Objective To establish a rapid diagnostic method for the detection of marine vibrios,and then construct a new technology platform for the clinical diagnosis of marine vibrio infection.Methods A pair of PCR primers and a sequencing primer based on the whA gene of V.vulnificus and the toxR genes of V.parahemolyticus and V.alginolyticus were designed respectively,and then the specific DNA fragments were amplified.Next,the single-stranded DNA templates were prepared for pyrosequencing.The obtained base sequence was validated by NCBI alignment.In addition,the 16S rRNA genotyping of V.vulnificus was also performed.Results The PCR primers and sequencing primer of V.vulnificus showed good specificity,and a 167 bp DNA fragment was amplified from 4 strains of V.vulnificus.The pyrosequencing results completely matched with the whA gene sequence of V.vulnificus.Meanwhile,the control strains were negative.A 105 bp DNA fragment and a 134 bp DNA fragment were amplified from 11 strains of V.parahemolyticus and V.alginolyticus,respectively,and the pyrosequencing results were consistent with the expected sequence.In addition,one of 4 strains of V.vulnificus was identified as 16S rRNA-A type,and the other 3 as 16S rRNA-B type.Conclusion The PCR-pyrosequencing method established in this study is a new method for the real-time detection of short nucleotide sequences.It has some advantages such as high throughput,high precision and simple operation,and may be applied to the fast and accurate identification of marine and terrestrial pathogenic bacteria.

Objective To study the mechanism of SW480 cell line death caused by transfection of retroviral vector-G1CEACDNa.Methods Plasmid G1CEACDNa was transferred into the SW480 cell line using liposomes method. RT-PCR was performed to examin the expression of CD gene indrectly.The cell growth curve was measured by means of cell counting. When the CD+SW480 cells were exposed to 5-FC (1mmol/L), the growth inhibition rate and the "bystander effect" were detected by MTT method.The ultrastructure was observed by electron microscope.Apoptosis was verified by flow cytometer .Results The product of RT-PCR showed a band at 1.5kb on the photo of electrophoresis. The growth of CD+ SW480 cells was inhibited 24h after administrating 5-FC,and the inhibition rates at 72h,96h,120h were 30.0%,50.0% and 80.0%,respectively.Apoptosis cells in different phases and apoptotic bodies in the field of electron microscope were observed. Meanwhile ,a few cells showed necrosis.Flow cytometer verified that a few cells appearred apoptosis 48h after exposed to 5-FC (1mmol/L), the apoptosis rate and the necrosis rate at 72h,96h were 20.2%,30.7% and 19.6%,21.1% respectively.Conclusions The death mechanism of SW480 cells transfected with G1CEACDNa followed by 5-FC treatment includes both necrosis and apoptosis.Apoptosis is possibly the bystander effect.

Objective To investigate the effect of bortezomib with arsenic trioxide different concentration on the cell cycle and apoptosis of Raji cells. Methods Flow cytometry analysis showed that the relative number of cells in different phases and the percentages of cells calculated in G1 and S phase of the cell cycle and apoptosis were assessed after treatment with As2O3 and BOR or in combination with BOR in different concentration at indicated time (24 h, 48 h, 72 h). Results Flow cytometric analysis showed that the cell cycle was arrested at G1 phase, the number of cells G1 period increased significantly, and S phase decreased on Raji cells after As2O3 treatment. The relationship between the cellular DNA contents and the concentration of As2O3 showed a dose-and time-dependent manner (P ＜0.0001). But it was found that BOR had no effect on Raji cell cycle, but, in two drugs combination, cell apoptosis rate significantly increased from 16.98 % to 45.84 %. Conclusion The results show that As2O3 exerted variable and definite effects on lymphoma Raji cells, which indicated that As2O3 might induce apoptosis and arrest cell cycle. The combination of two drugs had a effective and synergistic effect on apoptosis.

Objective:To analyze the clinical characteristics of patients with nocardiosis, so as to improve the diagnosis and treatment of nocardia infection in the future.Methods:From May 2016 to October 2020, 24 patients with nocardiosis in Xiangya Hospital, Central South University were enrolled, and their clinical data including clinical features, laboratory examinations, imaging findings, diagnosis and treatment process, and outcome were retrospectively analyzed.Results:Among the 24 patients with nocardiosis, 18 cases (75.0%) were males, and the median age was 54.5 years.Twenty-three patients had underlying diseases, of which the most common disease was antineutrophil cytoplasmic antibody-related vasculitis (16.7%(4/24)). Of nine species of Nocardia identified from the 24 patients, Nocardia farcinica was the most common species (seven cases). The lesion sites were mainly lungs (70.8%(17/24)), skin and soft tissues (42.0%(10/24)), brain (25.0%(6/24)) and blood system (17.0%(4/24)). There were 12 cases (50.0%) of patients with more than two lesion sites. The clinical manifestations, imaging examinations and laboratory tests of the 24 patients were not specific. The diagnosis depended on the etiology. Nineteen patients received trimethoprim-sulfamethoxazole-based combination therapy, and two were discontinued due to adverse reactions of sulfa drugs. After treatment, 19 cases (79.2%) were improved and five cases (20.8%) died. Conclusions:Patients with nocardiosis often have atypical clinical manifestations, and multiple organs are easily affected.Early and accurate identification and rapid and effective anti-biotic therapy are the keys to improve the overall prognosis of these patients.