A novel loop-mediated isothermal amplification-lateral flow dipstick method for Helicobacter pylori detection

Liu, Wenwen; lu, gang; Wang, Yu; Chen, Zhenghong; Gao, Yunyun; Yin, Zhipeng; Wu, Yi; Lv, Xiaoqian; Guo, Pengbo; Zhao, Yinghui

doi:10.3389/fmicb.2023.1094600

Cited by 3 publications

References 32 publications

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RETRACTED: Development and application of a rapid screening method for Rodentibacter heylii and Rodentibacter pneumotropicus in laboratory animals

Yan,

Chen,

Xing

et al. 2024

Preprint

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Background: Rodentibacter pneumotropicus and Rodentibacter heylii (formerly known as [Pasteurella pneumotropica]) are pathogens that must be excluded from specific pathogen-free (SPF)-level laboratory animals. Currently, the isolated culture is troublesome. The loop-mediated isothermal amplification (LAMP) with lateral flow dipstick (LFD) is simple, fast, specific, sensitive, and the closed device to prevent aerosol contamination. Methods: In this study, the deoxynucleotide triphosphate (dNTPs), magnesium (Mg2+), and temperature were set to various settings to determine the reaction system. The specificity and sensitivity studies were used to confirm the accuracy of the method. Multiple polymerase chain reaction, Real-time Quantitative polymerase chain reaction (qPCR), and LAMP-LFD were used to compare and analyze the trachea and lung tissues. 842 SPF-level laboratory animal samples were detected with LAMP-LFD. Results: The optimal concentrations of dNTPs, Mg2+, and reaction temperature for Rodentibacter pneumotropicus and Rodentibacter heylii were 1.6 mM, 6 mM, 63°C and 64°C, respectively. The lower limit of detection for LAMP-LFD can reach 1×10-5 ng/μL, which can detect Rodentibacter pneumotropicus and Rodentibacter heylii, and it has a sensitivity of 10 orders of magnitude higher than PCR. LAMP-LFD had no cross-reaction with 15 strains of common pathogenic bacteria in laboratory animals. Among the 28 infected samples analyzed, Multiple PCR, qPCR, and LAMP-LFD demonstrated the ability to detect Rodentibacter pneumotropicus and Rodentibacter heylii in 21, 23, and 19 positive samples, respectively. Furthermore, the isolated culture, Multiple PCR, qPCR, and LAMP-LFD identified 2, 23, 41, and 37 positive cases of Rodentibacter pneumotropicus and Rodentibacter heylii in a total of 842 clinical samples. Conclusion: In conclusion, LAMP-LFD was established together with an integrated detection device which can accurate identification of R. heylii and R. pneumotropicus. This simple and practical method greatly facilitated the detection efforts of grassroots organizations.

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RETRACTED: Development and application of a rapid screening method for Rodentibacter heylii and Rodentibacter pneumotropicus in laboratory animals

Yan,

Chen,

Xing

et al. 2024

Preprint

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Multiple cross displacement amplification-based lateral flow biosensor for rapid and sensitive detection of Helicobacter pylori

Chen,

Zhou,

Wang

et al. 2024

Front. Cell. Infect. Microbiol.

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Helicobacter pylori (H. pylori, HP), recognized globally as one of the most widespread bacteria, serves as primary etiological agent for numerous gastroduodenal diseases, highlighting the urgent need to develop rapid and sensitive diagnostic method for H. pylori infection. Here, we devised a new diagnostic test that merged multiple cross displacement amplification (MCDA) with nanoparticle‐based lateral flow biosensor (LFB), termed HP‐MCDA‐LFB, to facilitate the rapid and sensitive detection of H. pylori. The whole detection workflow, which includes stages such as DNA template extraction (~15 min), MCDA pre-amplification (~40 min), and result readout (~2 min), was efficiently completed within 1 h. After optimization, the HP-MCDA-LFB assay demonstrated remarkable sensitivity in detecting H. pylori, with a detection threshold as low as 60 fg of genomic DNA (~56 copies) per microliter. Furthermore, the HP-MCDA-LFB assay also achieved a perfect specificity rate of 100%, exhibiting no cross-reactivity with non-Helicobacter isolates. Particularly, the clinical feasibility of HP-MCDA-LFB assay was validated using 40 antral mucosa samples, among which 17 tested positive for H. pylori, which was in complete agreement with the results obtained from the rapid urease test. In conclusion, the HP‐MCDA‐LFB method developed in this study is a rapid, sensitive, and specific method for diagnosing H. pylori infection, indicating great potential for H. pylori eradication therapy.

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Application of Loop-Mediated Isothermal Amplification Assay Combined with Lateral Flow Dipstick (LAMP-LFD) for Specific and Sensitive Detection of Acidovorax citrulli (Schaad et al.) Causing Bacterial Fruit Blotch in Cucurbit Plants

Lan,

Luo,

Gan

et al. 2024

Agronomy

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Acidovorax citrulli (Ac) is an important pathogenic bacterium causing bacterial fruit blotch (BFB) in Cucurbitaceae plants and is an important quarantine pest in China. This study was conducted to establish a rapid, convenient, and accurate visual method for detecting A. citrulli. A. citrulli-specific primers and a prober were designed based on the conserved region of the YD-repeat protein gene. Loop-mediated isothermal amplification combined with lateral flow dipstick (LAMP-LFD) was used to establish an assay for the rapid visual detection of A. citrulli by optimizing the reaction temperature and time. The specificity, sensitivity, and performance of the optimized LAMP-LFD assay were evaluated using the genomic DNA of the tested isolates, A. citrulli pure culture, infested seeds, commercial seeds, and leaf samples. The optimal assay temperature and time were 64 °C and 60 min, respectively. The assay specifically detected A. citrulli, and no cross-reactions were observed with the genomic DNA of other closely related species. The detection sensitivity of the LAMP-LFD for detecting pure genomic DNA, the bacterial suspension, bacterial amount on seeds (colony-forming units (CFU)·g−1), and infection rate of seeds (%) were 1 fg·μL−1, 8 CFU·mL−1, 5 CFU·g−1, and 0.05% infestation per reaction, respectively. The positive detection rate of the LAMP-LFD assay was 20–100% in seed samples (n = 1000 seeds) with 0.05–0.1% infestation. The LAMP-LFD assay rapidly and accurately detected A. citrulli in seeds and leaf tissues carrying pathogens. This assay thus offers the advantages of easy operation, rapidity, high specificity and sensitivity, low cost (no need for complex and expensive precision instruments), visualization of detection results, good stability, and strong applicability, which can be used for epidemiological studies and disease management.

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A novel loop-mediated isothermal amplification-lateral flow dipstick method for Helicobacter pylori detection

Cited by 3 publications

References 32 publications

RETRACTED: Development and application of a rapid screening method for Rodentibacter heylii and Rodentibacter pneumotropicus in laboratory animals

RETRACTED: Development and application of a rapid screening method for Rodentibacter heylii and Rodentibacter pneumotropicus in laboratory animals

Multiple cross displacement amplification-based lateral flow biosensor for rapid and sensitive detection of Helicobacter pylori

Application of Loop-Mediated Isothermal Amplification Assay Combined with Lateral Flow Dipstick (LAMP-LFD) for Specific and Sensitive Detection of Acidovorax citrulli (Schaad et al.) Causing Bacterial Fruit Blotch in Cucurbit Plants

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