the
survey and bibliographic researches with the actual
phytosanitary status of Italian hop cultivations, a
monitoring of plant health status with preliminary
observations on the varieties’ susceptibility to the
relevant diseases was carried out, by field inspections.
From mid-April 2017 to the harvest of the cones, surveys
in the field, inspections and sampling were carried
out for a first pest’s overview of sample areas in
Central and Northern Italy. Results of the field surveys
pointed out that there are several phytosanitary
concerns that could compromise the health of Italian
hops. Besides, by means of field inspections and samplings,
the taxonomic identification of organisms and
microorganisms and crop damage assessment were
allowed, in order to apply proper control strategies
and containment protocols. Regarding arthropods,
mites (Tetranychus urticae) and Lepidoptera (Ostrinia
nubilalis) infestations resulted of particular interest,
while for nematodes, the presence of plant parasitic
specie belonging to family Longidoridae (Xiphinema
diversicaudatum), recognized vector of Arabis mosaic
virus on hop, need further researches. The survey
revealed a wide spread presence of HLVd in all the
investigated commercial cultivars, in single or mixed
infection with ApMV and/or viruses of the genus
Carlavirus, while one of the most widespread diseases observed on monitored hop has been downy mildew
caused by the oomycete Pseudoperonospora humuli.
This study represents a first step to support the development
of an appropriate Integrated Pest
Management strategy for Italian hop productions, but
in order to be able to approach the protection strategy
correctly, it is necessary to overcome the lack of hopregistered
products in Italy.
... In addition, Saaz belongs to varieties that are very sensitive to infection (Matoušek et al., 1997). Patzak et al. (2001) reports a reduction of alpha acid content by up to 40% in infected Saaz mericlones (31,72,114). The influence of the age of the plants on the content of alpha acids ( Table 2) was also manifested in the Saaz hops in relatively young hop growths planted after 2017. ...
The course of weather conditions during the growing season, but also beyond it, decides the economics of hop cultivation every year. Ongoing climate changes manifested by more frequent occurrence of compound hot and dry waves affect hop cultivation not only in the Czech Republic, but in the entire region of Central Europe. The paper evaluates the harvest results (yields, alpha acid content) of four Czech hop varieties in the period 2019–2022 with a focus on the weather diametrically opposed vintages 2021 and 2022 on a specific agricultural farm in the Žatec (Saaz) hop-growing region. While the 2021 vintage was rich in precipitation in the decisive vegetation phase, the 2022 vintage was very dry and hot. The age of the plants significantly influenced the alpha acid content of the varieties Saaz, Sládek and Premiant independently of the vintage. Young growths of the Saaz variety contained at least 60% rel. more alpha acids than older growths (more than 5 years old), while the increase in the Sládek variety was 30–40% rel. and in the Premiant variety 20–25% rel. In the case of the variety Agnus, the age of the plants did not have a demonstrable effect on the content of alpha acids. In 2021 the yields of all varieties were negatively affected by the calamitous occurrence of downy mildew. In 2022, the rainfall deficit was so large that it could not be fully compensated even by the maximum use of drip irrigation. Of the evaluated varieties, the Agnus variety coped best with the different weather conditions of the assessed vintages. This shows that the cultivation of climate resistant varieties is one of the promising but time-consuming ways to face the expected climate changes. Other options include regular renewal of hop growths, consistent regionalization, i.e. planting hop in locations that are suitable for the given variety and efficient use of irrigation systems.
... The viroid is widely distributed in hop gardens in many countries, including the USA (Nelson et al., 1997;Eastwell and Nelson, 2007), and the impact of the viroid on hop yield is relatively small on most cultivars, especially in comparison to other hop-infecting viroids. Nevertheless, the viroid can cause a serious reduction in cone yield (Barbara et al., 1990) in certain cultivars and alter the content of secondary metabolites (e.g., αand β-acids) associated with flavor (Barbara et al., 1990;Patzak et al., 2001). It is noteworthy that HLVd has recently been reported in Cannabis sativa, which, like hop, belongs to the Cannabaceae family, exhibiting various symptoms such as stunting, leaf malformation/ chlorosis, increased stem brittleness, and yield reduction in California (Bektaş et al., 2019;Warren et al., 2019). ...
Viroids are highly structured, small circular RNAs that do not encode proteins but can replicate and cause diseases in plants. Viroids were not discovered until the beginning of 1970 even though several major diseases including the spindle tuber disease in potatoes, sunblotch disease in avocados, and scar skin in apples, had been known for decades prior. In this chapter, viroids found in North America to date are reviewed according to their occurrence in host species. Also discussed are viroids’ geographic distribution, economic significance, and regulatory status in North American Plant Protection Organization countries.
... If present in any planting materials, the viroid could be transmitted during transplanting on equipment and, once established in a field, transmission could occur through leaf-to-leaf contact among plants or when weeding and rogueing activities occur (Pethybridge et al. 2008). In hops, this viroid can deleteriously impact the production of important organic acids and oils for brewing (Patzak et al. 2001), but it is unknown how HLVd influences acid and oil production in hemp. ...
Hemp (Cannabis sativa) acreage in Oregon has increased by approximately 240 times in the last five years and a greater number of hemp diseases have been observed. This special report documents pathogens, particularly those causing virus and virus-like diseases, that have been detected from field and greenhouse-grown hemp crops in Oregon, based on plant samples submitted to the Hermiston Agricultural Research and Extension Center Plant Clinic of Oregon State University in 2019 and 2020. Symptoms and signs were used to evaluate disease types and determine diagnostic assays used on each submission. Plants with signs or symptoms of fungal or oomycete infection were cultured to isolate pathogenic organisms and plants with symptoms suspected to be caused by virus infection were assayed for the presence of Beet curly top virus (BCTV), viroids, and phytoplasmas using polymerase chain reaction (PCR), or reverse transcription (RT)-PCR. Diseases with fungal or oomycete, and virus causes accounted for 26.5%, and 42.9% of submissions, respectively; co-infection of viral and fungal or oomycete pathogens were detected from 6.1% of submissions between 2019 and 2020. BCTV, a curtovirus, and hop latent viroid (HLVd) were the predominant pathogens detected from field and indoor grown hemp. Worland-like strains of BCTV represented 93% of all curtovirus detections. Eighty percent of HLVd detections occurred from plants that originated from indoor growing facilities. Based on BCTV vector, beet leafhopper, prevalence, field-grown hemp in western production regions may be affected by curly top and increasing hemp acreage in the landscape may have potential implications on other crops affected by curtoviruses. Virus and virus-like diseases could be a limiting factor for hemp production in some regions of the United States.
... According to our recent results, all these hop viroids are compatible and can infect hop with variable stability under experimental conditions ). The HLVd is present in all world hop production areas and does not produce any visible symptoms on most varieties, although it has a negative impact on the yield and quality of the crop (Adams et al. 1991; Barbara et al. 1990;Eastwell and Nelson 2007;Patzak et al. 2001;Pethybridge et al. 2008). The host range of HLVd has been limited to hop and stinging nettle (Urtica dioica L.), and the viroid has a relatively conserved sequence with a small number of sequence variants (Eastwell and Nelson 2007;Pethybridge et al. 2008;Puchta et al. 1988). ...
Hop (Humulus lupulus L.) plants are hosts to several viroids; some of them can be highly aggressive, and their infection can manifest as complete plant dieback. Since molecular detection of multiple viroids can be time consuming and cost inefficient, a reliable one-step multiplex RT-PCR (mRT-PCR) was developed to detect simultaneously all four viroids infecting hops: Hop latent viroid (HLVd), Hop stunt viroid (HSVd), Apple fruit crinkle viroid (AFCVd) and Citrus bark cracking viroid (CBCVd). Several primer pairs were tested on different viroid variants from hops, citruses and grapevines, and from among them, specific primer pairs for detection of hop viroids were selected and confirmed in a single-tube assay. To improve mRT-PCR reliability and validate its effectiveness, nad5 and DRH1 genes were included as an internal control. The specificities of single and mRT-PCR assays for all four viroids were comparable. The sensitivity of mRT-PCR was compared with that of dot-blot hybridization and single RT-PCR assay on biolistically infected hop plants. The results show mRT-PCR to be more sensitive than the dot-blot and slightly less sensitive than the single RT-PCR assay. Furthermore, mRT-PCR was validated using field samples and a group of 135 hop plants, which are used in the certification scheme for planting material propagation, and the method proved to be robust, rapid and simple. Additionally, this approach can be applicable to similar methods of systematic surveys of emerging diseases and epidemiological studies.
Hemp ( Cannabis sativa L.) is an emerging industrial crop in the United States. In recent surveys of hemp growers, disease identification and management were determined to be significant priority areas for future research. From previous studies on disease identification in Western U.S. hemp production systems, hop latent viroid (HLVd), beet curly top virus (BCTV), Cannabis sativa mitovirus 1 (CasaMV1), and citrus yellow vein associated virus (CYVaV) were detected at high incidences. In the present study, a multiplex reverse transcription PCR protocol was developed to detect this viroid and viruses from hemp leaf samples simultaneously. Using the developed protocol, the multiplex reverse transcription PCR could detect each virus and viroid specifically in 10 ⁻¹ -diluted cDNA. The protocol was validated on 6 hemp metavirome leaf samples from field-grown hemp grown in 2019 and 23 uncharacterized leaf samples in 2021 from Colorado. Of the samples tested, 95% tested positive for CasaMV1 in 2021. This was greater than infections of BCTV (4%), HLVd (0%), and CYVaV (0%). Coinfections accounted for 4% of samples in 2021. This method offers sensitive, specific, and fast simultaneous detection of three viruses and one viroid from hemp, contributing to large-scale pathogen-free hemp certification schemes.
[Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .
Hop latent viroid (HLVd), a subviral pathogen from the family Pospiviroidae, is a major threat to the global cannabis industry and is the causative agent for “dudding disease”. Infected plants can often be asymptomatic for a period of growth and then develop symptoms such as malformed and yellowing leaves, as well as stunted growth. During flowering, HLVd-infected plants show reduced levels of valuable metabolites. This study was undertaken to expand our basic knowledge of HLVd infectivity, transmission, and host range. HLVd-specific primers were used for RT-PCR detection in plant samples and were able to detect HLVd in as little as 5 picograms of total RNA. A survey of hemp samples obtained from a diseased production system proved sole infection of HLVd (72%) with no coexistence of hop stunt viroid. HLVd was infectious through successive passage assays using a crude sap or total RNA extract derived from infected hemp. HLVd was also highly transmissible through hemp seeds at rates of 58 to 80%. Host range assays revealed new hosts for HLVd: tomato, cucumber, chrysanthemum, Nicotiana benthamiana, and Arabidopsis thaliana (Col-0). Sequence analysis of 77 isolates revealed only 3 parsimony-informative sites, while 10 sites were detected among all HLVd isolates available in the GenBank. The phylogenetic relationship among HLVd isolates allowed for inferring two major clades based on the genetic distance. Our findings facilitate further studies on host–viroid interaction and viroid management.
The female inflorescence of hops is traditionally used in the brewing industry to impart bitterness, aroma, flavour and longer shelf life to beer, where the bittering acids are the main compounds that enhance the quality of the beer. In recent years, a wide variety of bioactive compounds have been reported to be present in female flowers. Due to the growing consumer interests in natural ingredients, a great deal of research has been carried out in the last few years to find new sources of functional biomolecules. This paper reviews the recent applications of hops in the food, pharmaceutical and cosmetic industries and provides an overview of the bioactive potential of hops. In addition, the analytical techniques of hops compound are reviewed. The biological activities of hops compound such as antibacterial, antifungal, cardioprotective, antioxidant, anti‐inflammatory and anticancer are reviewed and the recent effects of hop bittering derivatives on beer‐related microbial activity are also presented. It is thus clear that hops have a high potential as bioactive components for use as preservatives in fresh foods to extend shelf life and as cosmetic formulations for skin care products as well as nutraceutical and health implications. The female inflorescence of hops is recognised in the brewing industry as a bittering agent and is traditionally used in the brewing industry to impart bitterness, aroma, flavour and longer shelf life to beer, where the bittering acid is the main compound that en‐hances the quality of the beer. The biological activities of hops extracts such as antibacterial, antifungal, cardioprotective, antioxidant, anti‐inflammatory and anticancer are reviewed and the recent effects of hops and their derivatives on beer‐related microbial activity are presented.
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.
Methods of hop propagation by means of isolated top meristems and clonal propagation were prepared and tested in conditions in vitro. To cultivate top meristems and subsequent organogenesis accompanied with formation of multiple stems, a medium (Murashige, Skoog, 1962) was applied and tested. The medium was supplied with 10 μM per Iitre of BAP, 1 μM per litre of lBA, 1 μM per Iitre of GA3, 40 g per Iitre of glucose, and 0.8% agar (pH of medium was modified to 5.7-5.8). The medium containing a half of all components of the given medium with no growth regulators was found to be the best to root of the removed formed stems and to subsequent clonal propagation. Formed stems in vitro could be removed, propagated, rooted and subsequently planted in to nonsterile conditions and cultivated in the glasshouse for the whole year.
Analysis of hop viroids in the most important commercial hop clones confirmed a high incidence (approaching 100%) of hop latent viroid (HLVd). Hop stunt viroid (HSVd) was not found in these Bohemian hops. The mean value of HLVd amount in leaf tissues of hop garden-grown hop plants was 4.3 and 2.4 pg/mg fresh mass for variety Osvald's clones 31 and72, respectively. Electrophoretic analysis of HLVd showed homogeneityof isolated HLVd samples and did not reveal any differences between HLVd isolated from different hop clones. Plants derived from meristem cultures (mericlones) of variety Osvald's clone 72, which were includedin the programme of virus eradication and subjected to thermotherapy,were found to be virus and HLVd-free. On the other hand, HLVd concentration in ten newly established mericlones grown in in vitro conditions was about eightfold higher than in field-grown plants of the original variety Osvald's clone 31 and reached 31.1 pg/mg fresh weight on average. While a gradient of viroid concentration was observed in field-grown plants, and the highest viroid level was detected in the lower leaves and roots, no such gradient was observed for in vitro mericlones. In these plants a higher concentration was found in shoots than in roots
The resin and essential oil contents of hops (Humulus lupulus L., var., Pride of Ringwood) together with cone weight were studied over a period of 4 weeks prior to and 1 week subsequent to the estimated time of normal harvesting. The petrol ether- and methanol-soluble fractions of the hop cone showed no significant change during this period. Essential oil and α-acid contents increased gradually over the sampling period, whilst cone weight increased up to the estimated time of normal harvesting and decreased thereafter. The results indicate that, with the particular variety studied, harvesting could have been conducted over a 4-week period without significantly affecting α-acid or hop cone yield. The advantages of harvesting high-yielding hops over a 4-week period rather than planting an earlier maturing but lower-yielding variety are discussed.
A viroid-like RNA was detected in low molecular weight RNAs extracted from vilus-free hops by two-dimensional polyacrylamide gel electrophoresis (PAGE). It migrated considerably faster than hop stunt viroid in 5% PAGE containing 8M urea under a denaturing condition. A cDNA fragment was amplified from the low molecular weight RNAs containing the viroid-like RNA by reverse transcription and polymerase chain reaction (RT-PCR) using a pair of hop latent viroid (HLVd) specific primers. The amplified cDNA product of expected size was detected by agarose gel electrophoresis. The full-length cDNA of the viroid-like RNA in RT-PCR products was cloned and sequenced. The established nucleotide sequence of the inserted cDNA was completely identical to that previously reported for HLVd. The total nucleic acids were extracted from hops cultivated in Japan, and the presence of HLVd were examined by dot blot hybridization and RT-PCR. In a hop sample, HLVd was not detected by dot blot hybridization, but detected by RT-PCR. The result indicates that RT-PCR is potentially more sensitive than dot blot hybridization using a radioactive probe. HLVd was detected in all mother cultivars tested. However, the two virus-free clones obtained by meristem culture showed negative reaction in both dot blot hybridization and RT-PCR detection methods. It is also shown that HLVd probably can be eliminated by meristem culture as indicated in several plant viruses and viroids.
Controlled degradation of myrcene, the major monoterpene hydrocarbon of hop oil, is discussed with special emphasis on the roie of myrcene as a precursor for the formation of flavor compounds occurring in naturally aging hop oil. Products are isolated and identified by means of gas chroma-tographic, mass spectral, and infrared analyses. Related possible degradation pathways are suggested for a number of these second- and third-order terpenoid constituents, and spectral data for several terpene alcohols, oxides, ketones, and polymerization products are reported.
The detectability of hop latent viroid (HLVd) was investigated in field-grown hop (Humulus lupulus L.; an herbaceous perennial in which all the aerial parts die at the onset of winter) plants, using dot-blot hybridisation. The viroid was readily detected in all aerial tissues in the second half of the growing season but it could not be detected very early in the season. Between early- and mid-season, HLVd was first detected at the base of the new stems and then apparently spread up them as they grew but only became detectable near the tips of the shoots at mid-season, approximately at the time most elongation growth ended and flowering began. Petioles were the most convenient tissues to test, being easy to collect and, relative to leaf lamina tissue, low in inhibitors. Both dot-blot and in situ hybridisation failed to detect HLVd in shoot tips taken from plants grown at two ‘low’ temperatures (10°C and 15°C). Failure to produce any viroid-free plants by in vitro culture from such tips suggested that they did contain viroid but at levels too low to detect by either method. Lower temperatures and smaller explants are now being investigated as means of producing viroid-free plants.
The surface topography of lupulin glands was studied at various stages of maturity using scanning electron microscopy. Maximum gland filling coincided with maximum alpha-acid concentration and dry matter yield. The percentage of the different components of oil remained constant after maximum gland filling had occurred. Dry matter content of hops at maximum gland filling varied from year to year and was not an accurate guide to optimum harvest date.
Two aspects of hop latent viroid (HLVd) relevant to control were examined: the production of viroid-free plants from infected material and transmission of HLVd in the field. Plants free from HLVd were obtained by a combination of storing infected source plants at low temperature (2–4oC in the dark) for several months followed by meristem culture using small explants. A total of 77 plants of six cultivars and male pollinator clones were grown from meristems and 28 of these were free from HLVd. Tests showed that the cutting of stems (mimicking the use of tools) was more effective than abrasion (mimicking natural plant to plant contact) for the mechanical transmission of HLVd between hop plants. When field-grown test plants were inoculated, infection occurred more commonly in May before plants had grown large enough for significant contact between neighbouring plants than later in the season. The aphid Phorodon humuli could not be shown to transmit HLVd.
These results indicate that all hop varieties and pollinator clones can be made available to the industry free from HLVd and that the chances of infection can be reduced by avoiding early-season cultural operations that cut into hop shoots.