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Rice (Oryza sativa L.) is a vital crop feeding more than half of the world's population, with production occurring predominantly in Asian countries. However, rice cultivation faces challenges from various fronts, including biotic stresses intensified by climate change. False smut, caused by Ustilaginoidea virens , has emerged as a significant threat to rice production globally. The application of curative fungicides after symptom appearance has limited scope in managing this disease since the infection process usually starts during the early flowering stage of rice crops. This study investigates the utilization of spore-trapping technology coupled with Loop-Mediated Isothermal Amplification (LAMP) assay for monitoring airborne U. virens inocula in rice fields. For early detection and quantification of U. virens , sampling rods coated with silicone grease were deployed to collect airborne spores, and DNA extraction was performed using a modified method. Both PCR and LAMP assays were employed for detection, with LAMP offering advantages of rapidity, sensitivity, and simplicity. The study demonstrated the superior sensitivity of LAMP compared to PCR, detecting U. virens DNA at concentrations as low as 100 femtograms. Continuous monitoring of U. virens inoculum using spore trapping revealed the spatio-temporal dynamics of U. virens dispersal, providing valuable insights for disease management. Implementing a fungicidal application schedule based on airborne inoculum detection led to significant reductions in both false smut incidence and severity and improved crop yield. The meteorological parameters including minimum temperature, relative humidity in the morning and evening, sunshine, and solar radiation were found to be correlated with disease incidence. Multi-operator validation confirmed the robustness and specificity of the LAMP assay. Overall, this integrated approach offers a proactive strategy for monitoring and managing false smut disease, enhancing sustainable rice production and food security. [ABSTRACT FROM AUTHOR]

Rice (Oryza sativa L.) is a vital crop feeding more than half of the world’s population, with production occurring predominantly in Asian countries. However, rice cultivation faces challenges from various fronts, including biotic stresses intensified by climate change. False smut, caused by Ustilaginoidea virens, has emerged as a significant threat to rice production globally. The application of curative fungicides after symptom appearance has limited scope in managing this disease since the infection process usually starts during the early flowering stage of rice crops. This study investigates the utilization of spore-trapping technology coupled with Loop-Mediated Isothermal Amplification (LAMP) assay for monitoring airborne U. virens inocula in rice fields. For early detection and quantification of U. virens, sampling rods coated with silicone grease were deployed to collect airborne spores, and DNA extraction was performed using a modified method. Both PCR and LAMP assays were employed for detection, with LAMP offering advantages of rapidity, sensitivity, and simplicity. The study demonstrated the superior sensitivity of LAMP compared to PCR, detecting U. virens DNA at concentrations as low as 100 femtograms. Continuous monitoring of U. virens inoculum using spore trapping revealed the spatio-temporal dynamics of U. virens dispersal, providing valuable insights for disease management. Implementing a fungicidal application schedule based on airborne inoculum detection led to significant reductions in both false smut incidence and severity and improved crop yield. The meteorological parameters including minimum temperature, relative humidity in the morning and evening, sunshine, and solar radiation were found to be correlated with disease incidence. Multi-operator validation confirmed the robustness and specificity of the LAMP assay. Overall, this integrated approach offers a proactive strategy for monitoring and managing false smut disease, enhancing sustainable rice production and food security.

Plant disease management requires early intervention, which has traditionally required extensive scouting and surveillance to guide decision-making. Visual inspection, unfortunately, requires conditions for disease symptom expression and expertise in identifying causal organisms and may delay management strategies that prevent losses associated with a given plant pathogen. Advances in spore trap technology and molecular diagnostic technologies provide opportunities for greater confidence in pathogen detection and implementation by scientists, farmers, and regulatory agencies. [Figure: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Currently, the ash dieback causal agent Hymenoscyphus fraxineus is an established invasive pathogen in most European countries. Its potential to spread quickly among invaded forests is based on its propagules: airborne inoculum composed mainly of ascospores originated in apothecia growing on leaf litter infected during the previous vegetation season. The spread of the inoculum by air masses to distant areas is probable and depends on the availability of the ascospores in higher levels of air. Our study aimed to detect the inoculum in an infected area at heights of more than 20 meters. Our study was conducted in a municipal locality (Boršov nad Vltavou) with tens of infected ash trees (Fraxinus excelsior) in South Bohemia (SW Czechia). The infected trees surround an agricultural silo where five rotating arm spore traps (rotorods) were mounted for ten consequent 48h samplings during the peak of the sporulating season (17th July to 6th August 2020). The spore traps were mounted 48, 37, 25, 14 and 0,3 meters above ground. Samples were quantified by qPCR. Results clearly proved the ability of the spores to reach a height of 48 meters. Furthermore, H. fraxineus DNA was detected from all five spore traps during all ten samplings. Mostly, the amount of detected spores showed a decreasing trend with height, and varied a lot. During some of the samplings, higher spore concetrations were achieved at the top than at the lower traps, which can be explained by horizontal air transfer of the inoculum from other infected areas. Based on GLM analyses, higher spore concentrations were achieved during days without rain, lower air temperatures, after cloudy, humid and rainy weather without strong winds. A combination of rotorod ROTTRAP 52 with qPCR quantification proved to be an efficient technology for a study focused on the vertical spread of H. fraxineus propagules. [ABSTRACT FROM AUTHOR]

Cercospora leaf spot is an important foliar disease in sugar beet caused by Cercospora beticola. Tolerant cultivars are available, but application of fungicides is still mandatory for disease control. The timing of the fungicide application is crucial as it determines the outcome of disease epidemiology. A disease incidence (DI) of 5% is widely used as a threshold for fungicide application. Recently a method was developed that allows the quantification of aerial spore dispersal of C. beticola for measuring spore flight intensity. It was aimed in this study to prove if fungicide application based on spore flight might improve disease control compared to DI. In a field trial with artificial inoculation, a single fungicide application at the onset of spore flight slowed down disease development as indicated by reduced disease severity and aerial spore dispersal. However, it did not provide sufficient control in terms of sugar yield. Only a second fungicide application based on spore flight detection achieved an efficacy similar to two fungicide applications based on DI. In contrast, a single fungicide application based either on spore flight or DI was sufficient in two on-farm trials under natural infection with moderate disease pressure. This highlights the necessity of an early timed first fungicide application followed by a second application under high disease pressure induced by artificial inoculation. Although fungicide application based on spore flight achieved sufficient control success in on-farm trials, it seems not to improve disease control compared to the usage of DI as threshold. [ABSTRACT FROM AUTHOR]

Frosty pod rot, caused by Moniliophthora roreri, is the most damaging disease of cacao in Latin America and, to better comprehend its epidemiology, we must understand its dissemination and proliferation. However, we do not know how M. roreri spores loads fluctuate in time and space due to the lack of a reliable technique to quantify M. roreri spores in the fields. Therefore, we developed a method that relies on spore traps and qPCR to detect and quantify M. roreri spore loads. This study demonstrated that the qPCR protocol can detect down to 0.025 ng of M. roreri DNA and quantify between 0.006 ng and 60 ng. Moreover, it demonstrated that qPCR protocol can detect and quantify DNA extracted from spore suspension and spore traps containing at least 2.9 × 104 M. roreri spores. However, the variability of the estimates for spore samples was high. Finally, we described a spore-trap device designed to carry spore traps in the field. The qPCR protocol and spore-trap device here developed will help in the understanding of the M. roreri dissemination patterns since they can be used to assess the environmental loads of M. roreri spore in cacao fields. [ABSTRACT FROM AUTHOR]

Prophylactic stump treatments against the conifer root rot fungus Heterobasidion annosum s.l. should take into account the airborne inoculum density of the pathogen, in order to be economically feasible. Our objective was to test if an automatic volumetric spore trap (AVST) could be used as a sensitive alternative of passive traps for quantifying Heterobasidion airspora in forest stands. The routinely used wood disc exposure method (WDE) was implemented simultaneously with AVST in five coniferous monocultures and three near-natural stands without silvicultural management in Czechia. Air sampling took place for 24-h intervals in five months of the main sporulation period. The atmospheric concentration of H. annosum s.l. spores (cHa) was determined by qPCR with generic and species-specific primers. AVST detected more target species than WDE on 77% of sampling occasions. When comparing the relative abundance of the three European H. annosum s.l. species detected by AVST and WDE, H. parviporum and H. abietinum were found to be overrepresented on wood discs. cHa in November was significantly higher than in May and June, confirming the seasonal pattern typical for temperate Europe. For an accurate and rapid estimation of Heterobasidion primary infection risk, the usage of AVST is highly recommended. [ABSTRACT FROM AUTHOR]

In Sweden, reforestation of managed forests relies predominantly on planting nursery-produced tree seedlings. However, the intense production using containerized cultivation systems (e.g., high seedling density, irrigation from above, regular fertilization) creates favorable conditions for fungal infections. Despite the harmful role of diseases in forest nurseries, the origin and dispersal factors of fungal pathogens remain largely unknown. A better understanding of the airborne spread of pathogens could improve the prediction of fungal infection, ultimately optimizing preventative methods and decreasing the use of fungicides. This study investigated the temporal dynamics of airborne fungi in forest nurseries, with a focus on fungal pathogens. Airborne fungi were monitored in four Swedish forest nurseries over two growing seasons using spore traps and high-throughput sequencing. Fungal pathogens were identified using bioinformatics and quantified with quantitative PCR. Results showed strong temporal shifts of airborne fungal diversity and community composition following the growing seasons. The airborne spread included high abundances of important fungal pathogens (e.g., Cladosporium sp., Botrytis cinerea , Alternaria sp., Sydowia polyspora , and Melampsora populnea ) with individual temporal and spatial variations. In general, the deposited spore loads of nursery pathogens correlated positively with increased temperature and negatively with higher precipitation. This was expressed the strongest for Cladosporium sp., Alternaria sp., and M. populnea , which suggests a higher availability of fungal inoculum in warm and dry periods. This study highlights the influence of seasonality on the temporal dynamics of economically important fungal pathogens in Swedish forest nurseries, which should be considered in the development of a local decision support system.IMPORTANCEFungal diseases in forest nurseries have significant environmental and economic impacts on the tree seedling production. This study highlights the role of seasonality in the airborne spread of fungal pathogens in Swedish forest nurseries. By analyzing airborne fungal spores using advanced sequencing and monitoring techniques, key fungal pathogens and their dispersal patterns over two growing seasons were identified. The findings indicate that warmer, drier periods may increase the spread of fungal pathogens, emphasizing the need for targeted preventative measures. Understanding these temporal dynamics can help optimize the use of fungicides in forest nurseries, thereby promoting more sustainable and environmentally friendly management practices. This research provides valuable insights for improving disease management in forest nurseries, ultimately supporting sustainable tree seedling production.

Wheat is one of the main staple food crops, and 775 million tonnes of wheat were produced worldwide in 2022. Fungal diseases such as Fusarium head blight, Septoria tritici blotch, spot blotch, tan spot, stripe rust, leaf rust, and powdery mildew cause serious yield losses in wheat and can impact quality. We aimed to investigate the incidence of spores from major fungal pathogens of cereals in the field by comparing microscopic and metagenomic based approaches for spore identification. Spore traps were set up in four geographically distinct UK wheat fields (Carnoustie, Angus; Bishop Burton, Yorkshire; Swindon, Wiltshire; and Lenham, Kent). Six major cereal fungal pathogen genera (Alternaria spp., Blumeria graminis, Cladosporium spp., Fusarium spp., Puccinia spp., and Zymoseptoria spp.) were found using these techniques at all sites. Using metagenomic and BLAST analysis, 150 cereal pathogen species (33 different genera) were recorded on the spore trap tapes. The metagenomic BLAST analysis showed a higher accuracy in terms of species-specific identification than the taxonomic tool software Kraken2 or microscopic analysis. Microscopic data from the spore traps was subsequently correlated with weather data to examine the conditions which promote ascospore release of Fusarium spp. and Zymoseptoria spp. This revealed that Zymoseptoria spp. and Fusarium spp. ascospore release show a positive correlation with relative humidity (%RH). Whereas air temperature (°C) negatively affects Zymoseptoria spp. ascospore release. [ABSTRACT FROM AUTHOR]

Wheat is one of the main staple food crops, and 775 million tonnes of wheat were produced worldwide in 2022. Fungal diseases such as Fusarium head blight, Septoria tritici blotch, spot blotch, tan spot, stripe rust, leaf rust, and powdery mildew cause serious yield losses in wheat and can impact quality. We aimed to investigate the incidence of spores from major fungal pathogens of cereals in the field by comparing microscopic and metagenomic based approaches for spore identification. Spore traps were set up in four geographically distinct UK wheat fields (Carnoustie, Angus; Bishop Burton, Yorkshire; Swindon, Wiltshire; and Lenham, Kent). Six major cereal fungal pathogen genera (Alternaria spp., Blumeria graminis, Cladosporium spp., Fusarium spp., Puccinia spp., and Zymoseptoria spp.) were found using these techniques at all sites. Using metagenomic and BLAST analysis, 150 cereal pathogen species (33 different genera) were recorded on the spore trap tapes. The metagenomic BLAST analysis showed a higher accuracy in terms of species-specific identification than the taxonomic tool software Kraken2 or microscopic analysis. Microscopic data from the spore traps was subsequently correlated with weather data to examine the conditions which promote ascospore release of Fusarium spp. and Zymoseptoria spp. This revealed that Zymoseptoria spp. and Fusarium spp. ascospore release show a positive correlation with relative humidity (%RH). Whereas air temperature (°C) negatively affects Zymoseptoria spp. ascospore release.

Cherry spruce rust causes huge yield losses in Norway spruce seed production in Fennoscandia. The causal agent, Thekopsora areolata, has three types of spores that disperse during spring: basidiospores are produced on basidia that grow out from teliospores in overwintered bird cherry leaf litter to infect new pistillate spruce cones, aeciospores are released from old diseased spruce cones to infect bird cherry leaves, and urediniospores are produced from new bird cherry leaves for reinfection. No study has examined the dispersal of T. areolata spores, including the basidiospores that cause primary infection in spruce cones. In this study, teliospores of T. areolata were germinated in the laboratory and the morphology of basidiospores was described. T. areolata spores were sampled in Ultuna, Sweden and Joutsa, Finland with 21 spore traps at each site. Peaks in aeciospores were observed from 11 to 25 May and from 2 to 8 June at the Finnish site, and from 4 to 18 May at the Swedish site. Urediniospores were first observed 2–3 weeks after the peaks in aeciospores and they were mainly distributed within 10 m from the bird cherry trees. Peaks of 1–2 weeks in basidiospore detection coincided with multiple rain events. The basidiospore peak overlapped with the spruce pollen peak in Finland but not in Sweden. The quantities of basidiospores from different spore traps within 100 m from the spore source had no gradient. Information on spatial and temporal spore release is important for making decisions on disease management strategies. [ABSTRACT FROM AUTHOR]

Pestalotiopsis blight and leaf spot in macadamia are caused by species of Pestalotiopsis and Neopestalotiopsis. Conidia produced by these pathogens are mostly dispersed by wind and rain splash and are the primary infecting propagules. Rapid detection and quantification of airborne conidia are essential to determine the seasonal dynamics of inoculum and critical infection periods in macadamia orchards. A quantitative PCR (qPCR) using a new primer pair and a hydrolysis probe was developed to detect and quantify airborne conidia of Pestalotiopsis and Neopestalotiopsis species. The amplification efficiency of the qPCR was 96.4% and the assay had a limit of detection of 400 fg of Pestalotiopsis/Neopestalotiopsis gDNA, which corresponds to approximately 10 conidia. The qPCR assay coupled with spore trapping was used to monitor airborne conidia dispersal under field conditions. The results showed that the assay is specific and sensitive for detecting and quantifying Pestalotiopsis/Neopestalotiopsis conidia in macadamia orchards. The detection and quantification of the pathogen inoculum will improve our understanding of disease epidemiology and the ability to manage these diseases in macadamia. [ABSTRACT FROM AUTHOR]

Cercospora beticola is one of the most important fungal pathogens of sugar beet, causing cercospora leaf spot (CLS) disease. Due to the decreasing efficacy of various fungicides caused by resistance traits, the development of a sustainable disease management strategy has become more important. Therefore, detailed knowledge about the epidemiology of the pathogen is crucial. Until now, little was known about the spatiotemporal dispersal of C. beticola spores from the primary inoculum source. Rapid detection of C. beticola spores could facilitate a more precise and targeted disease control. Therefore, a TaqMan real‐time PCR assay for detection and quantification of C. beticola spores caught with Rotorod spore traps was established. In 2016 and 2017, field trials were conducted to monitor C. beticola aerial spore dispersal and disease development within an inoculated field and in the adjacent noninoculated area. With the established detection method, C. beticola spores were successfully quantified and used as a measure for aerial spore dispersal intensity. The analysis of the spatiotemporal spread of C. beticola spores revealed a delay and decrease of aerial spore dispersal with increasing distance from the inoculated area. Consequently, disease incidence and severity were reduced in a similar manner. These results imply that spore dispersal occurs mainly on a small scale within a field, although long distances can be overcome by C. beticola spores. Moreover, secondary aerial spore dispersal from sporulating leaf spots seems to be the main driver for CLS disease development. These results provide an important basis for further improvement of CLS control strategies. [ABSTRACT FROM AUTHOR]

Forest diseases caused by invasive fungal pathogens are becoming more Fungicommon, sometimes with dramatic consequences to forest ecosystems. The development of early detection systems is necessary for efficient surveillance anc! to mitigate the impact of invasive pathogens. Windbome spores are an important pathway for introduction of fungal pathogens into new areas; the design of spore trapping devices adapted to forests, capable of collecting different types of spores, and aligned with development of efficient molecular methods for detection of the pathogen, should help forest managers anticipate new disease outbreaks. Two types of Rotorod samplers were evaluated for the collection of airborne inoculum of forest fungal pathogens with a range of spore sizes in five forest types. Detection was by specific quantitative PCR qPCR) and by high-throughput sequencing (HTS) of amplified internal transcribed spacer sequences using a new bioinformatic pipeline. Fungicommon, Search, developed for diagnostic purposes. Validation of the pipeline was conducted on mock communities of 10 fungal species belonging to di fferent taxa, Although the sensitivity of the new HTS pipeline was lower than the specific qPCR, it was able to detect a wide variety of fungal pathogens. FungiSearch is easy to use. and the reference database is updatable, making the tool suitable for rapid identification of new pathogens. This new approach combining spore trapping and HTS detection is promising as a diagnostic tool for invasive fungal pathogens. [ABSTRACT FROM AUTHOR]

An increasing number of researchers are looking to understand the factors affecting microbial dispersion but are often limited by the costs of commercially available air samplers. Some have reduced these costs by designing self-made versions; however, there are no published sampler designs, and there is limited information provided on the actual construction process. Lack of appropriate reference material limits the use of these self-made samplers by many researchers. This manuscript provides a guide to designing and constructing rotating-arm impaction air samplers by covering (i) environmental considerations, (ii) construction materials and equipment, (iii) the construction process, and (iv) air sampler deployment. Information regarding how to calculate rotational velocity, motor speed, and power supply requirements and to troubleshoot common issues is presented in an approachable format for individuals without experience in electronics or machining. Although many of the components discussed in this guide may change in their availability or be updated over time, this document is intended to serve as a "builder's guide" for future research into air sampling technology for phytopathology research., Competing Interests: The author(s) declare no conflict of interest.

Grapevine downy mildew (DM) is caused by the dimorphic oomycete Plasmopara viticola, which incites epidemics through primary and secondary infection cycles that occur throughout the season. The secondary infection cycles are caused by the sporangia produced on DM lesions. The current research examined the relationship between numbers of airborne sporangia and DM development on grape leaves to determine whether spore sampler data can be useful to predict the potential for secondary infections of P. viticola. Three years (2015–2017) of spore sampler data confirmed that sporangia are a common component of the airborne microflora in a DM-infested vineyard and that their numbers depend on weather conditions. For a total of 108 days, leaf samples were collected from the vineyard at 2- to 3-day intervals and incubated under optimal conditions for P. viticola infection. The numbers of airborne sporangia sampled on 1 to 7 days before leaf sampling were significantly correlated with the numbers of DM lesions on the leaves. The best correlation (r=0.59), however, was found for the numbers of viable airborne sporangia (SPV), which were assessed by using equations driven by the vapour pressure deficit. In Bayesian and ROC curve analyses, SPV was found to be a good predictor of P. viticola infection of grape leaves, with AUROC=0.821 and false positive predictions mainly occurring at low SPV. A binary logistic regression showed that a threshold of 2.52 viable sporangia m-3 air day-1 enables a prediction of no infection with a posterior probability of 0.870, which was higher than the prior probability of 0.574. Numbers of viable sporangia in the vineyard air is therefore a useful predictor of infection and especially of no infection. The predictor missed some observed infections, but these infections were not severe (they accounted for only 31 of 374 DM lesions).

Forest disease management relies principally on a preventive approach in which epidemiological surveillance plays a crucial role. However, efficient and cost‐effective surveillance methods are not currently available for large spatial scales. Nevertheless, aerobiological networks have been set up for several decades in many countries to monitor pollen dispersal and provide real‐time assessments of allergenic risk. Here, we suggest that the same approach could be used for the surveillance of forest pathogens. Using molecular methods, we analysed samples from 12 sites of the French aerobiological network, at different dates. Both metabarcoding by high‐throughput sequencing (using two markers and two different bioinformatics approaches) and real‐time PCR targeting eight important forest pathogens were conducted. To validate the approach, temporal and spatial trends of spore detection were compared with field disease data. The metabarcoding approach demonstrated that many fungal plant pathogens could be found in aerobiological samples. Moreover, five of the eight targeted forest pathogens were detected by real‐time PCR, with temporal and spatial trends of spore capture consistent with field data. In particular, Hymenoscyphus fraxineus was detected at high frequency in aerobiological samples in the areas where ash dieback has been present for the longest period of time, and at lower frequency in areas with more recent invasion. Spore detection of seasonal pathogens showed a temporal pattern similar to that of disease reports. Overall, our study provides a proof of concept that permanent aerobiological networks combined with molecular methods may provide a useful tool for large‐scale surveillance of forest pathogens. [ABSTRACT FROM AUTHOR]

The airborne mycobiota has been understudied in comparison with the mycobiota present in other agricultural environments. Traditional, culture-based methods allow the study of a small fraction of the organisms present in the atmosphere, thus missing important information. In this study, the aerial mycobiota in a rice paddy has been examined during the cropping season (from June to September 2016) using qPCRs for two important rice pathogens (Pyricularia oryzae and Bipolaris oryzae) and by using DNA metabarcoding of the fungal ITS region. The metabarcoding results demonstrated a higher alpha diversity (Shannon--Wiener diversity index H' and total number of observed species) at the beginning of the trial (June), suggesting a higher level of community complexity, compared with the end of the season. The main taxa identified by HTS analysis showed a shift in their relative abundance that drove the cluster separation as a function of time and temperature. The most abundant OTUs corresponded to genera such as Cladosporium, Alternaria, Myrothecium, or Pyricularia. Changes in the mycobiota composition were clearly dependent on the average air temperature with a potential impact on disease development in rice. In parallel, oligotyping analysis was performed to obtain a sub-OTU identification which revealed the presence of several oligotypes of Pyricularia and Bipolaris with relative abundance changing during monitoring. [ABSTRACT FROM AUTHOR]

The seasonal abundance of Diatrypaceae spores was studied in southern California's desert table grape-growing region of Coachella Valley. Glass microscope slides covered with petroleum jelly were placed in a mature cv. Mid-Night Beauty® vineyard, and collected weekly from September 2006 to May 2009 (1400 samples for 140 consecutive weeks). Overall, Diatrypaceae-like colonies were recorded from Petri plates after processing spore traps in 93 (66%) of 140 weeks. Phylogenetic analyses showed Eutypella citricola and Eutypella microtheca to be the Diatrypaceae spp. captured from the spore traps. Though spores were captured throughout each year, their incidence varied among the different seasons. The greatest number of Eutypella spores were captured in autumn (38.7% of the total) followed by winter (30.6%), summer (19.7%), and spring (11%). The greatest numbers of spores were captured in October each year (15.7%) and least in June (1%). Eutypella spore release was correlated with rainfall only in 26 (28%) of the 93 weeks that spores were captured during the study. Analysis of diseased samples collected from the cv. Mid-Night Beauty® vineyard showed that E. citricola, E. microtheca and E. scoparia were the most prevalent fungi isolated from cankers. A pathogenicity study showed that E. citricola and E. microtheca isolates collected from spore traps caused larger vascular necrosis than the non-inoculated controls, indicating their role as pathogens on grapevines. This study has demonstrated Eutypella spp. to play an important role in grapevine health under desert growing conditions of southern California. In addition, and contrary to what it is largely accepted, results from this research suggest that Diatrypaceae spore release can occur in the absence of precipitation. This study expands current knowledge on epidemiology of Diatrypaceae spp. other than Eutypa lata, and provides important information for enhancing control strategies against grapevine trunk diseases under desert growing conditions. [ABSTRACT FROM AUTHOR]

Citrus black spot, caused by Phyllosticta citricarpa, is characterized by fruit blemishes and premature fruit drop, resulting in significant economic losses in summer rainfall areas. The pathogen forms both conidia and ascospores during its life cycle. However, the occurrence of these spores and their contributions to infection of fruit in field conditions are not well understood. Our research using direct leaf litter monitoring and volumetric spore trapping in Queensland orchards revealed that pseudothecia and ascospores in leaf litter as well as trapped ascospores had low abundance, while pycnidia and conidia were highly abundant. Both P. citricarpa and endophytic Phyllosticta spp. were identified, with P. citricarpa being dominant. In replicated field trials, we determined that infection of Imperial mandarin fruit by P. citricarpa occurred from fruit set until week 20 of fruit development, with the key infection events taking place between weeks 4 and 16 in Queensland subtropical conditions. These results demonstrate that protecting fruit during weeks 4 to 16 significantly reduced P. citricarpa infection. We found no significant correlation between the disease incidence in fruit and P. citricarpa conidial abundance in leaf litter or ascospore abundance measured by volumetric spore trapping. Therefore, it is suggested that inoculum sources in the tree canopy other than those detected by spore trapping and direct leaf litter monitoring may play a major role in the epidemiology of citrus black spot. Improved knowledge regarding epidemiology of P. citricarpa and an understanding of propagules causing infection may aid in development of more effective disease management strategies. [ABSTRACT FROM AUTHOR]

Grapevine downy mildew (DM) is caused by the dimorphic oomycete Plasmopara viticola , which incites epidemics through primary and secondary infection cycles that occur throughout the season. The secondary infection cycles are caused by the sporangia produced on DM lesions. The current research examined the relationship between numbers of airborne sporangia and DM development on grape leaves to determine whether spore sampler data can be useful to predict the potential for secondary infections of P. viticola. Three years (2015–2017) of spore sampler data confirmed that sporangia are a common component of the airborne microflora in a DM-infested vineyard and that their numbers depend on weather conditions. For a total of 108 days, leaf samples were collected from the vineyard at 2- to 3-day intervals and incubated under optimal conditions for P. viticola infection. The numbers of airborne sporangia sampled on 1 to 7 days before leaf sampling were significantly correlated with the numbers of DM lesions on the leaves. The best correlation (r=0.59), however, was found for the numbers of viable airborne sporangia (SPV), which were assessed by using equations driven by the vapour pressure deficit. In Bayesian and ROC curve analyses, SPV was found to be a good predictor of P. viticola infection of grape leaves, with AUROC=0.821 and false positive predictions mainly occurring at low SPV. A binary logistic regression showed that a threshold of 2.52 viable sporangia m-3 air day-1 enables a prediction of no infection with a posterior probability of 0.870, which was higher than the prior probability of 0.574. Numbers of viable sporangia in the vineyard air is therefore a useful predictor of infection and especially of no infection. The predictor missed some observed infections, but these infections were not severe (they accounted for only 31 of 374 DM lesions). [ABSTRACT FROM AUTHOR]

Plant-associated microorganisms play a crucial role in plant health and productivity. Belowground microbial diversity is widely reported as a major factor in determining the composition of the plant microbiome. In contrast, much less is known about the role of the atmosphere in relation to the plant microbiome. The current study examined the hypothesis that the atmospheric microbiome influences the composition of fungal communities of the aboveground organs (flowers, fruit, and leaves) of table grape and vice versa. The atmosphere surrounding grape plantings exhibited a significantly higher level of fungal diversity relative to the nearby plant organs and shared a higher number of phylotypes (5,536 OTUs, 40.3%) with the plant than between organs of the same plant. Using a Bayesian source tracking approach, plant organs were determined to be the major source of the atmospheric fungal community (92%). In contrast, airborne microbiota had only a minor contribution to the grape microbiome, representing the source of 15, 4, and 35% of the fungal communities of leaves, flowers, and fruits, respectively. Moreover, data indicate that plant organs and the surrounding atmosphere shared a fraction of each other’s fungal communities, and this shared pool of fungal taxa serves as a two-way reservoir of microorganisms. Microbial association analysis highlighted more positive than negative interactions between fungal phylotypes. Positive interactions were more common within the same environment, while negative interactions appeared to occur more frequently between different environments, i.e., atmosphere, leaf, flower, and fruit. The current study revealed the interplay between the fungal communities of the grape phyllosphere with the surrounding air. Plants were identified as a major source of recruitment for the atmospheric microbiome, while the surrounding atmosphere contributed only a small fraction of the plant fungal community. The results of the study suggested that the plant–air interface modulates the plant recruitment of atmospheric fungi, taking a step forward in understanding the plant holobiont assembly and how the atmosphere surrounding plants plays a role in this process. The impact of plants on the atmospheric microbiota has several biological and epidemiological implications for plants and humans.

Botryosphaeria dieback, caused by a group of pathogens in the Botryosphaeriaceae family, is one of the most common grapevine trunk disease complexes (GTDs) found in Oregon vineyards. To understand the period of spores released by Botryosphaeria spp. in Oregon vineyards, four Burkard 7-day recording volumetric spore traps were placed in vineyard blocks in northern and southern Oregon. Each trap was placed near a younger (<10 years) and older (>30 years) block in both regions. Spore traps were deployed at the beginning of December 2019 and continued until March 2021. Between these timeframes, 475 and 477 days of samples were collected from each spore trap in northern and southern Oregon, respectively. DNA extraction was performed from individual day samples and followed by qPCR analysis of Botryosphaeria spores trapped in each tape sample. Weather data such as temperature, precipitation, relative humidity (RH), and wind speed were collected from nearby weather stations. Association between these data and number of spores detected were analyzed using Pearson correlation analysis. In northern Oregon, the detection occurred between December and February, and the first spore detection occurred when cumulative growing degree day (GDD) totaled to 4,357 and 4,351 units (T Base = 0°C, biofix date = January 1) during the first and second seasons, respectively. Similarly, in southern Oregon, the detection occurred between November and January, and the first spore detection occurred when cumulative GDD was 4,405 units during the second season. Hours of continuous RH >86% was significantly associated with number of spores released ( P = 0.026; r = 0 . 42). During the spore detected dates, the RH was >86% for at least 19 consecutive hours. These findings provide an important implication to manage Botryosphaeria dieback by protecting pruning wounds during the most-spore-released periods. Furthermore, the knowledge of weather variables and their possible association with spore detection provides important information towards developing predictive models in future studies., Competing Interests: The author(s) declare no conflict of interest.

Plant-associated microorganisms play a crucial role in plant health and productivity. Belowground microbial diversity is widely reported as a major factor in determining the composition of the plant microbiome. In contrast, much less is known about the role of the atmosphere in relation to the plant microbiome. The current study examined the hypothesis that the atmospheric microbiome influences the composition of fungal communities of the aboveground organs (flowers, fruit, and leaves) of table grape and vice versa. The atmosphere surrounding grape plantings exhibited a significantly higher level of fungal diversity relative to the nearby plant organs and shared a higher number of phylotypes (5,536 OTUs, 40.3%) with the plant than between organs of the same plant. Using a Bayesian source tracking approach, plant organs were determined to be the major source of the atmospheric fungal community (92%). In contrast, airborne microbiota had only a minor contribution to the grape microbiome, representing the source of 15, 4, and 35% of the fungal communities of leaves, flowers, and fruits, respectively. Moreover, data indicate that plant organs and the surrounding atmosphere shared a fraction of each other's fungal communities, and this shared pool of fungal taxa serves as a two-way reservoir of microorganisms. Microbial association analysis highlighted more positive than negative interactions between fungal phylotypes. Positive interactions were more common within the same environment, while negative interactions appeared to occur more frequently between different environments, i.e., atmosphere, leaf, flower, and fruit. The current study revealed the interplay between the fungal communities of the grape phyllosphere with the surrounding air. Plants were identified as a major source of recruitment for the atmospheric microbiome, while the surrounding atmosphere contributed only a small fraction of the plant fungal community. The results of the study suggested that the plant–air interface modulates the plant recruitment of atmospheric fungi, taking a step forward in understanding the plant holobiont assembly and how the atmosphere surrounding plants plays a role in this process. The impact of plants on the atmospheric microbiota has several biological and epidemiological implications for plants and humans. [ABSTRACT FROM AUTHOR]

With the aim of unravelling the role of airborne Fusarium graminearum inoculum in the epidemic of Fusarium head blight (FHB) caused by this species in wheat spikes, a network of Burkard air samplers was set up in five wheat fields distributed in Belgium from 2011 to 2013. Each year from April to July, the daily amounts of F. graminearum inoculum above the wheat canopy were quantified using a newly developed TaqMan qPCR assay. The pattern of spore trapping observed was drastically different per year and per location with a frequency of detection between 9 and 66% and a mean daily concentration between 0.8 and 10.2 conidia-equivalent/m3. In one location, air was sampled for a whole year. Inoculum was frequently detected from the wheat stem elongation stage until the end of the harvesting period, but high inoculum levels were also observed during the fall. Using a window-pane analysis, different periods of time around wheat flowering (varying in length and starting date) were investigated for their importance in the relation between airborne inoculum and FHB parameters (FHB severity, frequency of F. graminearum infection and DON). For almost all the combinations of variables, strong and significant correlations were found for multiple window lengths and starting times. Inoculum quantities trapped around flowering were highly correlated with F. graminearum infection (up to R = 0.84) and DON (up to R = 0.9). Frequencies of detection were also well correlated with both of these parameters. DON concentrations at harvest could even be significantly associated with the F. graminearum inoculum trapped during periods finishing before the beginning of the anthesis (R = 0.77). Overall, these results highlight the key role of the airborne inoculum in F. graminearum epidemics and underline the importance of monitoring it for the development of disease forecasting tools. [ABSTRACT FROM AUTHOR]

The airborne inoculum of Fusarium circinatum Nirenberg & O'Donnell, the fungal pathogen causing Pine Pitch Canker (PPC), is one of the main means of spread of the disease in forest stands and forest nurseries. Since this world-wide known pathogen was introduced in Europe, its biology in this newly infested area still remains scarcely known. To shedmore light on this topic, we set up an experiment on a naturally PPC infested forest of Monterey pine in Galicia (NWSpain) with the following two goals: (i) to describe the seasonal spore dispersal pattern during one year of regular sampling and (ii) to assess the spatial spore dispersal pattern around the infested plot. Portable rotating arm spore traps were used and complemented with meteorologicalmeasurements. The abundance of F. circinatum spores in the samples was assessed by quantitative PCR (qPCR) with a hydrolysis probe. The results showed almost permanent occurrence of the air inoculum throughout the whole year, being detected in 27 of the 30 samplings. No clear temporal trends were observed, but a higher air inoculum was favoured by previous lower air temperatures and lower leaf wetness. Conversely, neither rainfall nor air humidity seemed to have any significant importance. The spatial spread of the inoculumwas noted to be successful up to a distance of 1000 m in the wind direction, even with winds of just 5 m·s-1. Our study shows that rotating arm spore traps combined with qPCR may be an efficient tool for F. circinatum detection. [ABSTRACT FROM AUTHOR]

We investigated the ability of Hymenoscyphus fraxineus mitovirus 1 (HfMV1) to spread in nature through the airborne inoculum of its fungal host, the devastatingHymenoscyphus fraxineus. Eight rotating-arm spore traps were installed to collect the airborne spores (presumably ascospores) ofH. fraxineusin four ecologically different ash forests in the Czech Republic. Spore trap samples were subjected to direct DNA and RNA extractions and subsequent qPCR and mitovirus-specific reverse transcription (RT) qPCR for detection and quantification ofH. fraxineusand HfMV1, respectively. The prevalence of HfMV1 was confirmed in 57% ofH. fraxineussampled inocula, showing that its occurrence is significantly correlated with its fungal hostH. fraxineus. Furthermore, the ratio of mitovirus carrying spores was shown to be constant at any level ofH. fraxineusinfection pressure independently of the studied locality. Temperatures of 19–21°C at the time of spore trapping correlated with the highest levels of both HfMV1 andH. fraxineus. Our data confirmH. fraxineusairborne inoculum as an efficient transport means for HfMV1 in nature. Moreover, this research uses a new technology to directly study a mycovirus through its host aerospora. [ABSTRACT FROM PUBLISHER]

In this study, a real-time PCR assay was established to quantify airborne inoculum of Blumeria graminis f. s p. tritici ( Bgt), the causal agent of wheat powdery mildew. Burkard 7-day recording spore traps were used to collect Bgt spores in air in Langfang City, Hebei Province, China. A linear relationship between number of spores counted with a compound microscope and those determined with the real-time PCR assay was obtained in artificially inoculated spore trap tape sections. The dynamics of airborne conidia concentrations of Bgt were similar. There were significant correlations ( P < 0.01) between spore concentrations by using a compound microscope and the real-time PCR assay in three growing seasons. The results demonstrated a potential method to quantitatively determine spore inoculum potential in wheat fields by using a real-time PCR assay. [ABSTRACT FROM AUTHOR]

To elucidate the early epidemic stages of septoria tritici blotch, especially the relationship between the onset of epidemics, the local availability of primary inoculum, and the presence of wheat debris, the early disease dynamics and airborne concentration in Zymoseptoria tritici ascospores were concomitantly assessed at a small spatiotemporal scale and over two years, using spore traps coupled with a qPCR assay. One plot, with the crop debris left, provided a local source of primary inoculum, while the other plot, without debris, lacked any. According to the assay's limits of detection, daily spore trap samples were classified as not detectable or not quantifiable, detectable, and quantifiable. The proportions of samples assigned to the different classes and numbers of spores in samples classified as quantifiable were significantly different between years, time periods (from November to March) and spore trap location (field with or without debris). The effect of year on the airborne ascospore concentration was high: 22 daily peaks with more than 230 ascospores m−3 day−1 were identified in the autumn of 2012/13, but none in the autumn of 2011/12. The local presence of wheat debris had no obvious effect on the amount of airborne ascospores or on the earliness of the two epidemics, especially in the year with high inoculum pressure (2012/13). These results suggest that the amount of primary airborne inoculum available in a wheat crop is not a limiting factor for the onset of an epidemic. [ABSTRACT FROM AUTHOR]

Hymenoscyphus fraxineus is an invasive fungal species causing the most serious disease of ashes (Fraxinus spp.) in Europe-ash dieback. The biology of this fungus is not totally elucidated, neither its relation to the saprophytic species Hymenoscyphus albidus, native in Europe. Our study is focused on the description of seasonal spore dispersal of both fungi and its relation to meteorological conditions, which is needed for more precise and effective control of the disease. For this experiment one long time infected mixed forest in the SE Czech Republic was chosen. A seven-day automatic volumetric spore trap and a weather station were installed to continuously sample the aerospora from April to October 2014. In seven periods a rotating arm spore trap was also used to obtain 48-h air samples to compare the efficiency of these two types of air samplers. Air samples were evaluated solely by qPCR with a very low detection limit. Results show co-occurrence of inoculum of both fungi throughout the entire sampling period with peak levels in August. The origin of the inoculum sampled in the periods without apothecia is discussed. Air-inoculum occurrence of both fungi is significantly correlated with each other, suggesting their coexistence in this forest. [ABSTRACT FROM AUTHOR]

Monitoring airborne inoculum is gaining interest as a potential means of giving growers an earlier warning of disease risk in a management unit or region. This information is sought by growers to aid in adapting to changes in the management tools at their disposal and the market-driven need to reduce the use of fungicides and cost of production. To effectively use inoculum monitoring as a decision aid, there is an increasing need to understand the physics of particle transport in managed and natural plant canopies to effectively deploy and use near-ground aerial inoculum data. This understanding, combined with the nuances of pathogen-specific biology and disease epidemiology, can serve as a guide to designing improved monitoring approaches. The complexity of any pathosystem and local environment are such that there is not a generalized approach to near-ground air sampler placement, but there is a conceptual framework to arrive at a "semi-optimal" solution based on available resources. This review is intended as a brief synopsis of the linkages among pathogen biology, disease epidemiology, and the physics of the aerial dispersion of pathogen inoculum and what to consider when deciding where to locate ground-based air samplers. We leverage prior work in developing airborne monitoring tools for hops, grapes, spinach, and turf, and research into the fluid mechanics governing particle transport in sparse canopies and urban and forest environments. We present simulation studies to demonstrate how particles move in the complex environments of agricultural fields and to illustrate the limited sampling area of common air samplers.

The aim of this paper was to study the effect of potassium phosphite and potassium silicate, alone or in combination with fungicide, for the control of asian soybean rust. The cultivar CD 214RR was used, with 0.5 m row spacing and 15 plants per linear meter. It was used the spore trap Siga to detect the rust spores in the air before the effective infection on the plants, doing preventive applications for the products that were tested. The experimental design was in randomized block with eight treatments and four replications. The treatments were: fungicide; fungicide + potassium phosphite; fungicide + potassium silicate; fungicide + potassium phosphite + potassium silicate; potassium phosphite; potassium silicate; potassium phosphite + potassium silicate; and untreated control. The dosage of the fungicide was 0.3 L ha-1 (cyproconazol + azoxistrobin), and the dosage of phosphite and silicate of potassium was 2 L ha-1. The spore trap Siga detects spores of Phakopsora pachyrhizi in the air before the symptoms. Applications of the products were made three days after the detection of the spores. It was verified that the use of fungicide reduced the severity of the disease, the apparent infection rate of the asian soybean rust, the defoliation and productivity losses, independently of the association with potassium phosphite and/or potassium silicate. The association of potassium phosphite and potassium silicate reduced the rust severity at the end of the development of the soybean crop, in relation to the untreated control, not providing, however, reduction on the losses of productivity. [ABSTRACT FROM AUTHOR]

A method based on real-time polymerase chain reaction ( PCR) and the use of rotating-arm spore traps was developed for quantifying airborne Hymenoscyphus pseudoalbidus ascospores. The method was sensitive and reproducible, and the collection efficiency was 10% of the spores present in the air. The temporal ascospore dispersal pattern was studied over 3 years by collecting spores every 15 days for a 24 h air-sampling period during the ash-growing season. The highest production was detected from the end of June to the beginning of September. The overall ascospore production did not differ significantly among stands within a specific year but there were differences from year to year. There was a positive correlation between air temperature and the number of ascospores trapped, with most of the positive samples being observed at temperatures above 12°C. The vertical profile of ascospore dispersal showed a strong decrease in ascospore density within a height of 3 m, regardless of date of collection. An analysis of the spore traps installed at increasing distances from an infected stand showed that most of the ascospores were deposited downwind within 50 m of the stand. These data are discussed in context of the epidemiology of the disease. [ABSTRACT FROM AUTHOR]

Airborne sexual spores of Leptosphaeria maculans and L. biglobosa are primary inoculum of fungi causing phoma stem canker of oilseed rape. In this study ascospore release of these two species was compared between Lithuania (of autumn 2010 and 2011) and Poland (autumns of 2009 till 2012), using identical equipment and methods. Dynamics of L. maculans and L. biglobosa ascospore dispersal was investigated using volumetric samplers followed by microscopic and molecular methods of detection. In Lithuania, the earliest detection of Leptosphaeria ascospores, using microscopy was on 1st September (2010) and on 2nd September (2011), whereas in Poland the earliest detection was on 5th September (2009 and 2010) and the latest on 11th October (2011), which demonstrates that differences between the seasons may exceed a month (36 days). In the case of molecular detection performed for the Polish samples, the dates of the earliest and the latest release of the first ascospores ranged from 5th September to 15th October (40 days). The number of days with ascospores detected in the air samples ranged from 6 (2012) to 25 (2009). At earliest, the detection of the highest concentration of ascospores was on 6th September (2010) and the latest - on 28th October (2009). These results demonstrate big differences between the results of spore monitoring between the years and the importance of aerobiological studies to follow the development of the pathogen in natural infection. The fluctuations in quantities of spores and fungal deoxyribonucleic acid (DNA) strongly depended on the weather conditions, mainly rainfall, air temperature and relative humidity. Ascospore release was observed immediately after the rainfall events, but not later than two days after the rainfall. On days without rain, spores were present in the air samples when average relative air humidity exceeded 90% and day temperature was below 15°C. In 2009, molecular detection revealed the presence of both fungal species in the air samples, whereas in 2010-2012 only L. biglobosa DNA was found. Molecular detection with a quantitative real-time polymerase chain reaction (qPCR) was identified as a fast and accurate method of pathogen detection and identification from air samples. The highest coefficient of correlation between microscopic and molecular detection of Leptosphaeria species was 0.828 (2009) and the lowest was 0.483 (2011); in all years both techniques of detection were statistically correlated. We have demonstrated that the correlation coefficient highly depended on the number of ascospores in air samples. Based on the studies performed in Lithuania and Poland, we have shown that climatic conditions in the northern part of Central Europe are favourable for the spread of Leptosphaeria spp. by ascospores; however, the patterns of ascospore release greatly differ between the both countries. [ABSTRACT FROM AUTHOR]