Huge Excess Mortality due to the delta strain of COVID-19 in Japan in August, 2021 BackgroundNo remarkable mortality attributable to COVID-19 confirmed by PCR test has been observed in Japan.

ObjectWe sought to quantify excess mortality using the National Institute of Infectious Diseases (NIID) model.

MethodWe applied the NIID model to deaths of all causes from 1987 up through October, 2021 for the whole of Japan and up through August, 2021 for Tokyo.

ResultsResults in Japan show huge number of excess mortality, up to 10 thousands in the two months, in August and September, 2021. On the other hand, in Tokyo, 1323 excess mortality was detected

Discussion and ConclusionWe detected substantial excess mortality in Japan in August and September, 2021 and in Tokyo in August, 2021. It might be important to continue to monitor excess mortality of COVID-19 carefully hereafter.
Emulator-based Bayesian optimization for efficient multi-objective calibration of an individual-based model of malaria Individual-based models have become important tools in the global battle against infectious diseases, yet model complexity can make calibration to biological and epidemiological data challenging. We propose a using a Bayesian optimization framework employing Gaussian process or machine learning emulator functions to calibrate a complex malaria transmission simulator. We demonstrate our approach by optimizing over a high-dimensional parameter space with respect to a portfolio of multiple fitting objectives built from datasets capturing the natural history of malaria transmission and disease progression. Our approach quickly outperforms previous calibrations, yielding an improved final goodness of fit. Per-objective parameter importance and sensitivity diagnostics provided by our approach offer epidemiological insights and enhance trust in predictions through greater interpretability.
Survey of Contaminated Percutaneous Injuries in Anesthesia Practitioners BackgroundAnesthesia practitioners are at inherent risk for percutaneous injuries by blood-contaminated needles and sharp objects. These exposures may result in transmission of HIV and hepatitis viruses. Data about this occupational hazard from contaminated needles and sharp devices is limited and decades old. We conducted a web-based survey to assess the occurrence, reporting, characteristics, and outcome of contaminated percutaneous injuries (CPI) in anesthesia residents, fellows, and attendings.

MethodsAfter institutional research board approval, an email was sent to 217 anesthesia practitioners requesting their participation in an online survey about contaminated percutaneous injuries. Responses were collected from February through March 2020. Results are reported as absolute numbers and proportions with 95% confidence interval (CI).

ResultsThe overall survey response rate was 51% (110/217). 59% (65/110) (95% CI, 50-68) of participants reported having one or more contaminated percutaneous injury during their years of anesthesia practice (42% (21/50) of residents, 50% (4/8) of fellows, 77% (40/52) of anesthesia attendings). Prevalence of injuries related to attendings years of anesthesia practice was 69% (95% CI, 44-94) for 5-10 years, 62.5% (95% CI, 29-96) for 10-15 years, and 79% (95% CI, 63- 95) for greater than 15 years of practice.

35% (95% CI, 26-44) of participants reported having one or more CPI within the last 5 years (40% of residents, 50% of fellows, 29% of attendings). Occurrence of CPI within the last 5 years based on attending anesthesiologist years of practice was 57% for less than 5 years, 37.5% for 10-15 years, and 20% for 15-20 years of practice. 75% (95% CI, 65-85) reported the incident at the time of injury. 59% (95% CI, 48-70) of injuries were due to hollow bore needles. 50% (95% CI, 39-61) of total injuries were high risk. 26% of injured anesthesia practitioners received post-exposure prophylaxis and there were zero seroconversions.

ConclusionMost anesthesiologists will sustain a contaminated percutaneous injury during their careers. Incidence of these injuries decreases with years of practice. Occurrence of these injuries is high among anesthesia residents, with the majority reporting their injuries. Half of the injuries are high risk with a quarter requiring postexposure prophylaxis. More education and interventions are needed to reduce percutaneous injuries and improve reporting. Estimating the impact of Angelman syndrome on parental productivity in Australia using productivity-adjusted life years Angelman syndrome (AS) is a severe neurodevelopmental condition associated with a significant socioeconomic impact. This is the first study to estimate the impact of AS on parental productivity in Australia using cost-of-illness modelling with simulated follow-up over a 10-year period using 2019 as the baseline year. Key data were obtained from a prospective cohort of AS families, peer-reviewed literature, and publicly-available sources. The base-case productivity burden borne by the estimated 330 living parents of the 428 prevalent-persons with AS totalled AUD$1.25 million, corresponding to a loss of 38.42% of PALYs per-parent. This shows that caring for a child with AS has a significant impact on the productivity of affected parents, with a large associated impact on the broader Australian economy. Cohort profile: The Australian Parkinson's Genetics Study (APGS) - pilot (N=1,532) PurposeParkinsons disease (PD) is a neurodegenerative disorder associated with progressive disability. While the precise aetiology is unknown, there is evidence of significant genetic and environmental influences on individual risk. The Australian Parkinsons Genetics Study (APGS) seeks to study genetic and patient-reported data from a large cohort of individuals with PD in Australia to understand the sociodemographic, genetic, and environmental basis of PD susceptibility, symptoms and progression.

ParticipantsIn the pilot phase reported here, 1,819 participants were recruited through assisted mailouts facilitated by Services Australia based on having three or more prescriptions for anti-PD medications in their Pharmaceutical Benefits Scheme (PBS) records. The average age at the time of the questionnaire was 64 {+/-} 6 years. We collected patient-reported PD information and socio-demographic variables via an online (93% of the cohort) or paper-based (7%) questionnaire. One thousand five hundred thirty-two participants (84.2%) met all inclusion criteria, and 1,499 provided a DNA sample via traditional post.

Findings to date65% of participants were male, and 92% identified as being of European descent. A previous traumatic brain injury was reported by 16% of participants and was correlated with a younger age of symptom onset. At the time of the questionnaire, constipation (36% of participants), depression (34%), anxiety (17%), melanoma (16%) and diabetes (10%) were the most reported comorbid conditions.

Future plansWe plan to recruit sex- and age-matched unaffected controls, genotype all participants, and collect non-motor symptoms and cognitive function data. Future work will explore the role of genetic and environmental factors in the aetiology of PD susceptibility, onset, symptoms, and progression, including as part of international PD research consortia.

Article summaryO_ST_ABSStrengths and limitations of this studyC_ST_ABSO_LIWe used a time- and cost-effective recruitment method that enabled us to reach out to a random sample of individuals who have been prescribed medications for Parkinsons disease across all over Australia to invite them to participate in this study.
C_LIO_LIThe identities of letter recipients remained private and confidential and were not shared with the researchers. However, those recipients who were interested and willing to participate were directed to a website where they could sign up and provide informed consent.
C_LIO_LIThe source database only captures individuals who have been prescribed medications to treat Parkinsons disease in Australia and who are eligible for Medicare. Those without an official diagnosis, not receiving treatment, or not eligible for government subsidies are not included.
C_LIO_LIWe collected a wide range of patient-reported variables relevant to disease onset, diagnosis, symptoms, medical comorbidities, lifestyle, and family history in a large cohort of participants. However, some variables might not be as accurate as when measured by a specialist clinician.
C_LIO_LIGiven the 9% response rate to our single-letter invitation, there is a substantial risk of self-selection bias. Thus, patient characteristics for this cohort might differ from those of the typical population of individuals with Parkinsons disease in Australia.
C_LI The Ophthalmic Surgical Backlog caused by the COVID-19 Pandemic: A population-based and microsimulation modelling study BackgroundJurisdictions worldwide ramped down ophthalmic surgeries to mitigate the effects of coronavirus disease 2019 (COVID-19), creating a global surgical backlog. We sought to predict the long-term impact of COVID-19 on ophthalmology surgical care delivery.

MethodsThis is a population-based and a microsimulation modelling study. Provincial administrative data from January 2019 to May 2021 was used to estimate the backlog size and wait-times following the COVID-19 pandemic. For the post-pandemic recovery phase, we estimated the resources required to clear the backlog of patients accumulated on the waitlist during the pandemic.

ResultsA total of 56,923 patients were on the waitlist in the province of Ontario awaiting non-emergency ophthalmic surgery as of March 15, 2020. The number of non-emergency surgeries performed in the province decreased by 45-98% from March to May 2020, and 48-80% from April to May 2021 compared to the same months in 2019. By 2 years and 3 years, the overall estimated number of patients awaiting surgery grew by 129% and 150%, respectively. The estimated mean wait-time for patients for all subspecialty surgeries increased to 282 (SD 91) in March 2023 compared to 94 days (SD 97) in 2019. The provincial monthly additional resources required to clear the backlog by March 2023 was estimated to be a 34% escalation from the pre-pandemic volumes (4,626 additional surgeries).

InterpretationThe magnitude of the ophthalmic surgical backlog from COVID-19 has important implications for the recovery phase. The estimates from this microsimulation modelling can be adapted to other jurisdictions to assist with recovery planning for vision saving surgeries. Brazilian model estimation for SARS-CoV-2 peak contagion (BMESPC) With newer data for SARS-CoV-2 and entering the second wave of contagion required the improvement of the forecasting model, structuring its model to forecast the peak of the first and second contagion wave in Brazil. The Brazilian model estimation for SARS-CoV-2 peak contagion (BMESPC) was structured, capable of estimating the peak of contagion for SARS-CoV-2 in the first and second waves, as the main objective of this work. Using the BMESPC model, it was possible to estimate, with a certain reliability degree, the peak of contagion for the first and second waves in Brazil, with one day difference from the real to the forecast. While at the state level, the calculated confidence interval proved to be more accurate. In this way, it is possible to use BMESPC to forecast the peak of contagion for several regions, provided that the necessary structure and calibration are respected. Pulsed electromagnetic fields may be effective for the management of primary osteoporosis: a systematic review and meta-analysis ObjectiveTo investigate the effectiveness of pulsed electromagnetic fields (PEMFs) for the management of primary osteoporosis in older adults.

DesignSystematic review and meta-analysis.

Data SourcesMEDLINE, EMBASE, Web of Science, CENTRAL and CCTR, Physiotherapy Evidence Database, CNKI, VIP, Wan Fang, ClinicalTrials.gov and Current controlled trials from the inception dates to April 30, 2021.

Eligibility criteria for study selectionRandomised controlled trials or quasi-randomised trials examining the effects of PEMFs compared to placebo or sham or other agents for the management of primary osteoporosis (including those with previous fractures).

Data extraction and synthesisTwo independent reviewers extracted data. Primary outcomes were bone mass and number of incident fractures. Secondary outcomes were functional assessments, quality of life, and adverse events. Risk of bias was assessed with the Cochrane Collaborations tool and certainty of evidence with the grading of recommendations assessment, development and evaluation (GRADE) framework. A random effects model was used to calculate mean differences and 95% confidence intervals.

ResultsEight trials including 396 participants met the inclusion criteria. Low certainty evidence showed that PEMFs was non-inferior to conventional pharmacological agents in preventing the decline of Bone Mineral Density (BMD) at the lumbar (MD 0.01; CI -0.04 to 0.06) and femur neck (MD 0.01; CI -0.02 to 0.04), and improving balance function measured by Berg Balance Scale (BBS) (MD 0.01; CI -0.09 to 0.11) and Timed Up and Go test (MD -0.04; CI -0.80 to 0.72), directly after intervention. The similar effects were observed in BMD and BBS at 12- and 24-weeks follow-up from baseline with moderate certainty evidence. Very low certainty evidence showed that PEMFs (versus exercise) had small but significant effect on BMD at the femur neck (MD 0.10; CI 0.01 to 0.20), and no effect on BMD at the lumbar (MD 0.15; CI -0.04 to 0.35).

ConclusionPEMFs had positive effects non-inferior to first-line treatment on BMD and balance function in older adults with primary osteoporosis, but with low to very low certainty evidence and short-term follow-ups. There is a need for high-quality randomised controlled trials evaluating PEMFs for the management of primary osteoporosis.

RegistrationPROSPERO CRD42018099518. Decision curve analysis to identify optimal candidates of liver resection for intermediate-stage hepatocellular carcinoma with hepatitis B cirrhosis：A cohort study BackgroundThe selection criterion for liver resection (LR) in intermediate-stage (IM) hepatocellular carcinoma (HCC) is still controversial. This study aims to compare LR and transarterial chemoembolization in the range of predicted death risk

MethodsThe multivariable Cox regression model (MVR) was estimated to predict mortality at 5yr. The cut-off values were determined by a two-piece-wise linear regression model, decision curve analysis with MVR model, and hazard ratio curve for treatment plotted against the predicted mortality.

Results825 IM-HCC with hepatitis B cirrhosis were included for analysis (TACE, n=622; LR, n=203). The 5-yr overall survival rate of LR patients was higher than the TACE group (52.8% vs. 20.8%; P<0.0001). The line of LR and TACE were crossing with predicted death risk at 100% (P for interaction =0.008). The benefit of LR versus TACE decreased progressively as predicted death risk>0.55 (95%CI: 0.45,0.62). When predicted death risk over 0.7, decision curve analysis suggested that LR and TACE did not increase net benefit. Patients were then divided into four subgroups by the cut-off values (<0.45, 0.45[&ge;] /<0.62, 0.62[&ge;] /<0.7, [&ge;] 0.7). The stratified analysis of treatment in different subgroups, hazard ratios were 0.39 (95%CI: 0.27, 0.56), 0.36 (95%CI: 0.23, 0.56), 0.51 (95%CI: 0.27, 0.98), and 0.46 (95%CI: 0.27, 0.80), respectively.

ConclusionsLR reached the maximal relative utility in the interval of 0.45 to 0.62, and both LR and TACE did not increase net benefit at the 5-yr death risk over 0.7. Preterm outcomes following COVID-19 lockdowns, Melbourne, Australia BackgroundCommunity lockdowns during the COVID-19 pandemic may influence preterm birth rates, but mechanisms are unclear.

MethodsWe compared neonatal outcomes of preterm infants born to mothers exposed to community lockdowns in 2020 (exposed group) to those born in 2019 (control group). Main outcome studied was composite of significant neonatal morbidity or death.

ResultsMedian gestational age was 35+4 weeks (295 infants, exposed group) vs. 35+0 weeks (347 infants, control group) (p = 0.108). The main outcome occurred in 36/295 (12.2%) infants in exposed group vs. 46/347 (13.3%) in control group (p = 0.69). Continuous positive airway pressure (CPAP) use, jaundice requiring phototherapy, hypoglycaemia requiring treatment, early neonatal white cell and neutrophil counts were significantly reduced in the exposed group.

ConclusionsCOVID-19 community lockdowns did not alter composite neonatal outcomes in preterm infants, but reduced rates of some common outcomes, and early white cell and neutrophil counts. Effectiveness of the Covid-19 vaccines in preventing infection in dental practitioners: results of a cross-sectional questionnaire based survey India started its vaccination program at the beginning of 2021, the main beneficiaries being health workers and frontline workers including police, paramilitary forces, sanitation workers, and disaster management volunteers in the first phase. By the time, the second wave of Covid-19 impacted India, approximately 14 million healthcare and frontline workers, including dentists had been vaccinated.

AimTo study the effectiveness of vaccination on a subset of high-risk healthcare workers i.e. dentists in preventing Covid-19 during the second wave of the pandemic.

Study designA questionnaire based pan-India online survey was carried out to record the Covid-related experiences of dentists prior to and after vaccination.

ResultThe sample size for this survey was 4493 respondents from across India. During the second wave, 9.18% (n=364) respondents became positive in spite of the vaccine, while 14.69%(n=78) became positive in the unvaccinated group. A chi-square test of independence was performed to examine the relation between vaccination and the Covid positivity rate in all age groups. The relation between these variables was highly significant, [X2 (1, N = 4493) = 15.9809, p=.000064].

ConclusionOur pan-India online survey inferred that vaccination has a definitive role to play in reducing the positivity rate amongst dentists during the second wave of the pandemic across all age groups. Prime hAd5 Spike plus Nucleocapsid Vaccination Induces Ten-Fold Increases in Mean T-Cell Responses in Phase 1 Subjects that are Sustained Against Spike Variants In response to the need for a safe, efficacious vaccine that elicits vigorous T cell as well as humoral protection against SARS-CoV-2 infection, we have developed a dual-antigen COVID-19 vaccine comprising both the viral spike (S) protein modified to increase cell-surface expression (S-Fusion) and nucleocapsid (N) protein with an Enhanced T-cell Stimulation Domain (N-ETSD) to enhance MHC class I and II presentation and T-cell responses. The antigens are delivered using a human adenovirus serotype 5 (hAd5) platform with E1, E2b, and E3 regions deleted that has been shown previously in cancer vaccine studies to be safe and effective in the presence of pre-existing hAd5 immunity. The findings reported here are focused on human T-cell responses due to the likelihood that such responses will sustain efficacy against emerging variants, a hypothesis supported by our in silico prediction of T-cell epitope HLA binding for both the first-wave SARS-CoV-2  A strain and the B.1.351 strain K417N, E484K, and N501Y spike and T201I N variants. We demonstrate the hAd5 S-Fusion + N-ETSD vaccine antigens expressed by previously SARS-CoV-2-infected patient dendritic cells elicit Th1 dominant activation of autologous patient T cells, indicating the vaccine antigens have the potential to elicit immune responses in previously infected patients. For participants in our open-label Phase 1b study of the vaccine (NCT04591717; https://clinicaltrials.gov/ct2/show/NCT04591717), the magnitude of Th-1 dominant S- and N-specific T-cell responses after a single prime subcutaneous injection were comparable to T-cell responses from previously infected patients. Furthermore, vaccinated participant T-cell responses to S were similar for A strain S and a series of spike variant peptides, including S variants in the B.1.1.7 and B.1.351 strains. The findings that this dual-antigen vaccine elicits SARS-CoV-2-relevant T-cell responses and that such cell-mediated protection is likely to be sustained against emerging variants supports the testing of this vaccine as a universal booster that would enhance and broaden existing immune protection conferred by currently approved S-based vaccines. Sensitivity of Estimated Tacrolimus Population Pharmacokinetic Profile to Inaccurate Assumptions about Dose Timing and Absorption: An Investigation in Real-World and Simulated Data A population pharmacokinetic (PK) study with 363 subjects was performed using real-world data extracted from electronic heath records (EHRs) to estimate the tacrolimus population PK profile. As population PK studies for oral medications performed using EHR data often assume a regular dosing schedule as prescribed without incorporating exact dosing time, we assessed the sensitivity of the PK parameter estimates to assumptions about dose timing using last-dose times extracted by our own natural language processing system, medExtractR. We also investigated the sensitivity of estimations to absorption rate constants that are often fixed at a published value in tacrolimus population PK analysis. There was no appreciable difference in parameters estimates with vs. without last-dose time incorporated in the data and our sensitivity analysis revealed little difference between parameters estimated assuming a range of absorption rate constants. We also conducted simulation studies to investigate how drug PK profiles and experimental designs such as concentration measurements affects sensitivity to incorrect assumptions about dose timing and absorption rates. Our findings suggest that drugs with a slower elimination rate (or a longer half-life) are less sensitive to dose timing errors and that experimental designs which only allow for trough blood concentrations are usually insensitive to deviation in absorption rate. Effects of repeated injection of 1% lidocaine vs. radial extra-corporeal shock wave therapy for treating myofascial trigger points: a randomized controlled trial Background and ObjectivesThis study tested the hypothesis that treatment of myofascial trigger points (MTrPs) in the upper trapezius muscle (UTM) with repeated injection of 1% lidocaine results in better alleviation of muscular stiffness and soreness as well as improved metabolism in the hypercontracted MTrP area than treatment with radial extracorporeal shock wave therapy (rESWT).

Materials and MethodsA single-blinded, prospective, randomized controlled trial was conducted on patients suffering from MTrPs in the UTM. Thirty patients were treated with repeated injection of 2 ml of 1% lidocaine (three injections; one injection per week). Another 30 patients were treated with rESWT (three treatment sessions; one treatment session per week; 2000 radial extracorporeal shock waves per treatment session; positive energy flux density = 0.10 mJ/mm2). The primary outcome measure was pain severity using the VAS score. The secondary outcome measures included muscle elasticity index, pressure pain threshold and neck disability index. Evaluation was performed at baseline (T1), 15-30 minutes after the first treatment in order to register immediate treatment effects (T2), before the second treatment (i.e., one week after baseline) (T3) and one week after the third treatment (i.e., four weeks after baseline) (T4).

ResultsThere were no statistically significant differences in the primary and secondary outcome measures between the patients in the lidocaine arm and the patients in the rESWT arm at T1 and T4. Within the arms, the mean differences of all outcomes were statistically significant (P < 0.001) when comparing the data obtained at T1 with the data obtained at T3 and the data obtained at T4.

ConclusionsThe results of this pilot study suggest that the use of rESWT in patients with MTrPs in the UTM is safe and leads to reduced pain and improved muscle elasticty, pressure pain threshold and neck disability index, without adverse effects. Larger trials are necessary to verify this. Clinicians should consider rESWT instead of injections of lidocaine in the treatment of MTrPs in the UTM. Preexisting diabetes and COVID-associated hyperglycaemia in patients with COVID-19 pneumonia AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSAimC_ST_ABSThe aim of the current study was to compare clinical characteristics, laboratory findings and major outcomes of patients hospitalized for COVID-19 pneumonia with COVID-associated hyperglycaemia or preexisting diabetes.

MethodsA cohort of 176 adult patients with a diagnosis of pre-existing diabetes (n=112) or COVID-associated hyperglycaemia (n=55) was studied. Clinical outcomes and laboratory findings were analysed according to the presence of the two conditions. The time to viral clearance was assessed during the follow-up after hospital discharge.

ResultPatients with COVID-associated hyperglycaemia had lower BMI, significantly less comorbidities and higher levels of inflammatory markers and indicators of multi-organ injury than those with preexisting diabetes. No differences between preexisting diabetes and COVID-associated hyperglycaemia were evident for symptoms at admission, humoral response against SARS-CoV-2 or autoantibodies to glutamic acid decarboxylase or interferon alpha-4. COVID-associated hyperglycaemia was independently associated with the risk of adverse clinical outcome defined as ICU admission or death (HR 2.11, 95% CI 1.34-3.31; p=0.001), even after adjustment for age, sex and other selected variables associated with COVID-19 severity. Furthermore, we documented a negative association (HR 0.661, 95% CI 0.43-1.02; p=0.063) between COVID-associated hyperglycaemia and the time to swab negativization.

ConclusionsThe recognition of hyperglycaemia as a specific clinical entity associated with COVID-19 pneumonia is relevant for early and appropriate patient management and close monitoring for the progression of disease severity. A trans-ancestry genome-wide association study of unexplained chronic ALT elevation as a proxy for nonalcoholic fatty liver disease with histological and radiological validation. Nonalcoholic fatty liver disease (NAFLD) is a growing cause of chronic liver disease. Using a proxy NAFLD definition of chronic alanine aminotransferase elevation (cALT) without other liver diseases, we performed a trans-ancestry genome-wide association study in the Million Veteran Program including 90,408 cALT cases and 128,187 controls. In the Discovery stage, seventy-seven loci exceeded genome-wide significance - including 25 without prior NAFLD or ALT associations - with one additional locus identified in European-American-only and two in African-American-only analyses (P<5x10-8). External replication in cohorts with NAFLD defined by histology (7,397 cases, 56,785 controls) or liver fat extracted from radiologic imaging (n=44,289) validated 17 SNPs (P<6.5x10-4) of which 9 were novel (TRIB1, PPARG, MTTP, SERPINA1, FTO, IL1RN, COBLL1, APOH, and IFI30). Pleiotropy analysis showed that 61 of 77 trans-ancestry and all 17 validated SNPs were jointly associated with metabolic and/or inflammatory traits, revealing a complex model of genetic architecture. Our approach integrating cALT, histology and imaging reveals new insights into genetic liability to NAFLD. Deceased donor kidney degradomics indicates cytoskeletal proteolytic alterations impacting post-transplant function. BackgroundIn brain death, cerebral injury contributes to systemic biological dysregulation, causing significant cellular stress in donor kidneys that adversely impacts the quality of grafts. Here, we hypothesized that DBD kidneys may undergo proteolytic processes that renders grafts susceptible to post-transplant dysfunction.

Material & MethodsUsing mass spectrometry and immunoblotting analyses, we profiled degradation patterns of cytoskeletal proteins in deceased (n=55) and living (n=10) donor kidneys.

ResultsWe found that in DBD kidneys, key podocyte cytoskeletal proteins had been proteolytically cleaved. Generated degradation profiles were not associated to donor related demographic and clinical factors but were associated to suboptimal post-transplant function. Strikingly, -actinin -4 and Talin-1 degradation profiles were not observed in circulatory-death or living-donor kidneys. As Talin-1 is a specific proteolytic target of Calpain-1, we investigated a potential trigger of Calpain activation and Talin-1 degradation using ex-vivo precision-cut human kidney slices and in-vitro immortalised human podocytes. Notably, we found that Transforming-Growth Factor-{beta} (TGF-{beta}) activated Calpain-1 and proteolytically cleaved Talin-1 to generate distinct peptide fragments. These peptide fragments were of similar size and matched the degradation patterns observed in DBD kidneys. Talin-1 degradation was prevented in vitro by Calpain-1 inhibition.

ConclusionsHere, we provide initial evidence that DBD kidneys are susceptible to cytoskeletal protein degradation that impacts posttransplant kidney function. Subsequent studies should aim to further investigate the link between brain death and activation of proteolytic pathways exploring new therapeutic opportunities. Substance Use Onset in High-Risk 9-13 Year-Olds in the ABCD Study AimA key aim of the Adolescent Brain Cognitive DevelopmentSM (ABCD) Study is to document substance use onset, patterns, and sequelae across adolescent development. However, substance use misreporting obscures accurate drug use characterization. Hair toxicology tests provide objective historical substance use data, but are rarely investigated in youth. Here, we compare objective hair toxicology results with self-reported substance use in youth.

MethodsA literature-based substance use risk algorithm identified 696 ABCD Study(R) participants for hair sample collections between baseline and 2-year follow-up (spanning ages 9-13) for laboratory analysis. Chi-square and t-tests assessed differences between participants demographics, positive and negative hair tests, risk algorithm scores, and self-reported substance use.

ResultsHair testing confirmed that 17% of at-risk 9-13 year-olds had evidence of past 3-month use of one (n=99), two (n=17), three (n=3), or four (n=2) drug classes. After considering prescribed medication use, 11% had a positive test incongruent with self- or parent-report. No participant with a positive result self-reported substance use consistent with their toxicology results. Participants with positive tests under-reported use (p<.001), reported less sipping of alcohol (p<.001), and scored higher on the risk algorithm (p<.001) than those with negative hair toxicology.

ConclusionsAn alarming 11% of tested samples in at-risk 9-13 year-olds were positive for at least one unreported substance, suggesting underreporting in this population when participating in a national healthy development study. The degree of underreporting cannot yet be calculated, as at-risk samples were prioritized for assays. Expanded toxicology testing is key to characterize substance use in youth. The kidney releases a non-polymerizing form of Uromodulin in the urine and circulation that retains the external hydrophobic patch domain Uromodulin (Tamm-Horsfall protein, THP) is a glycoprotein uniquely produced in the kidney. It is released by cells of the thick ascending limbs (TAL) apically in the urine, and basolaterally in the renal interstitium and systemic circulation. Processing of mature urinary THP, which polymerizes into supra-molecular filaments, requires cleavage of an external hydrophobic patch (EHP) at the C terminus. However, THP in the circulation is not polymerized, and it remains unclear if non-aggregated forms of THP exist natively in the urine. We propose that an alternative processing path, which retains the EHP domain, can lead to a non-polymerizing form of THP. We generated an antibody that specifically recognizes THP with retained EHP (THP+EHP) and established its presence in the urine in a non-polymerized native state. Proteomic characterization of urinary THP+EHP revealed its C-terminus to end at F617. In the human kidney, THP+EHP was not only detected in TAL cells, but also diffusely in the renal parenchyma. Using immunoprecipitation followed by proteomic sequencing and immunoblotting, we then demonstrated that serum THP has also retained EHP. In a small cohort of patients at risk for acute kidney injury (AKI), admission urinary THP+EHP was significantly lower in patients who subsequently developed AKI during hospitalization. Our findings uncover novel insights into uromodulin biology by establishing the presence of an alternative path for cellular processing, which could explain the release of non-polymerizing THP in the circulation. Larger studies are needed to establish the utility of urinary THP+EHP as a sensitive biomarker of kidney health and susceptibility to injury. Hyperreflective foci, OCT progression indicators in age-related macular degeneration, include transdifferentiated retinal pigment epithelium PurposeBy optical coherence tomography (OCT) imaging, hyperreflective foci (HRF) indicate progression risk for advanced age-related macular degeneration (AMD) and are in part attributable to ectopic retinal pigment epithelium (RPE). We hypothesized that ectopic RPE are molecularly distinct from in-layer cells and that their cross-retinal course follows Muller glia.

MethodIn clinical OCT (61 eyes, 44 AMD patients, 79.4 {+/-} 7.7 years; 29 female; follow-up = 4.7 {+/-} 0.9 years) one HRF type, RPE plume, was reviewed. Twenty eyes of 20 donors characterized by ex vivo OCT were analyzed by histology (normal, 4; early/intermediate AMD, 7; geographic atrophy (GA), 6; 3 neovascular AMD (nvAMD). Cryosections were stained with antibodies to retinoid (RPE65, CRALPB) and immune (CD68, CD163) markers. In published RPE cellular phenotypes, red immunoreactivity was assessed semi-quantitatively by one observer (none, some cells, all cells).

ResultsTrajectories of RPE plume and cellular debris paralleled Muller glia, whether ordered or subsident, near atrophy borders. RPE corresponding to HRF lost immunoreactivity for retinoid markers and gained immunoreactivity for immune markers. Aberrant immunoreactivity appeared in individual in-layer RPE cells and extended to all abnormal phenotypes. Muller glia remained CRALBP-positive. Plume cells approached and contacted retinal capillaries.

ConclusionGain- and loss-of-function starts with individual in-layer RPE cells and extends to all abnormal phenotypes. Down-regulated RPE retinoid handling may impair rod vision while Muller glia sustain cone vision. Evidence for RPE transdifferentiation, possibly due to ischemia, supports a proposed process of epithelial-mesenchyme transition. Data can propel new biomarkers and therapeutic strategies for AMD.

PrecisHyperreflective foci (HRF) are OCT progression risk indicators in age-related macular degeneration. Abnormal RPE cells including some that correspond to HRF lose immunoreactivity for retinoid markers and gain immunoreactivity for immune markers, indicating molecular transdifferentiation. Intrahost evolution of the HIV-2 capsid correlates with progression to AIDS BackgroundHIV-2 infection will progress to AIDS in most patients without treatment, albeit at approximately half the rate of HIV-1 infection. HIV-2 p26 amino acid variations are associated with lower viral loads and enhanced processing of T cell epitopes, which may lead to protective Gag-specific CTL responses common in slower disease progressors. Lower virus evolutionary rates, and positive selection on conserved residues in HIV-2 env have been associated with slower progression to AIDS. We therefore aimed to determine if intrahost evolution of HIV-2 p26 is associated with disease progression.

MethodsTwelve treatment-naive, HIV-2 mono-infected participants from the Guinea-Bissau Police cohort with longitudinal CD4+ T cell data and clinical follow-up were included in the analysis. CD4% change over time was analysed via linear regression models to stratify participants into relative faster and slower disease progressor groups. Gag amplicons of 735 nucleotides which spanned the p26 region were amplified by PCR and sequenced. We analysed p26 sequence diversity evolution, measured site-specific selection pressures and evolutionary rates, and determined if these evolutionary parameters were associated with progression status. Amino acid polymorphisms were mapped to existing p26 protein structures.

ResultsIn total, 369 heterochronous HIV-2 p26 sequences from 12 male patients with a median age of 30 (IQR: 28-37) years at enrolment were analysed. Faster progressors had lower CD4% and faster CD4% decline rates. Median pairwise sequence diversity was higher in faster progressors (5.7x10-3 versus 1.4x10-3 base substitutions per site, P<0.001). p26 evolved under negative selection in both groups (dN/dS=0.12). Virus evolutionary rates were higher in faster than slower progressors - synonymous rates: 4.6x10-3 vs. 2.3x10-3; and nonsynonymous rates: 6.9x10-4 vs. 2.7x10-4 substitutions/site/year, respectively. Virus evolutionary rates correlated negatively with CD4% change rates ({rho} = -0.8, P=0.02), but not CD4% level. However, Bayes factor (BF) testing indicated that the association between evolutionary rates and CD4% kinetics was supported by weak evidence (BF=0.5). The signature amino acid at p26 positions 6, 12 and 119 differed between faster (6A, 12I, 119A) and slower (6G, 12V, 119P) progressors. These amino acid positions clustered near to the TRIM5/p26 hexamer interface surface.

ConclusionsFaster p26 evolutionary rates were associated with faster progression to AIDS and were mostly driven by synonymous substitutions. Nonsynonymous evolutionary rates were an order of magnitude lower than synonymous rates, with limited amino acid sequence evolution over time within hosts. These results indicate the HIV-2 p26 may be an attractive vaccine or therapeutic target. "What do you mean I can't have a doctor? This is Canada!" - A Qualitative Study of the Myriad Consequences for Unattached Patients Awaiting Primary Care Attachment BackgroundPatient access to primary healthcare (PHC) is the foundation of a strong healthcare system and healthy populations. Attachment to a regular PHC provider, a key to healthcare access, has seen a decline in some jurisdictions. This study explored the consequences of unattachment from a patient perspective, an under-studied phenomenon to date.

MethodsA realist-informed qualitative study was conducted with unattached patients in Nova Scotia, Canada. Semi-structured interviews with nine participants were conducted and transcribed for analysis. The framework method was used to carry out analysis, which was guided by Donabedians model of assessing healthcare access and quality.

ResultsFive key findings were noted in this study: 1) Participants experienced a range of consequences from not having a regular PHC provider. Participants used creative strategies to 2) attempt to gain attachment to a regular PHC provider, and, to 3) address their health needs in the absence of a regular PHC provider. 4) Participants experienced negative feelings about themselves and the healthcare system, and 5) stress related to the consequences and added work of being unattached and lost care.

ConclusionsUnattached patients experienced a burden of care related to lost care and managing their own health and related information, due to the download of medical record management and system navigation to them. These findings may underestimate the consequences for further at-risk populations who would not have been included in our recruitment. This may result in poorer health outcomes, which could be mitigated by interventions at the structural level, such as enhanced centralized waitlists to promote attachment. Such waitlists may benefit from a triage approach to appropriately attach patients based on need. The Addiction Risk Factor: A Unitary Genetic Vulnerability Characterizes Substance Use Disorders and Their Associations with Common Correlates Substance use disorders commonly co-occur with one another and with other psychiatric disorders. They share common features including high impulsivity, negative affect, and lower executive function. We tested whether a common genetic factor undergirds liability to problematic alcohol use (PAU), problematic tobacco use (PTU), cannabis use disorder (CUD), and opioid use disorder (OUD) by applying genomic structural equation modelling to genome-wide association study summary statistics for individuals of European ancestry (Total N = 1,019,521; substance specific Ns range: 82,707-435,563), while adjusting for the genetics of substance use (Ns = 184,765-632,802). We also tested whether shared liability across SUDs is associated with behavioral constructs (risk taking, executive function, neuroticism; Ns = 328,339-427,037) and non-substance use psychopathology (psychotic, compulsive, and early neurodevelopmental disorders). Shared genetic liability to PAU, PTU, CUD, and OUD was characterized by a unidimensional addiction risk factor (termed The Addiction-Risk-Factor, independent of substance use. OUD and CUD demonstrated the largest loadings, while problematic tobacco use showed the lowest loading. The Addiction-Risk-Factor was associated with risk taking, neuroticism, executive function, and non-substance psychopathology, but retained specific variance before and after accounting for genetics of substance use. Thus, a common genetic factor partly explains susceptibility for alcohol, tobacco, cannabis, and opioid use disorder. The Addiction-Risk-Factor has a unique genetic architecture that is not shared with normative substance use or non-substance psychopathology, suggesting that addiction is not the linear combination of substance use and psychopathology. Molecular analysis suggests oligoclonality and metastasis of endometriosis lesions across anatomically defined subtypes. Endometriosis symptoms are heterogeneous with controversy on whether it constitutes a single disease or multiple distinct types. Our previous work found recurrent somatic cancer-driver alterations in endometriosis; however, these have not been found ubiquitously. A handful of cases spread across studies also suggest mutations might be shared (clonal) between lesions of the same type. As current classification systems correlate poorly with symptoms or outcomes, somatic genomics may improve the current system. Here, we investigate heterogeneity of somatic cancer-driver mutations within patients and across endometriosis types. We examined anatomically distinct types of endometriosis (ovarian, deep infiltrating, and superficial endometriosis) in 27 individual patients all of whom had at least two types of endometriosis. Specimens were analyzed using high-sensitivity targeted sequencing with orthogonal validation from droplet digital PCR and mutation-surrogate immunohistochemistry. Results found 13/27 patients had informative somatic driver mutation in endometriosis, 9/13 had identical mutations across distinct lesions. Endometriomas tended to have a higher mutational complexity, with functionally redundant driver mutations in same gene and within the same lesions.

Our data are consistent with clonality across endometriosis lesions regardless of subtype. Further the finding of redundancy in mutations with the same gene and lesions is also consistent with endometriosis representing an oligoclonal disease with dissemination likely to consist of multiple epithelial clones travelling together. This suggests the current anatomically defined classification of endometriosis does not fully recognize the etiology of the disease. A novel classification should take into account genomic and other molecular features. These findings could further contribute to development of a more personalized endometriosis diagnosis and care.