Development of a multienzyme isothermal rapid amplification and lateral flow dipstick combination assay for bovine coronavirus detection

Ji, Chengyuan; Feng, Yiqiu; Sun, Ruini; Gu, Qibing; Zhang, Yao; Ma, Jiale; Pan, Zihao; Yao, Huochun

doi:10.3389/fvets.2022.1059934

Cited by 10 publications

(13 citation statements)

References 35 publications

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“…However, only 16.6% of samples from A State tested positive for the tlh gene using the PCR assay, and all samples from the other three States tested negative. Our results suggest that the MIRA-LFD assay is as sensitive as the qPCR assay, taken together with previous studies [31,38,41]. Interestingly, all oyster samples from all four States tested negative for the tdh and trh genes using the MIRA-LFD, PCR, and qPCR assays.…”

Section: Detection Of Tlh Trh and Tdh Genes From Oyster Samplessupporting

confidence: 80%

Development of Multienzyme Isothermal Rapid Amplification (MIRA) Combined with Lateral-Flow Dipstick (LFD) Assay to Detect Species-Specific tlh and Pathogenic trh and tdh Genes of Vibrio parahaemolyticus

Park,

Zhang

2024

Pathogens

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Vibrio parahaemolyticus causes severe gastroenteritis in humans after consuming contaminated raw or undercooked seafood. A species-specific marker, the thermolabile hemolysin (tlh) gene, and two pathogenic markers, thermostable-related hemolysin (trh) and thermostable-direct hemolysin (tdh) genes, have been used to identify V. parahaemolyticus and determine its pathogenicity using both PCR and qPCR assays. To enable testing in field conditions with limited resources, this study aimed to develop a simple and rapid method to detect the species-specific (tlh) and pathogenic (trh and tdh) genes of V. parahaemolyticus using multienzyme isothermal rapid amplification (MIRA) combined with a lateral-flow dipstick (LFD). The amplification of the tlh, trh, and tdh genes could be completed within 20 min at temperatures ranging from 30 to 45 °C (p < 0.05). The test yielded positive results for V. parahaemolyticus but produced negative results for nine Vibrio species and eighteen foodborne pathogenic bacterial species. MIRA-LFD could detect 10 fg of DNA and 2 colony-forming units (CFU) of V. parahaemolyticus per reaction, demonstrating a sensitivity level comparable to that of qPCR, which can detect 10 fg of DNA and 2 CFU per reaction. Both MIRA-LFD and qPCR detected seven tlh-positive results from thirty-six oyster samples, whereas one positive result was obtained using the PCR assay. No positive results for the trh and tdh genes were obtained from any oyster samples using MIRA-LFD, PCR, and qPCR. This study suggests that MIRA-LFD is a simple and rapid method to detect species-specific and pathogenic genes of V. parahaemolyticus with high sensitivity.

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Section: Detection Of Tlh Trh and Tdh Genes From Oyster Samplessupporting

confidence: 80%

Development of Multienzyme Isothermal Rapid Amplification (MIRA) Combined with Lateral-Flow Dipstick (LFD) Assay to Detect Species-Specific tlh and Pathogenic trh and tdh Genes of Vibrio parahaemolyticus

Park,

Zhang

2024

Pathogens

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“…Interestingly, CkChpV alone can be precisely detected, with no cross-reaction with other common viruses, thereby allowing the accurate interpretation of data. MIRA combined with LFD can completely replace conventional PCR and only requires a constant temperature instrument in areas with poor research conditions for food safety, pathogen detection, and other application related to nucleic acid testing ( Ji et al, 2022 ).…”

Section: Discussionmentioning

confidence: 99%

Multienzyme isothermal rapid amplification and lateral flow dipstick combination assay for visible detection of chicken chaphamaparvovirus

Cui,

Xu,

et al. 2023

Poultry Science

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“…This event occurs at both the forward and reverse end of the DNA. Once the primer recombinase complex binds to the ssDNA, DNA polymerase synthesizes the new complementary DNA strand [ 69 ]. The use of helicase (gp41) and recombinases ( Streptomyces coelicolor recA, SC-recA) allows for rapid amplification that can happen between 10–30 min at a constant temperature between 35–42 °C [ 68 , 69 ].…”

Section: Raa and Mira With Paper Microfluidicsmentioning

confidence: 99%

“…Once the primer recombinase complex binds to the ssDNA, DNA polymerase synthesizes the new complementary DNA strand [ 69 ]. The use of helicase (gp41) and recombinases ( Streptomyces coelicolor recA, SC-recA) allows for rapid amplification that can happen between 10–30 min at a constant temperature between 35–42 °C [ 68 , 69 ]. MIRA has also been demonstrated with paper strips and paper microfluidic platforms [ 68 , 69 , 70 , 71 , 72 , 73 , 74 , 75 ].…”

Section: Raa and Mira With Paper Microfluidicsmentioning

confidence: 99%

“…The use of helicase (gp41) and recombinases ( Streptomyces coelicolor recA, SC-recA) allows for rapid amplification that can happen between 10–30 min at a constant temperature between 35–42 °C [ 68 , 69 ]. MIRA has also been demonstrated with paper strips and paper microfluidic platforms [ 68 , 69 , 70 , 71 , 72 , 73 , 74 , 75 ]. Like the RAA, all paper-based MIRA assays utilized LFIA with colorimetric detection.…”

Section: Raa and Mira With Paper Microfluidicsmentioning

confidence: 99%

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Recent Uses of Paper Microfluidics in Isothermal Nucleic Acid Amplification Tests

Reynolds,

Loeffler,

Leigh

et al. 2023

Biosensors

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Isothermal nucleic acid amplification tests have recently gained popularity over polymerase chain reaction (PCR), as they only require a constant temperature and significantly simplify nucleic acid amplification. Recently, numerous attempts have been made to incorporate paper microfluidics into these isothermal amplification tests. Paper microfluidics (including lateral flow strips) have been used to extract nucleic acids, amplify the target gene, and detect amplified products, all toward automating the process. We investigated the literature from 2020 to the present, i.e., since the onset of the COVID-19 pandemic, during which a significant surge in isothermal amplification tests has been observed. Paper microfluidic detection has been used extensively for recombinase polymerase amplification (RPA) and its related methods, along with loop-mediated isothermal amplification (LAMP) and rolling circle amplification (RCA). Detection was conducted primarily with colorimetric and fluorometric methods, although a few publications demonstrated flow distance- and surface-enhanced Raman spectroscopic (SERS)-based detection. A good number of publications could be found that demonstrated both amplification and detection on paper microfluidic platforms. A small number of publications could be found that showed extraction or all three procedures (i.e., fully integrated systems) on paper microfluidic platforms, necessitating the need for future work.

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Development of a multienzyme isothermal rapid amplification and lateral flow dipstick combination assay for bovine coronavirus detection

Cited by 10 publications

References 35 publications

Development of Multienzyme Isothermal Rapid Amplification (MIRA) Combined with Lateral-Flow Dipstick (LFD) Assay to Detect Species-Specific tlh and Pathogenic trh and tdh Genes of Vibrio parahaemolyticus

Development of Multienzyme Isothermal Rapid Amplification (MIRA) Combined with Lateral-Flow Dipstick (LFD) Assay to Detect Species-Specific tlh and Pathogenic trh and tdh Genes of Vibrio parahaemolyticus

Multienzyme isothermal rapid amplification and lateral flow dipstick combination assay for visible detection of chicken chaphamaparvovirus

Recent Uses of Paper Microfluidics in Isothermal Nucleic Acid Amplification Tests

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