A B S T R A C TCanine parvovirus type 2 (CPV-2) infection is the most lethal disease of dogs with higher mortality in puppies worldwide. In today's world, dogs are an integral part of our communities as well as dogs breeding and rearing has become a lucrative business. Therefore, a fast, accurate, portable, and costeffective CPV-2 detection method with the ability for on-site detection is highly desired. In this study, we for the first time proposed a nanosystem for CPV-2 DNA detection with RNA-guided RNA endonuclease Cas13a, which upon activation results in collateral RNA degradation. We expressed LwCas13a in prokaryotic expression system and purified it through nickel column. Activity of Cas13a was verified by RNA-bound fluorescent group while using a quenched fluorescent probe as signals. Further Cas13a was combined with Recombinase polymerase amplification (RPA) and T7 transcription to establish molecular detection system termed specific high-sensitivity enzymatic reporter un-locking (SHERLOCK) for sensitive detection of CPV-2 DNA. This nanosystem can detect 100 amol/L CPV-2 DNA within 30 min. The proposed nanosystem exhibited high specificity when tested for CPV-2 and other dog viruses. This CRISPR-Cas13a mediated sensitive detection approach can be of formidable advantage during CPV-2 outbreaks because it is time-efficient, less laborious and does not involve the use of sophisticated instruments.

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Nucleolus, which participates in many crucial cellular activities, is an ideal target for evaluating the state of a cell or an organism. Here, bright red‐emissive carbon dots (termed CPCDs) with excitation‐independent/polarity‐dependent fluorescence emission are synthesized by a one‐step hydrothermal reaction between congo red and p‐phenylenediamine. The CPCDs can achieve wash‐free, real‐time, long‐term, and high‐quality nucleolus imaging in live cells, as well as in vivo imaging of two common model animals—zebrafish and Caenorhabditis elegans (C. elegans). Strikingly, CPCDs realize the nucleolus imaging of organs/flowing blood cells in zebrafish at a cellular level for the first time, and the superb nucleolus imaging of C. elegans suggests that the germ cells in the spermatheca probably have no intact nuclei. These previously unachieved imaging results of the cells/tissues/organs may guide the zebrafish‐related studies and benefit the research of C. elegans development. More importantly, a novel strategy based on CPCDs for in vivo toxicity evaluation of materials/drugs (e.g., Ag+), which can visualize the otherwise unseen injuries in zebrafish, is developed. In conclusion, the CPCDs represent a robust tool for visualizing the structures and dynamic behaviors of live zebrafish and C. elegans, and may find important applications in cell biology and toxicology.

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Establishing antimicrobial photodynamic therapy (PDT) approaches to enhance antimicrobial activities of insoluble photosensitizers in terms of their solubility, stability, and safety is highly desirable yet challenging. Here, we reported super-stable carboxymethyl chitosan-polyethyleneimine-protoporphyrin IX nanoparticles (CMCS-PEI-PpIX NPs) for effective photodynamic inactivation of bacteria and fungi with excellent biocompatibility. CMCS-PEI-PpIX self-assembled into NPs in solution with significantly improved singlet oxygen generation. Consequently, NPs exhibited exceptional photoinactivation toward a broad range of microbes with record low minimal bactericidal concentrations. NPs were highly stable after 28 days of storage. Furthermore, NPs exhibited negligible cytotoxicity without hemolysis on red blood cells. NPs can efficiently attach to the microbial cell through electrostatic interaction to generate singlet oxygen with light irradiation, which caused severe damage to the cell membrane, causing microbial cell death. Overall, the development of CMCS-PEI-PpIX NPs provides a route for improving the microbicidal PDT efficacy, benefiting the clinic treatment of microbial infections.