Polymerase Chain Reaction (PCR) is a laboratory technique that identifies infectious diseases by finding a pathogen’s genetic material (DNA or RNA) in a sample. Unlike tests that look for the body’s immune response, PCR directly targets the microbe’s genetic signature. This allows for detection even in the earliest stages of an infection when only a small amount of the pathogen is present.
The power of PCR lies in its ability to amplify, or make many copies of, a specific segment of DNA. This accuracy and speed allow healthcare providers to make timely treatment decisions, which is important in controlling the spread of infectious diseases.
The PCR Process Explained
The PCR process is automated in a machine called a thermal cycler, which precisely controls a series of temperature changes that drive the reaction’s key steps.
The process consists of three main steps repeated in cycles. The first step is denaturation, where the reaction is heated to around 95°C (203°F) to separate the double-stranded DNA into two single strands. Next is annealing, where the temperature is lowered, allowing short DNA pieces called primers to bind to specific locations on the single-stranded DNA templates. These primers are designed to flank the genetic sequence of the targeted pathogen.
The final step is an extension, where the temperature is raised again, and an enzyme called DNA polymerase adds nucleotides to extend the primers, creating a new complementary strand of DNA. This three-step cycle is repeated 25 to 35 times, with each cycle doubling the amount of the target DNA. For RNA viruses like influenza or coronaviruses, an additional initial step called reverse transcription is needed. This step uses an enzyme to convert the virus’s RNA into DNA, which can then be amplified; this is known as RT-PCR.
Pathogens Identified by PCR
PCR technology can identify a wide range of pathogens, including viruses, bacteria, fungi, and parasites. Its high sensitivity allows for detection from various samples, such as blood, mucus, or tissue.
In virology, RT-PCR is the standard method for diagnosing many viruses. It is also used to detect and monitor viral loads for chronic infections. Viral pathogens identifiable through PCR include:
- Respiratory viruses like influenza, respiratory syncytial virus (RSV), and coronaviruses (including SARS-CoV-2)
- Chronic infections such as Human Immunodeficiency Virus (HIV) and hepatitis B and C
- Herpes viruses
- Human papillomavirus (HPV)
PCR has also replaced slower methods for identifying bacterial infections and can detect fungal pathogens and parasites. It can rapidly identify bacteria that cause illnesses such as:
- Strep throat (Streptococcus pyogenes)
- Lyme disease
- Chlamydia
- Gastroenteritis from sources like Campylobacter, Salmonella, and certain strains of E. coli
Comparing PCR to Other Diagnostic Tests
PCR tests are often compared to antigen tests, antibody tests, and traditional microbial cultures, each with distinct functions.
A primary distinction exists between PCR and antigen tests. PCR tests detect a pathogen’s specific genetic material and are highly sensitive, capable of identifying an infection in its earliest stages. Antigen tests, on the other hand, detect specific proteins on the surface of a virus. While antigen tests provide rapid results, they are less sensitive than PCR tests and may not detect low levels of the virus, potentially leading to false negatives.
PCR tests also differ from antibody, or serology, tests. A PCR test determines if there is an active infection by detecting the pathogen’s genetic code. In contrast, an antibody test detects the body’s immune response to a past infection by identifying antibodies in the blood. Therefore, an antibody test cannot determine if a person is currently sick, only that they were exposed in the past.
Compared to traditional culture methods, which involve growing microbes in a lab, PCR offers a substantial speed advantage. A culture can take days or weeks to yield a result, whereas a PCR test can provide answers within hours. Additionally, some pathogens are difficult to grow in a lab setting, but PCR can still detect their genetic material, making it a more reliable option.
Understanding Test Accuracy and Results
Test reliability is measured by sensitivity and specificity. Sensitivity is a test’s ability to correctly identify individuals with a disease, avoiding false negatives. Specificity is the ability to correctly identify those without the disease, avoiding false positives. PCR tests are recognized for high sensitivity and specificity, often approaching 100% under ideal conditions.
Despite this high accuracy, limitations can lead to incorrect results. A false negative, where an infected person tests negative, can occur for several reasons. These include collecting the sample too early in the infection, improper sample collection, or issues with sample handling and shipping.
A false positive, where a non-infected person tests positive, is less common but possible, often due to laboratory contamination. Because PCR is so sensitive, it can also detect fragments of a pathogen’s genetic material even after the person is no longer contagious. This means a positive result does not always indicate that someone can transmit the disease.