It's a Trap! Part I: Exploring the Applications of Rotating-Arm Impaction Samplers in Plant Pathology.

Although improved knowledge on the movement of airborne plant pathogens is likely to benefit plant health management, generating this knowledge is often far more complicated than anticipated. This complexity is driven by the dynamic nature of environmental variables, diversity among pathosystems that are targeted, and the unique needs of each research group. When using a rotating-arm impaction sampler, particle collection is dependent on the pathogen, environment, research objectives, and limitations (monetary, environmental, or labor). Consequently, no design will result in 100% collection efficiency. Fortunately, it is likely that multiple approaches can succeed despite these constraints. Choices made during design and implementation of samplers can influence the results, and recognizing this influence is crucial for researchers. This article is for beginners in the art and science of using rotating-arm impaction samplers; it provides a foundation for designing a project, from planning the experiment to processing samples. We present a relatively nontechnical discussion of the factors influencing pathogen dispersal and how placement of the rotating-arm air samplers alters propagule capture. We include a discussion of applications of rotating-arm air samplers to demonstrate their versatility and potential in plant pathology research as well as their limitations.

Seasonal periodicity of the airborne spores of fungi causing grapevine trunk diseases: an analysis of 247 studies published worldwide.

Grapevine trunk diseases (GTDs) are among the most devastating grapevine diseases globally. GTDs are caused by multiple fungi from various taxa, which release spores into the vineyard and infect wood tissue, mainly through wounds caused by viticultural operations. The timing of operations to avoid infection is critical concerning the periodicity of GTD spores in vineyards, and many studies have been conducted in different grape-growing areas worldwide. However, these studies provide conflicting and fragmented information. To synthesize current knowledge, we performed a systematic literature review, extracted quantitative data from published papers, and used these data to identify trends and knowledge gaps to be addressed in future studies. Our database included 26 papers covering 247 studies and 3,529 spore sampling records concerning a total of 29 fungal taxa responsible for Botryosphaeria dieback (BD), Esca complex (EC), and Eutypa dieback (ED). We found a clear seasonality in the presence and abundance of BD spores, with a peak from fall to spring, more in the northern than in the southern hemisphere, but not for EC and ED. Spores of these fungi were present throughout the growing season in both hemispheres, possibly due to higher variability in spore types, sporulation conditions, and spore release mechanisms in EC and ED fungi compared to BD. Our analysis has limitations due to knowledge gaps and data availability for some fungi (e.g., Basidiomycetes, causing EC). These limitations are discussed to facilitate further research.

Influence of climatic factors on dry flower, grey and green mould diseases of macadamia flowers in Australia.

AIMS: Flower blights (grey mould, green mould and dry flower) are important diseases of macadamia. Lack of information on pathogen biology and disease epidemiology in macadamia has hampered control options. Effects of climatic variables including temperature, relative humidity (RH) and vapour pressure deficit (VPD) on the abundance, germination and growth of conidia of four fungal pathogens that cause various flower blights in macadamia were studied. METHODS AND RESULTS: Mycelial growth, sporulation, conidial germination and germ tube growth for five isolates each of Botrytis cinerea, Cladosporium cladosporioides, Pestalotiopsis macadamiae and Neopestalotiopsis macadamiae, at eight temperatures, seven RH and the corresponding VPD regimes were determined in vitro. The optimal climatic range of each of the four pathogens was validated during macadamia flowering periods in the 2019 and 2020 seasons by conidia detected and quantified using quantitative PCR. Several growth models were fitted to the data with high significance; predicted optima from these models ranged from 0.9 to 1.1 kPa VPD for P. macadamiae and N. macadamiae and <0.6 kPa VPD for B. cinerea and C. cladosporioides. CONCLUSIONS: This study showed that VPD, as a determinant of the fecundity and growth of the four fungal pathogens, was predictive of flower blight incidence in macadamia. The importance of temperature, RH and, thus, VPD for defining the conditions for infection and flower blight epidemics was established. SIGNIFICANCE AND IMPACT OF THE STUDY: This information provides a firm basis for the development of prediction tools for flower blights in macadamia.

The Role of Ascospores and Conidia, in Relation to Weather Variables, in the Epidemiology of Stemphylium Leaf Blight of Onion.

Stemphylium leaf blight (SLB), caused by Stemphylium vesicarium, is an important foliar disease of onion in northeastern North America. The pathogen produces conidia and ascospores, but the relative contributions of these spore types to epidemics in onion is not known. Determining the abundance of ascospores and conidia during the growing season could contribute to a disease forecasting model. Airborne ascospores and conidia of S. vesicarium were trapped during the growing season of 2015 and 2016 at an onion trial in southern Ontario, Canada, using a Burkard 7-day volumetric sampler. Meteorological data were recorded hourly. Ascospore numbers peaked before the crop was planted and declined rapidly with time and at daily mean air temperatures >15°C. Conidia were present throughout the growing season and appear to be closely related to the development of SLB on onion. Daily spore concentrations were variable, but 59 to 73% of ascospores and approximately 60% of conidia were captured between 0600 and 1200 h. Spore concentrations increased 24 to 72 h after rainfall and precipitation and leaf wetness duration were consistently and positively associated with increases in numbers of conidia and subsequent SLB incidence. The first symptoms of SLB coincided with high numbers of conidia, rainfall, leaf wetness duration ≥8 h, and days with average daily temperature ≥18°C. The number of airborne ascospores was very low by the time SLB symptoms were observed. Ascospores may initiate infection on alternative hosts in early spring, while conidia are the most important inoculum for the epidemic on onions.

Effect of Climatic Variables on Abundance and Dispersal of Lecanosticta acicola Spores and Their Impact on Defoliation on Eastern White Pine.

The disease complex white pine needle damage (WPND), first reported in 2006, has now escalated to an epidemic state across the northeastern United States. Although this complex is composed of several fungal species, Lecanosticta acicola is considered to be the primary causal agent. Knowledge regarding the epidemiology, specific climatic factors that affect the spread of L. acicola on eastern white pine (Pinus strobus) in natural forest settings, and potential risks repeated defoliation may have on tree health is limited. Therefore, this study examined how climatic variables affect the abundance and distance of spore dispersal of L. acicola and compared litterfall caused by defoliation versus natural needle abscission. Conidia were observed on spore traps from May through August, with a peak in abundance occurring in June, corresponding to the defoliation of second- and third-year foliage measured in litter traps. During peak spore production, relative humidity and the occurrence of rainfall was found to have the greatest influence on spore abundance. Our results will aid managers in determining how far from infected trees natural regeneration will likely be affected and predicting future disease severity based on climatic conditions.

New species of Phaeomoniellales from a German vineyard and their potential threat to grapevine (Vitis vinifera) health.

Recently, the order Phaeomoniellales was established that includes fungi closely related to Phaeomoniella chlamydospora, a phytopathogen assumed to be the main causal agent of the two most destructive grapevine trunk diseases, Petri disease and esca. Other species of this order are reported as pathogens of other economically important crops, like olive, peach, apricot, cherry, plum, rambutan, lichee or langsat. However, they are rarely isolated and hence, little is known about their ecological traits and pathogenicity. During a 1-yr period of spore trapping in a German vineyard divided in minimally and intensively pruned grapevines, 23 fungal strains of the Phaeomoniellales were collected. Based on morphological and molecular (ITS, LSU and tub2) analyses the isolated strains were assigned to eight different species. Two species were identified as P. chlamydospora and Neophaeomoniella zymoides, respectively. The remaining six species displayed morphological and molecular differences to known species of the Phaeomoniellales and are newly described, namely Aequabiliella palatina, Minutiella simplex, Moristroma germanicum, Mo. palatinum, Neophaeomoniella constricta and N. ossiformis. A pathogenicity test conducted in the greenhouse revealed that except for P. chlamydospora, none of the species of the Phaeomoniellales isolated from spore traps is able to induce lesions in grapevine wood.

Aptamer-Based Trapping: Enrichment of Bacillus cereus Spores for Real-Time PCR Detection.

Aptamer-based trapping techniques are in general suitable to replace common antibody-based enrichment approaches. A time-consuming isolation or clean-up is often necessary during sample preparation, e.g. for the detection of spores. For the development of bioanalytical routine approaches, aptamers with a high affinity to B. cereus spores were applied for the establishment and validation of an aptamer-based trapping technique in milk with fat contents between 0.3 and 3.5%. Thereby, enrichment factors of up to sixfold were achieved. The combination of an aptamer-based enrichment by magnetic separation and the subsequent specific real-time PCR detection represents a reliable and rapid detection system.

Food Sensing: Aptamer-Based Trapping of Bacillus cereus Spores with Specific Detection via Real Time PCR in Milk.

Aerobic spores pose serious problems for both food product manufacturers and consumers. Milk is particularly at risk and thus an important issue of preventive consumer protection and quality assurance. The spore-former Bacillus cereus is a food poisoning Gram-positive pathogen which mainly produces two different types of toxins, the diarrhea inducing and the emetic toxins. Reliable and rapid analytical assays for the detection of B. cereus spores are required, which could be achieved by combining in vitro generated aptamers with highly specific molecular biological techniques. For the development of routine bioanalytical approaches, already existing aptamers with high affinity to B. cereus spores have been characterized by surface plasmon resonance (SPR) spectroscopy and fluorescence microscopy in terms of their dissociation constants and selectivity. Dissociation constants in the low nanomolar range (from 5.2 to 52.4 nM) were determined. Subsequently, the characterized aptamers were utilized for the establishment and validation of an aptamer-based trapping technique in both milk simulating buffer and milk with fat contents between 0.3 and 3.5%. Thereby, enrichment factors of up to 6-fold could be achieved. It could be observed that trapping protocol and characterized aptamers were fully adaptable to the application in milk. Due to the fact that aptamer selectivity is limited, a highly specific real time PCR assay was utilized following trapping to gain a higher degree of selectivity.

Identification and Quantification of Ascospores as the Primary Inoculum for Collar Rot of Greenhouse-Produced Tobacco Seedlings.

Collar rot, caused by Sclerotinia sclerotiorum, is a severe disease of tobacco seedlings grown in greenhouses. A semiselective medium was adapted and used to detect the presence and quantity of ascospores in commercial greenhouses. Petri dishes of the semiselective medium were placed inside and outside of greenhouses in four counties during the transplant production period in 1995 and 1996. Ascospores were present throughout the production period each year (February to April) and were confirmed to be the primary inoculum for the disease. Significant differences were observed in the number of ascospores trapped within and between counties. Peak numbers of ascospores were trapped between 10 and 12 a.m., and higher numbers of ascospores were trapped outside than inside houses. In general, distribution of ascospores inside houses was uniform unless a high concentration of apothecia was present very close to one section of the greenhouse. The semiselective medium and trapping technique used in this study may allow development of a forecasting system for collar rot of tobacco based on the presence and level of pathogen inoculum.

Effects of Shredding or Treating Apple Leaf Litter with Urea on Ascospore Dose of Venturia inaequalis and Disease Buildup.

Ascospores produced on diseased leaves in the leaf litter constitute the primary inoculum causing scab in commercial apple orchards in the northeastern United States. Two sanitation practices, shredding the leaf litter with a flail mower and urea application, were evaluated for their potential to reduce the risk of primary scab. Three measures of a treatment's potential were made: leaf litter density was evaluated to determine reduction of the source or primary inoculum; trapped ascospores were counted to measure the reduction of primary inoculum; and scab lesions on trees and seedlings were counted to determine reduction in disease incidence and severity on leaves and fruit. The results show that in the northeastern United States, shredding the leaf litter in November or April will reduce the risk of scab by 80 to 90% if all of the leaf litter is shredded. If 10 to 35% of the leaf litter cannot be shredded because of the limited offset of the flail mower and spread of the tree canopy, then the risk of scab is reduced by 50 to 65%. Urea applied to the leaf litter in November (when approximately 95% of the leaves had fallen) reduced the number of ascospores trapped by 50%. Urea applied to the leaf litter in April (before bud break) reduced the number of ascospores trapped by 66%. The reductions are discussed in relation to a sanitation action threshold that links sanitation to a strategy that reduces the fungicide dose to control primary scab in an orchard assessed with a moderate amount of foliar scab the previous autumn.