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Development of a PCR Lateral Flow Assay for Rapid Detection of Bacillus anthracis, the Causative Agent of Anthrax

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Abstract

Bacillus anthracis, the causative agent of anthrax is one of the most potent listed biological warfare agents. The conventional microbiological methods of its detection are labor intensive and time consuming, whereas molecular assays are fast, sensitive and specific. PCR is one of the most reliable diagnostic tools in molecular biology. The combination of PCR with lateral flow strips can reduce the diagnostic/detection time. It gives an alternative to gel electrophoresis and offers easy and clear interpretation of results. In the present study, a PCR Lateral flow (PCR-LF) assay targeting cya gene present on pXO1 plasmid of B. anthracis has been developed. The forward and reverse primers were tagged with 6-carboxyflourescein (6-FAM) and biotin, respectively, at 5′ end. The dual labeled PCR products were detected using lateral flow (LF) strips developed in this study. The PCR-LF assay could detect ≥ 5 pg of genomic DNA and ≥ 500 copies of target DNA harboured in a recombinant plasmid. The assay was able to detect as few as 103 and 10 CFU/mL of B. anthracis Sterne cells spiked in human blood after 6 and 24 h of enrichment, respectively.

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Acknowledgements

The authors are thankful to Director, DRDE, Gwalior for providing necessary facilities for this study (Acc No HR-2020-81). SB is thankful to Department of Biotechnology (DBT), Ministry of Science & Technology, Government of India for providing Senior Research Fellowship.

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Authors and Affiliations

  1. Bioprocess Technology Division, Defence Research and Development Establishment, Jhansi Road, Gwalior, 474002, India

    Swati Banger, Vijai Pal, N. K. Tripathi & A. K. Goel

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  1. Swati Banger
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  2. Vijai Pal
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  3. N. K. Tripathi
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  4. A. K. Goel
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Correspondence to A. K. Goel.

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The authors declare that they have no conflict of interest.

Ethical Approval

All the animal experiments were performed according to the Institutional Animal Ethics Committee (IAEC) vide registration number 37/1999/CPCSEA. The study was also approved by Institutional Biosafety Committee of Defence Research and Development Establishment, DRDO, Ministry of Defence, Government of India wide protocol no: BPT-02/58/AKG.

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Banger, S., Pal, V., Tripathi, N.K. et al. Development of a PCR Lateral Flow Assay for Rapid Detection of Bacillus anthracis, the Causative Agent of Anthrax. Mol Biotechnol 63, 702–709 (2021). https://doi.org/10.1007/s12033-021-00335-6

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  • DOI: https://doi.org/10.1007/s12033-021-00335-6

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