Accelerated Molecular Detection of Chlamydia trachomatis (CT)

Chlamydia trachomatis (CT) infections happen more often than we think and patients can also be asymptomatic (over 64.000 positive test results in The Netherlands in 2022, RIVM STD report 2022). Molecular diagnostics are the gold standard for diagnosing CT. However, as samples have to be sent to the laboratory, it can take a few days before the results are reported back to the patient. This means that the patient has to travel to the general practitioner or sexual health center at least twice and cannot be treated immediately. In some cases, it might happen that a doctor feels the need to prescribe antibiotics before knowing the test results.

• Project needs and goals
• Methods 
• Results 

Project goals 

The different bottlenecks that Chlamydia infection presents can be divided into two different parts, which determine the two different goals of the project:

• The need for a fast diagnosis for individual patients.
• The need of estimating infection rates in regions to screen communities of specific locations (through wastewater).

Methods

Clinical samples were collected and prepared for detection in 96 well plates, which were sealed in the Plate sealer and incorporated into the NextGenPCR. It is an instrument developed by Molecular Biology Systems (MBS), which elimiantes ramping rates when changing temperatures. Therefore, the instrument goes from melting to annealing temperatures in less than 0.1 second. As a consequence, the higher speed enables to process more samples with the best solution for high throughput. Furthermore, it consumes only 25 watts, so helps researchers to save energy costs.

NextGenPCR is used in the project for the amplification of CT genetic material. The amplification signal is read with the FLUOstar plate reader from BMG Labtech and later analysed with Excel.  

Results

Due to NextGenPCR, the time-to-result for CT from patient material has been accelerated compared to conventional real-time PCR. By using exactly the same protocol as the conventional real-time PCR on the NextGenPCR platform results were available 30% faster. This can be explained by the fact that the NextGenPCR system (shown in Figure 1) transfers the PCR plates to different heating blocks at set temperatures (no heating and cooling is necessary).

Nevertheless, after the optimization of NextGenPCR protocols for CT,  an additional decrease was accomplished resulting in CT results after approximately 17 minutes. The findings highlighted above show that NextGenPCR can provide PCR results for CT faster. It could accelerate the diagnosis of CT infections in patients, and create a more patient-friendly process. Patients have to see the doctor only once, reducing travel but also time-to-results. This can possibly result in prescribing (correct) treatment after short waiting times at a sexual health center).

With reducing PCR cycles from 45 to 30, and therefore having shorter protocol time, RFU remains high significantly. NTC= No template control.