A CRISPR/Cas12a-based photothermal platform for the portable detection of citrus-associated Alternaria genes using a thermometer.

The outbreak of citrus brown spot because of Alternaria is one of the most destructive citrus diseases. Additionally, Alternaria species produce highly toxic mycotoxins. Mass screening is a valid method to control the spread of Alternaria. Morphological analysis and polymerase chain reaction combined with gene-sequencing technique are the most commonly used techniques for detecting Alternaria. However, they are limited by either low convenience and accuracy or low instrument accessibility and high cost. To balance the convenience, accuracy, test availability, and low cost, we develop a CRISPR/Cas12a-based photothermal platform for the portable detection of Alternaria genes using a thermometer. Using this platform, the Alternaria genes from the synthetic sequences and cultured fungus of citrus, tomato, and apple can be detected using a thermometer with a detection limit of 1.5 pM. With the aid of the CRISPR/Cas12a system, citrus-associated Alternaria can be specifically differentiated from other citrus disease-associated microorganisms. When the photothermal platform is applied to analyze the citrus fruit samples collected in the field, good-consistency results are obtained with the gene-sequencing technology. The excellent performance of this portable method shows that it can be applied to screen for Alternaria in resource-poor settings.