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Highly Efficient Capture and Quantification of the Airborne Fungal Pathogen Sclerotinia sclerotiorum Employing a Nanoelectrode-Activated Microwell Array.

ABSTRACT: In this study, we present a microdevice for the capture and quantification of Sclerotinia sclerotiorum spores, pathogenic agents of one of the most harmful infectious diseases of crops, Sclerotinia stem rot. The early prognosis of an outbreak is critical to avoid severe economic losses and can be achieved by the detection of a small number of airborne spores. However, the current lack of simple and effective methods to quantify fungal airborne pathogens has hindered the development of an accurate early warning system. We developed a device that remedies these limitations based on a microfluidic design that contains a nanothick aluminum electrode structure integrated with a picoliter well array for dielectrophoresis-driven capture of spores and on-chip quantitative detection employing impedimetric sensing. Based on experimental results, we demonstrated a highly efficient spore trapping rate of more than 90% with an effective impedimetric sensing method that allowed the spore quantification of each column in the array and achieved a sensitivity of 2%/spore at 5 kHz and 1.6%/spore at 20 kHz, enabling single spore detection. We envision that our device will contribute to the development of a low-cost microfluidic platform that could be integrated into an infectious plant disease forecasting tool for crop protection.

SUBMITTER: Duarte PA

PROVIDER: S-EPMC8756577 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Sclerotinia sclerotiorum and Botrytis cinerea are closely related necrotrophic plant pathogenic fungi notable for their wide host ranges and environmental persistence. These attributes have made these species models for understanding the complexity of necrotrophic, broad host-range pathogenicity. Despite their similarities, the two species differ in mating behaviour and the ability to produce asexual spores. We have sequenced the genomes of one strain of S. sclerotiorum and two strains of B. cinerea. The comparative analysis of these genomes relative to one another and to other sequenced fungal genomes is provided here. Their 38-39 Mb genomes include 11,860-14,270 predicted genes, which share 83% amino acid identity on average between the two species. We have mapped the S. sclerotiorum assembly to 16 chromosomes and found large-scale co-linearity with the B. cinerea genomes. Seven percent of the S. sclerotiorum genome comprises transposable elements compared to <1% of B. cinerea. The arsenal of genes associated with necrotrophic processes is similar between the species, including genes involved in plant cell wall degradation and oxalic acid production. Analysis of secondary metabolism gene clusters revealed an expansion in number and diversity of B. cinerea-specific secondary metabolites relative to S. sclerotiorum. The potential diversity in secondary metabolism might be involved in adaptation to specific ecological niches. Comparative genome analysis revealed the basis of differing sexual mating compatibility systems between S. sclerotiorum and B. cinerea. The organization of the mating-type loci differs, and their structures provide evidence for the evolution of heterothallism from homothallism. These data shed light on the evolutionary and mechanistic bases of the genetically complex traits of necrotrophic pathogenicity and sexual mating. This resource should facilitate the functional studies designed to better understand what makes these fungi such successful and persistent pathogens of agronomic crops.

Project description:Endornaviruses are capsidless linear (+) ssRNA viruses in the family Endornaviridae. In this study, Scelrotinia sclerotiorum endornavirus 11 (SsEV11), a novel endornavirus infecting hypovirulent Sclerotinia sclerotiorum strain XY79, was identified and cloned using virome sequencing analysis and rapid amplification of cDNA ends (RACE) techniques. The full-length genome of SsEV11 is 11906 nt in length with a large ORF, which encodes a large polyprotein of 3928 amino acid residues, containing a viral methyltransferase domain, a cysteine-rich region, a putative DEADc, a viral helicase domain, and an RNA-dependent RNA polymerase (RdRp) 2 domain. The 5' and 3' untranslated regions (UTR) are 31 nt and 90 nt, respectively. According to the BLAST result of the nucleotide sequence, SsEV11 shows the highest identity (45%) with Sclerotinia minor endornavirus 1 (SmEV1). Phylogenetic analysis based on amino acid sequence of RdRp demonstrated that SsEV11 clusters to endornavirus and has a close relationship with Betaendornavirus. Phylogenetic analysis based on the sequence of endornaviral RdRp domain indicated that there were three large clusters in the phylogenetic tree. Combining the results of alignment analysis, Cluster I at least has five subclusters including typical members of Alphaendornavirus and many unclassified endornaviruses that isolated from fungi, oomycetes, algae, and insects; Cluster II also has five subclusters including typical members of Betaendornavirus, SsEV11, and other unclassified viruses that infected fungi; Cluster III includes many endorna-like viruses that infect nematodes, mites, and insects. Viruses in Cluster I and Cluster II are close to each other and relatively distant to those in Cluster III. Our study characterized a novel betaendornavirus, SsEV11, infected fungal pathogen S. sclerotiorum, and suggested that notable phylogenetic diverse exists in endornaviruses. In addition, at least, one novel genus, Gammaendornavirus, should be established to accommodate those endorna-like viruses in Cluster III.

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Highly Efficient Capture and Quantification of the Airborne Fungal Pathogen Sclerotinia sclerotiorum Employing a Nanoelectrode-Activated Microwell Array.

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