BackgroundThe small brown planthopper (SBPH) is an important pest of cereal crops and acts as a transmission vector for multiple RNA viruses. Rapid diagnosis of virus in the vector is crucial for efficient forecast and control of viral disease. Reverse transcription polymerase chain reaction (RT-PCR) is a rapid, sensitive and reliable method for virus detection. The traditional RT-PCR contains a RNA isolation step and is widely used for virus detection in insect. However, using the traditional RT-PCR for detecting RNA virus in individual SBPHs becomes challenging because of the expensive reagents and laborious procedure associated with RNA isolation when processing a large number of samples.ResultsWe established a simplified RT-PCR method without RNA isolation for RNA virus detection in a single SBPH. This method is achieved by grinding a single SBPH in sterile water and using the crude extract directly as the template for RT-PCR. The crude extract containing the virus RNA can be prepared in approximately two minutes. Rice stripe virus (RSV), rice black streaked dwarf virus (RBSDV) and Himetobi P virus (HiPV) were successfully detected using this simplified method. The detection results were validated by sequencing and dot immunobinding assay, indicating that this simplified method is reliable for detecting different viruses in insects. The evaluation of the sensitivity of this method showed that both RSV and HiPV can be detected when the cDNA from the crude extract was diluted up to 103 fold. Compared to the traditional RT-PCR with RNA isolation, the simplified RT-PCR method greatly reduces the sample processing time, decreases the detection cost, and improves the efficiency by avoiding RNA isolation.ConclusionsA simplified RT-PCR method is developed for rapid detection of RNA virus in a single SBPH without the laborious RNA isolation step. It offers a convenient alternative to the traditional RT-PCR method.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-017-0732-6) contains supplementary material, which is available to authorized users.
Shiraia bambusicola is a rare medicinal fungus found in China that causes bamboo plants to decay and die with severe infection. Hypocrellin, its main active ingredient, is widely used in several fields, such as medicine, agriculture, and food industry. In this study, to clarify the genomic components, taxonomic status, pathogenic genes, secondary metabolite synthesis pathways, and regulatory mechanisms of S. bambusicola, whole-genome sequencing, assembly, and functional annotation were performed using high-throughput sequencing and bioinformatics approaches. It was observed that S. bambusicola has 33 Mb genome size, 48.89% GC content, 333 scaffolds, 2590 contigs, 10,703 genes, 82 tRNAs, and 21 rRNAs. The total length of the repeat sequence is 2,151,640 bp. The annotation of 5945 proteins was obtained from InterProScan hits based on the Gene Ontology database. Phylogenetic analysis showed that S. bambusicola belongs to Shiraiaceae, a new family of Pleosporales. It was speculated that there are more than two species or genus in Shiraiaceae. According to the annotation, 777 secreted proteins were associated with virulence or detoxification, including 777 predicted by the PHI database, 776 by the CAZY and Fungal CytochromeP450 database, and 441 by the Proteases database. The 252 genes associated with the secondary metabolism of S. bambusicola were screened and enriched into 28 pathways, among which the terpenoids, staurosporine, aflatoxin, and folate synthesis pathways have not been reported in S. bambusicola. The T1PKS was the main gene cluster among the 28 secondary metabolite synthesis gene clusters in S. bambusicola. The analysis of the T3PKS gene cluster related to the synthesis of hypocrellin showed that there was some similarity between S. bambusicola and 10 other species of fungi; however, the similarity was very low wherein the highest similarity was 17%. The genomic information of S. bambusicola obtained in this study was valuable to understand its genetic function and pathogenicity. The genomic information revealed that several enzyme genes and secreted proteins might be related to their host interactions and pathogenicity. The annotation and analysis of its secondary metabolite synthesis genes and gene clusters will be an important reference for future studies on the biosynthesis and regulation mechanism of the secondary metabolites, contributing to the discovery of new metabolites and accelerating drug development and application.
To clarify the control effects of Trichoderma hamatum strain MHT1134 on Fusarium wilt in continuous pepper cropping fields and its regulatory effects on soil microecology, the physical and chemical properties, enzyme activities, community structures of soil samples from five field types were analysed. Samples were taken from fields that had been continuously planted for 1, 5, 9 years, and applied the strain MHT1134 for 1 and 2 years. The MHT1134 control effects on pepper wilt after application 1 year and 2 years were 63.03% and 70.21%, respectively. 4 kinds of physical and chemical indexes and 6 kinds of enzyme activities in soil were increased. With the continuous cropping years increased, the microbial abundance and diversity decreasing significantly. The relative abundances of Fusarium, Gibberella increased along with the planting years, but decreased after the MHT11134 application. However, the relative abundances of Trichoderma and Chaetomium significantly increased. Additionally, as the cropping years increased, the soil abundance of Actinobacteria gradually decreased, but it significantly increased from 17.56 to 22.44% after the MHT1134 application. Thus, strain MHT1134 effectively improved the microbial community structure of the soil, and it also positively affected soil quality. A continuous application may improve the control effect.
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