**1. Introduction**

Infectious diseases (ID) are caused by pathogenic microorganisms, such as bacteria, viruses, parasites, or fungi, and the diseases can be spread, directly or indirectly, from one person to another. Scientific advances in the biomedical area since the first half of last century, represented by the development of therapeutic drugs, vaccines, and advanced sanitation technologies, were carried as result of the control or prevention of infectious diseases. These diseases are considered, at the global level, as some of the most common public health problems. The relevance of these pathologies is evidenced by the number of individuals reached, the lack of knowledge about the infectious agents, their socioeconomic impact, the deepening of the molecular studies involving a precise and fast diagnosis, and appeals public health agency studies aimed at the development of diagnostic techniques for the early detection of symptomatic carriers as well as the asymptomatic carriers of these infections.

Changes in society, technology, and the microorganisms themselves are contributing to the emergence of new diseases, the reemergence of diseases once controlled, and to the development of antimicrobial resistance. According to the World Health Organization (WHO), the IDs constitute a significant proportion of all human diseases known, and at least 25% of about 60 million deaths that occur worldwide each year are estimated to be due to infectious diseases [1, 2].

Scientific studies of infectious agents and diseases provide a knowledge for the development of diagnostic tests for such diseases, drugs to treat, and vaccines for prevention. Earlier, specific and effective diagnosis is one of the most appropriate forms and strategies for managing. According to [3], in American hospitals, about 5 million cases of infectious disease are reported annually. Besides that, most of the cases are unreported, resulting in substantial morbidity and mortality [3]. In the last 20 years, analysis methods based on the detection and sequencing of 16S rDNA have been widely used in place of conventional culture methods.

Unfortunately, despite in clinical laboratories, the diagnosis of infectious diseases is directly associated with time of pathogen identification by conventional culture methods as these tests suffer from long turnaround times, from hours to days. A technical difficulty encountered in these methods performed before the advent of the molecular techniques was that not all pathogens are cultivable, and culture conditions ordinarily are not known. Other limitations of the traditional diagnostic include requirements for additional testing and wait times for characterizing detected pathogens (i.e., discernment of species, virulence factors, and antimicrobial resistance) [3, 4].

Nucleic acid amplification is one of the most valuable tools in virtually all life science fields, including application-oriented fields such as clinical medicine, for diagnosis of infectious diseases.

The serological methods are limited by the cross-reactions between the types of agents and by the fact that some infectious agents have no clearly identified epitopes that are sufficiently specific, requiring them as a stringent clinic diagnostic. In addition, the specific IgM antibodies are detected only in acute phase of infection, and for detecting infections, the serological tests are inaccurate, labor-intensive, and unreliable. In the last 20 years, analysis methods based on the nucleic acid amplification have been widely used in all life science fields as a new way for the diagnosis of human pathogens like virus, bacteria, and parasites.

The use of amplification techniques such as Polymerase Chain Reaction (PCR) has long been used to detection, genotyping, and quantification of virus and bacteria in various clinical specimens, such as serum, plasma, urine, semen, and liquid cerebrospinal fluid (CSF).

PCR-based diagnostics have been effectively developed for a wide range of microorganisms. Due to its incredible sensitivity, specificity, reproducibility, broad dynamic range, and speed of amplification, PCR has been championed by infectious disease experts for identifying organisms that cannot be grown in vitro, or in instances where existing culture techniques are insensitive and/or need prolonged incubation times [5].

Advances in development of molecular technology and diagnostics have enhanced understanding IDs' etiology, pathogenesis, and molecular epidemiology, which provide basis for appropriate detection, quantification, prevention, and control measures as well as rational design of vaccine, by which some diseases have been successfully eliminated.

Since 1985, many PCR amplification-based techniques have been designed for detection and identification, including: multiplex PCR (M-PCR), LAMP-PCR, digital PCR (dPCR), and real-time PCR.
