Methods for Preparation of MS2 Phage-Like Particles and Their Utilization as Process Control Viruses in RT-PCR and qRT-PCR Detection of RNA Viruses From Food Matrices and Clinical Specimens.

Abstract

RNA viruses are pathogenic agents of many serious infectious diseases affecting humans and animals. The detection of pathogenic RNA viruses is based on modern molecular methods, of which the most widely used methods are the reverse transcription polymerase chain reaction (RT-PCR) and the real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). All steps of RT-PCR and qRT-PCR should be strictly controlled to ensure the validity of obtained results. False-negative results may be caused not only by inhibition of RT or/and PCR steps but also by failure of the nucleic acid extraction step, particularly in the case of viral RNA extraction. The control of nucleic acid extraction generally involves the utilization of a non-pathogenic virus (process control virus) of similar structural properties to those of the target virus. Although in clinical samples the use of such process control virus is only recommended, in other kinds of settings such as food matrices its use is necessary. Currently, several different process control viruses are used for these purposes. Process control viruses can also be constructed artificially using technology for production of MS2 phage-like particles, which have many advantages in comparison with other used controls and are especially suited for controlling the detection and quantification of certain types of RNA viruses. The technology for production of MS2 phage-like particles is theoretically well established, uses the knowledge gained from the study of the familiar bacteriophage MS2 and utilizes many different approaches for the construction of the various process control viruses. Nevertheless, the practical use of MS2 phage-like particles in routine diagnostics is relatively uncommon. The current situation with regard to the use of MS2 phage-like particles as process control viruses in detection of RNA viruses and different methods of their construction, purification and use are summarized and discussed in this review.