scholarly journals Molecular Methods to Detect and Quantify Botryosphaeriaceae Inocula Associated With Grapevine Dieback in Australia

Plant Disease ◽  
2018 ◽  
Vol 102 (8) ◽  
pp. 1489-1499 ◽  
Author(s):  
Regina Billones-Baaijens ◽  
José Ramón Úrbez-Torres ◽  
Meifang Liu ◽  
Matthew Ayres ◽  
Mark Sosnowski ◽  
...  
Keyword(s):  
Fungal Species ◽  
Molecular Tools ◽  
Spore Dispersal ◽  
Dispersal Patterns ◽  
Spore Trap ◽  
Synthetic Dna ◽  
Qpcr Method ◽  
Botryosphaeria Dieback ◽  
Grapevine Dieback ◽  
Rain Splash

Botryosphaeria dieback, caused by species of Botryosphaeriaceae, is an important grapevine trunk disease in Australia. Inocula produced by the pathogens are primarily dispersed by rain splash and wind and infect pruning wounds leading to cankers, dieback, and eventually death of vines. The objective of this study was to develop molecular tools to detect and quantify Botryosphaeriaceae inocula from the environment. These tools are essential for investigating spore dispersal patterns of Botryosphaeriaceae pathogens in Australian vineyards. DNA extraction protocols were evaluated and one modified protocol was found suitable for extracting Botryosphaeriaceae DNA from artificially and naturally inoculated Burkard volumetric spore sampler tapes. Multispecies primers and a hydrolysis probe for quantitative PCR (qPCR) were further developed to detect and quantify Botryosphaeriaceae inocula from environmental samples. Specificity tests showed that the multispecies primers were able to amplify the DNA of 10 Botryosphaeriaceae species (58 isolates) found in Australia while none of the 27 nontarget fungal species (90 isolates) tested were amplified. The qPCR assay was suitable for amplifying purified DNA, synthetic DNA fragments (gBlocks), and mixed DNA from spore trap tapes. The qPCR method developed in this study was shown to be rapid and sensitive in detecting Botryosphaeriaceae inocula from the environment using spore traps.

scholarly journals Monitoring spore dispersal and early infections of Diplocarpon coronariae causing apple blotch using selected spore traps and a new qPCR method

2021 ◽  
Author(s):  
Clémence Boutry ◽  
Anne Bohr ◽  
Sascha Buchleither ◽  
Mathias Ludwig ◽  
Thomas Oberhänsli ◽  
...  
Keyword(s):  
Crop Protection ◽  
Late Summer ◽  
Early Summer ◽  
Apple Orchard ◽  
Spore Dispersal ◽  
Spore Trap ◽  
Disease Progress ◽  
Marssonina Coronaria ◽  
Qpcr Method ◽  
The Usa

Apple blotch (AB) is a major disease of apples in Asia and recently also emerging in Europe and the USA. It is caused by the fungus Diplocarpon coronariae (Dc) (formerly: Marssonina coronaria; teleomorph: Diplocarpon mali) and leads to severe defoliation of apple trees in late summer and thus to reduced yield and fruit quality. To develop effective crop protection strategies, a sound knowledge of the pathogen's biology is crucial. However, especially data on the early phase of disease development is scarce, and no data on spore dispersal for Europe is available. In this study, we assessed different spore traps for their capacity to capture Dc spores, and we developed a highly sensitive TaqMan qPCR method to quantify Dc conidia in spore trap samples. With these tools, we monitored the temporal and spatial spore dispersal and disease progress in spring and early summer in an extensively managed apple orchard in Switzerland in 2019 and 2020. Our results show that Dc overwinters in leaf litter and that spore dispersal and primary infections occur already in late April and beginning of May. We provide the first results on early-season spore dispersal of Dc, which, combined with the observed disease progress, helps to understand the disease dynamics and improve disease forecast models. Using the new qPCR method, we finally detected Dc in buds, on bark and on fruit mummies, suggesting that these apple organs may serve as additional overwintering habitats for the fungus.

scholarly journals Fungal Functional Genomics: Tunable Knockout-Knock-in Expression and Tagging Strategies

2009 ◽  
Vol 8 (5) ◽  
pp. 800-804 ◽  
Author(s):  
Luis F. Larrondo ◽  
Hildur V. Colot ◽  
Christopher L. Baker ◽  
Jennifer J. Loros ◽  
Jay C. Dunlap
Keyword(s):  
Functional Genomics ◽  
Target Gene ◽  
High Efficiency ◽  
Expression System ◽  
Gene Replacement ◽  
Fungal Species ◽  
Homologous Gene ◽  
Molecular Tools ◽  
Phenotypic Information ◽  
Endogenous Loci

ABSTRACT Strategies for promoting high-efficiency homologous gene replacement have been developed and adopted for many filamentous fungal species. The next generation of analysis requires the ability to manipulate gene expression and to tag genes expressed from their endogenous loci. Here we present a suite of molecular tools that provide versatile solutions for fungal high-throughput functional genomics studies based on locus-specific modification of any target gene. Additionally, case studies illustrate caveats to presumed overexpression constructs. A tunable expression system and different tagging strategies can provide valuable phenotypic information for uncharacterized genes and facilitate the analysis of essential loci, an emerging problem in systematic deletion studies of haploid organisms.

scholarly journals Temporal and Spatial Dispersal of Cladobotryum Conidia in the Controlled Environment of a Mushroom Growing Room

2006 ◽  
Vol 72 (11) ◽  
pp. 7212-7217 ◽  
Author(s):  
Bruce Adie ◽  
Helen Grogan ◽  
Simon Archer ◽  
Peter Mills
Keyword(s):  
Alternative Methods ◽  
Controlled Environment ◽  
Spore Dispersal ◽  
Direct Response ◽  
Physical Disturbance ◽  
Spore Trap ◽  
Tissue Paper ◽  
Temporal And Spatial ◽  
Cobweb Disease ◽  
Dust Extractor

ABSTRACT Cladobotryum spp. are responsible for cobweb disease of mushrooms. In two commercial and one experimental mushroom-growing room, Cladobotryum conidia were released into the air in direct response to physical disturbance of disease colonies during either crop watering or treatment by covering with salt to 10 mm. Conidia were detected using a Burkard spore trap or agar-based trap plates. A maximum concentration of ∼25,000 conidia m−3 was recorded in a small (75-m3) experimental growing room in the hour following the salting of 16 cobweb patches (0.55 m2). Concentrations of 100 and 40 conidia m−3 were recorded in the two larger commercial growing rooms in the hour following the salting of 18 and 11 patches of cobweb (diameter, approximately 50 to 200 mm), respectively. In controlled experiments, disturbed conidia were dispersed rapidly throughout a small growing room, with 91 to 97% of conidia settling out within 15 min. Eighty-five percent of conidia settled out within a 0.5-m radius when air-conditioning fans were switched off, consistent with airborne spore dispersal. Alternative methods for treating diseased areas to minimize conidial release and distribution were investigated and included covering disease colonies with damp paper tissue prior to salt application (tissue salting) and holding a dust extractor above disease colonies during salt application. Both methods resulted in no detectable airborne conidia, but the tissue paper salting technique was more convenient. Prevention of airborne conidial release and distribution is essential to avoid mushroom spotting symptoms, secondary colonies, and early crop termination.

Temporal spore dispersal patterns of grapevine trunk pathogens in South Africa

2010 ◽  
Vol 127 (3) ◽  
pp. 375-390 ◽  
Author(s):  
Jan M. van Niekerk ◽  
Frikkie J. Calitz ◽  
Francois Halleen ◽  
Paul H. Fourie
Keyword(s):  
South Africa ◽  
Spore Dispersal ◽  
Dispersal Patterns

scholarly journals Blowin’ in the Wind: Wind Dispersal Ability of Phytopathogenic Fusarium in a Wind Tunnel Experiment

Atmosphere ◽  
2021 ◽  
Vol 12 (12) ◽  
pp. 1653
Author(s):  
Annika Hoffmann ◽  
Roger Funk ◽  
Marina E. H. Müller
Keyword(s):  
Wind Tunnel ◽  
Fusarium Species ◽  
Empirical Studies ◽  
Dispersal Ability ◽  
Wind Dispersal ◽  
Spore Dispersal ◽  
Abiotic Conditions ◽  
Wind Velocities ◽  
Rain Splash ◽  
Wind Tunnel Study

Dispersal processes play an essential role in cereal diseases caused by phytopathogenic Fusarium. However, most empirical studies of Fusarium spore dispersal have focused on vertical transport by rain splash, while wind dispersal has been mostly neglected. Our objective was to determine the ability of Fusarium conidiospores to disperse via wind under controlled conditions in a wind tunnel study. Ten Fusarium species with diverse spore varieties were studied by placing them in the wind stream at wind velocities of 5 and 8 m s−1 and collecting them after 6 m and a period of 1 h using a newly developed air sampling box. Although spore concentrations were high in the releasing Petri Dishes, the tested isolates were recaptured in only 18 of 78 runs. F. equiseti and F. cerealis were the most frequently recovered species. Changing abiotic conditions, wind speed, and spore shapes had no significant effect on Fusarium spore recapture rates. Another experiment showed that conidiospores were rarely released from the grown mycelium. Therefore, the importance of wind alone as a dispersal medium for Fusarium conidiospores may have been overestimated so far. Further studies should investigate the importance of carrier media or mobile linkers combined with the wind dispersal of spores.

scholarly journals Spore Dispersal Patterns of Fusarium Circinatum on an Infected Monterey Pine Forest in North-Western Spain

2017 ◽  
Author(s):  
Miloň Dvořák ◽  
Patrik Janoš ◽  
Leticia Botella ◽  
Gabriela Rotková ◽  
Rafael Zas
Keyword(s):  
Temporal Trends ◽  
Fusarium Circinatum ◽  
Spore Dispersal ◽  
Dispersal Pattern ◽  
Dispersal Patterns ◽  
Spatial Spread ◽  
Monterey Pine ◽  
Airborne Inoculum ◽  
Air Temperatures ◽  
Forest Nurseries

The airborne inoculum of Fusarium circinatum, 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 infected area still remains scarcely known. To shed more light on this topic, we set an experiment on a naturally PPC infected forest of Monterey pine in Galicia (NW Spain) 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 meteorological measurements. The abundance of F. circinatum spores in the samples was evaluated by quantitative PCR (qPCR) with 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 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 inoculum was 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.

scholarly journals Settleable Dust and Bioburden in Portuguese Dwellings

2020 ◽  
Vol 8 (11) ◽  
pp. 1799
Author(s):  
Carla Viegas ◽  
Marta Dias ◽  
Beatriz Almeida ◽  
Estela Vicente ◽  
Liliana Aranha Caetano ◽  
...  
Keyword(s):  
Passive Sampling ◽  
Culture Media ◽  
Fungal Species ◽  
Winter Period ◽  
Azole Resistance ◽  
Resistance Screening ◽  
Molecular Tools ◽  
Fungal Contamination ◽  
Loading Rates ◽  
Dust Loading

Monitoring campaigns in several buildings have shown that occupants exposed to contaminated indoor air generally exhibit diverse health symptoms. This study intends to assess settleable dust loading rates and bioburden in Portuguese dwellings by passive sampling onto quartz fiber filters and electrostatic dust cloths (EDCs), respectively. Settled dust collected by EDCs was analyzed by culture-based methods (including azole-resistance screening) and qPCR, targeting four different toxigenic Aspergillus sections (Flavi, Fumigati, Circumdati, and Nidulantes). Dust loading rates and bioburden showed higher variability in the summer season. In both seasons, Penicillium sp. was the one with the highest prevalence (59.1% winter; 58.1% summer), followed by Aspergillus sp. in winter (13.0%). Fungal contamination increased in the winter period, while bacterial counts decreased. Aspergillus sections Circumdati and Nidulantes, detected in voriconazole supplemented media, and Aspergillus sections Fumigati and Nidulantes, detected by molecular tools, were found in the winter samples. This study reinforces the importance of applying: (a) Passive sampling methods in campaigns in dwellings; (b) two different culture media (MEA and DG18) to assess fungi; (c) in parallel, molecular tools targeting the most suitable indicators of fungal contamination; and (d) azole resistance screening to unveil azole resistance detection in fungal species.

Where do spores go? Rain-splash patterns using surrogate dye.

2017 ◽  
Vol 70 ◽  
pp. 328
Author(s):  
R.E. Campbell
Keyword(s):  
Ground Level ◽  
Tree Canopy ◽  
Dispersal Patterns ◽  
Release Point ◽  
Splash Dispersal ◽  
Data Points ◽  
Point Data ◽  
Rain Splash ◽  
Monitoring Protocols ◽  
Zero Counts

Microorganisms are incredibly difficult to trap, identify and enumerate efficiently and quickly. This makes it difficult to study incursions of new pathogens and the spread of existing ones effectively. Finding efficient ways of overcoming these difficulties is essential to guide monitoring protocols, control or mitigate spread, or find potential areas for eradication after incursions. We investigated the use of a fluorescent dye, PTSA (1,3,6,8-pyrenetetrasulfonic acid), to explore patterns of rain-splash dispersal of Neonectria ditissima spores. Spores mixed with PTSA dye were released in pear and apple trees within orchards and in an artificial setting using a marquee. Spores and dye were released from a central point source 2.5 m above ground and recaptured in a number of rain traps at ground level and within the tree canopy. It was often very difficult to detect low numbers of recaptured spores, with zero counts found at any distance from the release point. Data points were highly variable, as expected given the range of field conditions, but a clear relationship between dye and spores caught in rain traps under various scenarios was obtained. These results show the merit of PTSA tracer dye as a tool to quantify potential dispersal patterns of microorganisms in an actual landscape of interest with various rainfall scenarios.

scholarly journals Cultivar architecture modulates spore dispersal by rain splash: A new perspective to reduce disease progression in cultivar mixtures

PLoS ONE ◽  
2017 ◽  
Vol 12 (11) ◽  
pp. e0187788 ◽  
Author(s):  
Tiphaine Vidal ◽  
Pauline Lusley ◽  
Marc Leconte ◽  
Claude de Vallavieille-Pope ◽  
Laurent Huber ◽  
...  
Keyword(s):  
Disease Progression ◽  
Spore Dispersal ◽  
Rain Splash ◽  
New Perspective ◽  
Cultivar Mixtures

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

2018 ◽  
Vol 108 (3) ◽  
pp. 374-383 ◽  
Author(s):  
S. A. Wyka ◽  
C. D. McIntire ◽  
C. Smith ◽  
I. A. Munck ◽  
B. N. Rock ◽  
...  
Keyword(s):  
Climatic Factors ◽  
Fungal Species ◽  
Climatic Conditions ◽  
Climatic Variables ◽  
Spore Dispersal ◽  
Pine Needle ◽  
White Pine ◽  
Eastern White Pine ◽  
Potential Risks ◽  
Lecanosticta Acicola

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.

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