Rapid assays allowing for on-site or in-field testing can enable us to detect food contamination in a timely manner, thus facilitating the insurance of food safety. Isothermal nucleic acid ...amplification has emerged as a tool for rapid and high-efficiency signal amplification for both nucleic acid and non-nucleic acid targets. In this review, we outline how the features of isothermal amplification techniques have been leveraged for food safety analysis, and present the principles of isothermal amplification techniques. We then introduce isothermal amplification-based approaches for foodborne pathogen and authenticity analysis by detecting nucleic acid markers. Further, the techniques for heavy metal, mycotoxin and antibiotic detection by isothermal amplification combined with functional nucleic acids were reviewed. And particularly, imaging methods enabled by isothermal amplification for in situ analyzing foodborne pathogens are highlighted. Current limitations of these technologies, such as robustness and enzyme preservation, that hinder broad application, are also discussed.
•Isothermal amplification promises sensitive on-site food safety analysis.•Biomolecular affinities broaden target scope of isothermal amplification.•Food safety analysis enabled by isothermal amplification was reviewed.•Isothermal amplification-based in situ analyzing foodborne pathogens was highlighted.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
N6-methyladenosine (m6A) is the most abundant post-transcriptional modification in RNA and has important implications in physiological processes and tumor development. However, sensitive and specific ...quantification of locus-specific m6A modification levels remains a challenging task. In the present work, a novel m6A-sensitive DNAzyme was utilized to directly detect m6A by coupling with a three-way junction-mediated isothermal exponential CRISPR amplification reaction for the first time. This method was built on the fact that the binding arm of the DNAzyme bound to the specific site and its core structure catalyzed the selective cleavage of unmodified adenine instead of methylated adenines. Subsequently, the intact RNA was identified by the proximity effect of the three-way junction. Enormous amounts of single-stranded DNA products were generated through a combination of SDA and EXPAR for signal amplification. The specific real-time curve of products was recorded through detecting the fluorescence intensity triggered by CRISPR Cas12a. As a result, methylation target of abundance down to 1% was successfully identified. In addition, this strategy could be used for the analysis of cell RNA extracts. Combined with an electrochemical sensor for quantitative detection of RNA methylation, we demonstrated the generality of as-proposed strategy. We envision the present method would provide a new platform for the analysis of m6A in RNA and promote its application in clinical diseases.
•DNAzyme and the three-way junction were firstly used to specifically identify m6A RNA.•The integration of EXPAR and CRISPR Cas12a assisted exponential amplification realized sensitive detection of m6A modifications down to 1% abundance.•This strategy was easily elaborated into multiple reactions for different detection purposes.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The recent pneumonia outbreak caused by a novel coronavirus (SARS-CoV-2) is posing a great threat to global public health. Therefore, rapid and accurate identification of pathogenic viruses plays a ...vital role in selecting appropriate treatments, saving people’s lives and preventing epidemics. It is important to establish a quick standard diagnostic test for the detection of the infectious disease (COVID-19) to prevent subsequent secondary spread. Polymerase chain reaction (PCR) is regarded as a gold standard test for the molecular diagnosis of viral and bacterial infections with high sensitivity and specificity. Isothermal nucleic acid amplification is considered to be a highly promising candidate method due to its fundamental advantage in quick procedure time at constant temperature without thermocycler operation. A variety of improved or new approaches also have been developed. This review summarizes the currently available detection methods for coronavirus nucleic acid. It is anticipated that this will assist researchers and clinicians in developing better techniques for timely and effective detection of coronavirus infection.
The recent pneumonia outbreak caused by a novel coronavirus (SARS-CoV-2) in China, poses a great threat to global public health. Therefore, rapid and accurate identification of pathogenic viruses plays a vital role in selecting appropriate treatments, saving people’s lives and preventing epidemics. This review summarizes the currently available detection methods for coronavirus nucleic acid. It is anticipated that this will assist researchers and clinicians in developing better techniques for timely and effective detection of coronavirus infection. Display omitted
•This review summarizes the currently available detection methods for coronavirus nucleic acid.•It will assist researchers in developing better techniques for timely and effective detection of coronavirus infection.•It will help the establishment of SARS-CoV-2 RNA detection method which is useful for the early diagnosis of COVID-19.
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FFLJ, GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, ODKLJ, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Influenza A epidemics, which occur annually in varying degrees worldwide, is a global challenge to healthcare facilities owing to several limitations of the current detection methods. Therefore, the ...development of a rapid, convenient, and economical method for the early diagnosis of influenza A will aid clinical treatment and epidemic control. Currently, most of the commonly used clinical rapid tests utilize colloidal gold test strips that detect specific influenza virus antigens but are limited by low sensitivity. Therefore, this study combined catalytic hairpin assembly (CHA) with colloidal gold immunochromatographic assay (GICA) to develop a highly sensitive and visual CHA-GICA test strip. Clinical sample analysis revealed that the sensitivity of the assay was 81.8% and 74% under optimal (35 °C) and room temperature (25 °C) conditions, respectively. In conclusion, this study developed a rapid nucleic acid assay for detecting influenza A virus with high sensitivity and specificity, which can improve the clinical detection of influenza A.
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•A new rapid detection method for influenza A virus.•Clinical sample analysis revealed that the sensitivity and specificity of the assay were 81.8% and 100%.•This method can be applied to point-of-care testing.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Among the ultimate requirements of biosensor technology, sensitivity, stability, and specificity are undoubtedly crucial features to enter the market. Actually, sensitivity is a major drawback for ...biosensors, especially where there is a low abundance of targets. To address this concern, biosensor technology has been supported by effective methodologies for the target isothermal amplification, successfully applied to sensitively detect nucleic acids of viral and bacterial sources, capable to operate in low-resource settings and instrumentations, an important prerequisite for microfluidics and point-of-care diagnostics. This review of 133 refs provides an in-depth description of the available methods for isothermal amplification of nucleic acids, with pros and cons, and their integration into different detection systems. The potential of combining the amplification-powered biosensor with technological advances in the branches of nanotechnology and paper microfluidics is also proved by excellent examples from the literature.
•The final requirement of biosensor technology for ultra-trace concentrations is undoubtedly sensitivity.•Isothermal amplification assisted biosensors are successfully applied to provide increased sensitivity.•An in-depth description of the available amplification methods is provided with integration into different configurations.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Infectious diseases are a serious global problem, which not only take an enormous human toll but also incur tremendous economic losses. In combating infectious diseases, rapid and accurate diagnostic ...tests are required for pathogen identification at the point of care (POC). In this review, investigations of diagnostic strategies for infectious diseases that are based on aptamers, especially nucleic acid aptamers, oligonucleotides that have high affinities and specificities toward their targets, are described. Owing to their unique features including low cost of production, easy chemical modification, high chemical stability, reproducibility, and low levels of immunogenicity and toxicity, aptamers have been widely utilized as bio-recognition elements (bio-receptors) for the development of infection diagnostic systems. We discuss nucleic acid aptamer-based methods that have been developed for diagnosis of infections using a format that organizes discussion according to the target pathogenic analytes including toxins or proteins, whole cells and nucleic acids. Also included is, a summary of recent advances made in the sensitive detection of pathogenic bacteria utilizing the isothermal nucleic acid amplification method. Lastly, a nucleic acid aptamer-based POC system is described and future directions of studies in this area are discussed.
•Aptamer-based diagnostic strategies for infectious diseases are summarized.•Novel, homogenous, aptamer-based nucleic acid sensing strategies are described.•A description is given of recent advances for sensitive pathogen detection utilizing isothermal nucleic acid amplification.•Aptamer-based POC diagnostic systems as well as future directions of studies in this area are discussed.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Global food safety stands out as a prominent public concern, affecting populations worldwide. The recurrent challenge of food safety incidents reveals the need for a robust inspection framework. In ...recent years, the integration of isothermal nucleic acid amplification with CRISPR-Cas12a techniques has emerged as a promising tool for molecular detection of food hazards, presenting next generation of biosensing for food safety detection. This paper provides a comprehensive review of the current state of research on the synergistic application of isothermal nucleic acid amplification and CRISPR-Cas12a technology in the field of food safety. This innovative combination not only enriches the analytical tools, but also improving assay performance such as sensitivity and specificity, addressing the limitations of traditional methods. The review summarized various detection methodologies by the integration of isothermal nucleic acid amplification and CRISPR-Cas12a technology for diverse food safety concerns, including pathogenic bacterium, viruses, mycotoxins, food adulteration, and genetically modified foods. Each section elucidates the specific strategies employed and highlights the advantages conferred. Furthermore, the paper discussed the challenges faced by this technology in the context of food safety, offering insightful discussions on potential solutions and future prospects.
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BFBNIB, GIS, IJS, KISLJ, NUK, PNG, UL, UM, UPUK
As meat adulteration issue is becoming increasingly prominent worldwide, it is crucial to realize on-site detection with simple equipment. Conventional nucleic acid amplification methods are reliable ...but the requirement of complex equipment, skilled technicians and long operation time limit their on-site use. Here, a simple denaturation bubble-mediated strand exchange amplification method (SEA) requiring only a pair of primers and one polymerase was first reported for identifying adulteration of pork source by targeting the specie-specific mitochondrial DNA sequence. The SEA method displayed good specificity for pork and could detect as low as 30 pg/μL pork DNA. In binary mixtures, the SEA method could detect 1% pork meat total DNA by both colorimetric and fluorescence determination, fulfilling the requirement of artificial meat adulteration. Excitedly, the whole detection process could be finished within 1 h by coupling with fast tissue DNA extraction method, only requiring a simple heating block. Therefore, with simplicity and rapidity, SEA method will be suitable for on-site meat species identification.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Because of high sensitivity and specificity, isothermal nucleic acid amplification are widely applied in many fields. To facilitate and improve their performance, various nanomaterials, like ...nanoparticles, nanowires, nanosheets, nanotubes, and nanoporous films are introduced in isothermal nucleic acid amplification. However, the specific application, roles, and prospect of nanomaterials in isothermal nucleic acid amplification have not been comprehensively reviewed. Here, the application of different nanomaterials (0D, 1D, 2D, and 3D) in isothermal nucleic acid amplification is comprehensively discussed and recent progress in the field is summarized. The nanomaterials are mainly used for reaction enhancer, signal generation/amplification, or surface loading carriers. In addition, 3D nanomaterials can be also functioned as isolated chambers for digital nucleic acid amplification and the tools for DNA sequencing of amplified products. Challenges and future recommendations are also proposed to be better used for recent covid‐19 detection, point‐of‐care diagnostic, food safety, and other fields.
The application of different nanomaterials (0D, 1D, 2D, and 3D) in isothermal nucleic acid amplification is comprehensively discussed. The nanomaterials are mainly used for reaction enhancer, signal generation/amplification, or surface loading carriers. And 3D nanomaterials can be also functioned as isolated chambers for digital nucleic acid amplification and the tools for DNA sequencing.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Mobile microbiology is an evolving concept that has the potential to reduce morbidity and mortality associated with infectious diseases on a global level. Molecular methods used in the context of ...mobile microbiology ensure rapid and accurate aetiological diagnostics and allow timely initiation of clinical care. The great majority of published data regarding molecular diagnostics in mobile laboratories have focused on emerging viral infections and using laboratory-developed assays. Use of clinically validated and commercially available molecular diagnostic instruments in routine diagnostics of infectious diseases in mobile laboratories has received only limited attention in the field.
This review summarizes the suitability of a range of portable diagnostic molecular instruments for application in mobile laboratories by taking into account the instruments' analytical concepts, technical features and environmental requirements, as well as results of major validation studies.
Data on technical features of selected portable instruments were mainly extracted from manufacturers' websites. Information on validation studies of various molecular assays developed for the selected instruments was extracted from peer-reviewed publications searched for through PubMed.
Eight portable diagnostic molecular instruments (Alere q, GeneXpert Edge, GeneXpert Omni, Genedrive, PanNAT, Revogene, cobas Liat and ID Now) that are commercially available or in the launching stage are presented and evaluated in the context of the mobile microbiology concept, with particular emphasis on technical features and environmental requirements. Both the cobas Liat and the Alere i assays have been extensively validated in a variety of studies carried out in both adult and paediatric patients from various settings (ranging from primary care to emergency care departments in tertiary centres). Most studies showed comparable performance of cobas Liat and Alere i molecular assays with the standard-of-care in vitro diagnostics molecular assays routinely performed in dedicated/centralized molecular diagnostics laboratories. In addition, acceptable performance of Alere q and Genedrive instruments has been shown in implementations studies for early infant diagnosis of children born to human immunodeficiency virus-positive mothers and detection of hepatitis C virus RNA, respectively. Additional validation studies on existing (GeneXpert Edge, PanNAT, Revogene) and emerging (GeneXpert Omni) technologies are warranted.
Several portable molecular diagnostic platforms reviewed are suitable for mobile microbiology applications. Further development in this field should be directed toward providing a broader range of assays per instrument, multiplexing, reducing the frequency of invalid results, and price cutting.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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