An isolate of rice stripe virus (designated as RSV-YL) was purified. The particles showed to be pleomorphisms under electron microscope, mainly branched filaments of about 80-250 nm in length and about 8 nm in width. There are also some open circular filaments of 3 nm and 8 nm in width, and some filaments of 13 nm in width and 130-190 nm in length. The basic morphism of RSV particles should be filaments of 3 nm in width and various length. By SDS-PAGE analysis, the molecular weight of disease-specific protein (SP) encoded by vRNA4 was 19.9 kDa and that of coat protein (CP) encoded by vcRNA3 was 33.6 kDa. When nucleic acid extracted from the purified RSV was electrophoresed under nondenaturing condition, the size of four dsRNAs (designated as dsRNA1-4 in order of decreasing size) was 4.9×106,2.7×106,2.0×106 and 1.7×106 Da, respectively, and that of four ssRNAs (designated as ssRNA1-4 in order of decreasing size) was 3.0×106,1.2×106,0.9×106 and 0.8×106 Da, respectively. A fifth segment with a size of 0.58×106 Da identified as ssRNA5 associated with the purified virus sometimes. The antiserum against the coat protein further purified by preparative electrophoresis was raised and used to investigate the serological relationships between RSV-CP and RSV-SP, CP and SP of rice grassy stunt virus (RGSV) which is also a member of Tenuivirus. The results showed that RSV-CP had no serological reaction with SP of RSV and PGSV, but could weakly react with antiserum of RGSV-CP, which confirmed that there is distantly evolutionary relationship between RGSV and RSV.

Objective:This study is aimed to investigate the value of absolute lymphocyte count (ALC) in predicting the clinical prognosis of patients with myelodyplastic syndrome(MDS).Methods:245 patients with MDS who diagnosed in our hospital from 2009 to 2019 were analyzed retrospectively, re-diagnosed according to WHO 2016 standard, and 208 patients with intact IPSS-R were risk-stratified, all of the patients′ peripheral blood ALC were collected and analyzed, through the time dependent receiver operating characteristic curve (ROC) analysis in Survival ROC package of R language, the optimal threshold value of ALC was 1.0×10 9/L. The patients of MDS were divided into normal ALC group (ALC ≥1.0×10 9/L) and low ALC group (ALC<1.0×10 9/L). Pearson χ 2 test and Mann-Whitney U test was used to analyze the differences in general data between the two groups. The overall survival (OS) curve and leukemia-free survival (LFS) were plotted by Kaplan-Meier method and compared by Long-rank test. Factors influencing the prognosis of MDS were analyzed by Cox Regression Model. Results:There were 97 cases in low ALC group and 148 cases in normal ALC group. The low ALC group had lower OS (15 months vs 60 months, P<0.000 1) and higher IPSS-R score (5.0 vs 3.75, P = 0.001). Multivariate analysis showed that ALC (<1.0×10 9/L) (HR:0.374,95% CI:0.153-0.917, P = 0.032) was independent risk factor of OS in IPSS-R-intermediate-risk MDS patients. Conclusion:This study shows that ALC in peripheral blood is an independent risk factor in IPSS-R-intermediate-risk MDS patients, which provides clinical evidence for the influence of body immunity on the development of MDS.

Objective To develop a fluorescent recombinase-aided isothermal amplification (RAA)-based nucleic acid assay for detection of Leshimania. Methods Specific primers and probes were designed targeting Leishmania internal transcribed spacer 1 (ITS1) gene for RAA assay, and a fluorescent RAA assay was developed for detection of Leishmania following screening of primer pairs and optimization of primer and probe concentrations. The sensitivity of RAA assay for detection of Leishmania was evaluated using recombinant plasmid containing Leishmania ITS1 gene sequences at different copies and Leshimania genomic DNA at different concentrations as templates, and the specificity of RAA assay for detection of Leishmania was evaluated using the genomic DNA of transfusion-transmitted parasites, including Babesia microti, Toxoplasma gondii, Plamodium vivax, P. ovale, P. falciparum, P. malariae, L. donovani and L. infantum. Results After the optimal primer pair was screened from 9 pairs of primer combinations, the final primer and probe concentrations were optimized as 0.3 μmol/L and 0.08 μmol/L, respectively. Nucleic acid detection of Leishmania was completed by the fluorescent RAA assay at an isothermal temperature of 39 °C within 20 min. Remarkable florescent signals were seen within 5 min following RAA detection of genomic DNA of L. donovani and L. infantum, and no cross-reactions were observed with B. microti, T. gondii, P. vivax, P. ovale, P. falciparum or P. malariae. The lowest limitation of detection of the fluorescent RAA assay was 10 copies/μL recombinant plasmid containing Leishmania ITS1 gene sequences and 1 fg/μL Leishmania genomic DNA. Conclusions A rapid, simple, sensitive and specific fluorescent RAA assay is successfully developed for detection of L. donovani and L. infantum, which is effective for field screening of leishmaniasis.

Objective To establish a nucleic acid assay for detection of Paragonimus skrjabini based on the recombinase-aided isothermal amplification (RAA) technique, and to preliminarily evaluate its detection efficiency. Methods The metacercariae of P. skrjabini, P. westermani and Euparagonimus cenocopiosus were isolated from crabs, and genomic DNA was extracted for molecular characterization. The cytochrome coxidase 1 (cox1) gene sequence of P. skrjabini was selected as the target gene fragment, and the primers and probes were designed, screened and synthesized for RAA assay. The genomic DNA of P. skrjabini metacercariae from Jiyuan City and Yiyang County of Luoyang City, Henan Province were used as templates for verification of the fluorescent RAA assay. The fluorescent RAA assay was performed to detect different concentrations of plasmids containing target gene fragment and P. skrjabini metacercariae genomic DNA to determine the sensitivity. Fluorescent RAA assay was performed with recombinant plasmids containing P. skrjabini cox1 gene sequences at different concentrations and P. skrjabini genomic DNA as templates to evaluate its sensitivity, and the genomic DNA of P. westermani, E. cenocopiosus, Clonorchis sinensis and Schistosoma japonicum was detected with fluorescent RAA assay to evaluate its specificity. Results P. skrjabini, P. westermani and E. cenocopiosus metacercariae were isolated from crabs, respectively. Molecular characterization and phylogenetic analysis confirmed their homology with the genes sequences of standard Paragonimus strains in GenBank. A fluorescent RAA assay was successfully established for nucleic acid detection of P. skrjabini, and the genomic DNA of P. skrjabini metacercariae from Jiyuan City and Yiyang County of Luoyang City, Henan Province was amplified using the fluorescent RAA assay within 5 min, while the negative control was not amplified. If the recombinant plasmid containing P. skrjabini cox1 gene sequences was used as templates, the fluorescent RAA assay showed the lowest detection limit of 10 copies/μL, and positive amplification was observed within 5 min. If genomic DNA was used as templates, the fluorescent RAA assay showed the lowest detection limit of 10 pg/μL, and all positive amplifications were found within 5 to 10 min. In addition, the fluorescent RAA assay was tested negative for P. westermani, E. cenocopiosus, C. sinensis and S. japonicum. Conclusions A rapid, sensitive and specific fluorescent RAA assay is successfully established for nucleic acid detection of P. skrjabini, which has potential values in rapid field detection and species identification in freshwater crabs in areas endemic for P. skrjabini.