Elucidating the Role of CRISPR/Cas in Single-Step Isothermal Nucleic Acid Amplification Testing Assays.

Developing assays that combine CRISPR/Cas and isothermal nucleic acid amplification has become a burgeoning research area due to the novelty and simplicity of CRISPR/Cas and the potential for point-of-care uses. Most current research explores various two-step assays by appending different CRISPR/Cas effectors to the end of different isothermal nucleic acid amplification methods. However, efforts in integrating both components into more ideal single-step assays are scarce, and poor-performing single-step assays have been reported. Moreover, lack of investigations into CRISPR/Cas in single-step assays results in incomplete understanding. To fill this knowledge gap, we conducted a systematic investigation by developing and comparing assays that share the identical recombinase polymerase amplification (RPA) but differ in CRISPR/Cas12a. We found that the addition of CRISPR/Cas12a indeed unlocks signal amplification but, at the same time, impedes RPA and that CRISPR/Cas12a concentration is a key parameter for attenuating RPA impediment and ensuring assay performance. Accordingly, we found that our protospacer adjacent motif (PAM)-free CRISPR/Cas12a-assisted RPA assay, which only moderately impeded RPA at its optimal CRISPR/Cas12a concentration, outperformed its counterparts in assay design, signal, sensitivity, and speed. We also discovered that a new commercial Cas12a effector could also drive our PAM-free CRISPR/Cas12a-assisted RPA assay and reduce its cost, though simultaneously lowering its signal. Our study and the new insights can be broadly applied to steer and facilitate further advances in CRISPR/Cas-based assays.

Dental students' knowledge of and attitudes towards prescribing medication in Western Australia.

INTRODUCTION: Dentists can prescribe medications for prophylactic and therapeutic purposes. In Australia, dental graduates can autonomously practise within the scope of their qualifications without needing to undertake an internship post-graduation. Although previous research has identified knowledge gaps amongst dental students in Australia on pharmacology and pharmacotherapeutic knowledge, there has been no published research that qualitatively highlights dental students' knowledge of medication prescribing in Australia. This study aimed to undertake a qualitative analysis of Western Australian dental students' attitudes towards and knowledge of medication prescribing. METHODS: This qualitative case study design employed semi-structured interviews as means of collecting data. It utilised a purposive sampling in penultimate and final year dental students at the University of Western Australia. RESULTS: Twenty dental students participated in the study. Five key themes were identified in the data: current and previous education, application when delivering pharmacology and pharmacotherapeutics, teaching to ensure patient-centred care, supervision and referral to relevant resources. Participants agreed that having previous knowledge in the area provides students with confidence regarding medication prescribing. Furthermore, students in this study valued having guidance when prescribing medications. CONCLUSIONS: This study highlights the complexity of medication prescribing for dental students. It also highlights their experience with the current pharmacology and pharmacotherapeutics curriculum. All students interviewed valued application-based teaching, making it specific to dentistry. If curriculum were to be redeveloped, considering their perceptions may be a valuable tool.

Highly Sensitive Serum Protein Analysis Using Magnetic Bead-Based Proximity Extension Assay.

Many protein biomarkers are present in biofluids at a very low level but may play critical roles in important biological processes. The fact that these low-abundance proteins remain largely unexplored underscores the importance of developing new tools for highly sensitive protein detection. Although digital enzyme-linked immunosorbent assay (ELISA) has demonstrated ultrahigh sensitivity compared with conventional ELISA, the requirement of specialized instruments limits the accessibility and prevents the widespread implementation. On the other hand, proximity ligation assays (PLA) and proximity extension assays (PEA) show sensitive and specific protein detection using regular laboratory setups, but their sensitivity needs to be further improved to match digital ELISA. To achieve highly sensitive protein detection with minimal accessibility limitation, we develop a magnetic bead-based PEA (magPEA), which posts triple epitope recognition requirement and enables extensive washing for improved sensitivity and enhanced specificity. We demonstrate that the incorporation of magnetic beads into PEA workflow facilitates orders of magnitude sensitivity improvement compared with conventional ELISA, homogeneous PEA, and solid-phase PLA and achieves limits of detection close to that of digital ELISA when using IL-6, IL-8, and GM-CSF as validation. Our magPEA provides a simple approach for highly sensitive protein detection that can be readily implemented to other laboratories and will thus ultimately accelerate the study of the low abundance protein biomarkers in the future.

Australian dental students' knowledge on antibiotics prophylaxis for dental procedures.

BACKGROUND: Prescribing medicine is integral to clinical dentistry. Infective endocarditis may be rare but fatal if left untreated. As a result, judicious prescribing of antibiotics should be implemented due to potential. To our knowledge, no Australian study has examined dental students' knowledge and perceptions about antibiotic prophylaxis for dental procedures. METHODS: Australian dental students were invited to undertake the survey comprising case vignettes to investigate their medication knowledge. A total of 117 responses were received. The questions were 12 clinically relevant questions and three perception-based questions. Results were analysed using descriptive statistics as well as the chi-squared test. RESULTS: The 117 respondents had a mean correct response of 7.34 ± 2.64 (range 3-12 out of 12). Out of 117 students, 89 (76%) answered more than half of the questions correctly. Only three students (3%) answered all the questions correctly. Nearly two-thirds felt that they knew about antibiotic prophylaxis used for dental procedures. CONCLUSION: Most respondents answered more than half, but not all, of the clinical questions correctly. It is crucial to highlight that dental student may never receive any more training on antimicrobial stewardship (AMS) at any point in their future careers. It may be ideal that this issue is addressed at the dental school. One way to target this is to potentially nationalised teaching delivery of dental AMS across Australia.

Ligation-Enabled Fluorescence-Coding PCR for High-Dimensional Fluorescence-Based Nucleic Acid Detection.

Polymerase chain reaction (PCR) is by far the most commonly used method of nucleic acid amplification and has likewise been employed for a plethora of diagnostic purposes. Nonetheless, multiplexed PCR-based detection schemes have hitherto been largely limited by technical challenges associated with nonspecific interactions and other limitations inherent to traditional fluorescence-based assays. Here, we describe a novel strategy for multiplexed PCR-based analysis called Ligation-eNabled fluorescence-Coding PCR (LiNC PCR) that exponentially enhances the multiplexing capability of standard fluorescence-based PCR assays. The technique relies upon a simple, preliminary ligation reaction in which target DNA sequences are converted to PCR template molecules with distinct endpoint fluorescence signatures. Universal TaqMan probes are used to create target-specific multicolor fluorescence signals that can be readily decoded to identify amplified targets of interest. We demonstrate the LiNC PCR technique by implementing a two-color-based assay for detection of 10 ovarian cancer epigenetic biomarkers at analytical sensitivities as low as 60 template molecules with no detectable target cross-talk. Overall, LiNC PCR provides a simple and inexpensive method for achieving high-dimensional multiplexing that can be implemented in manifold molecular diagnostic applications.

Comparing the outcomes of gold-standard dental examinations with photographic screening by mid-level dental providers.

OBJECTIVES: To determine the detectability of the intra-oral photographic method in comparison to the baseline, comprehensive dental examination (CDE), in children when performed by different levels of dental practitioners. MATERIALS AND METHODS: As part of a quality assurance program, intra-oral photographs were obtained from 77 patients (2-18 years) as part of a CDE before undertaking dental treatment under general anaesthesia. A DSLR camera was utilised to acquire images which were subsequently uploaded to a cloud-based server. The baseline for each participant was established during the CDE, utilising both visual and radiographic examination, which was then compared to the assessment made by a mid-level dental practitioners (MLDP). The evaluation was based on utilising an odontogram where the teeth were charted either as decayed or filled. RESULTS: Specificity (95%) was higher than sensitivity (61.5%) when comparing the photographic assessment with the benchmark CDE assessment. The inter-rater reliability between the two methods of assessment was substantial, with a kappa score of 0.62. The photographic assessment method underestimated the decayed and filled teeth, as observed by the caries experience (dft/DFT scores) (CDE = 7.01 vs. photographic assessment = 5.22). There were lower levels of diagnostic detection in the posterior teeth as compared to the anterior teeth assessments. CONCLUSIONS: Although the CDE is still considered to be the gold-standard, this study found that the photographic caries assessment by MLDP produced an acceptable diagnostic level of detection particularly for the anterior teeth. CLINICAL RELEVANCE: The photographic method could offer a potential cost-saving and user-friendly screening.

Efficacy of Alternative Medicaments for Pulp Treatment in Primary Teeth in the Short Term: A Meta-analysis.

OBJECTIVE: This systematic review and meta-analysis aimed to evaluate the overall, clinical, and radiographical success rates of alternative pulpotomy medicaments in primary teeth. METHODS: A systematic search of five databases was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Prospective clinical trials using alternative pulpotomy medicaments in children were included. The outcome measures were overall, clinical, and radiographic success, expressed in percentages and converted to odds ratios. Fifteen articles were included in the meta-analysis. RESULTS: Combined odds ratios for overall, clinical, and radiographic success was 0.55 (95% confidence interval [CI]: 0.12-2.41; P = .42; I2 = 76%), 1.03 (95% CI: 0.57-1.86; P = .92; I2 = 0%), and 0.84 (95% CI: 0.54-1.47; P = .66; I2 = 34%), respectively. The results suggest an inconclusive outcome in the success rate of alternative medicaments. CONCLUSIONS: There is insufficient evidence to support the efficacy of alternative pulpotomy medicaments for use in primary teeth. Further robust studies are required before such alternative medicaments should be used in clinical practice.

Oral health-related quality of life changes in children following dental treatment under general anaesthesia: a meta-analysis.

OBJECTIVE: To conduct a meta-analysis of studies that have employed the Early Childhood Oral Health Impact Scale (ECOHIS) and Child Oral Health-Related Quality of Life (COHRQoL) instruments, to evaluate the oral health-related quality of life (OHRQoL) changes in children following dental treatment under general anaesthesia (DGA). METHOD: A systematic search of 5 databases was conducted in accordance with the PRISMA guidelines. The inclusion criteria were use of ECOHIS and COHRQoL, pre-and post-operative assessments, patients aged between 0 and 16 years, no restrictions on the follow-up period and DGA. The primary outcome measure was changes in quality of life for both the children, which was based on mean difference (MD). Twenty-two articles were included in the meta-analysis. RESULTS: A favourable outcome in OHRQoL was identified in all studies. The combined MD for ECOHIS and COHRQoL were 1.62 [95% CI 1.52-1.71; P < 0.00001; I2 = 0%] and 0.86 [95% CI 0.74-0.99; P < 0.00001; I2 = 0%], respectively, both with no evidence of heterogeneity. CONCLUSION: There is evidence to support that the OHRQoL of children was improved, with large effect size, in the short-term following DGA. CLINICAL RELEVANCE: Dental treatment under GA significantly improved the OHRQoL of children.

The family impact of dental general anaesthesia in children: A meta-analysis.

BACKGROUND: The health and well-being of children are intimately linked to their parents' physical, emotional, and social health and social circumstances. AIM: To conduct a meta-analysis of studies that have used the Early Childhood Oral Health Impact Scale (ECOHIS) and Child Oral Health-Related Quality of Life (COHRQoL) instruments, to evaluate the family impact scale (FIS) changes following their children's dental treatment under general anaesthesia (DGA). METHODOLOGY: A systematic search was undertaken using the PRISMA guidelines. The inclusion criteria consisted of patients below 16 years of age, DGA, pre- and post-operative assessments, and the use of ECOHIS and COHRQoL. FIS changes were the primary outcome measure, where the mean difference (MD) was calculated. RESULTS: After initial search of 105 studies in the database, twenty-one articles were included in the analysis. A positive outcome in the FIS changes was identified in all studies. The combined MD for FIS using ECOHIS and COHRQoL was 1.52 [95% CI: 1.15-1.89; P < 0.00001; I2 =87%] and 1.10 [95% CI: 0.68-1.52; P < 0.00001; I2 =79%], respectively. CONCLUSIONS: Dental treatment with general anaesthesia for children had a significant positive impact on parental emotions, activity, and conflict. Following DGA, there was significant improvement in the FIS, with large MD.

Risk factors of atopic dermatitis in Korean schoolchildren: 2010 international study of asthma and allergies in childhood.

BACKGROUND & OBJECTIVE: We aimed to analyse the risk factors of atopic dermatitis (AD) in Korean schoolchildren in 2010. METHODS: A nationwide, cross-sectional study was conducted in children aged 6-7 years and adolescents aged 12-13 years who were randomly selected. Information was obtained through a Korean version of the International Study of Asthma and Allergies in Childhood questionnaire (ISAAC), and skin prick tests were performed. AD-diagnosed children were selected for risk factor analysis by using logistic regression. RESULTS: We enrolled 4,003 children (M/F = 2,021/ 1,982) in aged 6-7 years and 4,112 children (M/F = 2,029/2,083) in 12-13 years. In children aged 6-7 years, the lifetime prevalence of AD diagnosis was 35.6% (N = 1,424). On the other hand, in the 12 to 13 year age group, the lifetime prevalence of AD diagnosis was 24.2% (N = 981). In the univariate logistic regression analysis in 6-7 year-old children, possible risk factors were atopy, a parental history of allergic disease, the use of antibiotics during infancy, a history of having moved into a newly built house during infancy, the presence of visible mould in the house, and remodelling of house within 12 months. The statistical significance persisted after adjustment. However, antibiotic use during infancy and remodelling within 12 months showed no statistical significance as a risk factor for AD. In contrast, multivariate logistic regression analysis in adolescents demonstrated that female sex, atopy, a parental history of allergic diseases, the presence of visible mould in the house, and a history of having moved into a newly built house during infancy was associated with AD. There was no significant association between AD and other risk factors. CONCLUSION: In Korean schoolchildren, risk factors such as atopy, the presence of parental allergic diseases, moving into a newly built house during infancy and visible mould in the house were associated with AD.

Gait changes after using a temporomandibular joint exerciser in patients who underwent lower limb joint surgery.

[Purpose] The improvements in gait of the patients with lower limb disease who used a temporomandibular joint (TMJ) exerciser were verified. [Subjects and Methods] Eleven subjects were included. Their mean age was 53.2 years. The lower limb joint angles before and after using the TMJ exerciser were measured using a gait analyzer. Before the gait experiment, the TMJ exerciser setting process and one-leg stance balance test (OLST) were repeated until the balance maintenance time improved. [Results] Because of the OLST, the mean change in the body center point after the subjects used the exerciser improved from 5.76 mm to 4.20 mm. When the TMJ exerciser was used, the joint angle range of the subjects approached that of the normal individuals. [Conclusion] According to the gait experiments, the angles of the subjects' hips, knees, and ankle joints approached to those of the normal individuals after the subjects used the TMJ exerciser; however, the results did not completely match. The changes in the hip, knee, and ankle joint angles were statistically significant, which confirm the usefulness of the TMJ exerciser.

Comparison of femoral tunnel geometry, using in vivo 3-dimensional computed tomography, during transportal and outside-in single-bundle anterior cruciate ligament reconstruction techniques.

PURPOSE: To compare the transportal (TP) and outside-in (OI) techniques regarding femoral tunnel position and geometry after anatomic single-bundle (SB) anterior cruciate ligament (ACL) reconstruction. METHODS: This study included 51 patients who underwent anatomic SB ACL reconstruction with the TP (n = 21) or OI (n = 30) technique. All patients underwent 3-dimensional computed tomography 3 days after the operation. The femoral tunnel position (quadrant method), femoral graft bending angle, femoral tunnel length, and posterior wall breakage were assessed by immediate postoperative 3-dimensional computed tomography with OsiriX imaging software. RESULTS: The OI technique had a shallower femoral tunnel position (arthroscopic position) than did the TP technique (P = .005). The mean femoral graft bending angle was significantly more acute with the OI technique (101.3° ± 8.2°) than with the TP technique (107.9° ± 10.0°) (P = .02). The mean femoral tunnel length was significantly greater with the OI technique (33.0 ± 3.5 mm) than with the TP technique (29.6 ± 3.9 mm) (P = .003). Posterior wall breakage occurred in 7 cases (33.3%) with the TP technique and 1 case (3.3%) with the OI technique (P = .02). CONCLUSIONS: The mean femoral tunnel position was significantly shallower (arthroscopic position) with the OI technique than with the TP technique. The OI technique resulted in a more acute femoral graft bending angle, longer femoral tunnel length, and lower incidence of posterior wall breakage than did the TP technique. These results might be helpful for anatomic SB ACL reconstruction using TP and OI techniques. LEVEL OF EVIDENCE: Level III, retrospective comparative study.

Mechanism of DNA adsorption and desorption on graphene oxide.

Graphene oxide (GO) adsorbing a fluorophore-labeled single-stranded (ss) DNA serves as a sensor system because subsequent desorption of the adsorbed probe DNA from GO in the presence of complementary target DNA enhances the fluorescence. In this study, we investigated the interaction of single- and double-stranded (ds) DNAs with GO by using a fluorescently labeled DNA probe. Although GO is known to preferentially interact with ssDNA, we found that dsDNA can also be adsorbed on GO, albeit with lower affinity. Furthermore, the status of ssDNA or dsDNA previously adsorbed on the GO surface was investigated by adding complementary or noncomplementary DNA (cDNA or non-cDNA) to the adsorption complex. We observed that hybridization occurred between the cDNA and the probe DNA on the GO surface. On the basis of the kinetics driven by the incoming additional DNA, we propose a mechanism for the desorption of the preadsorbed probe DNA from the GO surface: the desorption of the GO-adsorbed DNA was facilitated following its hybridization with cDNA on the GO surface; when the GO surface was almost saturated with the adsorbed DNA, nonspecific desorption dominated the process through a simple displacement of the GO-adsorbed DNA molecules by the incoming DNA molecules because of the law of mass action. Our results can be applied to design appropriate DNA probes and to choose proper GO concentrations for experimental setups to improve specific signaling in many biosensor systems based on the GO platform.

Refining the CRiSPHe (checklist for reporting research using a simulated patient methodology in Health): a Delphi study.

OBJECTIVES: A recent applicability study highlighted the need for the existing checklist for reporting research using a simulated patient methodology (CRiSP) to be clearer and user-friendly. The aim of this study was to update the checklist to address these concerns. METHODS: A fourth round of the Delphi consensus study, used in the original checklist development work, was conducted. Previous participants, who had expertise in SP methodology, were invited to complete a questionnaire including a list of 13 checklist items developed in the previous study and revised following applicability testing. Closed questions were analysed for frequency. Consensus was predefined as >80% agreement. All items were discussed in a roundtable meeting and further modified as necessary. Responses to open questions were content analysed. KEY FINDINGS: Twenty-one authors participated. There was a statistical consensus in 12 out of 13 modified checklist items. CONCLUSIONS: A final reporting checklist for studies in health research using SP methodology has been developed using a consensus approach. Further refinements may be needed to increase the generalizability of the checklist in different contexts.

Electronic health records and e-prescribing in Australia: An exploration of technological utilisation in Australian community pharmacies.

OBJECTIVE: This study aimed to assess the utilisation, benefits, and challenges associated with Electronic Health Records (EHR) and e-prescribing systems in Australian Community Pharmacies, focusing on their integration into daily practice and the impacts on operational efficiency, while also gathering qualitative insights from community pharmacists. METHODS: A mixed-methods online survey was carried out among community pharmacists throughout Australia to assess the utilisation of EHR and e-prescribing systems, including the benefits and challenges associated with their use. Data was analysed based on pharmacists' age, gender, and practice location (metropolitan vs. regional). The chi-square test was applied to examine the relationship between these demographic factors and the utilisation and operational challenges of EHR and e-prescribing systems. RESULTS: The survey engaged 120 Australian community pharmacists. Of the participants, 67 % reported usability and efficiency issues with EHR systems. Regarding e-prescribing, 58 % of pharmacists faced delays due to slow software performance, while 42 % encountered errors in data transmission. Despite these challenges, the benefits of e-prescribing were evident, with 79 % of respondents noting the elimination of illegible prescriptions and 40 % observing a reduction in their workload. Issues with prescription quantity discrepancies and the reprinting process were highlighted, indicating areas for improvement in workflow and system usability. The analysis revealed no significant statistical relationship between the utilisation and challenges of EHR and e-prescribing systems with the demographic variables of age, gender and location (p > 0.05), emphasising the necessity for healthcare solutions that address the needs of all pharmacists regardless of specific demographic segments. CONCLUSION: In Australian community pharmacies, EHR and e-prescribing may enhance patient care but come with challenges such as data completeness, technical issues, and usability concerns. Implementing successful integration relies on user-centric design, standardised practices, and robust infrastructure. While demanding for pharmacists, the digital transition improves efficiency and quality of care. Ensuring user-friendly tools is crucial for the smooth utilisation of digital health.

Prevalence and Factors Influencing Post-Operative Complications following Tooth Extraction: A Narrative Review.

Background: Complications from dental extractions may result in multiple post-operative visits and adversely affect the patient's life. Preventing complications may decrease post-operative morbidity for the individual as well as lower societal costs, such as lost time from work and healthcare costs. Objectives: This narrative review aims to assess the prevalence and factors influencing post-operative complications following tooth extraction, helping clinicians minimise the risk. Data Sources. Cross-sectional studies. Study Eligibility and Participants. Patients undergoing dental extractions. Our exclusion criteria included in vitro studies, animal studies, terminally ill patients, and tooth loss not due to dental extraction. Literature was collected from "PubMed" and "Web of Science" through search criteria based on the "PICO" framework. Twenty articles were used to formulate a prevalence table, and 156 articles were included for the factors influencing complications. Study Appraisal and Synthesis Methods. This narrative review was reported using the SANRA (a scale for the quality assessment of narrative review articles) checklist. Due to the scope of our narrative review and its associated objectives, the quality of cross-sectional studies (AXIS) will be conducted from the studies outlining the prevalence. Results: Alveolar osteitis appears to be the most prevalent post-operative complication following tooth extraction. Predisposing factors can be significant in their ability to alter the risk of postoperative complications, and clinicians should provide patient-centred care to mitigate this risk. Limitations. Due to the breadth of context, a systematic review was not feasible, as it may have introduced heterogeneity. Conclusion: This narrative review has highlighted an array of factors which can influence the prevalence of post-operative complications. Future research would benefit from individually reporting post-operative complications, reducing the heterogeneity in definitions of the complications, and including greater detail on the predisposing factors studied.

The influence of patients' beliefs about medicines and the relationship with suboptimal medicine use in community-dwelling older adults: a systematic review of quantitative studies.

BACKGROUND: Medication use in older adults is increasing, therefore, reducing the risk of suboptimal medicine use is imperative in achieving optimal therapeutic outcomes. Research suggests that factors such as personal beliefs and beliefs about medicines may be associated with non-adherence and inappropriate medicine use. AIM: To systematically review and identify quantitative research on the influence of beliefs about medicines and the relationship with suboptimal medicine use in older adults. METHOD: Searches were conducted on PubMed, EMBASE, CINAHL, and PsycINFO for quantitative studies (inception to March 2023). INCLUSION CRITERIA: (1) exposure: participants' beliefs (personal, cultural, and medication-related), (2) outcomes: polypharmacy, potentially inappropriate medicines use, or non-adherence, and (3) participants: community-dwelling adults 65 years or above. Study selection, data extraction and quality appraisal (Joanna Briggs Institute critical appraisal checklist) were completed independently by two investigators. Data were combined in a narrative synthesis and presented in a summary of findings table. RESULTS: Nineteen articles were included: 15 cross-sectional and four cohort studies. Outcomes of included papers were as follows; adherence (n = 18) and potentially inappropriate medicine use (n = 1). Ten studies found stronger beliefs in the necessity of medicines and/or fewer concerns led to better adherence, with one paper contradicting these findings. Three studies did not find associations between adherence and beliefs. One study confirmed an association between unnecessary drug use and a lack of belief in a "powerful other" (e.g. doctor). CONCLUSION: Further investigation is necessary to (1) ascertain the importance of necessity or concern beliefs in fostering adherence and, (2) examine the influence of beliefs on polypharmacy and inappropriate medicine use.

Digitized Kinetic Analysis Enhances Genotyping Capacity of CRISPR-Based Biosensing.

CRISPR/Cas systems have been widely employed for nucleic acid biosensing and have been further advanced for mutation detection by virtue of the sequence specificity of crRNA. However, existing CRISPR-based genotyping methods are limited by the mismatch tolerance of Cas effectors, necessitating a comprehensive screening of crRNAs to effectively distinguish between wild-type and point-mutated sequences. To circumvent the limitation of conventional CRISPR-based genotyping, here, we introduce Single-Molecule kinetic Analysis via a Real-Time digital CRISPR/Cas12a-assisted assay (SMART-dCRISPR). SMART-dCRISPR leverages the differential kinetics of the signal increase in CRISPR/Cas systems, which is modulated by the complementarity between crRNA and the target sequence. It employs single-molecule digital measurements to discern mutations based on kinetic profiles that could otherwise be obscured by variations in the target concentrations. We applied SMART-dCRISPR to genotype notable mutations in SARS-CoV-2, point mutation (K417N) and deletion (69/70DEL), successfully distinguishing wild-type, Omicron BA.1, and Omicron BA.2 SARS-CoV-2 strains from clinical nasopharyngeal/nasal swab samples. Additionally, we introduced a portable digital real-time sensing device to streamline SMART-dCRISPR and enhance its practicality for point-of-care settings. The combination of a rapid and sensitive isothermal CRISPR-based assay with single-molecule kinetic analysis in a portable format significantly enhances the versatility of CRISPR-based nucleic acid biosensing and genotyping.

Exploring the usability of simulated patient methodology in dental clinics in Western Australia: A pilot survey.

OBJECTIVES: This study aimed to explore the dental staff knowledge of simulated patient methodology and support for its use to investigate dental staffs' triaging ability. MATERIAL AND METHODS: Staff at dental practices in Western Australia were invited to participate in a cross-sectional online questionnaire, consisting of demographic questions, questions on triaging, and knowledge of simulated patient methodology. Descriptive and parametric tests were undertaken for quantitative data; qualitative responses were thematically analyzed. RESULTS: Of the 100 participants, most were female (71%), aged 25-39 years (57%), dentists (46%), and worked in private practices (60%). While 82% of participants triaged dental appointment enquiries, only 26% had heard of simulated patient studies. The majority (66%) of participants spent 1-5 min when triaging appointments and less than half (29%) asked about medical history, aggravating or alleviating factors. Although there was a general positive attitude toward use of simulated patient methodology to investigate practice, some concerns were identified. CONCLUSIONS: The findings of our exploratory study suggests that there may be a potential for utilizing simulated patient studies to improve the care of patients by dental receptionists in general dental practices.

Desorption of single-stranded nucleic acids from graphene oxide by disruption of hydrogen bonding.

Graphene oxide (GO) is known to interact with single-stranded nucleic acids through pi-stacking interactions and hydrogen bonds between the nucleobases and the hexagonal cells of GO. It also quenches the fluorescence when the fluorophore comes near to the GO mesh. When single-stranded (ss) regions of either DNA or RNA are present, those regions were adsorbed onto the surface of GO with a quenching of fluorescence located proximally to the GO surface. We demonstrated that bound single-stranded nucleic acids can be readily dissociated from GO by disrupting hydrogen bonding with urea, which was confirmed with fluorescence measurement and gel electrophoresis. Hydrogen bonding mainly contributes to the interaction between GO and single-stranded nucleic acids such as ssDNA and RNA. The GO-coated mesoporous silica nanoparticles (GO-MSNs) were synthesized for better separation of RNAs from cells. Cellular RNAs were readily adsorbed and eluted with ease by using GO-MSN and urea, respectively, demonstrating that GO-MSN and urea elution is a facile RNA extraction method.