Polymerase chain reaction in the diagnosis of infectious diseases

PCR is one of the methods of DNA diagnostics that allows to increase the number of copies of the detected region of the genome( DNA) of bacteria or viruses by millions of times using the DNA polymerase enzyme. The segment of the nucleic acid tested for a given genome is multiplied( amplified) many times, which allows it to be identified. First, the DNA molecule of bacteria or viruses is divided into two chains by heating, then, in the presence of synthesized DNA primers( the nucleotide sequence is specific for the genome being determined), they bind to complementary regions of DNA, a second chain of nucleic acid is synthesized after each primer in the presence of a thermostable DNA polymerase. Two DNA molecules are obtained. The process is repeated many times. For diagnosis, one molecule of DNA, that is, one bacterium or a viral particle, is sufficient. The introduction of an additional step in the reaction - the synthesis of DNA on the RNA molecule by the enzyme reverse transcriptase - made it possible to test RNA viruses, for example, the HCV virus. PCR is a three-step process, repeated cyclically: denaturation, primer annealing, DNA synthesis( polymerization).The synthesized amount of DNA is identified by ELISA or by electrophoresis.

PCR can use a variety of biological material - serum or blood plasma, scraping from the urethra, biopsy, pleural fluid, cerebrospinal fluid, etc. First of all, PCR is used to diagnose infectious diseases, such as HBV, HCV, HBV, CMV infection, sexually transmitted infections( gonorrhea, chlamydia, plasma mycoplasma, ureaplasma infection), tuberculosis, HIV infection,e.

Advantage of PCR in the diagnosis of infectious diseases before other methods of research is as follows:

■ The causative agent of infection can be found in any biological environment of the body, including the material obtained by biopsy;

■ Diagnosis of infectious diseases at the earliest stages of the disease;

■ quantitative evaluation of the results of studies( how many viruses or bacteria are contained in the material under investigation);

■ high sensitivity of the method;for example, the sensitivity of PCR to detect DNA of HBV virus in the blood is 0.001 pg / ml( approximately 4x102 copies / ml), while the sensitivity of the DNA hybridization method using branched probes is 2.1 pg / ml( approximately 7x105 copies / ml).