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Plant pathogens are causing substantial economic losses and thus became a significant threat to global agriculture. Effective and timely detection methods are prerequisite for combating the damages caused by the plant pathogens. In the realm of plant pathogen detection, the isothermal amplification techniques, e.g., recombinase polymerase amplification (RPA) and loop-mediated isothermal amplification (LAMP), have emerged as a fast, precise, and most sensitive alternative to conventional PCR but they often comprise high rates of non-specific amplification and operational complexity. In recent advancements, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated nuclease Cas systems, particularly Cas12, have emerged as powerful tools for highly sensitive, specific, and rapid pathogen detection. Exploiting the collateral activities of Cas12, which selectively cleaves single-stranded DNA (ssDNA), novel detection platforms have been developed. The mechanism employs the formation of a triple complex molecule comprising guide RNA, Cas12 enzyme, and the substrate target nucleotide sequence. Upon recognition of the target, Cas12 indiscriminately cleaves the DNA strand, leading to the release of fluorescence from the cleaved ssDNA reporter. Integration of isothermal amplification methods with CRISPR/Cas12 enables one-step detection assays, facilitating rapid pathogen identification within 30 min at a single temperature. This integrated RPA-CRISPR/Cas12a approach eliminates the need for RNA extraction and cDNA conversion, allowing direct use of crude plant sap as a template. With an affordable fluorescence visualization system, this portable method achieves 100-fold greater sensitivity than conventional techniques. This review summarizes recent advances in RPA-CRISPR/Cas12a for detecting plant pathogens, covering primer design, field-level portability, and enhanced sensitivity., Competing Interests: Conflicts of interestThe authors have no conflicts of interest., (© King Abdulaziz City for Science and Technology 2024. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

In sugarcane ( Saccharum spp. hybrids) cultivation, viral diseases pose a great challenge across the globe. Yellow leaf (YL) disease is one of the important viral diseases caused by Sugarcane yellow leaf virus (ScYLV), a positive-sense ssRNA virus, genus Polerovirus , family Solemoviridae. The disease symptoms appear in later stages of crop growth during grand growth to maturity phase with intense midrib yellowing in the abaxial leaf surface. At present, this disease is managed through tissue (meristem) culture and healthy seed nurseries in India. However, the virus-free plants are infected quickly by secondary inoculum from aphid vectors in the field, which necessitates the importance of developing YL-resistant varieties. We screened about 600-625 sugarcane parental clones to identify true YL resistance based on 0-5 disease rating scale since 2015 and categorised them as resistant, moderately resistant, moderately susceptible, susceptible and highly susceptible. Leaf samples were collected from all these categories of plants during 2018-20 for the viral titre estimation through absolute quantification method (qRT-PCR assay). The viral load was invariably high in all categories of susceptible samples that ranged from 4.40 × 10 2 to 8.429 × 10 6 , whereas in YL-free asymptomatic clones, the viral load ranged from 82.35 ± 5.90 to 5.121 × 10 4 . The results clearly indicated that highest viral titre of 10 5 -10 7 copies was present in all the susceptible clones irrespective of their disease severity grades. Our results clearly established that about 22.85% of apparently resistant sugarcane clones remained free from YL symptoms with significantly low ScYLV titre although we could not find a significant correlation between virus titre and symptom expression. The identified resistant parents will serve as sources of YL resistance to develop virus resistant sugarcane varieties., Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03541-y., Competing Interests: Conflict of interestThe authors declared that there is no conflict of interest., (© King Abdulaziz City for Science and Technology 2023, Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)