The Detection And Risk Factor Analysis Of BK Virus Infection In Renal Transplant Recipients

Objection:The purpose of this research is to approach the detection methods of BK virus by the examination of urinary castoff cells,serum sample RT-PCR detection and urinary sample RT-PCR detection.As a result,we are able to get the message of the condition and charactesr of the BK virus infected people,sieve the possible risk factors and predict their long-term graft function.Methods:1,Fluorescent quantitation PCR ex-reference strain and standard curve: Design of the primer,determination of the reaction system,chemical synthesis of masculine template,PCR amplification and identification,extraction and depuration of the DNA fragment,combinational joining with P carrier,convertion into the Bacillus coli DH5αcompetent cell,masculine clone appreciation of the microbial population,extraction of the recombinant plasmid and gradient deliquation,PCR reaction and description of standard curve;2,We chose 113 people who received transplantation operation in 2005.1 to 2009.2 as the research objects.We made the examination of urinary castoff cells,obtained the peripheral serum and urine sample,extracted the serum and urinary DNA and took the message of the condition of the BKV infection by RT-PCR reaction.We divided the objects in three groups: Urine and Serum(-),Urine(+) but Serum(-),Urine(+) and Serum(+) by the result of PCR detection.All of the examination data was analyzed statistically.Results:According to the result of PCR and standard curve,we confirmed that the primer,probe and purpose DNA fragment could complement each other normally.The recombinant plasmid being enzimatic cut and attenuated into concentration gradient can obtain good standard curve image.The sensitivity low limit of PCR reaction is 2.14×10~2copis/ul template concentration.By the result of the examination of urinary castoff cells,serum sample RT-PCR detection and urinary sample RT-PCR detection of the 113 objects,the Decoy cell detection rate is 23.9%.Urinary BKV RT-PCR masculine rate is 22.12%,while the serum BKV RT-PCR masculine rate is 5.30%.Comparing the clinical data of three groups[Urine and Serum(-),Urine(+) but Serum(-),Urine(+) and Serum(+)],we got the conclusion that the Hematodialysis duration in Urine and Serum(-) is shorter than other two groups as while as the pulmonary infection incidence rate is lower.Among the three groups,the age,gender,cold ischemia time,cadaveric/living donor,DGF,induction therapy, immune suppression and acute rejection had no statistic differences.We calculated the 30dCRR of the three groups and made rank sum test of ranked data according to the standard whether 30dCRR is larger than 67%.There is no statistic difference among the three groups.Conclusion:Through the research we established and mastered the BKV detection methods and took the detection for some of the renal transplantation reciepients.The examination of urinary castoff cells can be used as a preliminary screening method of BKV infection.Its negative estimation rate is high,in contrast the positive estimation rate is low.to be used as the diagnosis method,for which can make the result false-negative.The urine sample and serum sample PCR detection can be used as a high sensitivity and specificity method to detect BKV infection while they can disclose the virus infection replication extent. In contrast with other two groups,the urine and serum(-) group had longer time in the hematodialysis duration before transplantation and lower incidence rate in the pulmonary infection after transplantation,both of which had statistic difference.The age,gender,cold ischemia time,cadaveric/living donor,DGF,induction therapy,immune suppression and acute rejection of the transplant recipients had no significant difference in the three groups.When there is no BKVN(BK virus nephritis),there is no evidence to prove that simple serum or urinary BKV PCR detection(+) may influence the long-term graft function.