Detection of STIs: Diagnosing Neisseria gonorrhoeae with RT-PCR

Gonorrhea is a sexually transmitted infection (STI) caused by Neisseria gonorrhoeae, a bacterium that can lead to serious complications if not detected and treated in time. It primarily affects young individuals between the ages of 15 and 24. Due to its high prevalence, accurate diagnostic methods are essential. RT-PCR is one of the most effective tests for identifying this bacterium, as it amplifies and detects specific fragments of its DNA.

How is the sample collected for diagnosis?

The diagnostic process begins with the collection of a sample from the affected area using a swab. This sample is then placed in a 2 mL tube and transported to the laboratory for analysis. Proper sample collection and handling are crucial to ensuring reliable results.

DNA extraction

Once in the laboratory, DNA extraction is performed, a process that varies depending on the extraction kit used but generally follows these steps:

• Cell lysis with specific reagents and centrifugation to separate cellular components.
• Addition of chloroform, incubation, and subsequent centrifugation to obtain the DNA-containing supernatant.
• DNA precipitation with isopropanol and removal of impurities through ethanol washes.
• Storage of the extracted DNA at -20°C for further analysis.

For sample centrifugation, a device such as the Bioprocen 22 R is used. This high-performance refrigerated centrifuge is designed for handling microtubes. Its versatility lies in the variety of available rotors, allowing it to adapt to different laboratory needs and ensuring efficient DNA separation at every stage of the process.

Follow this link to learn more about this equipment.

Amplification and detection with RT-PCR

The goal of RT-PCR is to amplify the isolated DNA. To achieve this, a set of “primers” targeting specific genes—those found in Neisseria gonorrhoeae but not in human DNA is added to the extracted DNA, along with an enzyme called RT-polymerase, which performs the amplification.

Simply put, these primers act as markers, binding to the genes of interest and directing RT-polymerase to amplify those specific genes.

The test results depend on the amount of genetic material present in the sample:

• Healthy individuals: No amplification occurs since Neisseria gonorrhoeae DNA is not detected.
• Asymptomatic patients: Minimal amplification is observed, indicating a low bacterial load.
• Symptomatic patients: High amplification is detected due to a large amount of bacterial DNA.

The diagnosis of gonorrhea using RT-PCR is a fundamental tool for detecting Neisseria gonorrhoeae, allowing for rapid and accurate identification. The combination of proper sample collection, the use of laboratory equipment such as the Bioprocen 22 R, and advanced genetic amplification technology ensures reliable results for timely treatment.