Hannanta-Anan, Pimkhuan
Loading...
Preferred name
Hannanta-Anan, Pimkhuan
Alternative Name
Hannanta-anan, Pimkhuan
Hannanta-Anan, P.
Main Affiliation
Email
pimkhuan.ha@kmitl.ac.th
Now showing 1 - 2 of 2
- Some of the metrics are blocked by yourconsent settings
Item type:Publication, An RT-RPA-Cas12a platform for rapid and sensitive detection of tilapia lake virus(2022-11-15); Kitthamarat, MintraTilapia lake virus (TiLV) or Tilapia tilapinevirus is a highly contagious pathogen that causes severe symptoms and massive mortalities in tilapia, affecting numerous tilapia-farming communities across the continents. Though TiLV has grown into a serious threat that could undermine the aquaculture-based economy and global food security, effective therapeutics and vaccines are not yet commercially available, rendering prompt detection and control the most practical strategy. Current TiLV diagnostic techniques, however, face a number of constraints that restrict their utility, including low throughput and high resource demands imposed upon users. To fill in this critical gap, we developed a novel CRISPR-Cas12a method coupled with reverse-transcriptase recombinase polymerase amplification (RT-RPA-Cas12a) for TiLV detection, using Segment 9 as the target. The platform could detect as few as 200 copies of RNA and produced no false positive results when tested with other fish pathogens. The assay could be carried out at a constant temperature between 37 and 42 °C in less than an hour, allowing naked-eye interpretation of the results through lateral flow and smartphone-based readouts. Taken together, our RT-RPA-Cas12a platform is accurate, streamlined, and user-friendly, and thus has the potential to be a valuable asset in combating the growing threat posed by TiLV. - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Smartphone-compatible, CRISPR-based platforms for sensitive detection of acute hepatopancreatic necrosis disease in shrimp(2022-12-01); Chaijarasphong, ThawatchaiAcute Hepatopancreatic Necrosis Disease (AHPND), caused by bacterial isolates expressing PirAB binary toxins, represents the severest and most economically destructive disease affecting penaeid shrimp. Its rapid disease progression and associated massive mortalities call for vigilant monitoring and early diagnosis, but molecular detection methods that simultaneously satisfy the requirements of sensitivity, specificity, and portability are still scarce. In this work, the CRISPR-Cas12a technology was harnessed for the development of two fluorescent assays compatible with naked-eye visualization. The first assay, AP4-Cas12a, was based on the OIE-recommended AP4 two-tubed nested PCR method and was designed to bypass the time-consuming and potentially hazardous agarose gel electrophoresis step. Using AP4-Cas12a, the detection limit of 10 copies per reaction could be achieved within less than 30 minutes post-PCR. The second assay, RPA-Cas12a, utilized recombinase polymerase amplification (RPA) to rapidly and isothermally amplify the target DNA, followed by amplicon detection by Cas12a, resulting in a protocol that can be completed in less than an hour at a constant temperature of 37°C. The detection limit of RPA-Cas12a is 100 copies of plasmid DNA or 100 fg of bacterial genomic DNA per reaction. Importantly, we validated that both assays are compatible with a previously reported smartphone-based device for facile visualization of fluorescence, thereby providing an affordable option that requires less consumables than lateral flow detection. Using this portable device for readouts, the AP4-Cas12a and RPA-Cas12a methods showed excellent concordance with the AP4-agarose gel electrophoresis approach in the evaluation of clinical samples. Therefore, the developed Cas12a assays have the potential to streamline both in-laboratory and onsite diagnosis of AHPND.