The infection of Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the main causes of economic losses in sericulture. Thus, it is essential to establish rapid and effective method for BmNPV detection. In the present study, we have developed a recombinase-aided amplification (RAA) to amplify the BmNPV genomic DNA at 37 °C within 30 min, and achieved a rapid detection method by coupling with a lateral flow dipstick (LFD). The RAA-LFD method had a satisfactory detection limit of 6 copies/μL of recombinant plasmid pMD19-T-IE1, and BmNPV infection of silkworm can be detected 12 h post-infection. This method was highly specific for BmNPV, and without cross-reactivity to other silkworm pathogens. In contrast to conventional polymerase chain reaction (PCR), the RAA-LFD assay showed higher sensitivity, cost-saving, and especially is apt to on-site detection of BmNPV infection in the sericulture production.
- MeSH
- Bombyx * virology MeSH
- DNA, Viral genetics MeSH
- Limit of Detection MeSH
- Nucleopolyhedroviruses * genetics isolation & purification MeSH
- Recombinases * metabolism genetics MeSH
- Sensitivity and Specificity MeSH
- Nucleic Acid Amplification Techniques * methods MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
Silkworm diseases caused by fungi infection occur frequently in sericulture and brought huge economic loss to sericulture. However, on the other hand, some fungi such as Beauveria bassiana, as an important entomological fungus, play an important role in biological control of insect pests. Here, two fungal pathogens causing yellow muscardine were isolated from the silkworm and named as SZY1 and SZY2. These two strains showed almost the same conidial morphology which were smooth, near-spherical, spherical, or ovoid and 2.7 ± 0.6 μm × 2.5 ± 0.9 μm in size, and the hyphal growth rate was also similar. However, the conidia production of SZY2 was almost twice as many as that of SZY1. The complete ribosomal RNA gene was sequenced and analyzed. As a result, the gene sequences of internal transcript space 1 (ITS1)-5.8S rRNA-internal transcript space 2 (ITS2) of SZY1 and SZY2 were identical in sequence and size, and for 18S rRNA, 28S rRNA, and intergenic spacer (IGS), the gene identity of SZY1 to SZY2 was 99%, 99%, and 98%, respectively. Results of phylogenetic analysis based on either ITS1-5.8S rRNA-ITS2 or 18S rRNA showed that both SZY1 and SZY2 were closely related to Beauveria bassiana. These results revealed that the pathogens of yellow muscardine SZY1 and SZY2 were identified as two different strains of Beauveria bassiana, which could provide diagnostic evidence for silkworm muscardine and was helpful for the research and development of novel Bombyx batryticatus and fungal biological insecticide.
- MeSH
- Beauveria * genetics MeSH
- Bombyx * genetics microbiology MeSH
- Phylogeny MeSH
- RNA, Ribosomal, 18S MeSH
- RNA, Ribosomal, 5.8S MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
In this study, NaYF4:20%Yb, 2%Er upconverting nanoparticles (UCNPs) were synthesized by solvothermal method and characterized by transmission electron microscopy and upconversion fluorescence spectrometry. The results showed that the UCNP particles present good dispersion and uniform spherical shape with a size of 29 ~ 42 nm. Hydroxyl UCNPs were converted to hydrophilic carboxylic acid-functionalized ones by ligand exchange, and the streptavidin was attached on the surface of carboxylic acid-functionalized UCNPs via amide bond. The DNA nanosensors based on UCNPs with DNA probes have been successfully developed. Only the genomic DNA of Nosema bombycis can be specifically detected by the DNA nanosensors when the DNA of Bombyx mori and its pathogens was used as target DNA. When the DNA nanosensors were used to detect the DNA of N. bombycis, a broad emission peak signal appeared at 580 nm. There is linear relationship between the signal intensity and DNA concentration of N. bombycis, I580/I545 (R2 = 0.820) and I545/I654 (R2 = 0.901). The detectable minimum concentration of genomic DNA of N. bombycis was 100 ng/μL while the tested concentrations of N. bombycis genomic DNA were 3000 ng/μL, 1500 ng/μL, 1000 ng/μL, 500 ng/μL, 250 ng/μL, and 100 ng/μL, respectively. The whole detection process for target DNA takes less than 60 min.
- MeSH
- DNA MeSH
- Carboxylic Acids MeSH
- Nanoparticles * chemistry MeSH
- Nosema MeSH
- Fluorescence Resonance Energy Transfer * MeSH
- Publication type
- Journal Article MeSH
Microsporidia are a group of obligate intracellular unicellular eukaryotes that can parasitize a wide variety of other eukaryotes ranging from protists to invertebrates and vertebrates. In this study, we examined the microsporidium Nosema sp. isolated from the mulberry pest, Hemerophila atrilineata Butler, 1881, named herein “Nosema sp. HA”. The fresh spores were long oval in shape, 3.8 ± 0.4 μm in length and 1.9 ± 0.3 μm in width. Analysis of tissue infection of silkworm, Bombyx mori Linnaeus, 1758, indicated that the midgut, Malpighian tubules, muscle, fat body, silk glands, hemocytes, nerve tissue and gonads of silkworm were infected with Nosema sp. HA. The complete rRNA gene sequence of this microsporidium contained 4 305 base pairs (GenBank Accession JN882299), including the large subunit rRNA (2 492 bp), the internal transcribed spacer (187 bp), the small subunit rRNA (1 232 bp), the intergenic spacer (279 bp) and the 5S region (115 bp). The organization of the rRNA gene is 5′-LSU-ITS-SSU-IGS-5S-3′. Phylogenetic analysis, comparison of sequence identities and the arrangement in the rRNA gene subunits suggested that this isolate is separate from other Nosema species.
- MeSH
- RNA, Fungal genetics MeSH
- Phylogeny MeSH
- Host-Parasite Interactions MeSH
- Microsporidia physiology genetics ultrastructure MeSH
- Moths parasitology MeSH
- RNA, Ribosomal genetics MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
