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We evaluated the effectiveness of three different treatment groups at managing apple shoot blight, and the resulting canker incidence and canker length on wood caused by Erwinia amylovora. Preventative foliar sprays or trunk injections of giant knotweed extract (Regalia), oxytetracycline (Arbor-OTC or FireLine + Regulaid), or streptomycin (Agri-mycin/FireWall + Regulaid) were applied to mature ‘Fuji’ trees. Regalia and oxytetracycline were ineffective at reducing shoot blight severity, showing poor disease reductions of 18.2% and 24.3% compared to untreated controls across both years. Streptomycin was effective at controlling shoot blight severity when applied as a spray application, reducing necrosis by up to 93.9% across both years. Canker incidence was also poorly reduced by Regalia and oxytetracycline with an average decrease of 33.3% and 52.4%, respectively. Again, spray applications of streptomycin were most effective at reducing canker incidence (95.2%). When present, canker length was best controlled by spray applications of streptomycin, showing an average reduction of 95.7%. The effectiveness of Regalia and oxytetracycline was poor, reducing canker length by only 30.4% and 43.5%, respectively. Trunk injections of Regalia were consistently less effective than spray applications. Compared to their spray application counterpart, Regalia injections were, on average, 12.5%, 26.3%, and 25.1% less effective at reducing shoot blight severity, canker incidence, and canker length, respectively. Injected Arbor-OTC was more effective than spray applications of oxytetracycline. On average, Arbor-OTC injections were up to 28.3%, 40.1%, and 30% more effective at reducing shoot blight severity, canker incidence, and canker length compared to spray applications. Overall, Regalia and oxytetracycline were not as effective as streptomycin at controlling fire blight. The search for organic antibiotic alternatives for shoot blight and canker control continues, as cankers are increasing in economic importance by causing bearing wood and young tree death.

The bitter rot of apple is caused by Colletotrichum spp. and is a serious pre-harvest disease that can manifest in postharvest losses on harvested fruit. In this study, we obtained genome sequences from four different species, C. chrysophilum, C. noveboracense, C. nupharicola, and C. fioriniae, that infect apple and cause diseases on other fruits, vegetables, and flowers. Our genomic data were obtained from isolates/species that have not yet been sequenced and represent geographic-specific regions. Genome sequencing allowed for the construction of phylogenetic trees, which corroborated the overall concordance observed in prior MLST studies. Bioinformatic pipelines were used to discover CAZyme, effector, and secondary metabolic (SM) gene clusters in all nine Colletotrichum isolates. We found redundancy and a high level of similarity across species regarding CAZyme classes and predicted cytoplastic and apoplastic effectors. SM gene clusters displayed the most diversity in type and the most common cluster was one that encodes genes involved in the production of alternapyrone. Our study provides a solid platform to identify targets for functional studies that underpin pathogenicity, virulence, and/or quiescence that can be targeted for the development of new control strategies. With these new genomics resources, exploration via omics-based technologies using these isolates will help ascertain the biological underpinnings of their widespread success and observed geographic dominance in specific areas throughout the country.

The genus Colletotrichum includes nine major clades with 252 species and 15 major phylogenetic lineages, also known as species complexes. Colletotrichum spp. are one of the top fungal plant pathogens causing anthracnose and pre- and postharvest fruit rots worldwide. Apple orchards are imperiled by devastating losses from apple bitter rot, ranging from 24 to 98%, which is a serious disease caused by several Colletotrichum species. Bitter rot is also a major postharvest rot disease, with C. fioriniae causing from 2 to 14% of unmarketable fruit in commercial apple storages. Dominant species causing apple bitter rot in the Mid-Atlantic United States are C. fioriniae from the Colletotrichum acutatum species complex and C. chrysophilum and C. noveboracense from the C. gloeosporioides species complex (CGSC). C. fioriniae is the dominant species causing apple bitter rot in the Northeastern and Mid-Atlantic states. C. chrysophilum was first identified on banana and cashew but has been recently found as the second most dominant species causing apple bitter rot in the Mid-Atlantic. As the third most dominant pathogen, C. noveboracense MB 836581 was identified as a novel species in the CGSC, causing apple bitter rot in the Mid-Atlantic. C. nupharicola is a sister group to C. fructicola and C. noveboracense, also causing bitter rot on apple. We deliver the resources of 10 new genomes, including two isolates of C. fioriniae, three isolates of C. chrysophilum, three isolates of C. noveboracense, and two isolates of C. nupharicola collected from apple fruit, yellow waterlily, and Juglans nigra. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Bitter rot of apple is an economically important worldwide disease caused by different Colletotrichum species, depending on many factors such as climate, geography, other hosts, and crop management practices. Culture, morphology, and single-locus sequencing-based methods for identifying the Colletotrichum species are severely limited in effectiveness, while the multilocus sequence typing methods available for delineating species are costly, time-intensive, and require high expertise. We developed species-specific hydrolysis probe real-time PCR assays for the following nine Colletotrichum species causing bitter rot in the Mid-Atlantic U.S.A.: C. fructicola, C. chrysophilum, C. noveboracense, C. gloeosporioides s.s., C. henanense, C. siamense and C. theobromicola from the C. gloeosporioides species complex, and C. fioriniae and C. nymphaeae from the C. acutatum species complex. After searching 14 gene regions, we designed primers and probes in 5 of them for the nine target species. Four primer–probe set pairs were able to be duplexed. Sensitivity tests showed as little as 0.5 pg DNA were detectable. These real-time PCR assays will provide rapid and reliable identification of these key Colletotrichum species and will be critically important for studies aiming to elucidate their biology, epidemiology, and management on apples as the number one produced and consumed tree fruit in the U.S.A.

Florida ranks among the most important citrus growing regions in the USA. The present study investigates the occurrence, diversity, and pathogenicity of fungal species associated with symptomatic sweet orange (Citrus sinensis) cv. Valencia plants and fruit. The survey was conducted on twigs and fruit collected in Southwest Florida during 2022. Based on morphological and molecular characteristics, the identified isolates belonged to the species Lasiodiplodia iraniensis, Diaporthe pseudomangiferae, and Diaporthe ueckerae. The pathogenicity of representative isolates was evaluated on citrus fruit and plants. Lasiodiplodia iraniensis was the most virulent on fruit and plants, followed by Diaporthe pseudomangiferae. Diaporthe ueckerae had the lowest virulence on fruit, and it was not pathogenic to plants. In vitro tests were performed to assess the effect of temperature on mycelial radial growth. The optimum temperature of growth ranged from 26.0 to 28.4 °C for all the evaluated species, and L. iraniensis showed the fastest mycelial growth. This study represents the first report of L. iraniensis as a causal agent of tree dieback and fruit stem-end rot on C. sinensis worldwide. Moreover, D. pseudomangiferae and D. ueckerae are reported here for the first time in association with citrus diseases worldwide.

PATHMAP (Pathogen And Tree fruit Health MAP) is a smartphone application (app) and interactive dashboard developed specifically for support specialists, extension personnel, and university scientists supporting the tree fruit industry. The PATHMAP app collects detailed information about observed diseases, insect pests, and disorders and provides the option to attach photos. The data are then visualized using a graphical interface dashboard displaying an interactive color-coded map. Prior to the development of PATHMAP, abundant tree fruit disorder data were collected each year, but a central interactive repository for archiving data and facilitating communication of field observations did not exist. PATHMAP has been beta tested by university extension personnel, private consultants, and university scientists to ensure usability and functionality. PATHMAP will be used within the tree fruit industry for monitoring known pest patterns, occurrences, and outbreaks of emerging pathogens. It will augment existing extension diagnosis listservs that have value in visual diagnosis but are cumbersome and have no archiving capabilities. Data obtained through the tool can be used in epidemiological meta-analyses and to develop new predictive models, and can serve as a platform to track emerging pathogens, insects, and disorders for a variety of cropping systems. [Figure: see text] The author(s) have dedicated the work to the public domain under the Creative Commons CC0 “No Rights Reserved” license by waiving all of his or her rights to the work worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law, 2022.

The bacterium Erwinia amylovora causes fire blight and continues to threaten global commercial apple and pear production. Conventional microbiology techniques cannot accurately determine the presence of live pathogen cells in fire blight cankers. Several factors may prevent E. amylovora from growing on solid culture media, including competing microbiota and the release of bacterial-growth-inhibitory compounds by plant material during sample processing. We previously developed a canker processing methodology and a chip-based viability digital PCR (v-dPCR) assay using propidium monoazide (PMA) to bypass these obstacles. However, sample analysis was still time-consuming and physically demanding. In this work, we improved the previous protocol using an automatic tissue homogenizer and transferred the chip-based v-dPCR to the BioRad QX200 droplet dPCR (ddPCR) platform. The improved sample processing method allowed the simultaneous, fast, and effortless processing of up to six samples. Moreover, the transferred v-ddPCR protocol was compatible with the same PMA treatment and showed a similar dynamic range, from 7.2 × 102 to 7.6 × 107 cells mL−1, as the previous v-dPCR. Finally, the improved protocol allowed, for the first time, the detection of E. amylovora viable but nonculturable (VBNC) cells in cankers and bark tissues surrounding cankers. Our v-ddPCR assay will enable new ways to evaluate resistant pome fruit tree germplasm, further dissect the E. amylovora life cycle, and elucidate E. amylovora physiology, epidemiology, and new options for canker management.

Erwinia amylovora causes fire blight, a disease responsible for enormous economic losses in the pome fruit-producing areas where it is present. Despite the abundant research on fire blight, information about E. amylovora population dynamics and survival in fire blight cankers and the plant defense responses to this pathogen in the infected bark are limited. In our study, we obtained fire blight cankers in apple, pear, and Asian pear cultivars showing differing resistance to the disease by shoot inoculation with E. amylovora. We collected cankers from irrigated and non-irrigated trees every 3 months in two independent field experiments and analyzed samples by viability digital PCR. We also assessed the expression of pathogenicity-related (PR) genes in the bark of selected apple and Asian pear cultivars. A logistic regression analysis revealed the impact of environmental and host factors on E. amylovora detection rates in cankers. The chances of detecting live E. amylovora cells in cankers increased significantly in those collected from irrigated trees, in July, and/or during an experiment performed in a year with an expected average rainfall when compared to samples from non-irrigated trees, collected in January, and/or during an experiment performed under environmental conditions dominated by drought. We found a positive correlation between the pathogen detection rates in cankers and the host resistance to fire blight that might be explained by lower E. amylovora survival rates in more damaged tissues of susceptible hosts. The genes PR-1, PR-2, PR-5, and PR-8 were induced in the bark surrounding apple and Asian pear fire blight cankers. Our study, involving the analysis of more than 800 canker samples, provides new knowledge about the fire blight disease cycle and lays the foundation for improved fire blight management and eradication strategies in pome fruit orchards.