Halo Blight Research Articles

Chemical management strategies for halo blight of hop and in vitro sensitivity of Diaporthe humulicola populations to various fungicide classes.

Halo blight of hop, caused by Diaporthe humulicola, has increased in eastern North America since 2018. When left untreated, the disease can cause yield loss ranging from 17-56%. Currently, there are no fungicides registered for use on halo blight of hop. From 2020 to 2022 field trials were conducted using 10 fungicides registered for use on powdery and downy mildew of hop to determine their efficacy against halo blight. To validate field results, the EC50 value was determined for each active ingredient including flutriafol, tebuconazole + fluopyram, cyflufenamid, and trifloxystrobin + salicylhydroxamic acid (SHAM). Each fungicide tested had an EC50 value less than 50 ppm. A discriminatory dose was used to test the sensitivity of 206 D. humulicola isolates collected from the eastern U.S. and Canada in a poison agar assay. Results showed that tebuconazole + fluopyram decreased the incidence and severity of halo blight in the field. Also, this fungicide combination had EC50 values of 2.26 x 10-1 ppm and significantly reduced the growth of most of the isolates tested. Trifloxystrobin + SHAM decreased the presence of halo blight in the field tiral, but some isolates were less sensitive in discriminatory dose testing. Our results show that fungicides in FRAC groups 3, 7, and 11 were the most effective to control halo blight. Analyses of field trials showed a positive correlation between the severity of early season downy mildew infections and late season halo blight infections.

First report of Diaporthe humulicola causing halo blight of hop (Humulus lupulus L.) in Prince Edward Island, Canada

This study documents the first report of halo blight of hop (Humulus lupulus L.), caused by the fungal pathogen Diaporthe humulicola, in Prince Edward Island (PE), Canada. This pathogen has been previously reported in Michigan, New York, Connecticut and Quebec. Disease symptoms were reported in hopyards in Harrington and Mount Albion, PE. Symptoms reported include both foliar lesions and severe necrosis of hop cones. Through a metagenomic approach using Nanopore sequencing of the full Internal Transcribed Spacer (ITS) region products, multiple fungal species were identified in infected hop cones. Many plant pathogens were present in the hop cones including species of Botrytis, Alternaria, Colletotrichum and Fusarium. Additionally, metagenomic sequencing identified reads that most closely matched D. humulicola. Symptomatic leaf tissue sections were cultured on amended PDA to isolate the pathogen, then colony PCR was conducted and ITS regions were sequenced, confirming the identity of D. humulicola. Pathogenicity was validated using Koch’s postulates on hop leaf disks, with lesion development occurring 7 days post-inoculation. Considering the humid conditions characteristic of eastern North American hop-growing zones, halo blight’s emergence in PE emphasizes the need for comprehensive monitoring and improved best management strategies to protect the PE hop industry.

Potential of Nettle Infusion to Protect Common Bean from Halo Blight Disease

The use of plant-based preparations to replace chemical pesticides is a challenge for sustainable agriculture. Preparations from nettle (Urtica dioica L.) are good candidates, as the treatment of common bean plants (Phaseolus vulgaris L.) with aqueous suspensions of nettle reduced symptoms of halo blight disease caused by Pseudomonas syringae pv. phaseolicola (Pph). The aim of the present work was to test if nettle infusions (Ui) have similar effects and find out what activity is behind this: antimicrobial, promotion of the natural defenses of plants, and/or antioxidant. To achieve this, Pph growth was tested in the presence of infusions of nettle leaves collected in two different years (Ui18 and Ui22), and we found that it was only weakly inhibited at high concentrations of Ui18. Interestingly, Ui22 promoted bacteria growth at all concentrations. Second, we estimated the production of reactive oxygen species (ROS) in response to flagellin22 (flg22) in common bean leaf discs, since recognition of this bacterial peptide usually leads to ROS accumulation in tissues as a plant immune response. However, leaf discs that were preincubated in Ui showed no accumulation of ROS after flg22 treatment, suggesting that Ui can neutralize ROS production. Finally, in a Pph inoculation experiment of common bean plants grown in vitro, we observed that pretreatment of plants with Ui drastically reduced foliar oxidative damage and disease symptoms 6 h after inoculation. This effect was more noticeable for Ui22, which was related to the higher antioxidant activity found in this extract in comparison with Ui18. These results suggest that the protective properties of Ui are mainly due to the content of antioxidant bioactive compounds.

Efficiency of inoculation methods for the assessment of bacterial halo blight in coffee seedlings

AbstractBacterial halo blight (BHB), caused by Pseudomonas coronafaciens pv. garcae (Pcg), is one of the most significant diseases affecting coffee trees worldwide, resulting in considerable damage and notable losses within coffee production systems. Further research into the epidemiology and aetiology of BHB in coffee plants is essential for developing effective management strategies. Understanding the efficiency of each inoculation methodology in inducing symptoms is crucial for advancing this research. Therefore, this study aimed to assess the efficiency of seven Pcg inoculation methods using five concentrations to enhance the highest BHB intensity. The experimental design was a complete randomized block design in a factorial (7 × 5) variance analysis with four replications. The first three pairs of leaves of coffee seedlings of the cultivar Catuaí Vermelho IAC‐99 were inoculated and evaluated for the incidence and severity of BHB. The inoculation methods of Pcg by injury provided higher BHB AUDPCI and AUDPCS values. Among these, the inoculum injection and multi‐needle wounding at inoculum concentrations of 1.6 × 109 CFU mL−1 demonstrated higher disease intensity.

Antibiotic Resistance in Plant Pathogenic Bacteria: Recent Data and Environmental Impact of Unchecked Use and the Potential of Biocontrol Agents as an Eco-Friendly Alternative.

The economic impact of phytopathogenic bacteria on agriculture is staggering, costing billions of US dollars globally. Pseudomonas syringae is the top most phytopathogenic bacteria, having more than 60 pathovars, which cause bacteria speck in tomatoes, halo blight in beans, and so on. Although antibiotics or a combination of antibiotics are used to manage infectious diseases in plants, they are employed far less in agriculture compared to human and animal populations. Moreover, the majority of antibiotics used in plants are immediately washed away, leading to environmental damage to ecosystems and food chains. Due to the serious risk of antibiotic resistance (AR) and the potential for environmental contamination with antibiotic residues and resistance genes, the use of unchecked antibiotics against phytopathogenic bacteria is not advisable. Despite the significant concern regarding AR in the world today, there are inadequate and outdated data on the AR of phytopathogenic bacteria. This review presents recent AR data on plant pathogenic bacteria (PPB), along with their environmental impact. In light of these findings, we suggest the use of biocontrol agents as a sustainable, eco-friendly, and effective alternative to controlling phytopathogenic bacteria.

Indole Alkaloid Production by the Halo Blight Bacterium Treated with the Phytoalexin Genistein.

When Pseudomonas savastanoi pv. phaseolicola, the bacterium that causes halo blight, induces hypersensitive immunity in common bean leaves, salicylic acid and phytoalexins accumulate at the site of infection. Both salicylic acid and the phytoalexin resveratrol exert antibiotic activities and toxicities in vitro, adversely disrupting the P. savastanoi pv. phaseolicola proteome and metabolism and stalling replication and motility. These efficacious properties likely contribute to the cessation of bacterial spread in beans. Genistein is an isoflavonoid phytoalexin that also accumulates during bean immunity, so we tested its antibiotic potential in vitro. Quantitative proteomics revealed that genistein did not induce proteomic changes in P. savastanoi pv. phaseolicola in the same way that salicylic acid or resveratrol did. Rather, a dioxygenase that could function to metabolize genistein was among the most highly induced enzymes. Indeed, high-throughput metabolomics provided direct evidence for genistein catabolism. Metabolomics also revealed that genistein induced the bacterium to produce indole compounds, several of which had structural similarity to auxin. Additional mass spectrometry analyses proved that the bacterium produced an isomer of the auxin indole-3-acetic acid but not indole-3-acetic acid proper. These results reveal that P. savastanoi pv. phaseolicola can tolerate bean genistein and that the bacterium likely responds to bean-produced genistein during infection, using it as a signal to increase pathogenicity, possibly by altering host cell physiology or metabolism through the production of potential auxin mimics.

Structural analysis and molecular substrate recognition properties of Arabidopsis thaliana ornithine transcarbamylase, the molecular target of phaseolotoxin produced by Pseudomonas syringae.

Halo blight is a plant disease that leads to a significant decrease in the yield of common bean crops and kiwi fruits. The infection is caused by Pseudomonas syringae pathovars that produce phaseolotoxin, an antimetabolite which targets arginine metabolism, particularly by inhibition of ornithine transcarbamylase (OTC). OTC is responsible for production of citrulline from ornithine and carbamoyl phosphate. Here we present the first crystal structures of the plant OTC from Arabidopsis thaliana (AtOTC). Structural analysis of AtOTC complexed with ornithine and carbamoyl phosphate reveals that OTC undergoes a significant structural transition when ornithine enters the active site, from the opened to the closed state. In this study we discuss the mode of OTC inhibition by phaseolotoxin, which seems to be able to act only on the fully opened active site. Once the toxin is proteolytically cleaved, it mimics the reaction transition state analogue to fit inside the fully closed active site of OTC. Additionally, we indicate the differences around the gate loop region which rationally explain the resistance of some bacterial OTCs to phaseolotoxin.

Inheritance of Resistance to Bacterial Halo Blight and Bacterial Leaf Spot in Arabica Coffee.

Developing coffee cultivars resistant to multiple diseases by combining resistance genes is a top priority in breeding programs. To create cultivars resistant to diseases and nematodes, we transferred genes for resistance to bacterial infections caused by Pseudomonas coronafaciens pv. garcae, which causes bacterial halo blight (BHB), and P. amygdali pv. tabaci, which causes bacterial leaf spots (BLS), into Arabica coffee. Genetic analyses were conducted on breeding populations to estimate the number and function of genes that confer resistance to BHB and BLS. In total, 2,109 plants in the F2 generation and reciprocal backcrosses were inoculated with P. coronafaciens pv. garcae, while 1,996 plants were inoculated with P. amygdali pv. tabaci. Results showed that resistance to both pathogens had a heritability of 0.99, and the segregations of resistance indicated that each disease was controlled by a single dominant gene. The analyses also revealed that the resistance genes for BHB and BLS were linked, with an average distance of 10.75 cM between them on the same chromosome.

Newly isolated phages preying on Pseudomonas syringae pv. garcae: In vitro and ex vivo inactivation studies in coffee plant leafs

Coffee canker, or bacterial halo blight (BHB) of coffee, is a disease caused by the phytopathogenic bacterium Pseudomonas syringae pv. garcae (Psg), having been found for the first time in 1955, in the Garça region (State of São Paulo), and which has stood out in the Brazilian coffee plantations in recent years, leading to severe economic losses that seriously affect coffee trade. The treatments available are still scarce, involving frequent spraying of coffee plantations with either copper derivatives or the antibiotic kasugamycin. However, these compounds should be avoided due to environmental toxicity and the development of bacterial resistances. Herein we report the isolation and physical/biological characterisation of two novel lytic phages and their efficacy in the control of Psg. Phages ph002F and ph004F were isolated from coffee plant leaves in Brazil (Sorocaba/SP and Itu/SP cities), using Psg IBSBF-158 as the host. According to the transmission electron microscopy analyses, both phages belong to the class Caudoviricetes and present myovirus-like morphotypes. Phages ph002F and ph004F showed eclipse times of 5 min and 20 min, respectively, and a burst size of 123 PFU/host cell and 12 PFU/host cell, respectively, allowing to conclude they replicate well in Psg IBSBF-158 with latency periods of 50 min. Phage ph002F (reduction of 4.59 log CFU/mL, compared to uninfected culture) was more effective in inactivating Psg than phage ph004F (reduction of 3.85 log CFU/mL) after 10 h of incubation at a MOI of 10. As a cocktail, the two phages were highly effective in reducing the bacterial load (reduction of 5.26 log CFU/mL at a MOI of 0.1 or reduction of 5.03 log CFU/mL at a MOI of 10, relative to untreated culture), after 12 h of treatment. This study provides evidence that the isolated phages are promising candidates against the causative agent of BHB in coffee plants.

Association of cropping practices, cropping areas, and foliar diseases of common bean (Phaseolus vulgaris L.) in Ethiopia

Common bean production is being constrained by a number of diseases. The survey was carried out to prioritize common bean diseases and their associations with cropping areas and cultural practices. A stratified random assessment on 114 farmer's fields was conducted in Ethiopia's four common bean producing zones (Metekel, Sidama, Wolaita, and Halaba). Data are collected randomly on disease severity, incidence levels, and agronomic practices. The results revealed that eleven foliar diseases were found to affect common bean production. Among them, the maximum disease severity and distribution was scored for angular leaf spot (Pseudocercospora griseola) (mean disease severity of 17.8%) and bean anthracnose (Colletotrichum lindemuthianum) (10.9%) followed by floury leaf spot (Mycovellosiella phaseoli), ascochyta blight (Ascochyta phaseolorum Sacc), and web blight (Thanatephorus cucumeris). A multiple correspondence analysis was conducted, and revealed a significant association between the diseases, cropping areas and cultural practices. Optimal crop density was found to be associated with lower bean anthracnose and mosaic virus severities, while being liked to high levels of angular leaf spot and frog-eye leaf spot. Additionally, optimum sowing dates were associated with low severities of bean anthracnose, angular leaf spot, frog-eye leaf spot, Halo Blight, and mosaic virus. On the other hand, a high weediness level was found to be associated with higher levels of web blight and bean anthracnose. Intercropping and row planting were obtained to reduce angular leaf spot and increase occurance of ascochyta blight compared to monocropping. Furthermore, row planting were found to decrease bean anthracnose, angular leaf spot, and cercospora leaf spot when compared to broadcasting. The application of fertilizer was found to decrease the occurances of bean anthracnose, halo blight, floury leaf spot, and angular leaf spot. Therefore, efforts should focus on the optimization and integration of cultural practices for managing disease and producing common bean sustainably.

First report of halo blight on hop (Humulus lupulus) caused by Diaporthe humulicola in Minnesota.

Halo blight, caused by Diaporthe humulicola, is an emerging issue in hop production in the Upper Midwestern and Eastern North America. Reports of halo blight thus far have included Connecticut, Michigan, New York, and Quebec (Allan-Perkins et al.; Hatlen et al. 2022; Higgins et al. 2021; Sharma et al. 2022). In August 2020, brownish-gray necrotic foliar lesions and damaged cones were observed in an experimental hopyard consisting of a breeding population of hop plants grown at the University of Minnesota - Southern Research and Outreach Center in Waseca, MN. The foliar lesions consisted of necrotic concentric circles with some possessing chlorotic halos. Damage to the cones often appeared as reddish brown as bands around cone midsections, scattered on bracts and bracteoles, and in severe cases near entire cones. Disease incidence within the experimental hopyard was observed on >50% of hop plants. No pycnidia were observed on leaves or cones following collection of samples. A total of eleven samples were obtained from diseased leaves or cones. Symptomatic plant tissue was surface-sterilized and sections excised from the leading edge of lesions were plated onto potato dextrose agar (PDA). Fungal growth was hyphal tipped and incubated at 22° C under a 12-h photoperiod for a period of 21 days (Hatlen et al 2022). Culture characteristics on PDA included raised white to light gray mycelium with irregular pycnidia distribution over the surface. DNA was extracted from mycelia using the MagMAX Plant DNA Isolation Kit (Applied Biosystems, Foster City, CA). A representative isolate (M4N) was selected for DNA amplification and bi-directional Sanger sequencing using the following primers ITS1/ITS4 (ITS) for the internal transcribed spacer, CYLH3F/H3-1b for histone 3 (HIS), and Ef1728f/EF1-986R for translation elongation factor 1-α (TEF) (Carbone and Kohn 1999; Glass and Donaldson 1995; White et al. 1990). Following amplification and sequencing, reads were trimmed and assembled using Geneious Prime (Biomatters, New Zealand). BLASTn analysis revealed that the ITS (GenBank Accession OQ144379), HIS (GenBank Accession OQ256246), and TEF (GenBank Accession OQ256245) were 99 - 100% identical to D. humulicola sequences (MN152927, MN180213, MN180207) infecting hop in other US regions (Allan-Perkins et al. 2019; Hatlen et al. 2022). To complete Koch's postulates, two sets each of six 3-month old plants of the hop cv. 'Chinook' were inoculated with either 50 mL of conidial suspension (6 x 105 conidia/mL) derived from pycnidia harvested from 28-day old cultures or with water as a negative control. Following inoculation, plants were then grown in a greenhouse at 100% relative humidity at 22°C with a 14-h photoperiod. Light brown lesions with concentric circles appeared on the adaxial side of the leaf after 3 weeks but were not observed on mock-inoculated plants. We subsequently re-isolated D. humulicola from 100% of infected leaves which was identified based upon colony and conidial morphology using descriptions from Higgins et al. (2021). alpha-conidia (n = 20) averaged 10.96 µm ± 1.12 in length and 5.11 µm ± 0.67 in width, were unicellular and hyaline. No beta-conidia were observed, consistent with previous reports of this pathogen. No disease symptoms appeared on mock-inoculated plants, and D. humulicola was not recovered from mock-inoculated plants. There is significant concern regarding the increasing prevalence of D. humulicola as an emerging pathogen affecting hop production across the Midwestern and Great Lakes region of North America. Future research is needed to determine differences in hop varietal susceptibility and fungicide efficacy for management of this disease.

First report in South Korea of a bacterial leaf blight of oats caused by Pseudomonas salomonii.

In March 2020, approximately 20% of leaves in a commercial oat field growing Avena sativa (cv. Samhan) in Jeongeup, Korea (35.3859°, 126.5607°), displayed leaf blight in the seedling stage of development. The lesions observed in 2020 were of a yellow discoloration that spread from the leaf tip downward with minimal brown delineation (Fig. S1A). These symptoms differed from the haloes delineated at the edges by extensive brown necrosis caused by Pseudomonas coronofaciens (Kim 2020a; Fig. S1C). To isolate the causal agent, 3-cm-long pieces of symptomatic leaves from ten different oat plants were disinfected by submersion in 70% ethanol for 5 min followed by immersion in 1% sodium hypochlorite for 5 min and rinsing extensively with sterile distilled water. The air-dried segments were transferred intact to nutrient agar, and only one colony type (yellow-colored, wet, with a shiny convex surface) was observed. After single colony isolation, three isolates from different 2020 field-grown diseased leaves, termed 2007, 2009, and 2011, were selected at random. The isolates produced fluorescent siderophores on King's medium B and triggered a hypersensitive response (HR) when infiltrated into tobacco (cv. Xanthi) leaves (Fig. S1D). Multilocus sequence typing analysis with four housekeeping genes was used for taxonomic identification of these isolates (Maiden et al. 1998). The 16S rRNA sequences were amplified with the 27F/1492R universal primers (Weisburg et al. 1991). The primers for three housekeeping genes were designed using genome sequence of P. coronaficiens X-1 causing halo blight disease in Korea (NZ_CP050260.1, Kim, 2020b). The products obtained had sizes of 640 bp for gltA (using primers F: 5'-CCT GGT AGC CAA GAT GCC GAC-3'; R: 5'-CAA AGA TCA CGG TGA ACA TGC TGG-3'), 710 bp for gyrB (with primers F: 5'-TCG GCA GCC GAG GTC ATC ATG AC-3'; R: 5'-TTG TCT TTG GTC TGC GAG CTG AA-3'), and 870 bp for rpoD ( with primers F: 5'-GTG AAG GCG AAA TCG AAA TCG-3'; R: 5'-CCG ATG TTG CCT TCC TGG ATC AG-3'). The concatenated sequences (2,353 bp/2,376 bp) had 99% identity with the gene sequences from P. salomonii type strain (AY091528.1, NZ_FNOX01000003, LC486864.1, LC486849.1), but lesser identity (90%) with P. coronafaciens (Fig. S2). Pathogenicity of the Korean isolates was confirmed by fulfilling Koch's postulates using leaves of 2 week-old greenhouse-grown 'Samhan' seedlings. Plants (n=50) were sprayed with 108 cfu/ml bacterial suspensions in water or with sterile water as controls. The plants were incubated for a week at 23 °C in 100% relative humidity under a 10 h light/14 h dark photoperiod. Five days after bacterial inoculation, yellow discoloration appeared at the leaf tips which progressed downward with time (Fig. S1B). Three bacterial isolates extracted from yellowed, inoculated leaves had 16S rRNA gene sequences identical to that of P. salomonii Korean isolates, 2007, 2009 and 2011, and they caused the anticipated symptoms when inoculated into oat leaves. These findings indicate that P. salomonii should be added to the potential pathogens of oats grown in Korea. Understanding whether spring weather conditions (warmth and humidity) boost this oat disease will help devise disease alert systems for farmers (Anderson 2004; Chakraborty 2005).

Evaluating Nicotinamide Adenine Dinucleotide for Its Effects on Halo Blight of Snap Bean.

Halo blight, caused by Pseudomonas syringae pv. phaseolicola, is one of the major bacterial diseases on snap bean in Florida, and the outbreaks of this disease have occurred more often in recent years. Current management of this disease primarily depends on application of fixed copper-based bactericides but climate change and resistance development in the pathogen populations still cause hardship for management of this disease, especially in south Florida. In this study, nicotinamide adenine dinucleotide (NAD+) was evaluated in the greenhouse for its potential to reduce halo blight on snap bean. When NAD+ at 5 mM was applied by soil drench, foliar spray, or leaf infiltration, NAD+ significantly (P < 0.05) reduced disease severity of halo blight on snap bean compared with the untreated control. When NAD+ was applied by leaf infiltration, among the tested concentrations, NAD+ at 0.5 to 1.0 mM was most effective in decreasing halo blight disease. NAD+ at 2.5 mM applied as a foliar spray in rotation with Kocide 3000 (copper hydroxide) at 0.5 mg/ml further reduced disease severity compared with Kocide 3000 alone. In the in vitro study, no inhibitory effects of NAD+ were detected on the bacterial pathogen P. syringae pv. phaseolicola. Results of real-time PCR showed that the defense-related genes PR1, AZI1, EDS1, SARD1, PDF1.2, and PAL1 were upregulated in the NAD+ treatment. Taken together, these data indicated that NAD+ significantly suppressed halo blight on snap bean, and application of NAD+ has the potential in management of this important disease.

First Report of Halo Blight on Hop Caused by Diaporthe humulicola in New York.

In late July and August 2015, foliar disease was observed in three hop (Humulus lupulus; unknown cultivars) yards in Ontario, Otsego, and Putnam counties, New York (NY). Disease incidence ranged between 70 and 90% of plants, and up to 25% of the leaves per plant were affected. Leaf symptoms were large, necrotic patches with a chlorotic halo (2 to 10 cm diam.). Leaves and dry, easily shattered cones were placed at high humidity for 10 days. Pycnidia were abundant in leaf lesions which extruded conidia. Pycnidia were also observed on cone bracts and bracteoles. Fifteen isolations were made from each yard by placing a pycnidium onto 2% water agar + 0.02% (w/v) ampicillin. Colonies were hyphal tipped and transferred to potato dextrose agar (PDA) before incubation at 20°C with a 12-h photoperiod. Colonies on PDA had flat mycelia and were white to cream in color. The isolation frequency was 100%. To induce sporulation, five isolates were grown on PDA with autoclaved alfalfa stems for 7 to 10 days. Alpha conidia were hyaline, and oval with obtuse ends. Mean alpha conidial dimensions were (n = 20): 9.1 m × 3.4 µm (BE1; Ontario Co.); 11.8 × 3.8 µm (BE34; Ontario Co.); 9.6 × 4.1 µm (BE10; Ontario Co.); 10.2 × 3.7 µm (BE52; Otsego Co.); and 10.3 × 3.6 µm (BE69; Putnam Co.). Beta conidia were not observed. DNA was extracted and PCR performed to amplify the internal transcribed spacer (ITS) region (primers ITS1/ITS4; White et al. 1990), translation elongation factor 1-α (TEF; EF1-728F/EF1-986R; Carbone and Kohn 1999), a partial region of β-tubulin (TUB; Bt2a/Bt2b; Glass and Donaldson 1995), a partial region of histone 3 (H3) (H3; CYLH3F/H3-1b Crous et al. 2004), and calmodulin (CAL; CAL-228F/CAL2Rd; Groenewald et al. 2013) genes. For all NY isolates, sequence similarity was >99% to D. humulicola CT2018-3 for the ITS region, and TEF, HIS, and CAL genes. Sequence similarity to CT2018-3 for the TUB region ranged from 86.96% (BE-1) to 96.15% (BE-10). . Analyses with the ITS, TEF, CAL, and HIS sequences supported our identification of the NY isolates as D. humulicola. Sequences were deposited in GenBank (OM370960 to OM370984). For pathogenicity testing, BE-34 and BE-69 were grown on PDA + autoclaved alfalfa stems at room temperature and a 12-h photoperiod for 10 days. Conidia were harvested by flooding the plate with sterile water. Conidial concentration was quantified, and the inoculum suspension diluted to ~5  105 (+ 0.01% polysorbate-20)/ml. Five cv. Cascade plants were sprayed with inoculum until run-off and covered with a plastic bag for 72 h. Non-inoculated control plants were sprayed with 0.01% polysorbate-20 and bagged. Plants were placed in a misting chamber and exposed to alternating 25°C light/18°C dark with a 16 h photoperiod. Mist was applied for 1 h daily. Necrotic lesions like the field specimens were observed on all inoculated plants after 28 days with no symptoms on control plants. Diseased leaves were detached and placed in a humid chamber for 2 days, and pycnidia observed in lesions. The reisolation frequency of D. humulicola was 100%. Conidia from the isolates had similar morphology to the original isolates. This is the first report of halo blight caused by D. humulicola on hop in NY. Halo blight has been reported on hop and associated with significant yield loss through cone shattering in MI (Higgins et al. 2021), CT (Allan-Perkins et al. 2020), and Quebec, Canada (Hatlen et al. 2021). Research is needed to determine if management is warranted.

Development and Validation of an Allele-Specific Marker for Resistance to Bacterial Halo Blight in Coffea arabica

Bacterial halo blight (BHB) is a bacterial disease, caused by Pseudomonas syringae pv. garcae, which has been gaining prominence in the main coffee-producing regions. Chemical control of this disease increases production costs and is environmentally undesirable. In this scenario, the development of new cultivars resistant to BHB is the most economical and sustainable alternative. Marker-Assisted Selection (MAS) is an appropriate strategy to assist breeding programs for resistant genotype selection. In a previous Genome-Wide Association Study (GWAS) for C. arabica and P. syringae pv. garcae interaction, we identified a locus, probably linked to qualitative resistance to the pathogen. In this work, we developed and validated a pair of Allele-Specific-Polymerase Chain Reaction (AS-PCR) primers for this locus in C. arabica breeding populations. This pair of AS-PCR primers, called Psg_QL1, was tested both in a backcross (BC) (n = 38) and in an F2 population (n = 138) segregating for resistance to BHB. The linkage between the Psg_QL1 marker and qualitative resistance showed an accuracy of 93.75%. Our results demonstrated that the Psg_QL1 marker can be applied in MAS in a robust, simple, fast, and low-cost way.

OxyR Positively Influences Phaseolotoxin Synthesis and Pyoverdin Production in Pseudomonas savastanoi pv. phaseolicola NPS3121.

Phaseolotoxin is a major virulence factor of the bean pathogen bacterium P. savastanoi pv. phaseolicola. This toxin plays a key role in the development of the halo blight disease in bean plants. So far, the signal transduction pathways involved in the synthesis of phaseolotoxin have not been elucidated. The influence of regulation mechanisms related to the oxidative stress response, in particular the OxyR protein, it has been suggested to be involved in this process.. In this study we evaluated the role of OxyR in P. savastanoi pv. phaseolicola, mainly compared to the synthesis of phaseolotoxin and the virulence of this phytopathogen. Generation of the oxyR-mutant, pathogenicity and virulence tests, and analyses of gene expression by RT-PCR assays were performed. The results showed that OxyR exerts an effect on the synthesis of phaseolotoxin and positively influences the expression of the Pht and Pbo cluster genes. Likewise, OxyR influences the production of pyoverdine by the control of the expression of the genes encoding the PvdS sigma factor, involved in the synthesis of this pigment. This study is the first report on members of the OxyR regulon of P. savastanoi pv. phaseolicola NPS3121.

Genome-wide association study for resistance to Pseudomonas syringae pv. garcae in Coffea arabica.

Bacteria halo blight (BHB), a coffee plant disease caused by Pseudomonas syringae pv. garcae, has been gaining importance in producing mountain regions and mild temperatures areas as well as in coffee nurseries. Most Coffea arabica cultivars are susceptible to this disease. In contrast, a great source of genetic diversity and resistance to BHB are found in C. arabica Ethiopian accessions. Aiming to identify quantitative trait nucleotides (QTNs) associated with resistance to BHB and the influence of these genomic regions during the domestication of C. arabica, we conducted an analysis of population structure and a Genome-Wide Association Study (GWAS). For this, we used genotyping by sequencing (GBS) and phenotyping for resistance to BHB of a panel with 120C. arabica Ethiopian accessions from a historical FAO collection, 11C. arabica cultivars, and the BA-10 genotype. Population structure analysis based on single-nucleotide polymorphisms (SNPs) markers showed that the 132 accessions are divided into 3 clusters: most wild Ethiopian accessions, domesticated Ethiopian accessions, and cultivars. GWAS, using the single-locus model MLM and the multi-locus models mrMLM, FASTmrMLM, FASTmrEMMA, and ISIS EM-BLASSO, identified 11 QTNs associated with resistance to BHB. Among these QTNs, the four with the highest values of association for resistance to BHB are linked to g000 (Chr_0_434_435) and g010741 genes, which are predicted to encode a serine/threonine-kinase protein and a nucleotide binding site leucine-rich repeat (NBS-LRR), respectively. These genes displayed a similar transcriptional downregulation profile in a C. arabica susceptible cultivar and in a C. arabica cultivar with quantitative resistance, when infected with P. syringae pv. garcae. However, peaks of upregulation were observed in a C. arabica cultivar with qualitative resistance, for both genes. Our results provide SNPs that have potential for application in Marker Assisted Selection (MAS) and expand our understanding about the complex genetic control of the resistance to BHB in C. arabica. In addition, the findings contribute to increasing understanding of the C. arabica domestication history.

Genome-wide meta-QTL analyses provide novel insight into disease resistance repertoires in common bean

BackgroundCommon bean (Phaseolus vulgaris) is considered a staple food in a number of developing countries. Several diseases attack the crop leading to substantial economic losses around the globe. However, the crop has rarely been investigated for multiple disease resistance traits using Meta-analysis approach.Results and conclusionsIn this study, in order to identify the most reliable and stable quantitative trait loci (QTL) conveying disease resistance in common bean, we carried out a meta-QTL (MQTL) analysis using 152 QTLs belonging to 44 populations reported in 33 publications within the past 20 years. These QTLs were decreased into nine MQTLs and the average of confidence interval (CI) was reduced by 2.64 folds with an average of 5.12 cM in MQTLs. Uneven distribution of MQTLs across common bean genome was noted where sub-telomeric regions carry most of the corresponding genes and MQTLs. One MQTL was identified to be specifically associated with resistance to halo blight disease caused by the bacterial pathogen Pseudomonas savastanoi pv. phaseolicola, while three and one MQTLs were specifically associated with resistance to white mold and anthracnose caused by the fungal pathogens Sclerotinia sclerotiorum and Colletotrichum lindemuthianum, respectively. Furthermore, two MQTLs were detected governing resistance to halo blight and anthracnose, while two MQTLs were detected for resistance against anthracnose and white mold, suggesting putative genes governing resistance against these diseases at a shared locus. Comparative genomics and synteny analyses provide a valuable strategy to identify a number of well‑known functionally described genes as well as numerous putative novels candidate genes in common bean, Arabidopsis and soybean genomes.

Breeding and Agronomic Evaluation of Jilv 20, a New Mungbean (Vigna radiata L.) Cultivar

Mungbean (Vigna radiata L.), one of the most widely grown edible legumes in Asia, plays important roles in the improvement of agricultural cultivation systems as well as human diets. As an understudied crop, however, most mungbean varieties are characterized by unstable yields, poor disease resistance, and unsuitability for mechanical harvesting, thereby leading to a low production income. We, therefore, developed Jilv 20, a new mungbean cultivar with a dull seed coat, by crossing Bao 942-34 with Weilv 9002-341. After 11 years of selection and evaluation, Jilv 20 has shown wide adaptability, early maturity, high yield, halo blight resistance, and suitability for mechanical harvesting. The growth period of Jilv 20 is 79.5 and 66.7 days in the spring and summer sowing seasons, respectively, and the average plant height is approximately 56.4 cm. In this study, average yields were 1737.9, 1532.3, and 2045.1 kg/hm2 in northern spring-sowing, northern summer-sowing, and southern sites, respectively; these values were respectively 27.83%, 28.48%, and 6.96% higher than those of control cultivar Zhonglv 5, which has been popular in past decades because of its wide adaptability. The average protein and starch contents of Jilv 20 seeds were 25.0% and 49.56%, respectively. Further application and extension of Jilv 20 in China should contribute to mungbean production, breeding, and industrial development.

Predictive modeling of Pseudomonas syringae virulence on bean using gradient boosted decision trees.

Pseudomonas syringae is a genetically diverse bacterial species complex responsible for numerous agronomically important crop diseases. Individual P. syringae isolates are assigned pathovar designations based on their host of isolation and the associated disease symptoms, and these pathovar designations are often assumed to reflect host specificity although this assumption has rarely been rigorously tested. Here we developed a rapid seed infection assay to measure the virulence of 121 diverse P. syringae isolates on common bean (Phaseolus vulgaris). This collection includes P. syringae phylogroup 2 (PG2) bean isolates (pathovar syringae) that cause bacterial spot disease and P. syringae phylogroup 3 (PG3) bean isolates (pathovar phaseolicola) that cause the more serious halo blight disease. We found that bean isolates in general were significantly more virulent on bean than non-bean isolates and observed no significant virulence difference between the PG2 and PG3 bean isolates. However, when we compared virulence within PGs we found that PG3 bean isolates were significantly more virulent than PG3 non-bean isolates, while there was no significant difference in virulence between PG2 bean and non-bean isolates. These results indicate that PG3 strains have a higher level of host specificity than PG2 strains. We then used gradient boosting machine learning to predict each strain’s virulence on bean based on whole genome k-mers, type III secreted effector k-mers, and the presence/absence of type III effectors and phytotoxins. Our model performed best using whole genome data and was able to predict virulence with high accuracy (mean absolute error = 0.05). Finally, we functionally validated the model by predicting virulence for 16 strains and found that 15 (94%) had virulence levels within the bounds of estimated predictions. This study strengthens the hypothesis that P. syringae PG2 strains have evolved a different lifestyle than other P. syringae strains as reflected in their lower level of host specificity. It also acts as a proof-of-principle to demonstrate the power of machine learning for predicting host specific adaptation.