Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy.

The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic, and pre-symptomatic carriers of the virus. CRISPR diagnostics can augment gold-standard PCR-based testing if they can be made rapid, portable, and accurate. Here, we report the development of an amplification-free CRISPR-Cas13a assay for direct detection of SARS-CoV-2 from nasal swab RNA that can be read with a mobile phone microscope. The assay achieved ∼100 copies/µL sensitivity in under 30 min of measurement time and accurately detected pre-extracted RNA from a set of positive clinical samples in under 5 min. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity and directly quantified viral load using enzyme kinetics. Integrated with a reader device based on a mobile phone, this assay has the potential to enable rapid, low-cost, point-of-care screening for SARS-CoV-2.

Accelerated RNA detection using tandem CRISPR nucleases.

Direct, amplification-free detection of RNA has the potential to transform molecular diagnostics by enabling simple on-site analysis of human or environmental samples. CRISPR-Cas nucleases offer programmable RNA-guided RNA recognition that triggers cleavage and release of a fluorescent reporter molecule, but long reaction times hamper their detection sensitivity and speed. Here, we show that unrelated CRISPR nucleases can be deployed in tandem to provide both direct RNA sensing and rapid signal generation, thus enabling robust detection of ~30 molecules per µl of RNA in 20 min. Combining RNA-guided Cas13 and Csm6 with a chemically stabilized activator creates a one-step assay that can detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA extracted from respiratory swab samples with quantitative reverse transcriptase PCR (qRT-PCR)-derived cycle threshold (Ct) values up to 33, using a compact detector. This Fast Integrated Nuclease Detection In Tandem (FIND-IT) approach enables sensitive, direct RNA detection in a format that is amenable to point-of-care infection diagnosis as well as to a wide range of other diagnostic or research applications.

Preparing small-dose red cell concentrates (RCCs) for neonatal and pediatric transfusions: Impact of RCC volume, storage, and irradiation.

BACKGROUND: Preparing small-dose red cell concentrates (RCCs) is a common practice for pediatric and neonatal transfusions. However, there is a lack of quality monitoring data to indicate that both the preparation and storage of small-dose RCCs does not alter in vitro red cell quality. The present study seeks to provide data to support this practice. MATERIALS AND METHODS: To evaluate quality of stored small aliquots, six ABO/Rh matched leukoreduced citrate phosphate-dextrose/saline-adenine-glucose-mannitol (LR CPD/SAGM) RCCs were pooled and split into 30 ml aliquots, 80 ml aliquots, and a standard 290 ml unit, with testing performed for up to 43 days post-collection. To evaluate the impact of irradiation on small-dose RCC preparation, a total of 48 independent LR CPD/SAGM RCCs were used (non-irradiated: n = 24; irradiated: n = 24). Aliquoting with/without irradiation was performed within 7 days of collection and baseline testing was performed within 24 h of aliquot production. RESULTS: Limited variability in hemolysis, mean cell volume, and extracellular potassium concentrations were seen between the different aliquot sizes throughout the 43-day storage period. Aliquot production did not accentuate damage based on any of these tested parameters in both the non-irradiated and irradiated subsets. A significant increase was seen in the potassium concentrations in the irradiated parent and aliquot samples relative to their non-irradiated counterparts. CONCLUSIONS: Non-irradiated small-aliquot dose RCCs meet in vitro quality criteria required for safe transfusion throughout the 42-day storage period. The same can be said for aliquots derived from irradiated units and tested within 24 h of aliquot production.

The costs of treating vaginal and vulval cancer in England (2009-2015).

BACKGROUND: Human papillomavirus (HPV) infection is a pre-requisite for cervical cancer, which represents the third most common cancer among women worldwide. A causal relationship also exists between HPV and cancer in other areas of the female reproductive system including the vagina and vulva. Whilst the incidence of vaginal cancer in the UK has remained relatively stable over the past 25 years, vulval cancer rates are increasing. A body of literature exists on the epidemiology and aetiology of vaginal and vulval cancer, but little is known about the economic burden. The objective of this study was to quantify the costs of treating these cancers on the National Health Service (NHS) in England. METHODS: Inpatient and outpatient episodes were derived from Hospital Episode Statistics (HES). Health Resource Group (HRG) tariffs and National Reference Costs were used to estimate the cost of treating pre-cancerous and invasive vaginal and vulval lesions in England. RESULTS: The study showed that for the 5 years from 2009/2010 to 2014/2015 the total cost associated with pre-cancerous and invasive vaginal and vulval lesions was over £14 million per year on average (95% of which was attributed to inpatient costs). Vulval cancer accounted for the largest proportion; an estimated 60% of the total cost (£8.82 million). On average 4316 patients per year in England were admitted to hospital and 912 patients attended outpatient settings for pre-cancerous and invasive disease of the vagina and vulva. CONCLUSION: The results indicate that vaginal and vulval cancer cost the English health care system over £14 million per year. Given the causal role of HPV in a proportion of these cancers, preventative measures such as the national HPV immunisation programme have the potential to reduce the economic burden. To ensure optimal use of NHS resources, it is important that future economic evaluations of such preventative measures consider the full burden of HPV related disease.

Comparison of the Graston Technique® With Instrument-Assisted Soft Tissue Mobilization for Increasing Dorsiflexion Range of Motion.

CONTEXT: Limited dorsiflexion (DF) range of motion (ROM) is commonly observed in both the athletic and general populations and is a predisposing factor for lower extremity injury. Graston Technique® (GT) is a form of instrument-assisted soft tissue mobilization (IASTM), used commonly to increase ROM. Evidence of the long-term effects of GT on ROM is lacking, particularly comparing the full GT protocol versus IASTM alone. OBJECTIVE: To evaluate the effectiveness of 6 sessions of the GT or IASTM compared with a control (CON) group for increasing closed-chain DF ROM. DESIGN: Cohort design with randomization. SETTING: Athletic training clinic. PATIENTS OR OTHER PARTICIPANTS: A total of 23 physically active participants (37 limbs) with <34° of DF. Participants' limbs were randomly allocated to the GT, IASTM, or CON group. INTERVENTION: Participants' closed-chain DF ROM (standing and kneeling) were assessed at baseline and 24-48 hours following their sixth treatment. Participants in the CON group were measured at baseline and 3 weeks later. The intervention groups received 6 treatments during a 3-week period, whereas the CON group received no treatment. The GT group received a warm-up, instrument application, stretching, and strengthening of the triceps surae. The IASTM group received a warm-up and instrument application. MAIN OUTCOME MEASURES: Closed-chain DF was assessed with a digital inclinometer in standing and kneeling. RESULTS: A significant difference between groups was found in the standing position (P = .03) but not in kneeling (P = .15). Post hoc testing showed significant improvements in DF in standing following the GT compared with the control (P = .02). CONCLUSIONS: The GT significantly increases ankle DF following 6 treatments in participants with DF ROM deficits; however, no differences were found between GT and IASTM. The GT may be an effective intervention for clinicians to consider when treating patients with DF deficits.

Penile cancer treatment costs in England.

BACKGROUND: Penile cancer is a rare malignancy in Western countries, with an incidence rate of around 1 per 100,000. Due to its rarity, most treatment recommendations are based on small trials and case series reports. Furthermore, data on the resource implications are scarce. The objective of this study was to estimate the annual economic burden of treating penile cancer in England between 2006 and 2011 and the cost of treating a single case based on a modified version of the European Association of Urology penile cancer treatment guidelines. METHODS: A retrospective (non-comparative) case series was performed using data extracted from Hospital Episode Statistics. Patient admission data for invasive penile cancer or carcinoma in situ of the penis was extracted by ICD-10 code and matched to data from the 2010/11 National Tariff to calculate the mean number of patients and associated annual cost. A mathematical model was simultaneously developed to estimate mean treatment costs per patient based on interventions and their associated outcomes, advised under a modified version of the European Association of Urologists Treatment Guidelines. RESULTS: Approximately 640 patients per year received some form of inpatient care between 2006 and 2011, amounting to an average of 1,292 spells of care; with an average of 48 patients being treated in an outpatient setting. Mean annual costs per invasive penile cancer inpatient and outpatient were £3,737 and £1,051 respectively, with total mean annual costs amounting to £2,442,020 (excluding high cost drugs). The mean cost per case, including follow-up, was estimated to be £7,421 to £8,063. Results were sensitive to the setting in which care was delivered. CONCLUSIONS: The treatment of penile cancer consumes similar levels of resource to other urological cancers. This should be factored in to decisions concerning new treatment modalities as well as choices around resource allocation in specialist treatment centres and the value of preventative measures.

The cost of anal cancer in England: retrospective hospital data analysis and Markov model.

BACKGROUND: Anal cancer requires a multidisciplinary approach to treatment with often complex interventions. Little is known regarding the associated costs and resource use. METHODS: Patient records were extracted from a national hospital database to estimate the number of patients treated for anal cancer in England. Identified resource use was linked to published UK cost estimates to quantify the reimbursement of treatment through the Payment by Results system. A mathematical model was developed simultaneously to validate findings and to calculate the average 10-year cost of treating a squamous cell anal carcinoma case from diagnosis. The model utilised data from the Association of Coloproctology of Great Britain and Ireland's anal cancer position statement. RESULTS: On average, 1,564 patients were admitted to hospital and 389 attended an outpatient facility per year. The average annual cost per inpatient and outpatient ranged from £4,562-£5,230 and £1,146-£1,335, respectively. Based on the model estimates, the inflated cost per case was between £16,470-£16,652. Results were most sensitive to the mode of admission for primary treatment and the costs of staging/diagnosis (inflated range: £14,309-£23,264). CONCLUSIONS: Despite limitations in the available data, these results indicate that the cost of treating anal cancer is significant. Further observational work is required in order to verify these findings.

Transference of outdoor gait-training to treadmill running biomechanics and strength measures: A randomized controlled trial.

Outdoor gait-training has been successful in improving pain and reducing contact time during outdoor running for runners with exercise-related lower leg pain (ERLLP). However, it is unclear if these adaptations translate to gold standard treadmill running and clinical strength assessments. The study purpose was to assess the influence of a 4-week outdoor gait-training intervention with home exercises (FBHE) on treadmill running biomechanics and lower extremity strength compared to home exercises alone (HE) among runners with ERLLP. Seventeen runners with ERLLP were randomly allocated to FBHE and HE groups (FBHE: 3 M, 6F, 23 ± 4 years, 22.0 ± 4.6 kg/m2; HE: 3 M, 5F, 25 ± 5 years, 24.0 ± 4.0 kg/m2). Both groups completed eight sessions of home exercises over 4 weeks. The FBHE group received gait-training through wearable sensors to reduce contact time. Treadmill running gait and clinical strength assessments were conducted at baseline and 4-weeks. Multivariate repeated measures analyses of variance were used to assess the influence of group and timepoint for all outcomes. The FBHE group demonstrated significantly decreased contact time at 4-weeks compared to baseline and the HE group (Mean Difference [MD] range: -42 ms - -39 ms; p-range: <0.001-0.02). The FBHE group had significantly increased cadence (MD: +21 steps/min; p = 0.003) and decreased loading impulse (MD: -51, p < 0.001) during treadmill running at 4-weeks compared to the HE group. Strength did not significantly differ adjusting for multiple comparisons (p > 0.007). The outdoor FBHE intervention transferred to favorable changes in treadmill running biomechanics. Clinicians treating runners with ERLLP patients should implement data-driven outdoor gait-training to maximize patient benefits across running locations.

Local microvascular tissue oxygenation of the intrinsic foot muscles in patients with diabetes: A cross-sectional case-comparison study.

BACKGROUND: Foot-related complications including impaired peripheral circulation and lower limb ulceration are severe consequences for those with diabetes mellitus. This study aimed to assess differences in tissue oxygenation and oxygen utilization of the plantar surface intrinsic foot muscles between diabetic participants and healthy comparisons following short foot exercise and a standard walking protocol. METHODS: Eighteen participants, 9 with diabetes and 9 healthy age- and sex-matched comparisons, completed two interventions in a randomized order. For the short foot exercise intervention, participants completed 5 sets of 15 intrinsic foot muscle contractions. For the walking intervention, participants completed a modified six-minute walk test. Tissue oxygenation variables including oxygenated hemoglobin, deoxygenated hemoglobin, and tissue saturation index were measured using near-infrared spectroscopy in quiet stance and during intrinsic foot muscle contraction cycles following each intervention. Means, standard deviations, 95 % confidence intervals, mean differences, and Cohen's d effect sizes were calculated for each tissue oxygenation variable. RESULTS: The results of this study indicated no significant group differences in quiet standing tissue oxygenation measures at baseline and following each intervention. Participants in the diabetic group had significantly less change in tissue saturation index during intrinsic foot muscle contractions compared to healthy participants after the short foot exercise intervention (ES= 4.00, P = .0002) and walking intervention (ES= 1.33, P = .015). CONCLUSIONS: By utilizing wireless NIRS and novel research methodology, this study was able to explore changes in plantar surface tissue oxygenation of the intrinsic foot muscles following a targeted short foot exercise intervention as well as a standard walking protocol in patients diagnosed with diabetes compared to age- and sex- matched individuals without diabetes. We identified that diabetic participants presented with less oxygen utilization during intrinsic foot muscle contractions performed following both exercise interventions compared to their healthy age- and sex- matched comparisons.

Clinical usage, application procedures, and perceived effectiveness of cupping therapy among healthcare professionals in the United States: A cross-sectional survey.

BACKGROUND: Despite its growing popularity since the mid-1900s, the application procedures and factors influencing the usage of cupping therapy among healthcare professionals in the United States remains unclear. Therefore, the purpose of this study was to investigate the clinical usage, application procedures, and perceived effectiveness of cupping therapy among healthcare professionals in the United States. MATERIALS AND METHODS: A convenience sample of 158 healthcare professionals (age: 29.36 ± 7.42 years) participated in this cross-sectional study (completion rate = 86.15%; n = 158/195). A custom web-based survey designed to assess respondents' clinical incorporation of cupping therapy, was disseminated through snowball sampling on various social media platforms. Descriptive statistics including means, standard deviations, and frequencies were calculated per survey item. RESULTS: Ninety-one percent of respondents reported working as an athletic trainer in either a secondary school (30%) or collegiate setting (48%). Dry cupping was the most commonly reported type of cupping therapy (99%), and 75% of respondents reported using both static and moving cupping techniques. Significant differences in cup application time and the amount of air extraction were noted between static and moving cupping. Cupping therapy was reported as most effective for treating muscle tightness, myofascial trigger points, and musculoskeletal pain. CONCLUSION: While the usage and application of cupping therapy may vary based on the type and style of cupping as well as the patient's condition, respondents reported cupping therapy to be an effective method for treating various musculoskeletal conditions.

Sensor-based gait training to reduce contact time for runners with exercise-related lower leg pain: a randomised controlled trial.

Objectives: To assess the effects of a 4-week randomised controlled trial comparing an outdoor gait-training programme to reduce contact time in conjunction with home exercises (contact time gait-training feedback with home exercises (FBHE)) to home exercises (HEs) alone for runners with exercise-related lower leg pain on sensor-derived biomechanics and patient-reported outcomes. Design: Randomised controlled trial. Setting: Laboratory and field-based study. Participants: 20 runners with exercise-related lower leg pain were randomly allocated into FBHE (4 male (M), 6 female (F), 23±4 years, 22.0±4.3 kg/m2) or HE groups (3 M, 7 F, 25±5 years, 23.6±3.9 kg/m2). Interventions: Both groups completed eight sessions of HEs over 4 weeks. The FBHE group received vibrotactile feedback through wearable sensors to reduce contact time during outdoor running. Primary and secondary outcome measures: Patient-reported outcome measures (PROMs) and outdoor gait assessments were conducted for both groups at baseline and 4 weeks. PROMs were repeated at 6 weeks, and feedback retention was assessed at 6 weeks for the FBHE group. Repeated measures analyses of variance were used to assess the influence of group and timepoint on primary outcomes. Results: The FBHE group reported increased function and recovery on PROMs beyond the HE group at 6 weeks (p<0.001). There was a significant group by time interaction for Global Rating of Change (p=0.004) and contact time (p=0.002); the FBHE group reported greater subjective improvement and reduced contact time at 4 and 6 weeks compared with the HE group and compared with baseline. The FBHE group had increased cadence (mean difference: 7 steps/min, p=0.01) at 4 weeks during outdoor running compared with baseline. Conclusion: FBHE was more effective than HE alone for runners with exercise-related lower leg pain, manifested with improved PROMs, reduced contact time and increased cadence. Trial registration number: NCT04270565.