Identifying the most at-risk age-group and longitudinal trends of drug allergy labeling amongst 7.3 million individuals in Hong Kong.

BACKGROUND: Incorrect drug 'allergy' labels remain a global public health concern. Identifying regional trends of drug allergy labeling can guide appropriate public health interventions, but longitudinal or population drug allergy studies remain scarce. We analysed the serial epidemiology of drug allergy labeling to identify specific subgroups at highest risk of drug allergy labeling for potential interventions. METHODS: Longitudinal, population-wide drug allergy labels and clinical data from over 7,337,778 individuals in Hong Kong between 2016 and 2021 were analysed. RESULTS: The absolute prevalence and incidence of documented drug allergy were 5.61% and 277/100,000 population, respectively. Annual incidence of new allergy labels was stable between 2016 and 2019, until a significant drop in 2020 (-16.3%) during the COVID19 pandemic. The most common allergy labels were anti-infectives (245,832 [44.5%]), non-steroidal anti-inflammatory (106,843 [19.3%]), and nervous system drugs (45,802 [8.3%]). The most common labeled culprits for the most severe immediate-type (anaphylaxis) and non-immediate-type (Stevens-Johnson syndrome) reactions were beta-lactams and nervous system drugs, respectively. For individuals at highest risk of labeling, there was significantly higher incidence of overall drug and beta-lactam allergy labeling amongst individuals aged > 40 years which contributed to the majority of newly labeled allergies (377,004, 68.2%). CONCLUSIONS: Contrary to traditional dogma, we identified disproportionately higher incidence of drug allergy labeling amongst older individuals, rather than the paediatric age group. We advocate for more population-wide drug allergy studies to investigate this phenomenon in other cohorts as well as future preventative and delabeling efforts focusing on the adult population.

Elucidating the structure of donor-acceptor conjugated polymer aggregates in liquid solution.

High-spin donor-acceptor conjugated polymers are extensively studied for their potential applications in magnetic and spintronic devices. Inter-chain charge transfer among these high-spin polymers mainly depends on the nature of the local structure of the thin film and π-stacking between the polymer chains. However, the microscopic structural details of high-spin polymeric materials are rarely studied with an atomistic force field, and the molecular-level local structure in the liquid phase remains ambiguous. Here, we have examined the effects of oligomer chain length, side chain, and processing temperature on the organization of the high-spin cyclopentadithiophene-benzobisthiadiazole donor-acceptor conjugated polymer in chloroform solvent. We find that the oligomers display ordered aggregates whose structure depends on their chain length, with an average π-stacking distance of 3.38 ± 0.03 Å (at T = 298 K) in good agreement with the experiment. Also, the oligomers with longer alkyl side chains show better solvation and a shorter π-stacking distance. Furthermore, the clusters grow faster at higher temperature with more ordered aggregation between the oligomer chains.

COVID-19 Vaccine Allergy Safety Track (VAS-Track) pathway: real-world outcomes on vaccination rates and antibody protection.

BACKGROUND: Misdiagnosed vaccine-related "allergies" lead to unnecessary vaccine deferrals and incomplete vaccinations, leaving patients unprotected against COVID-19. To overcome limitations and queues for Allergist assessment, the "VAS-Track" pathway was developed to evaluate patients via a multi-disciplinary triage model including nurses, non-specialists, and Allergists. OBJECTIVE: We assessed the effectiveness and safety of VAS-Track and evaluate its real-world impact in terms of vaccination rates and COVID-19 protection. METHODS: Patients referred to VAS-Track between September 2021 and March 2022 were recruited. Subgroup analysis was performed with prospective pre- and post-clinic antibody levels. RESULTS: Nurse-assisted screening identified 10,412 (76%) referrals as inappropriate. 369 patients were assessed by VAS-Track. Overall, 100% of patients were recommended to complete vaccination and 332 (90%) completed their primary series. No patients reported any significant allergic reactions following subsequent vaccination. Vaccination completion rates between patients seen by non-specialists and additional Allergist review were similar (90% vs. 89%, p = 0.617). Vaccination rates were higher among patients with prior history of immediate-type reactions (odds ratio: 2.43, p = 0.025). Subgroup analysis revealed that only 20% (56/284) of patients had seropositive COVID-19 neutralizing antibody levels (≥ 15 AU/mL) prior to VAS-Track, which increased to 92% after vaccine completion (pre-clinic antibody level 6.0 ± 13.5 AU/mL vs. post-clinic antibody level 778.8 ± 337.4 AU/mL, p > 0.001). CONCLUSIONS: A multi-disciplinary allergy team was able to streamline our COVID-19 VAS services, enabling almost all patients to complete their primary series, significantly boosting antibody levels and real-world COVID-19 protection. We propose similar multidisciplinary models to be further utilized, especially in the settings with limited allergy services.

The Role of the Allergist in Coronavirus Disease 2019 Vaccine Allergy Safety: A Pilot Study on a "Hub-and-Spoke" Model for Population-Wide Allergy Service.

BACKGROUND: Hong Kong started its coronavirus disease 2019 (COVID-19) vaccination program in February 2021. A territory-wide Vaccine Allergy Safety (VAS) clinic was set up to assess individuals deemed at "higher risk" of COVID-19 vaccine-associated allergies. A novel "hub-and-spoke" model was piloted to tackle the overwhelming demand of services by allowing nonallergists to conduct assessment. OBJECTIVE: To evaluate the outcomes of the VAS hub-and-spoke model for allergy assessment. METHODS: Records of patients attending the VAS hub-and-spoke Clinics between March and August 2021 were reviewed (n = 2725). We studied the overall results between the Hub (allergist led) and Spoke (nonallergist led) Clinics. The Hub and the Hong Kong West Cluster Spoke Clinic were selected for subgroup analysis as they saw the largest number of patients (n = 1411). RESULTS: A total of 2725 patients were assessed under the VAS hub-and-spoke model. Overall, 2324 patients (85.3%) were recommended to proceed with vaccination. Allergists recommended significantly more patients for vaccination than nonallergists (odds ratio = 21.58; P < .001). Subgroup analysis revealed that 881 of 1055 (83.5%) patients received their first dose of COVID-19 vaccination safely after assessment. Among those recommended vaccination, more patients assessed by allergists received their first dose of vaccination (odds ratio = 4.18; P < .001). CONCLUSION: The hub-and-spoke model has proven to be successful for the vaccination campaign. This study has illustrated the crucial role of allergists in countering vaccine hesitancy. Results from the study revealed considerable differences in outcomes between allergist-led and nonallergist-led clinics. Precise reasons for these differences warrant further evaluation. We are hopeful that the hub-and-spoke model can be similarly adapted for other allergist-integrative services in the future.

Regulation of quantum spin conversions in a single molecular radical.

Free radicals, generally formed through the cleavage of covalent electron-pair bonds, play an important role in diverse fields ranging from synthetic chemistry to spintronics and nonlinear optics. However, the characterization and regulation of the radical state at a single-molecule level face formidable challenges. Here we present the detection and sophisticated tuning of the open-shell character of individual diradicals with a donor-acceptor structure via a sensitive single-molecule electrical approach. The radical is sandwiched between nanogapped graphene electrodes via covalent amide bonds to construct stable graphene-molecule-graphene single-molecule junctions. We measure the electrical conductance as a function of temperature and track the evolution of the closed-shell and open-shell electronic structures in real time, the open-shell triplet state being stabilized with increasing temperature. Furthermore, we tune the spin states by external stimuli, such as electrical and magnetic fields, and extract thermodynamic and kinetic parameters of the transition between closed-shell and open-shell states. Our findings provide insights into the evolution of single-molecule radicals under external stimuli, which may proof instrumental for the development of functional quantum spin-based molecular devices.

Feasibility of a drug allergy registry-based excipient allergy database and call for universal mandatory drug ingredient disclosure: the case of PEG.

Background: Excipient allergy is a rare, but potentially lethal, form of drug allergy. Diagnosing excipient allergy remains difficult in regions without mandatory drug ingredient disclosure and is a significant barrier to drug safety. Objective: To investigate the feasibility of a drug allergy registry-based excipient database to identify potential excipient culprits in patients with history of drug allergy, using polyethylene glycol (PEG) as an example. Methods: An excipient registry was created by compiling the excipient lists pertaining to all available formulations of the top 50 most reported drug allergy culprits in Hong Kong. Availability of excipient information, and its relationship with total number of formulations of individual drugs were analysed. All formulations were checked for the presence or absence of PEG. Results: Complete excipient information was available for 36.5% (729/2,000) of all formulations of the top 50 reported drug allergy culprits in Hong Kong. The number of formulations for each drug was associated with proportion of available excipient information (ρ = 0.466, p = 0.001). Out of 729 formulations, 109 (15.0%) and 620 (85.0%) were confirmed to contain and not contain PEG, respectively. Excipient information was not available for the other 1,271 (63.6%) formulations. We were unable to confirm the presence or absence of PEG in any of the top 50 drug allergy culprits in Hong Kong. Conclusion: In countries without mandatory drug ingredient disclosure, excipient databases are unlikely able to identify potential excipient allergy in drug allergy patients. Legislations to enforce mandatory and universal ingredient disclosure are urgently needed.

Current microfluidic platforms for reverse engineering of cornea.

According to the World Health Organization, corneal blindness constitutes 5.1% of global blindness population. Surgical outcomes have been improved significantly in the treatment of corneal blindness. However, corneal transplantation is limited by global shortage of donor tissue, prompting researchers to explore alternative therapies such as novel ocular pharmaceutics to delay corneal disease progression. Animal models are commonly adopted for investigating pharmacokinetics of ocular drugs. However, this approach is limited by physiological differences in the eye between animals and human, ethical issues and poor bench-to-bedside translatability. Cornea-on-a-chip (CoC) microfluidic platforms have gained great attention as one of the advanced in vitro strategies for constructing physiologically representative corneal models. With significant improvements in tissue engineering technology, CoC integrates corneal cells with microfluidics to recapitulate human corneal microenvironment for the study of corneal pathophysiological changes and evaluation of ocular drugs. Such model, in complement to animal studies, can potentially accelerate translational research, in particular the pre-clinical screening of ophthalmic medication, driving clinical treatment advancement for corneal diseases. This review provides an overview of engineered CoC platforms with respect to their merits, applications, and technical challenges. Emerging directions in CoC technology are also proposed for further investigations, to accentuate preclinical obstacles in corneal research.

A Plus Shaped Cavity in Optical Fiber Based Refractive Index Sensor.

The optical fiber grating sensors have strong potential for the detection of biological samples. However, a careful effort is still in demand to enhance the performance of existing grating sensors especially in biological sensing. Therefore, in this work, we have introduced a novel plus shaped cavity (PSC) in optical fiber model and used it for the detection of haemoglobin (Hb) refractive index (RI). The numerical analysis of designed model is done by the testing of single and double vertical slots cavity in optical fiber core structure. The testing of designed sensor model is done at the wavelength of 800 nm at which the RI of oxygenated and deoxygenated Hb is 1.392 and 1.389, respectively. The analysis of reported PSC sensor model is done in the wide range of Hb RI from 1.333 to 1.392. The tested range of RI corresponds to the Hb concentration from 0 to 140 gl-1. The obtained results states that for the tested range of RI, the autocorrelation coefficient of R2 = 99.51 % is achieved. The analysis of projected work is done by using finite difference time domain (FDTD) method. The introduction of PSC can increase in sensitivity. In proposed PSC, the length and width of created slots are [Formula: see text] and [Formula: see text], respectively, which is quite enough to observe the response of analytes RI. This can minimize the creation of multiple gratings required for observing the analyte response.

Insights into Sorption and Molecular Transport of Aqueous Glucose into Zeolite Nanopores.

Liquid-phase heterogeneous catalysis using zeolites is important for biomass conversion to fuels and chemicals. There is a substantial body of work on gas-phase sorption in zeolites with different topologies; however, studies investigating the diffusion of complex molecules in liquid medium into zeolitic nanopores are scarce. Here, we present a molecular dynamics study to understand the sorption and diffusion of aqueous ß-d-glucose into ß-zeolite silicate at T = 395 K and P = 1 bar. Through 2-µs-long molecular dynamics trajectories, we reveal the role of the solvent, the kinetics of the pore filling, and the effect of the water model on these properties. We find that the glucose and water loading is a function of the initial glucose concentration. Although the glucose concentration increases monotonically with the initial glucose concentration, the water loading exhibits a nonmonotonic behavior. At the highest initial concentration (∼20 wt %), we find that the equilibrium loading of glucose is approximately five molecules per unit cell and displays a weak dependence on the water model. Glucose molecules follow a single-file diffusion in the nanopores due to confinement. The dynamics of glucose and water molecules slows significantly at the interface. The average residence time for glucose molecules is an order of magnitude larger than that in the bulk solution, while it is about twice as large for the water molecules. Our simulations reveal critical molecular details of the glucose molecule's local environment inside the zeolite pore relevant to catalytic conversion of biomass to valuable chemicals.

Alternative functions of CRISPR-Cas systems in the evolutionary arms race.

CRISPR-Cas systems of bacteria and archaea comprise chromosomal loci with typical repetitive clusters and associated genes encoding a range of Cas proteins. Adaptation of CRISPR arrays occurs when virus-derived and plasmid-derived sequences are integrated as new CRISPR spacers. Cas proteins use CRISPR-derived RNA guides to specifically recognize and cleave nucleic acids of invading mobile genetic elements. Apart from this role as an adaptive immune system, some CRISPR-associated nucleases are hijacked by mobile genetic elements: viruses use them to attack their prokaryotic hosts, and transposons have adopted CRISPR systems for guided transposition. In addition, some CRISPR-Cas systems control the expression of genes involved in bacterial physiology and virulence. Moreover, pathogenic bacteria may use their Cas nuclease activity indirectly to evade the human immune system or directly to invade the nucleus and damage the chromosomal DNA of infected human cells. Thus, the evolutionary arms race has led to the expansion of exciting variations in CRISPR mechanisms and functionalities. In this Review, we explore the latest insights into the diverse functions of CRISPR-Cas systems beyond adaptive immunity and discuss the implications for the development of CRISPR-based applications.

Virtual reality and augmented reality- emerging screening and diagnostic techniques in ophthalmology: A systematic review.

In health care, virtual reality (VR) and augmented reality (AR) have been applied extensively for many purposes. Similar to other technologies such as telemedicine and artificial intelligence, VR and AR may improve clinical diagnosis and screening services in ophthalmology by alleviating current problems, including workforce shortage, diagnostic error, and underdiagnosis. In the past decade a number of studies and products have used VR and AR concepts to build clinical tests for ophthalmology, but comprehensive reviews on these studies are limited. Therefore, we conducted a systematic review on the use of VR and AR as a diagnostic and screening tool in ophthalmology. We identified 26 studies that implemented a variety of VR and AR tests on different conditions, including VR cover tests for binocular vision disorder, VR perimetry for glaucoma, and AR slit lamp biomicroscopy for retinal diseases. In general, while VR and AR tools can become standardized, automated, and cost-effective tests with good user experience, several weaknesses, including unsatisfactory accuracy, weak validation, and hardware limitations, have prevented these VR and AR tools from having wider clinical application. Also, a comparison between VR and AR is made to explain why studies have predominantly used VR rather than AR.

CRISPRs in the human genome are differentially expressed between malignant and normal adjacent to tumor tissue.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) have been identified in bacteria, archaea and mitochondria of plants, but not in eukaryotes. Here, we report the discovery of 12,572 putative CRISPRs randomly distributed across the human chromosomes, which we termed hCRISPRs. By using available transcriptome datasets, we demonstrate that hCRISPRs are distinctively expressed as small non-coding RNAs (sncRNAs) in cell lines and human tissues. Moreover, expression patterns thereof enabled us to distinguish normal from malignant tissues. In prostate cancer, we confirmed the differential hCRISPR expression between normal adjacent and malignant primary prostate tissue by RT-qPCR and demonstrate that the SHERLOCK and DETECTR dipstick tools are suitable to detect these sncRNAs. We anticipate that the discovery of CRISPRs in the human genome can be further exploited for diagnostic purposes in cancer and other medical conditions, which certainly will lead to the development of point-of-care tests based on the differential expression of the hCRISPRs.

Reply to Fabre et al. Comment on "Tanmoy et al. CRISPR-Cas Diversity in Clinical Salmonella enterica Serovar Typhi Isolates from South Asian Countries. Genes 2020, 11, 1365".

We respectfully thank Fabre et al. [...].

An in-vitro study of subretinal perfluorocarbon liquid (PFCL) droplets and the physics of their retention and evacuation.

PURPOSE: To investigate the physics associated with the retention and removal of subretinal perfluorocarbon liquid (PFCL), as inspired by a series of anecdotal cases of spontaneous 'disappearance' of subretinal PFCL. METHODS: The profiles of subretinal PFCL in situ from published OCT images were studied and compared with that of PFCL droplets resting on a hydrophilic surface in vitro. A mathematical model based on Sampson's and Poiseuille's formula was developed to explain how evacuation of subretinal PFCL without aspiration could occur. RESULTS: The mathematical model suggested that in vivo subretinal PFCL can completely evacuate in less than a second via a 41-guage retinal hole. Perfluorocarbon liquid (PFCL) droplets in situ subretinally substantially varied in their aspect ratios (from 0.28 to 2.71) and their contact angles with the retinal pigment epithelium (from 98° to 155°). Conversely, PFCL in vitro had aspect ratios and contact angles close to 1 and 150° respectively. CONCLUSION: This study showed evidence that stretching of the retina to accommodate subretinal PFCL occurs, which might be responsible for the varied profile of the droplets and resultant forces that can cause retinal holes, and spontaneous evacuation of large PFCL droplets. By filling the vitreous cavity with PFCL, a small retinotomy alone might allow spontaneous evacuation without the need for aspiration.

Envelope Proteins Pertain with Evolution and Adaptive Mechanism of the Novel Influenza A/H1N1 in Humans.

The novel swine-origin influenza A/H1N1 virus (S-OIV) first detected in April 2009 has been identified to transmit from human to human directly and is the cause of currently emerged pandemic. In this study, nucleotide and deduced amino acid sequences of hemagglutinin (HA) and neuraminidase (NA) of the S-OIV and other influenza A viruses were analyzed through bioinformatic tools for phylogenetic analysis, genetic recombination and point mutation to investigate the emergence and adaptation of the S-OIV in human. The phylogenetic analysis showed that the HA comes from triple reassortant influenza A/H1N2 and the NA from Eurasian swine influenza A/H1N1 indicating HA and NA to descend from different lineages during the genesis of the S-OIV. Recombination analysis nullified the possibility of occurrence of recombination in HA and NA denoting the role of reassortment in the outbreak. Several conservative mutations are observed in the amino acid sequences of the HA and NA and this mutated residues are identical in the S-OIV. The results reported herein suggested the notion that the recent pandemic is the result of reassortment of different genes from different lineages of two envelope proteins, HA and NA which are responsible for antigenic activity of virus. This study further suggests that the adaptive capability of the S-OIV in human is acquired by the unique mutations generated during emergence.

Development of Dopamine Sensor Using Silver Nanoparticles and PEG-Functionalized Tapered Optical Fiber Structure.

This article presents a localized surface plasmon resonance (LSPR) phenomenon based optical fiber sensor (OFS) for the detection of dopamine (DA). DA functions as a hormone and a neurotransmitter in the human body and plays a crucial role in the peripheral system. To develop the OFS for DA detection, taper fiber probe was fabricated and immobilized with silver nanoparticles (AgNPs) and functionalized with Polyethylene glycol (PEG). The developed sensor shows the great selectivity in the presence of ascorbic acid (AA) oxidation due to PEG coating. The morphology of the AgNPs and uniformity of coating over the surface of sensing probe were confirmed with UV-visible spectrophotometer, transmission electron microscope (TEM), energy-dispersive X-ray spectroscopy (EDS), and scanning electron microscope (SEM). The calibration curve is found to be linear over the range of 10 nM-1 µM with the lowest detection limit of 0.058 µM. Also provides a wide dynamic range of detection (10 nm-100 µM). The parameters responsible for the performance of OFS, such as sensitivity, detection limit, and selectivity are greatly improved in the proposed sensor. The applicability of the proposed sensor has been validated and have the potential to use for routine diagnosis.

Campylobacter jejuni Cas9 Modulates the Transcriptome in Caco-2 Intestinal Epithelial Cells.

The zoonotic human pathogen Campylobacter jejuni is known for its ability to induce DNA-damage and cell death pathology in humans. The molecular mechanism behind this phenomenon involves nuclear translocation by Cas9, a nuclease in C. jejuni (CjeCas9) that is the molecular marker of the Type II CRISPR-Cas system. However, it is unknown via which cellular pathways CjeCas9 drives human intestinal epithelial cells into cell death. Here, we show that CjeCas9 released by C. jejuni during the infection of Caco-2 human intestinal epithelial cells directly modulates Caco-2 transcriptomes during the first four hours of infection. Specifically, our results reveal that CjeCas9 activates DNA damage (p53, ATM (Ataxia Telangiectasia Mutated Protein)), pro-inflammatory (NF-κB (Nuclear factor-κB)) signaling and cell death pathways, driving Caco-2 cells infected by wild-type C. jejuni, but not when infected by a cas9 deletion mutant, towards programmed cell death. This work corroborates our previous finding that CjeCas9 is cytotoxic and highlights on a RNA level the basal cellular pathways that are modulated.

CRISPR-Cas Diversity in Clinical Salmonella enterica Serovar Typhi Isolates from South Asian Countries.

Typhoid fever, caused by Salmonella enterica serovar Typhi (S. Typhi), is a global health concern and its treatment is problematic due to the rise in antimicrobial resistance (AMR). Rapid detection of patients infected with AMR positive S. Typhi is, therefore, crucial to prevent further spreading. Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated genes (CRISPR-Cas), is an adaptive immune system that initially was used for typing purposes. Later, it was discovered to play a role in defense against phages and plasmids, including ones that carry AMR genes, and, at present, it is being explored for its usage in diagnostics. Despite the availability of whole-genome sequences (WGS), very few studied the CRISPR-Cas system of S. Typhi, let alone in typing purposes or relation to AMR. In the present study, we analyzed the CRISPR-Cas system of S. Typhi using WGS data of 1059 isolates obtained from Bangladesh, India, Nepal, and Pakistan in combination with demographic data and AMR status. Our results reveal that the S. Typhi CRISPR loci can be classified into two groups: A (evidence level >2) and B (evidence level ≤2), in which we identified a total of 47 unique spacers and 15 unique direct repeats. Further analysis of the identified spacers and repeats demonstrated specific patterns that harbored significant associations with genotype, demographic characteristics, and AMR status, thus raising the possibility of their usage as biomarkers. Potential spacer targets were identified and, interestingly, the phage-targeting spacers belonged to the group-A and plasmid-targeting spacers to the group-B CRISPR loci. Further analyses of the spacer targets led to the identification of an S. Typhi protospacer adjacent motif (PAM) sequence, TTTCA/T. New cas-genes known as DinG, DEDDh, and WYL were also discovered in the S. Typhi genome. However, a specific variant of the WYL gene was only identified in the extensively drug-resistant (XDR) lineage from Pakistan and ciprofloxacin-resistant lineage from Bangladesh. From this work, we conclude that there are strong correlations between variations identified in the S. Typhi CRISPR-Cas system and endemic AMR positive S. Typhi isolates.

Tackling the chemical diversity of microbial nonulosonic acids - a universal large-scale survey approach.

Nonulosonic acids, commonly referred to as sialic acids, are a highly important group of nine-carbon sugars common to all domains of life. They all share biosynthetic and structural features, but otherwise display a remarkable chemical diversity. In humans, sialic acids cover all cells which makes them important for processes such as cellular protection, immunity and brain development. On the other hand, sialic acids and other nonulosonic acids have been associated with pathological processes including cancer and viral infections. In prokaryotes, nonulosonic acids are commonly associated with pathogens, which developed through molecular mimicry a strategy to circumvent the host's immune response. However, the remarkably large chemical diversity of prokaryotic nonulosonic acids challenges their discovery, and research on molecular characteristics essential for medical applications are often not feasible. Here, we demonstrate a novel, universal large-scale discovery approach that tackles the unmapped diversity of prokaryotic nonulosonic acids. Thereby, we utilize selective chemical labelling combined with a newly established mass spectrometric all-ion-reaction scanning approach to identify nonulosonic acids and other ulosonic acid-like sugars. In doing so, we provide a first molecular-level comparative study on the frequency and diversity across different phyla. We not only illustrate their surprisingly wide-spread occurrence in non-pathogenic species, but also provide evidence of potential higher carbon variants. Many biomedical studies rely on synthetic routes for sialic acids, which are highly demanding and often of low product yields. Our approach enables large-scale exploration for alternative sources of these highly important compounds.

Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA.

CRISPR-Cas9 systems are enriched in human pathogenic bacteria and have been linked to cytotoxicity by an unknown mechanism. Here, we show that upon infection of human cells, Campylobacter jejuni secretes its Cas9 (CjeCas9) nuclease into their cytoplasm. Next, a native nuclear localization signal enables CjeCas9 nuclear entry, where it catalyzes metal-dependent nonspecific DNA cleavage leading to cell death. Compared to CjeCas9, native Cas9 of Streptococcus pyogenes (SpyCas9) is more suitable for guide-dependent editing. However, in human cells, native SpyCas9 may still cause some DNA damage, most likely because of its ssDNA cleavage activity. This side effect can be completely prevented by saturation of SpyCas9 with an appropriate guide RNA, which is only partially effective for CjeCas9. We conclude that CjeCas9 plays an active role in attacking human cells rather than in viral defense. Moreover, these unique catalytic features may therefore make CjeCas9 less suitable for genome editing applications.