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An ERA-CRISPR/Cas12a Method for Highly Sensitive Detection of Human Adenovirus Type 55.

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Titel: An ERA-CRISPR/Cas12a Method for Highly Sensitive Detection of Human Adenovirus Type 55.
Autoren: Zhang, Letian, Luo, Zhenghan, Wang, Taiwu, Han, Yifang, Ye, Fuqiang, Wang, Chunhui, Chen, Yue, Zhang, Jinhai
Quelle: Diagnostics (2075-4418); Nov2025, Vol. 17 Issue 21, p2725, 13p
Schlagwörter: HUMAN adenoviruses, CRISPRS, DETECTION of microorganisms, STATISTICAL accuracy, NUCLEIC acids, DETECTION algorithms, SENSITIVITY & specificity (Statistics), LOOP-mediated isothermal amplification
Abstract: Background/Objectives: Human adenovirus 55 (HAdV55) is a notable pathogen causing community-acquired pneumonia; outbreaks occur frequently in military camps, hospitals, and schools, thereby posing a threat to public health security. This study aimed to develop a method for detecting HAdV55 nucleic acid by targeting the conserved region of the Hexon gene. The sequence was amplified using enzymatic recombination isothermal amplification (ERA) technology, in conjunction with CRISPR-Cas12a technology, to enhance the amplification signal. Methods: Optimized primer and crRNA sequences were selected through ERA isothermal amplification testing. The ERA-CRISPR/Cas12a detection method was completed within 30 min at a constant temperature of 42 °C. Results: Sensitivity was assessed by detecting standard plasmids and live strains at various dilution concentrations. The detection limits were determined to be 9 copies/reaction for standard plasmids and 2.5 copies/reaction for cultured HAdV55 strains. Specificity tests were conducted on positive samples for five common respiratory pathogens and five other adenovirus subtypes, all of which showed no cross-reactivity. Conclusions: A rapid ERA-CRISPR/Cas12a nucleic acid detection method for HAdV55 has been successfully developed, demonstrating high sensitivity and specificity without the need for expensive or complex instruments. This method holds promise for on-site pathogen screening and detection. [ABSTRACT FROM AUTHOR]
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Datenbank: Biomedical Index
Beschreibung
Abstract:Background/Objectives: Human adenovirus 55 (HAdV55) is a notable pathogen causing community-acquired pneumonia; outbreaks occur frequently in military camps, hospitals, and schools, thereby posing a threat to public health security. This study aimed to develop a method for detecting HAdV55 nucleic acid by targeting the conserved region of the Hexon gene. The sequence was amplified using enzymatic recombination isothermal amplification (ERA) technology, in conjunction with CRISPR-Cas12a technology, to enhance the amplification signal. Methods: Optimized primer and crRNA sequences were selected through ERA isothermal amplification testing. The ERA-CRISPR/Cas12a detection method was completed within 30 min at a constant temperature of 42 °C. Results: Sensitivity was assessed by detecting standard plasmids and live strains at various dilution concentrations. The detection limits were determined to be 9 copies/reaction for standard plasmids and 2.5 copies/reaction for cultured HAdV55 strains. Specificity tests were conducted on positive samples for five common respiratory pathogens and five other adenovirus subtypes, all of which showed no cross-reactivity. Conclusions: A rapid ERA-CRISPR/Cas12a nucleic acid detection method for HAdV55 has been successfully developed, demonstrating high sensitivity and specificity without the need for expensive or complex instruments. This method holds promise for on-site pathogen screening and detection. [ABSTRACT FROM AUTHOR]
ISSN:20754418
DOI:10.3390/diagnostics15212725