In order to reveal the phenotypic characteristics of 17 JE virus strains isolated from different years, plaque sizes, mice neurovirulence and mice neuroinvasiveness of the isolates were studied and compared. BHK21 cell monolayers were used for testing the plaque sizes. The virus neurovirulence was tested in 9-11g mice inoculated intracerebrally and the virus neuroinvasiveness was tested in 9-11g and 14-16g by subcutaneous inoculation. Results showed that all the viruses produced clear plaques on the BHK21 cell monolayers with different sizes and all the virus strains appeared high neurovirulence in the mice with higher than lg8. 0/0.03 mL virus titers, while no apparent difference among them. The neuroinvasiveness (subcutaneous virulence) tested in the 9-11g mice had shown a little difference, but when tested in the 12-14 g mice,the difference was apparent. The results demonstrated that JEV in nature were highly neurovirulent with no apparent difference. However the neuroinvasiveness of the JEV in nature was greatly different, which didn't relate to the years of isolation and genotypes, but most of the viruses isolated from patients showed higher neuroinvasiveness.
Animals ; Cell Line ; China ; Culicidae ; virology ; Encephalitis Virus, Japanese ; genetics ; isolation & purification ; pathogenicity ; Encephalitis, Japanese ; virology ; Genotype ; Humans ; Mice ; Phenotype ; Viral Plaque Assay ; Virulence

BACKGROUNDThe large phage antibody library is used to obtain high-affinity human antibody, and the Loxp/cre site-specific recombination system is a potential method for constructing a large phage antibody library. In the present study, a phage antibody library vector pDF was reconstructed to construct diabody more quickly and conveniently without injury to homologous recombination and the expression function of the vector and thus to integrate construction of the large phage antibody library with the preparation of diabodies.

METHODSscFv was obtained by overlap polymerase chain reaction (PCR) amplification with the newly designed VL and VH extension primers. loxp511 was flanked by VL and VH and the endonuclease ACC III encoding sequences were introduced on both sides of loxp511. scFv was cloned into the vector pDF to obtain the vector pDscFv. The vector expression function was identified and the feasibility of diabody preparation was evaluated. A large phage antibody library was constructed in pDscFv. Several antigens were used to screen the antibody library and the quality of the antibody library was evaluated.

RESULTSThe phage antibody library expression vector pDscFv was successfully constructed and confirmed to express functional scFv. The large phage antibody library constructed using this vector was of high diversity. Screening of the library on 6 antigens confirmed the generation of specific antibodies to these antigens. Two antibodies were subjected to enzymatic digestion and were prepared into diabody with functional expression.

CONCLUSIONSThe reconstructed vector pDscFv retains its recombination capability and expression function and can be used to construct large phage antibody libraries. It can be used as a convenient and quick method for preparing diabodies after simple enzymatic digestion, which facilitates clinical trials and application of antibody therapy.

Bacteriophages ; genetics ; Digoxin ; immunology ; Enzyme-Linked Immunosorbent Assay ; Genetic Vectors ; Immunoglobulin Fragments ; biosynthesis ; immunology ; Peptide Library

Urinary tract infection is the most common infectious complication after kidney transplantation.To further reduce the incidence of urinary tract infection after kidney transplantation,improve the diagnosis and treatment level of urinary tract infection after kidney transplantation in China,prevent the development of bacterial drug resistance and ensure the safety and effectiveness of drug use,Branch of Organ Transplantation of Chinese Medical Association organized experts in the fields of kidney transplantation and infectious diseases to consider clinical status of urinary tract infection after kidney transplantation in China,refer to"Diagnosis and Treatment of Urological and Andrological Diseases in China(2022 edition)"and"Urinary Tract Infection in Solid Organ Transplant Recipients in American Society of Transplantation Practical Guidelines for Infectious Diseases(2019 edition)",and formulate"Guidelines for Clinical Diagnosis and Treatment of Urinary Tract Infection in Kidney Transplant Recipients in China"from the perspectives of clinical classification and definition,epidemiology and etiology,diagnosis and treatment of urinary tract infection after kidney transplantation,respectively.