High-sensitivity and high-precision detection method of Helicobacter pylori

Li Zhiyuan's team from Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, combined with the latest CRISPR/Cas12 a technology, proposed a high-sensitivity detection method for highly pathogenic Helicobacter pylori strains through ring-mediated isothermal amplification (LAMP) combined with the latest CRISPR/Cas12 a technology.

This method only needs to detect saliva samples to quickly and accurately detect positive patients infected with this strain, and the relevant research results have recently been Harnessing enhanced CRISPR/Cas12a trans-cleavage activity with extended reporters and reductants for early diagnosis of Helicobacter pylori, the causative agent of peptic ulcers and stomach cancer, was published in
Biosensors and Bioelectronics, a prestigious academic journal in the field of international medical testing.

Helicobacter pylori infection is the main causative agent of chronic gastritis, peptic ulcer, and is closely related to gastric cancer, gastric mucosa-associated lymphoid tissue (MALT) lymphoma and other diseases, and is listed as a first-class biological carcinogen
by the World Health Organization (WHO).
Most patients with Helicobacter pylori infection can be cured with multiple therapies according to the doctor's advice, but the vast majority of infected patients are almost asymptomatic in the early stage and are easily overlooked
.
In addition, not all people with Helicobacter pylori infection develop the disease, and strains with cytotoxin-associated protein (CagA) and vacuolar toxin (VacA) expression are the strains
associated with gastric inflammation, ulceration, and gastric cancer.
Therefore, there is an urgent clinical need for a rapid, accurate, highly specific, and highly sensitive on-site detection method to help prevent the spread of highly pathogenic Helicobacter pylori and real-time monitoring and diagnosis of already ill populations
。

At present, clinical H.
pylori detection methods mainly rely on histological or Helicobacter pylori culture, UBT (13Cor 14Curea breath test) and serological detection, but these methods have obvious shortcomings that hinder their popularization
.
In contrast, the ring-mediated isothermal amplification (LAMP) technique used by Li's team has been shown to be more sensitive (100 times) than PCR, and the test results can be obtained quickly using a simple water bath at a constant temperature (65°C).

In addition, the combination of CRISPR/Cas technology can further improve its detection sensitivity and reduce non-specific amplification
。 Specifically, this method significantly improves the backcut activity of LbCas12a by optimizing the ssDNA reporter with a novel buffer system to optimize CEXTRAR.
And the detection sensitivity is improved by 16 times, and picomolar sensitivity can be achieved without target pre-amplification, which can be used for the early detection of highly pathogenic Helicobacter pylori in clinical saliva samples.

Combined with LAMP technology, CEXTRAR can successfully obtain extremely high sensitivity CagA and VacA using all three detection methods real-time fluorescence (43 aM and 96 aM), in-tube fluorescence (430 aM and 960 aM), and lateral lateral flow readings (4.
3 aM and 9.
6 aM),
respectively.
A comparative test of the CEXTRAR method by the traditional 13C-urea breath test and PCR showed that CEXTRAR was able to A false-negative result
of the 13C urea breath test was detected.

Compared with the traditional Helicobacter pylori detection method, this method has the characteristics of simpler, faster and cheaper, and has higher sensitivity and specificity, and its promotion and application will be used in the detection and treatment of highly pathogenic Helicobacter pylori in the general population and related diseases such as gastritis, gastric ulcer and gastric cancer plays an important role
in the occurrence and improvement of prognosis.

Jean de Dieu Habimana, a doctoral candidate at Guangzhou Health Institute, is the first author of the paper, and Dr.
Huang Rongqi and Li Zhiyuan are co-corresponding authors
.
The collaboration of this study is Dongguan Hospital
affiliated to Southern Medical University.
The research was supported by the Natural Science Foundation of Guangdong Province and the Key R&D Program of Hunan Province.

Paper link

Schematic of the operation of the CEXTRAR platform for real-time fluorescence and colorimetric monitoring of Helicobacter pylori