A CRISPR/Cas12a-Based System for Sensitive Detection of Antimicrobial-Resistant Genes in Carbapenem-Resistant Enterobacterales.

Antimicrobial-resistant (AMR) bacteria pose a significant global health threat, and bacteria that produce New Delhi metallo-ß-lactamase (NDM) are particularly concerning due to their resistance to most ß-lactam antibiotics, including carbapenems. The emergence and spread of NDM-producing genes in food-producing animals highlight the need for a fast and accurate method for detecting AMR bacteria. We therefore propose a PCR-coupled CRISPR/Cas12a-based fluorescence assay that can detect NDM-producing genes (blaNDM) in bacteria. Thanks to its designed gRNA, this CRISPR/Cas12a system was able to simultaneously cleave PCR amplicons and ssDNA-FQ reporters, generating fluorescence signals. Our method was found to be highly specific when tested against other foodborne pathogens that do not carry blaNDM and also demonstrated an excellent capability to distinguish single-nucleotide polymorphism. In the case of blaNDM-1 carrying E. coli, the assay performed exceptionally well, with a detection limit of 2.7 × 100 CFU/mL: 100 times better than conventional PCR with gel electrophoresis. Moreover, the developed assay detected AMR bacteria in food samples and exhibited enhanced performance compared to previously published real-time PCR assays. Thus, this novel PCR-coupled CRISPR/Cas12a-based fluorescence assay has considerable potential to improve current approaches to AMR gene detection and thereby contribute to mitigating the global threat of AMR.

Recent advances in CRISPR-based systems for the detection of foodborne pathogens.

There has long been a need for more advanced forms of pathogen detection in the food industry. Though in its infancy, biosensing based on clustered regularly interspaced short palindromic repeats (CRISPR) has the potential to solve many problems that cannot be addressed using conventional methods. In this review, we briefly introduce and classify the various CRISPR/Cas protein effectors that have thus far been used in biosensors. We then assess the current state of CRISPR technology in food-safety contexts; describe how each Cas effector is utilized in foodborne-pathogen detection; and discuss the limitations of the current technology, as well as how it might usefully be applied in other areas of the food industry. We conclude that, if the limitations of existing CRISPR/Cas-based detection methods are overcome, they can be deployed on a wide scale and produce a range of positive food-safety outcomes.

Development of triplex PCR for simultaneous detection of soybean and wheat.

There is a constant demand for an effective detection method of major allergens, such as soybean (Glycine max) and wheat (Triticum aestivum). In this study, a triplex polymerase chain reaction (PCR) method was developed for rapid detection of soybean and wheat in processed food products. This triplex PCR contains soybean- and wheat-specific primer pairs and universal primer pair for plant species. Each primer pair was applied to 22 different plant species and showed high specificity with no amplification in non-target species. The sensitivity of triplex PCR for soybean and wheat was 10 pg. The detection limit for soybean and wheat in pea mixture was 0.1%. The developed triplex PCR showed high sensitivity and specificity and was applied to 21 different commercial products. The results were in accordance with the label. Thus, this method is expected to be useful in preventing allergy-related issues via accurate labeling of major allergens in food.

Case Series Study on the Use of BU Pharmacopuncture Treatment in Patients with Acute Lumbar Sprain.

OBJECTIVE: The purpose of this study was to investigate the clinical effects of BU pharmacopuncture therapy consisting of bear's gall(fel ursi) and ox bezoar(bovis calculus) on acute lumbar sprain. METHODS: 12 patients diagnosed as acute lumbar sprain in 6 designated local Korean medicine clinics from October 2017 to February 2018 were treated by BU pharmacopuncture. Several acupoints in abdomen and lumbar region were selected by clinicians at their own discretion. The effectiveness of the therapy was evaluated using VAS and ODI. After that we reviewed the medical records of all these patients to evaluate the effectiveness and safety of the therapy. RESULTS: The VAS and ODI scales were significantly decreased after BU pharmacopuncture therapy. And no major complications and adverse effects were reported. CONCLUSION: BU pharmacopuncture showed rapid pain relief in patients with acute lumbar sprain. It is possible to shorten the treatment period of acute lumbar sprain and prevent progressing to chronic back pain in advance. To establish the effects of BU pharmacopuncture therapy, more succeeding clinical and laboratory studies are needed.

Measurement of activity distribution in an Am-241 disc source using peeled-off Gafchromic EBT3 films.

Commercial alpha-emitting sources are fabricated mainly in a disc type. An alpha particle irradiator in Radiation Bioengineering Laboratory at Seoul National University was installed with an Am-241 disc source. Commercial Am-241 disc sources are fabricated by incorporating the radioactive element into a thin substrate layer. Those disc sources are utilized assuming that the radioactive element is uniformly distributed in the active layer of disc sources. In this study, we employed peeled-off Gafchromic EBT3 films to investigate the uniformity of areal radioactivity density over the disc source and to measure the effect of non-uniform activity distribution on dose distribution at the bottom of the cell culture dish positioned in a varying distance from the source. The measurements with the peeled-off EBT3 films informed that the areal activity density in the disc source differed by up to approximately 45% from the average. However, the inhomogeneous Am-241 distribution in a disc source did not affect the radial distribution of fluence rate at the inner bottom of cell dish when the dish is apart from the source sufficiently. The dose distribution measured with an EBT3 film nearly accorded with that obtained by Monte Carlo simulation assuming the uniform Am-241 activity distribution in the active layer of the disc source. Finally, the dose to a single-cell layer of 5 µm in a nominal thickness was obtained by Monte Carlo simulation assuming a uniform Am-241 activity distribution in the disc source at distances of 20 and 30 mm from the source. The cellular dose estimates were higher than the film dose estimates at all radial distances. The cellular dose decreased with an increasing radial distance from the center to a smaller extent than the EBT3 film dose did.

The effects of n-type doping on lithium storage in TiO2.

In this report, we discuss the Li-storage performance of niobium-doped TiO2 nanostructures (Ti(1-y)Nb(y)O(2+δ)), with a special focus on the effects of ionic/electronic charge carrier concentration (defect chemistry) on Li storage and transport properties. By Nb-doping, Li storage kinetics of titania electrode material is significantly improved mainly due to the increased electronic charge carrier concentration (n-type doping). However, it was found that there is a maximum beyond which further doping is rather detrimental to Li diffusion kinetics. Defect chemical analysis indicates that this limited doping effect is due to the trapping of free lithium ions by the critical electron-ion association reactions at room temperature such as Li(i)(•) + e' ⇌ Li(i)(×) and Nb(Ti)(•) + e' ⇌ Nb(Ti)(×).