Method for diagnosis and monitoring of viral infection by analysis of viral transrenal nucleic acids in urine

Abstract

The present invention relates to methods for diagnosis or monitoring of viral infection by detecting the presence of transrenal viral nucleic acids or nucleic acids of viral origin in urine sample, with or without isolation of nucleic acids from a urine sample. The analysis of the nucleic acids is performed through hybridization of the nucleic acids with specific probes, or through a chain amplification reaction with specific primers. The methods are applicable to all viral pathogenic agents, including RNA, DNA, episomal, or integrated viruses.

1 . A method for detecting a viral infection in a subject, said method comprising
 detecting the presence of viral transrenal nucleic acids in a urine sample from said subject, said detecting comprising contacting the viral transrenal nucleic acids with a primer containing a complementary sequence and extending the primer to produce an extension product,   and determining a viral infection in said subject based on the presence of the extension product.   
     
     
         2 . The method according to  claim 1 , wherein said nucleic acids are deoxyribonucleic acid (DNA). 
     
     
         3 . The method according to  claim 1 , further comprising quantitating said viral transrenal nucleic acids. 
     
     
         4 . The method according to  claim 3 , wherein said quantitating is performed by a means selected from hybridization, PCR, nested PCR, SSCP, LCR, and SDA. 
     
     
         5 . The method according to  claim 1 , further comprising the step of separating the urine sample into a cell-free fraction containing said transrenal nucleic acids. 
     
     
         6 . The method according to  claim 5 , wherein said viral transrenal nucleic acids comprise DNA fragments of between about 100 and about 200 bp. 
     
     
         7 . The method according to  claim 1 , further comprising isolation or purification of said nucleic acids. 
     
     
         8 . The method according to  claim 7 , wherein said purification is performed through chemical or physical methods. 
     
     
         9 . The method according to  claim 1 , including a pretreatment of the urine sample with one or more agents that inhibit the degradation of the nucleic acids. 
     
     
         10 . The method according to  claim 9 , wherein said agents are selected from ion-chelating agents, denaturing agents, and ionic detergents. 
     
     
         11 . The method according to  claim 10 , wherein said ion-chelating agents are EDTA; said denaturing agents are guanidine HCl or guanidine isothiocyanate; and said ionic detergents are N-lauryl sarcosine or sodium dodecyl sulfate. 
     
     
         12 . The method according to  claim 1 , wherein said detecting the presence of said viral nucleic acids is performed through a technique selected from hybridization of the nucleic acids, a cycling probe reaction, a polymerase chain reaction, a nested polymerase chain reaction, single-strand conformation polymorphism, a ligase chain reaction, strand displacement amplification, and restriction fragments length polymorphism. 
     
     
         13 . The method according to  claim 1 , wherein said viral infection is hepatitis B virus (HBV) infection. 
     
     
         14 . A method for monitoring a viral infection in a subject, said method comprising:
 a) quantitating the amount of a viral transrenal nucleic acid in a first urine sample from said subject, in accordance with  claim 3 ;   b) quantitating the amount of said viral transrenal nucleic acid in a second urine sample from said subject, in accordance with  claim 3 ; and   c) comparing the amount of said viral transrenal nucleic acid in said first and said second urine sample from said subject, thereby monitoring said viral infection in said subject.   
     
     
         15 . The method according to  claim 14 , further comprising administering to said subject a compound that reduces or inhibits said viral infection. 
     
     
         16 . The method according to  claim 14 , wherein said subject is at risk of developing a recurring viral infection. 
     
     
         17 . The method according to  claim 14 , wherein said viral infection is hepatitis B virus (HBV) infection.