Search

Background and ObjectivesThere is increasing interest in characterizing the earliest phases of Parkinson disease (PD). However, few studies have investigated prediagnostic trajectories of cognition and function. Our objective was to describe prediagnostic cognitive and functional trajectories in PD in older women and men.MethodsWe studied 9,595 women and 5,795 men from 2 prospective cohort studies of community-dwelling elders followed up to 20 years. In individuals without prevalent PD, we estimated the associations of incident PD diagnosis with rates of change in cognition and function before and after diagnosis compared with healthy older adults using multivariate mixed-effects models.ResultsOver follow-up, 297 individuals developed incident PD. Interactions between the terms in our model and sex were statistically significant for the 3 outcomes (p< 0.001 for all), so we stratified results by sex. Compared with older men without PD, men who developed PD exhibited faster decline in global cognition (0.04 SD more annual change,p< 0.001), executive function (0.05 SD more annual change,p< 0.001), and functional status (0.06 SD more annual change,p< 0.001) in the prediagnostic period. Women who developed PD compared with women without PD displayed faster decline in executive function (0.02 SD more annual change,p= 0.006) and functional status in the prediagnostic period (0.07 SD more annual change,p< 0.001).DiscussionIndividuals with incident PD exhibit cognitive and functional decline during the prediagnostic phase that exceeds rates associated with normal aging. Better understanding heterogeneity in prodromal PD is essential to enable earlier diagnosis and identify impactful nonmotor symptoms in all subgroups.

Background/Objective: To date, a validated Chinese (Mandarin) six-minute walk test (6MWT) translated instruction is not available. Translation of the Chinese 6MWT instruction is done in an ad hoc manner within the Chinese-speaking populations. This study aimed to develop a set of valid and reliable Chinese (Mandarin) instructions of the 6MWT. Methods: Translation was performed from the original English instruction via the recommended “Process of translation and adaptation of instruments” by the World Health Organization to generate the Chinese instructions. The Chinese instructions were tested with 52 healthy adult participants for its validity. Each participant underwent three 6MWTs and a cardiopulmonary exercise test. Randomization allowed participants to undergo the walk test in both the original English and the new Chinese instructions. Face and content validity, intra-rater and inter-rater reliability of the Chinese instructions of the 6MWT were established through the translation process. Criterion validity was established by analyzing the results of the 6MWT and cardiopulmonary exercise test. Results: Intraclass correlation coefficient for inter-rater reliability was excellent ([Formula: see text], 95% confidence [Formula: see text]–1.000). Similarly, the intra-rater reliability across the three raters was high (R1: [Formula: see text], 95% confidence interval [Formula: see text]–1.000; R2: [Formula: see text], 95% [Formula: see text]–1.000; R3: [Formula: see text], 95% [Formula: see text]–1.000). The 6-min walk distances collected from the Chinese and English instructed trials correlated positively with the maximal oxygen consumption ([Formula: see text], [Formula: see text]; [Formula: see text], [Formula: see text]). Conclusion: This is the first study to develop and validate the Chinese (Mandarin) instructions of the 6MWT, and the translation is as reliable and valid as the original English instructions.

Background Juvenile idiopathic arthritis (JIA) is common and difficult to treat. Cannabidiol (CBD) is now widely available, but no studies to date have investigated the use of CBD for JIA. Methods We performed a chart review to identify patients with JIA at a Midwestern medical institution between 2017 and 2019. We surveyed primary caregivers of JIA patients using an anonymous, online survey with questions on caregiver knowledge and attitudes towards CBD. We compared respondents with no interest in CBD use vs. those contemplating or currently using CBD using descriptive statistics. Results Of 900 reviewed charts, 422 met inclusion criteria. Of these, 236 consented to be sent a survey link, and n=136 (58%) completed surveys. Overall, 34.5% (n=47) of respondents reported no interest in using a CBD product for their child’s JIA, while 54% (n=79) reported contemplating using CBD and 7% (n=10) reported currently giving their child CBD. Only 2% of respondents contemplating or actively using a CBD product learned about CBD from their child’s rheumatologist, compared with television (70%) or a friend (50%). Most respondents had not talked to their child’s rheumatologist about using CBD. Of those currently using CBD, most used oral or topical products, and only 10% of respondents (n=1) knew what dose they were giving their child. Conclusions Our results show infrequent use but a large interest in CBD among caregivers of children with JIA. Given CBD’s unknown safety profile in children with JIA, this study highlights a need for better studies and education around CBD for pediatric rheumatologists.

Objectives There are currently scarce data exploring ureteric reimplantation (UR) during cytoreductive surgery (CRS). Methods We identified patients undergoing CRS for peritoneal surface malignancies (PSM) of any origin at a single high-volume unit. UR was defined as ureteroureterostomy, transureterouretostomy, ureteroneocystostomy, ureterosigmoidostomy or ileal conduit performed during CRS. Peri-operative outcomes, long-term survival and risk factors for requiring UR were analysed. Results Seven hundred and sixty-seven CRSs were identified. Twenty-three (3.0%) procedures involved UR. Bladder resection and colorectal cancer (CRC) were associated with increased risk of UR (bladder resection: OR 12.90, 95% CI 4.91–33.90, p Conclusions UR during CRS does not increase major morbidity or mortality for carefully selected patients, and is associated with low rates of urologic complications. Whilst decreased survival was apparent in patients with HAMN undergoing UR, it is unclear whether this relationship is causal.

Background Antigen testing offers rapid and inexpensive testing for SARS-CoV-2 but concerns regarding performance, especially sensitivity, remain. Limited data exists for use of antigen testing in asymptomatic patients; thus, performance and reliability of antigen testing remains unclear. Methods 148 symptomatic and 144 asymptomatic adults were included. A nasal swab was collected for testing by Quidel Sofia SARS IFA (Sofia) as point of care. A nasopharyngeal swab was also collected and transported to the laboratory for testing by Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV RT-PCR (Cepheid). Results Overall, Sofia had good agreement with Cepheid (> 95%) in adults, however was less sensitive. Sofia had a sensitivity of 87.8% and 33.3% for symptomatic and asymptomatic patients, respectively. Among symptomatic patients, testing > 5 days post symptom onset resulted in lower sensitivity (82%) when compared with testing within 5 days of symptom onset (90%). Of the four Sofia false-negative results in the asymptomatic cohort, 50% went on to develop COVID-19 disease within 5 days of testing. Specificity in both symptomatic and asymptomatic cohorts was 100%. Conclusions Sofia has acceptable performance in symptomatic adults when tested 5 days of symptom onset and asymptomatic patients.

Background Friedreich's ataxia is an inherited, progressive, neurodegenerative disease that typically begins in childhood. Disease severity is commonly assessed with rating scales, such as the modified Friedreich's Ataxia Rating Scale, which are usually administered in the clinic by a neurology specialist. Objective This study evaluated the utility of home-based, self-administered digital endpoints in children with Friedreich's ataxia and unaffected controls and their relationship to standard clinical rating scales. Methods In a cross-sectional study with 25 participants (13 with Friedreich's ataxia and 12 unaffected controls, aged 6-15 years), home-based digital endpoints that reflect activities of daily living were recorded over 1 week. Domains analyzed were hand motor function with a digitized drawing, automated analysis of speech with a recorded oral diadochokinesis test, and gait and balance with wearable sensors. Results Hand-drawing and speech tests were easy to conduct and generated high-quality data. The sensor-based gait and balance tests suffered from technical limitations in this study setup. Several parameters discriminated between groups or correlated strongly with modified Friedreich's Ataxia Rating Scale total score and activities of daily living total score in the Friedreich's ataxia group. Hand-drawing parameters also strongly correlated with standard 9-hole peg test scores. Interpretation Deploying digital endpoints in home settings is feasible in this population, results in meaningful and robust data collection, and may allow for frequent sampling over longer periods of time to track disease progression. Care must be taken when training participants, and investigators should consider the complexity of the tasks and equipment used.

The literature provides various tools for identifying successful interventions for interfering behavior of individuals with intellectual or developmental disabilities, yet less literature is available pertaining to students who do not have cognitive impairments. The purpose of the current study was to investigate the correspondence between the results of contingency space analyses (CSA) and interventions for four middle-school boys who engaged in interfering behavior. Researchers directly observed natural interactions between each student, his teacher, and peers during three baseline sessions. The students' teacher then implemented three intervention conditions. Using direct observation data, researchers conducted a CSA of the baseline data and determined the natural contingencies between student behavior and social consequences. The results of the CSAs were then compared to the effects of the intervention conditions. For three students, results of the CSA were consistent with results of the intervention analysis. Results provide preliminary support for the CSA as a tool for identifying hypothesis-based interventions.

Background Hydroxychloroquine is a treatment for rheumatic disease and considered safe during pregnancy. Interest in hydroxychloroquine has increased as it is being examined as a potential treatment and prophylaxis for coronavirus disease 2019. Data on the risks of specific birth defects associated with hydroxychloroquine use are sparse. Methods Using data from two case–control studies (National Birth Defects Prevention Study and Slone Epidemiology Center Birth Defects Study), we described women who reported hydroxychloroquine use in pregnancy and the presence of specific major birth defects in their offspring. Cases had at least one major birth defect and controls were live‐born healthy infants. Women self‐reported medication use information in the few months before pregnancy through delivery. Results In total, 0.06% (19/31,468) of case and 0.04% (5/11,614) of control mothers in National Birth Defects Prevention Study, and 0.04% (11/29,838) of case and 0.05% (7/12,868) of control mothers in Birth Defects Study reported hydroxychloroquine use. Hydroxychloroquine users had complicated medical histories and frequent medication use for a variety of conditions. The observed birth defects among women taking hydroxychloroquine were varied and included nine oral cleft cases; the elevated observed:expected ratios for specific oral cleft phenotypes and for oral clefts overall had 95% confidence intervals that included 1.0. Conclusion While teratogens typically produce a specific pattern of birth defects, the observed birth defects among the hydroxychloroquine‐exposed women did not present a clear pattern, suggesting no meaningful evidence for the risk of specific birth defects. The number of exposed cases is small; results should be interpreted cautiously.

Robotic technology has reduced the errors of implant alignment in unicompartmental knee arthroplasty (UKA), but its impact on functional recovery after UKA is poorly defined. The purpose of this study was to compare early functional recovery, pain levels, and satisfaction in UKA performed with either robotic assistance or conventional methods. A retrospective analysis was performed on 89 matched consecutive patients who underwent outpatient UKA by a single physician using either conventional instruments (n = 39) or robotic methods (n = 50), with otherwise identical perioperative protocols. Outcomes studied included Lower Extremity Functional Score (LEFS), new Knee Society Score (KSS), Knee Injury and Osteoarthritis Outcome Score for Joint Replacement (KOOS-JR.), VR/SF-12, Visual Analog Scale (VAS) pain scores, and perioperative opioid consumption. Patients in the robotic cohort had superior early functional outcomes, with greater LEFS (conventional = 23; robotic = 31) at 1 week post-op p = 0.015 and KOOS-JR (conventional = 74; robotic = 81) at up to 6 months post-op p = 0.037 ; these two values remained statistically significant after mixed-model regression analysis p = 0.010 ; p = 0.023 , respectively. At 1 year post-op, expectations were more likely to be met in those who received robotic assistance p = 0.06 . No differences were reported with respect to postoperative opioid usage p = 0.320 , reoperations p = 1.00 , and complications p = 0.628 . Robotic-assisted UKA resulted in more rapid recovery and less early postoperative pain and were more likely to meet expectations than conventional UKA, although functional differences equilibrated by 1 year postoperatively. Further follow-up is necessary to determine if implant durability is impacted by robotics.

Eliglustat, an oral substrate reduction therapy, is approved for eligible adults with Gaucher disease type 1. In the Phase 3 ENGAGE trial of previously untreated adults with Gaucher disease type 1, eliglustat‐treated patients had statistically significant improvements in organ volumes and hematologic parameters compared with placebo in the 9‐month primary analysis. We report final outcomes by time on eliglustat among all patients who participated in the ENGAGE trial and extension. No patient deteriorated clinically or withdrew due to adverse events; 39/40 patients entered the open‐label extension period and 34/40 (85%) remained in the trial until completion or switching to commercial eliglustat after its approval (2.3–6 years). Clinically meaningful improvements in Gaucher disease manifestations were seen in all patients concomitant with reductions in pathological lipid substrate levels (glucosylceramide and glucosylsphingosine). Among patients with 4.5 years of eliglustat exposure, mean spleen volume decreased by 66% (from 17.1 to 5.8 multiples of normal [MN], n = 13), mean liver volume decreased by 23% (from 1.5 to 1.1 MN, n = 13), mean hemoglobin increased 1.4 g/dl (from 11.9 to 13.4 g/dl, n = 12), mean platelet count increased by 87% (from 67.6 to 122.6 × 109/L, n = 12), median chitotriosidase decreased by 82% (from 13 394 to 2312 nmol/h/ml, n = 11), median glucosylceramide decreased by 79% (from 11.5 to 2.4 μg/ml, n = 11), median glucosylsphingosine decreased by 84% (from 518.5 to 72.1 ng/ml, n = 10), and mean spine T‐score increased from −1.07 (osteopenia) to −0.53 (normal) (n = 9). The magnitude of improvement in Gaucher disease manifestations and biomarkers over time was similar among the full trial cohort. Eliglustat was well‐tolerated and led to clinically significant improvements in previously untreated patients with Gaucher disease type 1 during 4.5 years of treatment.

ObjectiveHealth care delivery systems transformed rapidly at the beginning of the coronavirus disease 2019 (COVID-19) pandemic to slow the spread of the virus while identifying novel methods for providing care. In many ways, the pandemic affected both persons with neurologic illness and neurologists. This study describes the perspectives and experiences of community neurologists providing care for patients with neurodegenerative illnesses during the COVID-19 pandemic.MethodsWe conducted a qualitative study with 20 community neurologists from a multisite comparative-effectiveness trial of outpatient palliative care from July 23, 2020, to November 11, 2020. Participants were interviewed individually about the impact of the coronavirus disease 2019 (COVID-19) pandemic on their professional and personal lives. Interviews were analyzed with matrix analysis to identify key themes.ResultsFour main themes illustrated the impact of the pandemic on community neurologists: (1) challenges of the current political climate, (2) lack of support for new models of care, (3) being on the frontline of suffering, and (4) clinician self-care. Taken together, the themes capture the unusual environment in which community neurologists practice, the lack of clinician trust among some patients, patient and professional isolation, and opportunities to support quality care delivery.ConclusionsThe COVID-19 pandemic and pandemic politics created an environment that made care provision challenging for community neurologists. Efforts to improve care delivery should proactively work to reduce clinician burnout while incorporating support for new models of care adopted due to the pandemic.Trial Registration InformationClinicalTrials.gov identifier: NCT03076671.

Hypertension and diabetes are the greatest factors influencing the progression of chronic kidney disease (CKD). Investigation into the role of nephron number in CKD alone or with hypertension has revealed a strong inverse relationship between the two; however, not much is known about the connection between nephron number and diabetic kidney disease. The heterogeneous stock-derived model of unilateral renal agenesis (HSRA) rat, a novel model of nephron deficiency, provides a unique opportunity to study the association between nephron number and hypertension and diabetes on CKD. HSRA rats exhibit failure of one kidney to develop in 50−75% of offspring, whereas the remaining offspring are born with two kidneys. Rats born with one kidney (HSRA-S) develop significant renal injury with age compared with two-kidney littermates (HSRA-C). The induction of hypertension as a secondary stressor leads to significantly more renal injury in HSRA-S compared with HSRA-C rats and nephrectomized HSRA-C (HSRA-UNX) rats. The present study sought to address the hypothesis that nephron deficiency in the HSRA rat would hasten renal injury in the presence of a secondary stressor of hyperglycemia. HSRA animals did not exhibit diabetes-related traits at any age; thus, streptozotocin (STZ) was used to induce hyperglycemia in HSRA-S, HSRA-C, and HSRA-UNX rats. STZ- and vehicle-treated animals were followed for 15 wk. STZ-treated animals developed robust hyperglycemia, but in contrast to the response to hypertension, neither HSRA-S nor HSRA-UNX animals developed proteinuria compared with vehicle treatment. In total, our data indicate that hyperglycemia from STZ alone does not have a significant impact on the onset or progression of injury in young one-kidney HSRA animals. NEW & NOTEWORTHY The HSRA rat, a novel model of nephron deficiency, provides a unique opportunity to study the association between nephron number and confounding cardiovascular complications that impact kidney health. Although hypertension was previously shown to exacerbate renal injury in young HSRA animals, diabetic hyperglycemia did not lead to worse renal injury, suggesting that nephron number has limited impact on kidney injury, at least in this model.

BackgroundThe recognition, prevention and treatment of venous thromboembolism (VTE) remains a major challenge in the face of the recent COVID-19 pandemic which has been associated with significant cardiovascular, renal, respiratory and hematologic complications related to hypercoagulability. There has been little literature thus far on the utility of screening ultrasound and the role of the clinical pharmacist in treating these patients.MethodsWe present a prospective pilot program of thirty-one consecutive COVID-19 patients who were provided four extremity screening ultrasounds for VTE on admission. This was coordinated by a clinical pharmacist as part of a multidisciplinary approach. Quantitative and qualitative data were recorded with the goal of describing the utility of the clinical pharmacist in ultrasound screening. Data collected include demographics, information on clinical symptoms or signs at presentation, and laboratory and radiologic results during the hospitalization from each individual electronic medical record.ResultsNine of the thirty-one patients presented with VTE. Of the nine patients, there were twenty-two total clotted vessels, all of which were asymptomatic. The clinical pharmacist, as the coordinator for a multidisciplinary COVID-19 associated coagulopathy management team, drafted a screening and treatment protocol for anticoagulation prophylaxis and therapy of VTE after ultrasound findings.ConclusionVTE screening of hospitalized COVID-19 patients reveals a significant number of asymptomatic VTEs and justifies diagnostic, prophylactic, and treatment measures coordinated by a clinical pharmacist.

While stress may be a potential mechanism by which childhood threat and deprivation influence mental health, few studies have considered specific stress‐related white matter pathways, such as the stria terminalis (ST) and medial forebrain bundle (MFB). Our goal was to examine the relationships between childhood adversity and ST and MFB structural integrity and whether these pathways may provide a link between childhood adversity and affective symptoms and disorders. Participants were young adults (n = 100) with a full distribution of maltreatment history and affective symptom severity. Threat was determined by measures of childhood abuse and repeated traumatic events. Socioeconomic deprivation (SED) was determined by a measure of childhood socioeconomic status (parental education). Participants underwent diffusion spectrum imaging. Human Connectome Project data was used to perform ST and MFB tractography; these tracts were used as ROIs to extract generalized fractional anisotropy (gFA) from each participant. Childhood threat was associated with ST gFA, such that greater threat was associated with less ST gFA. SED was also associated with ST gFA, however, conversely to threat, greater SED was associated with greater ST gFA. Additionally, threat was negatively associated with MFB gFA, and MFB gFA was negatively associated with post‐traumatic stress symptoms. Our results suggest that childhood threat and deprivation have opposing influences on ST structural integrity, providing new evidence that the context of childhood adversity may have an important influence on its neurobiological effects, even on the same structure. Further, the MFB may provide a novel link between childhood threat and affective symptoms., This study provides the first evidence that childhood threat and deprivation have opposing influences on stria terminalis generalized fractional anisotropy (gFA), and novel evidence that different dimensions of childhood adversity may have differential influences on structural integrity or underlying microstructure, even within the same region. Further, our results demonstrate that the medial forebrain bundle may provide a novel link between childhood threat and affective symptoms. Examining how these stress‐related, visceral circuits link childhood adversity to affective symptoms could improve our understanding of mechanisms underlying affective disorders and provide novel targets for intervention.

Introduction: We evaluated knowledge, attitudes, and practices (KAP) related to influenza and influenza vaccination among pregnant women in three selected countries. Methods: During 2017, pregnant women seeking antenatal care at hospitals at participating sites were enrolled. We described characteristics and responses to KAP questions. We also evaluated predictors associated with influenza vaccination during pregnancy at sites with substantial influenza vaccine uptake by multivariable logistic regression. Results: Overall, 4,648 pregnant women completed the survey. There were substantial differences among the three survey populations; only 8% of the women in Nagpur had heard of influenza, compared to 90% in Lima and 96% in Bangkok (p-value

Background The decision to initiate pharmacotherapy is integral in the care for pregnant women with gestational diabetes mellitus (GDM). We sought to compare pregnancy outcomes between two threshold percentages of elevated glucose values prior to initiation of pharmacotherapy for GDM. We hypothesized that a lower threshold at pharmacotherapy initiation will be associated with lower rates of adverse perinatal outcomes. Methods This was a retrospective cohort study of women with GDM delivering in a single tertiary care center. Pregnancy outcomes were compared using bivariable and multivariable analyses between women who started pharmacotherapy (insulin or oral hypoglycemic agent) after a failed trial of dietary modifications at two different ranges of elevated capillary blood glucose (CBG) values: Group 1 when 20–39% CBG values were above goal; Group 2 when ≥40% CBG values were above goal. The primary outcome was a composite GDM-associated neonatal adverse outcome that included: macrosomia, large for gestational age (LGA), shoulder dystocia, hypoglycemia, hyperbilirubinemia requiring phototherapy, respiratory distress syndrome, stillbirth, and neonatal demise. Secondary outcomes included cesarean delivery, preterm birth ( Results A total of 417 women were included in the study. In univariable analysis, the composite neonatal outcome was statistically significantly higher in Group 2 compared to Group 1 (47.9% vs. 31.4%, p = 0.001). In addition, rates of preterm birth (15.7% vs 7.4%, p = 0.011), NICU admission (11.7% vs 4.0%, p = 0.006), and LGA (21.2% vs 9.1% p = 0.001) were higher in Group 2. In contrast, higher rates of SGA were noted in Group 1 (8.0% vs. 2.9%, p = 0.019). There was no difference in cesarean section rates. These findings persisted in multivariable analysis after adjusting for confounding factors (composite neonatal outcome aOR = 0.50, 95%CI [0.31–0.78]). Conclusions Initiation of pharmacotherapy for GDM when 20–39% of CBG values are above goal, compared to ≥40%, was associated with decreased rates of adverse neonatal outcomes attributable to GDM. This was accompanied by higher rates of SGA among women receiving pharmacotherapy at the lower threshold. Additional studies are required to identify the optimal threshold of abnormal CBG values to initiate pharmacotherapy for GDM.

Background: Transgender and gender expansive (TGE) youth face a number of adversities that are associated with deleterious consequences, including mental health problems, substance misuse, suicide, and homelessness. However, there is evidence that TGE youth still demonstrate resilience under extraordinary conditions. Aims: Therefore, this study examined how TGE youth who are homeless and engaging in survival sex describe and understand the protective factors present in their lives. Methods: In-depth qualitative interviews were conducted with 57 TGE youth living in a large U.S. city. Participants were 17-26 years old (M = 19.88, SD = 1.55), and identified as transgender woman (53%), transgender man (16%), and other (32%; gender-queer, gender-fluid, androgynous, or non-binary). Youth identified as Black (40%), multiracial (35%), Latinx (16%), White (5%), and as another race (3%). Researchers conducted a secondary analysis of qualitative data using thematic analysis. Results: We identified three themes to capture youth’s understanding of their protective factors: relying on oneself and others to avoid violence, accessing gender-affirming health and mental health care, and proactively maintaining sexual health. Participants described strategies for mitigating risk of harm while attempting to meet their basic needs. Additionally, they discussed the importance of having gender-affirming health and mental health services. Participants also expressed keen awareness of their sexual health risks and making calculated, often proactive decisions about their sexual health. Conclusion: The protective factors described by TGE youth reflect the unique ways in which hyper-marginalized youth must navigate their own survival and well-being, and align with literature suggesting resilience is not mutually exclusive from risk. By focusing on TGE youth, this research privileges the experiences of individuals historically underrepresented in research and encourages future research to incorporate these perspectives into policies and programs.

ObjectivesTo investigate the impact of Alzheimer's disease (AD) co-pathology on an in vivo structural measure of neurodegeneration in Lewy body disorders (LBD).MethodsWe studied 72 LBD patients (Parkinson disease (PD)=2, PD-MCI=25, PD with dementia=10, dementia with Lewy bodies=35) with either CSF analysis or neuropathological examination and structural MRI during life. The cohort was divided into those harboring significant AD co-pathology, either at autopsy (intermediate/high AD neuropathologic change) or with CSF signature indicating AD co-pathology (t-tau/Aβ1-42 >0.3) (LBD+AD, N=19), and those without AD co-pathology (LBD-AD, N=53). We also included a reference group of 25 patients with CSF biomarker-confirmed amnestic AD. We investigated differences in MRI cortical thickness estimates between groups, and in the 21 autopsied LBD patients (LBD-AD=14, LBD+AD=7), directly tested the association between antemortem MRI and post-mortem burdens of tau, Aβ, and alpha-synuclein using digital histopathology in five representative neocortical regions.ResultsThe LBD+AD group was characterized by cortical thinning in anterior/medial and lateral temporal regions (P&nbsp

As stroke mortality rates decrease, there has been a surge of effort to study poststroke dementia (PSD) to improve long‐term quality of life for stroke survivors. Hippocampal volume may be an important neuroimaging biomarker in poststroke dementia, as it has been associated with many other forms of dementia. However, studying hippocampal volume using MRI requires hippocampal segmentation. Advances in automated segmentation methods have allowed for studying the hippocampus on a large scale, which is important for robust results in the heterogeneous stroke population. However, most of these automated methods use a single atlas‐based approach and may fail in the presence of severe structural abnormalities common in stroke. Hippodeep, a new convolutional neural network‐based hippocampal segmentation method, does not rely solely on a single atlas‐based approach and thus may be better suited for stroke populations. Here, we compared quality control and the accuracy of segmentations generated by Hippodeep and two well‐accepted hippocampal segmentation methods on stroke MRIs (FreeSurfer 6.0 whole hippocampus and FreeSurfer 6.0 sum of hippocampal subfields). Quality control was performed using a stringent protocol for visual inspection of the segmentations, and accuracy was measured as volumetric correlation with manual segmentations. Hippodeep performed significantly better than both FreeSurfer methods in terms of quality control. All three automated segmentation methods had good correlation with manual segmentations and no one method was significantly more correlated than the others. Overall, this study suggests that both Hippodeep and FreeSurfer may be useful for hippocampal segmentation in stroke rehabilitation research, but Hippodeep may be more robust to stroke lesion anatomy., In this study, we compared three automated hippocampal segmentation methods in a large stroke population in terms of quality control and segmentation accuracy compared to manual segmentations. While all three methods yielded similar volumes, new convolutional neural network based segmentation method Hippodeep had the lowest method‐wise quality control fail rate, suggesting it may be the most robust to post‐stroke anatomical distortions.

The American lobster, Homarus americanus, cardiac neuromuscular system is controlled by the cardiac ganglion (CG), a central pattern generator consisting of four premotor and five motor neurons. Here, we show that the premotor and motor neurons can establish independent bursting patterns when decoupled by a physical ligature. We also show that mRNA encoding myosuppressin, a cardioactive neuropeptide, is produced within the CG. We thus asked whether myosuppressin modulates the decoupled premotor and motor neurons, and if so, how this modulation might underlie the role(s) that these neurons play in myosuppressin’s effects on ganglionic output. Although myosuppressin exerted dose-dependent effects on burst frequency and duration in both premotor and motor neurons in the intact CG, its effects on the ligatured ganglion were more complex, with different effects and thresholds on the two types of neurons. These data suggest that the motor neurons are more important in determining the changes in frequency of the CG elicited by low concentrations of myosuppressin, whereas the premotor neurons have a greater impact on changes elicited in burst duration. A single putative myosuppressin receptor (MSR-I) was previously described from the Homarus nervous system. We identified four additional putative MSRs (MSR-II–V) and investigated their individual distributions in the CG premotor and motor neurons using RT-PCR. Transcripts for only three receptors (MSR-II–IV) were amplified from the CG. Potential differential distributions of the receptors were observed between the premotor and motor neurons; these differences may contribute to the distinct physiological responses of the two neuron types to myosuppressin. NEW & NOTEWORTHY Premotor and motor neurons of the Homarus americanus cardiac ganglion (CG) are normally electrically and chemically coupled, and generate rhythmic bursting that drives cardiac contractions; we show that they can establish independent bursting patterns when physically decoupled by a ligature. The neuropeptide myosuppressin modulates different aspects of the bursting pattern in these neuron types to determine the overall modulation of the intact CG. Differential distribution of myosuppressin receptors may underlie the observed responses to myosuppressin.

Using electrocochleography, the summating potential (SP) is a deflection from baseline to tones and an early rise in the response to clicks. Here, we use normal hearing gerbils and gerbils with outer hair cells removed with a combination of furosemide and kanamycin to investigate cellular origins of the SP. Round window electrocochleography to tones and clicks was performed before and after application of tetrodotoxin to prevent action potentials, and then again after kainic acid to prevent generation of an EPSP. With appropriate subtractions of the response curves from the different conditions, the contributions to the SP from outer hair cells, inner hair cell, and neural "spiking" and "dendritic" responses were isolated. Like hair cells, the spiking and dendritic components had opposite polarities to tones - the dendritic component had negative polarity and the spiking component had positive polarity. The magnitude of the spiking component was larger than the dendritic across frequencies and intensities. The onset to tones and to clicks followed a similar sequence; the outer hair cells responded first, then inner hair cells, then the dendritic component, and then the compound action potential of the spiking response. These results show the sources of the SP include at least the four components studied, and that these have a mixture of polarities and magnitudes that vary across frequency and intensity. Thus, multiple possible interactions must be considered when interpreting the SP for clinical uses.

Neuromodulation treatment effect size for bothersome tinnitus may be larger and more predictable by adopting a target selection approach guided by personalized striatal networks or functional connectivity maps. Several corticostriatal mechanisms are likely to play a role in tinnitus, including the dorsal/ventral striatum and the putamen. We examined whether significant tinnitus treatment response by deep brain stimulation (DBS) of the caudate nucleus may be related to striatal network increased functional connectivity with tinnitus networks that involve the auditory cortex or ventral cerebellum. The first study was a cross‐sectional 2‐by‐2 factorial design (tinnitus, no tinnitus; hearing loss, normal hearing, n = 68) to define cohort level abnormal functional connectivity maps using high‐field 7.0 T resting‐state fMRI. The second study was a pilot case–control series (n = 2) to examine whether tinnitus modulation response to caudate tail subdivision stimulation would be contingent on individual level striatal connectivity map relationships with tinnitus networks. Resting‐state fMRI identified five caudate subdivisions with abnormal cohort level functional connectivity maps. Of those, two connectivity maps exhibited increased connectivity with tinnitus networks—dorsal caudate head with Heschl's gyrus and caudate tail with the ventral cerebellum. DBS of the caudate tail in the case‐series responder resulted in dramatic reductions in tinnitus severity and loudness, in contrast to the nonresponder who showed no tinnitus modulation. The individual level connectivity map of the responder was in alignment with the cohort expectation connectivity map, where the caudate tail exhibited increased connectivity with tinnitus networks, whereas the nonresponder individual level connectivity map did not., Successful neuromodulation for bothersome tinnitus may more attainable by adopting a target selection approach guided by personalized striatal networks or functional connectivity maps. Here, we present a two‐pronged fMRI study (cohort and case–control) demonstrating increased caudate‐corticocerebellar connectivity in both a large cohort of tinnitus sufferers and a single patient that was a neuromodulatory DBS responder.

Background Financial incentives and audit/feedback are widely used in primary care to influence clinician behaviour and increase quality of care. While observational data suggest a decline in quality when these interventions are stopped, their removal has not been evaluated in a randomised controlled trial (RCT), to our knowledge. This trial aimed to determine whether chlamydia testing in general practice is sustained when financial incentives and/or audit/feedback are removed. Methods and findings We undertook a 2 × 2 factorial cluster RCT in 60 general practices in 4 Australian states targeting 49,525 patients aged 16–29 years for annual chlamydia testing. Clinics were recruited between July 2014 and September 2015 and were followed for up to 2 years or until 31 December 2016. Clinics were eligible if they were in the intervention group of a previous cluster RCT where general practitioners (GPs) received financial incentives (AU$5–AU$8) for each chlamydia test and quarterly audit/feedback reports of their chlamydia testing rates. Clinics were randomised into 1 of 4 groups: incentives removed but audit/feedback retained (group A), audit/feedback removed but incentives retained (group B), both removed (group C), or both retained (group D). The primary outcome was the annual chlamydia testing rate among 16- to 29-year-old patients, where the numerator was the number who had at least 1 chlamydia test within 12 months and the denominator was the number who had at least 1 consultation during the same 12 months. We undertook a factorial analysis in which we investigated the effects of removal versus retention of incentives (groups A + C versus groups B + D) and the effects of removal versus retention of audit/feedback (group B + C versus groups A + D) separately. Of 60 clinics, 59 were randomised and 55 (91.7%) provided data (group A: 15 clinics, 11,196 patients; group B: 14, 11,944; group C: 13, 11,566; group D: 13, 14,819). Annual testing decreased from 20.2% to 11.7% (difference −8.8%; 95% CI −10.5% to −7.0%) in clinics with incentives removed and decreased from 20.6% to 14.3% (difference −7.1%; 95% CI −9.6% to −4.7%) where incentives were retained. The adjusted absolute difference in treatment effect was −0.9% (95% CI −3.5% to 1.7%; p = 0.2267). Annual testing decreased from 21.0% to 11.6% (difference −9.5%; 95% CI −11.7% to −7.4%) in clinics where audit/feedback was removed and decreased from 19.9% to 14.5% (difference −6.4%; 95% CI −8.6% to −4.2%) where audit/feedback was retained. The adjusted absolute difference in treatment effect was −2.6% (95% CI −5.4% to −0.1%; p = 0.0336). Study limitations included an unexpected reduction in testing across all groups impacting statistical power, loss of 4 clinics after randomisation, and inclusion of rural clinics only. Conclusions Audit/feedback is more effective than financial incentives of AU$5–AU$8 per chlamydia test at sustaining GP chlamydia testing practices over time in Australian general practice. Trial registration Australian New Zealand Clinical Trials Registry ACTRN12614000595617, In a cluster randomized trial, Jane S Hocking and colleagues investigate the impact of removing financial incentives and/or audit and feedback on chlamydia testing in general practice in Australia., Author summary Why was this study done? Financial incentives and audit/feedback are widely used in primary care to influence clinician behaviour and increase quality of care. As healthcare costs continue to increase, governments and funding agencies are reassessing funding models for primary care, with widespread cuts to financial incentives. While observational data suggest a decline in quality when these interventions are stopped, their removal has not been evaluated in a randomised controlled trial (RCT). What did the researchers do and find? We conducted a 2 × 2 factorial cluster RCT in Australian general practices that aimed to determine the impact on chlamydia testing in general practice when incentive payments per activity and/or audit/feedback on activity performance were removed. Clinics were randomised into 1 of 4 groups: incentives removed but audit/feedback retained, audit/feedback removed but incentives retained, both removed, and both retained. The primary outcome was the annual chlamydia testing rate among 16- to 29-year-old patients. We found that removal of incentive payments had little impact on general practice chlamydia testing, but the removal of audit and feedback reduced testing. What do these results mean? Our payments were consistent with other incentives general practitioners (GPs) received at the time, suggesting that in the Australian general practice setting, incentive payments of this amount do not have a substantial impact on influencing GP preventive healthcare activities such as chlamydia testing. The removal of quarterly audit and feedback for GPs had a greater impact on testing rates, reflecting the importance of this strategy in influencing GP preventive healthcare activities. The provision of audit and feedback was costlier than the provision of financial incentives. However, using online video conferencing and fully automating the audit and feedback reports would reduce costs. Our results suggest that, in Australia at least, audit and feedback is more effective than incentive payments of AU$5 to AU$8 per activity at influencing GP behaviour.

Background: Persistent pain is a prevalent condition that negatively influences physical, emotional, social and family functioning in adolescents. Pain science education is a promising therapy for adults, yet to be thoroughly investigated for persistent pain in adolescents. There is a need to develop suitable curricula for adolescent pain science education. Methods: An interdisciplinary meeting of 12 clinicians and researchers was held during March 2018 in Adelaide, South Australia. An a priori objective of the meeting was to identify and gain consensus on key learning objectives for adolescent pain science education using a modified-Delphi process. Results and Conclusion: Consensus was reached via a modified Delphi process for seven learning objectives to form the foundation of a curriculum: 1) Pain is a protector; 2) The pain system can become overprotective; 3) Pain is a brain output; 4) Pain is not an accurate marker of tissue state; 5) There are many potential contributors to anyone’s pain; 6) We are all bioplastic and; 7) Pain education is treatment. Recommendations are made for promising areas for future research in adolescent pain science education.

Background Cardiovascular disease (CVD) is the leading cause of mortality in India. Social and behavioural factors are strongly interrelated in the prevention and control of CVD. The ability to make lifestyle changes to control hypertension and diabetes (major risk factors for CVD) is determined by factors such as education, gender, caste, poverty, and urbanicity. This study aimed to improve our understanding of the inter-relationship of social and behavioural factors in the management of elevated serum glucose and high blood pressure and co-morbid mental health conditions. Methods A population-based catchment area cross sectional survey was conducted in Kerala, India. Data were collected from residents aged over 30 years (n = 997) using standardized tools and clinical measures. We performed latent class analysis incrementally to extract homogeneous latent classes of individuals based on their responses to social and behavioural risk factors in the survey. Using structural equation models, we assessed the mediating effect of depression and anxiety, and social or behavioural risk factors, on management of high blood pressure and raised serum glucose levels. Results The prevalence of high blood pressure and blood glucose in the sample was 33 and 26% respectively. Latent class analysis found three clusters of risk factors. One had a predominance of behavioural characteristics, another of social risk factors and the third was a low risk group. Age, female sex, and marital status had an effect on high blood pressure and high glucose, though were mediated by mental health, social and behavioural risk factors. Conclusions Interventions to improve the management of risk factors for CVD need to address social risk factors and be sensitive to the needs of population sub-groups that may require additional support to access health services. An integration of social and health services may be required to achieve this.

Background For breast cancer patients undergoing neoadjuvant chemotherapy (NAC), pathologic complete response (pCR; no invasive or in situ) cannot be assessed non-invasively so all patients undergo surgery. The aim of our study was to develop and validate a radiomics classifier that classifies breast cancer pCR post-NAC on MRI prior to surgery. Methods This retrospective study included women treated with NAC for breast cancer from 2014 to 2016 with (1) pre- and post-NAC breast MRI and (2) post-NAC surgical pathology report assessing response. Automated radiomics analysis of pre- and post-NAC breast MRI involved image segmentation, radiomics feature extraction, feature pre-filtering, and classifier building through recursive feature elimination random forest (RFE-RF) machine learning. The RFE-RF classifier was trained with nested five-fold cross-validation using (a) radiomics only (model 1) and (b) radiomics and molecular subtype (model 2). Class imbalance was addressed using the synthetic minority oversampling technique. Results Two hundred seventy-three women with 278 invasive breast cancers were included; the training set consisted of 222 cancers (61 pCR, 161 no-pCR; mean age 51.8 years, SD 11.8), and the independent test set consisted of 56 cancers (13 pCR, 43 no-pCR; mean age 51.3 years, SD 11.8). There was no significant difference in pCR or molecular subtype between the training and test sets. Model 1 achieved a cross-validation AUROC of 0.72 (95% CI 0.64, 0.79) and a similarly accurate (P = 0.1) AUROC of 0.83 (95% CI 0.71, 0.94) in both the training and test sets. Model 2 achieved a cross-validation AUROC of 0.80 (95% CI 0.72, 0.87) and a similar (P = 0.9) AUROC of 0.78 (95% CI 0.62, 0.94) in both the training and test sets. Conclusions This study validated a radiomics classifier combining radiomics with molecular subtypes that accurately classifies pCR on MRI post-NAC.

Background HIV testing programs in the United States aim to reach ethnic minority populations who experience high incidence of HIV, yet 40% of African Americans have never been tested for HIV. The objective of this study is to identify community-based strategies to increase testing among African Americans in both urban and rural areas. Methods This study conducted focus group discussions (FGDs) informed by community-based participatory research principles to examine African American’s concerns and ideas around HIV testing and HIV self-testing. Participants included highly affected (i.e., PLWH, MSM, PWID, low-income, teens and young adults) populations from African American communities in North Carolina, aged 15 years and older. We digitally transcribed and analyzed qualitative data using MAXQDA and axial coding to identify emergent themes. Results Fifty-two men and women between 15 to 60 years old living in urban (n=41) and rural (n=11) areas of North Carolina participated in focus group discussions. HIV testing barriers differed by HIV testing setting: facility-based, community-based, and HIV self-testing. In community-based settings, barriers included confidentiality concerns. In facility-based settings (e.g., clinics), barriers included negative treatment by healthcare workers. With HIV self-testing, barriers included improper use of self-testing kits and lack of post-test support. HIV testing facilitators included partnering with community leaders, decentralizing testing beyond facility-based sites, and protecting confidentiality. Conclusions Findings suggest that HIV testing concerns among African Americans vary by HIV testing setting. African Americans may be willing to test for HIV at community events in public locations if client confidentiality is preserved and use HIV self-testing kits in private if post-test social support and services are provided. These community-identified facilitators may improve African American testing rates and uptake of HIV self-testing kits.

Community involvement may be essential for conservation programme success. We focus on farmers, asking how and why they believe conservation interventions will work, or not. Here we test models of folk theories of the human motivational factors required for behaviour change, in 3 rural central Chilean communities. We hypothesize that different models will be supported by farmers with different experiences with conservation programmes, and that socioeconomic and production system variation will explain further variation in who supports each working model. We use a multiple methods approach, combining a questionnaire with participant-observation. We find support for three of the working models of human behavioural change, among different socio-economic profiles of farmers. We believe that the schema of working models provides a boundary object to facilitate communication between conservationists and stakeholders, and can help improve conservation project design and implementation. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13280-019-01315-6) contains supplementary material, which is available to authorized users.

KCNMA1, encoding the voltage- and calcium-activated potassium channel, has a pivotal role in brain physiology. Mutations in KCNMA1 are associated with epilepsy and/or dyskinesia (PNKD3). Two KCNMA1 mutations correlated with these phenotypes, D434G and N999S, were previously identified as producing gain-of-function (GOF) effects on BK channel activity. Three new patients have been reported harboring N999S, one carrying a second mutation, R1128W, but the effects of these mutations have not yet been reported under physiological K+ conditions or compared to D434G. In this study, we characterize N999S, the novel N999S/R1128W double mutation, and D434G in a brain BK channel splice variant, comparing the effects on BK current properties under a physiological K+ gradient with action potential voltage commands. N999S, N999S/R1128W, and D434G cDNAs were expressed in HEK293T cells and characterized by patch-clamp electrophysiology. N999S BK currents were shifted to negative potentials, with faster activation and slower deactivation compared with wild type (WT) and D434G. The double mutation N999S/R1128W did not show any additional changes in current properties compared with N999S alone. The antiepileptic drug acetazolamide was assessed for its ability to directly modulate WT and N999S channels. Neither the WT nor N999S channels were sensitive to the antiepileptic drug acetazolamide, but both were sensitive to the inhibitor paxilline. We conclude that N999S is a strong GOF mutation that surpasses the D434G phenotype, without mitigation by R1128W. Acetazolamide has no direct modulatory action on either WT or N999S channels, indicating that its use may not be contraindicated in patients harboring GOF KCNMA1 mutations. NEW & NOTEWORTHY KCNMA1-linked channelopathy is a new neurological disorder characterized by mutations in the BK voltage- and calcium-activated potassium channel. The epilepsy- and dyskinesia-associated gain-of-function mutations N999S and D434G comprise the largest number of patients in the cohort. This study provides the first direct comparison between D434G and N999S BK channel properties as well as a novel double mutation, N999S/R1128W, from another patient, defining the functional effects during an action potential stimulus.

BACKGROUND: We determined the association between ratios of plasma ceramide species of differing fatty-acyl chain lengths and incident dementia and Alzheimers disease (AD) dementia in a large, community-based sample. METHODS: We measured plasma ceramide levels in 1892 [54% women, mean age 70.1 (SD 6.9) yr.] dementia-free Framingham Offspring Study cohort participants between 2005 and 2008. We related ratios of very long-chain (C24:0, C22:0) to long-chain (C16:0) ceramides to subsequent risk of incident dementia and AD dementia. Structural MRI brain measures were included as secondary outcomes. RESULTS: During a median 6.5year follow-up, 81 participants developed dementia, of whom 60 were diagnosed with AD dementia. In multivariable Cox-proportional hazards analyses, each standard deviation (SD) increment in the ratio of ceramides C24:0/C16:0 was associated with a 27% reduction in the risk of dementia (HR 0.73, 95% CI 0.56-0.96) and AD dementia (HR 0.73, 95% CI 0.53-1.00). The ratio of ceramides C22:0/C16:0 was also inversely associated with incident dementia (HR per SD 0.75, 95% CI 0.57-0.98), and approached statistical significance for AD (HR 0.73, 95% CI 0.53-1.01, P=0.056). Higher ratios of ceramides C24:0/C16:0 and C22:0/C16:0 were also cross-sectionally associated with lower white matter hyperintensity burden on MRI (-0.05±0.02, P=0.02; -0.06±0.02, P=0.003; respectively per SD increase), but not with other MRI brain measures. CONCLUSIONS: Higher plasma ratios of very long-chain to long-chain ceramides are associated with a reduced risk of incident dementia and AD dementia in our community-based sample. Circulating ceramide ratios may serve as potential biomarkers for predicting dementia risk in cognitively healthy adults.

Background: To evaluate the relationship between periconceptional (period before and/or after conception) substance use and unfavorable pregnancy contexts, including unintended pregnancy. Materials and Methods: This is a cross-sectional analysis of English- or Spanish-speaking women aged 16–44 years with pregnancies

Aim The purpose of this study was to explore the latent conditions of cooperation and conflict in intra‐hospital patient transfers (i.e. transfers of patients between units in a hospital). Design Secondary qualitative analysis of 28 interviews conducted with 29 hospital staff, including physicians (N = 13), nurses (N = 10) and support staff (N = 6) from a single, large academic tertiary hospital in the Northeastern United States. Methods A two‐member multidisciplinary team applied a directed content analysis approach to data collected from semi‐structured interviews. Results Three recurrent themes were generated: (a) patient flow policies created imbalances of power; (b) relationships were helpful to facilitate safe transfers; and (c) method of admission order communication was a source of disagreement. Hospital quality improvement efforts could benefit from a teaming approach to minimize unintentional power imbalances and optimize communicative relationships between units.

Background Citizen science is increasingly prevalent in the biomedical sciences, including the field of human genomics. Genomic citizen science initiatives present new opportunities to engage individuals in scientific discovery, but they also are provoking new questions regarding who owns the outputs of the research, including intangible ideas and discoveries and tangible writings, tools, technologies, and products. The legal and ethical claims of participants to research outputs become stronger—and also more likely to conflict with those of institution-based researchers and other stakeholders—as participants become more involved, quantitatively and qualitatively, in the research process. It is not yet known, however, how genomic citizen science initiatives are managing the interests of their participants in accessing and controlling research outputs in practice. To help fill this gap, we conducted an in-depth review of relevant policies and practices of U.S.-based genomic citizen science initiatives. Methods We queried the peer-reviewed literature and grey literature to identify 22 genomic citizen science initiatives that satisfied six inclusion criteria. A data collection form was used to capture initiative features, policies, and practices relevant to participants’ access to and control over research outputs. Results This analysis revealed that the genomic citizen science landscape is diverse and includes many initiatives that do not have institutional affiliations. Two trends that are in apparent tension were identified: commercialization and operationalization of a philosophy of openness. While most initiatives supported participants’ access to research outputs, including datasets and published findings, none supported participants’ control over results via intellectual property, licensing, or commercialization rights. However, several initiatives disclaimed their own rights to profit from outputs. Conclusions There are opportunities for citizen science initiatives to incorporate more features that support participants’ access to and control over research outputs, consistent with their specific objectives, operations, and technical capabilities.

Human immunodeficiency virus (HIV) infected adults are at a higher risk of pneumococcal colonisation and disease, even while receiving antiretroviral therapy (ART). To help evaluate potential indirect effects of vaccination of HIV-infected adults, we assessed whether HIV-infected adults disproportionately contribute to household transmission of pneumococci. We constructed a hidden Markov model to capture the dynamics of pneumococcal carriage acquisition and clearance observed during a longitudinal household-based nasopharyngeal swabbing study, while accounting for sample misclassifications. Households were followed-up twice weekly for approximately 10 months each year during a three-year study period for nasopharyngeal carriage detection via real-time PCR. We estimated the effect of participant’s age, HIV status, presence of a HIV-infected adult within the household and other covariates on pneumococcal acquisition and clearance probabilities. Of 1,684 individuals enrolled, 279 (16.6%) were younger children (, Author summary We assessed the contribution of HIV-infected adults to household pneumococcal transmission by applying a hidden Markov model to pneumococcal cohort data comprising 115,595 nasopharyngeal samples from 1,684 individuals in rural and urban settings in South Africa. We estimated 14.4% of sample misclassifications (false negatives), representing 85.6% sensitivity of a test that was used to detect pneumococcus. Pneumococcal carriage prevalence and acquisition rates, and average duration were usually higher in younger or older children than adults. The use of ART and antibiotics reduced the average carriage duration across all age and HIV groups, and carriage acquisition risks increased in larger household sizes. Despite the longer average carriage duration and higher median carriage density in HIV-infected than HIV-uninfected adults, we found similar carriage acquisition and onward transmission risks in the dual groups. These findings suggest that vaccinating HIV-infected adults on ART with PCV would reduce their risk for pneumococcal disease but may add little to the indirect protection against carriage of the rest of the population.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a historic pandemic of respiratory disease (coronavirus disease 2019 [COVID-19]), and current evidence suggests that severe disease is associated with dysregulated immunity within the respiratory tract. However, the innate immune mechanisms that mediate protection during COVID-19 are not well defined. Here, we characterize a mouse model of SARS-CoV-2 infection and find that early CCR2 signaling restricts the viral burden in the lung. We find that a recently developed mouse-adapted SARS-CoV-2 (MA-SARS-CoV-2) strain as well as the emerging B.1.351 variant trigger an inflammatory response in the lung characterized by the expression of proinflammatory cytokines and interferon-stimulated genes. Using intravital antibody labeling, we demonstrate that MA-SARS-CoV-2 infection leads to increases in circulating monocytes and an influx of CD45+ cells into the lung parenchyma that is dominated by monocyte-derived cells. Single-cell RNA sequencing (scRNA-Seq) analysis of lung homogenates identified a hyperinflammatory monocyte profile. We utilize this model to demonstrate that mechanistically, CCR2 signaling promotes the infiltration of classical monocytes into the lung and the expansion of monocyte-derived cells. Parenchymal monocyte-derived cells appear to play a protective role against MA-SARS-CoV-2, as mice lacking CCR2 showed higher viral loads in the lungs, increased lung viral dissemination, and elevated inflammatory cytokine responses. These studies have identified a potential CCR2-monocyte axis that is critical for promoting viral control and restricting inflammation within the respiratory tract during SARS-CoV-2 infection. IMPORTANCE SARS-CoV-2 has caused a historic pandemic of respiratory disease (COVID-19), and current evidence suggests that severe disease is associated with dysregulated immunity within the respiratory tract. However, the innate immune mechanisms that mediate protection during COVID-19 are not well defined. Here, we characterize a mouse model of SARS-CoV-2 infection and find that early CCR2-dependent infiltration of monocytes restricts the viral burden in the lung. We find that SARS-CoV-2 triggers an inflammatory response in the lung characterized by the expression of proinflammatory cytokines and interferon-stimulated genes. Using RNA sequencing and flow cytometry approaches, we demonstrate that SARS-CoV-2 infection leads to increases in circulating monocytes and an influx of CD45+ cells into the lung parenchyma that is dominated by monocyte-derived cells. Mechanistically, CCR2 signaling promoted the infiltration of classical monocytes into the lung and the expansion of monocyte-derived cells. Parenchymal monocyte-derived cells appear to play a protective role against MA-SARS-CoV-2, as mice lacking CCR2 showed higher viral loads in the lungs, increased lung viral dissemination, and elevated inflammatory cytokine responses. These studies have identified that the CCR2 pathway is critical for promoting viral control and restricting inflammation within the respiratory tract during SARS-CoV-2 infection.

Background The current project sought to evaluate the impact that white matter hyperintensities (WMH) have on executive function in cognitively normal Mexican Americans, an underserved population with onset and more rapid progression of dementia. Methods Data from 515 participants (360 female) enrolled in the Health and Aging Brain Study: Health Disparities project were analyzed. Participants underwent clinical evaluation, cognitive testing, and a brain MRI. Linear regression was used to predict the effect of total WMH volume on cognitive test scores. Age, sex, and education were entered as covariates. Results Regression analysis showed that WMH volume significantly predicted executive function. WMH also predicted global cognition and attention scores, although not significantly after adjusting for age. Conclusion In this sample of cognitively normal Mexican Americans, we found that WMH volume was associated with lower scores in a measure of executive function, after accounting for age, sex, and education.

Nonalcoholic fatty liver disease (NAFLD) is a risk factor for liver cancer and prevalence varies by ethnicity. Along with genetic and lifestyle factors, the gut microbiome (GM) may contribute to NAFLD and its progression to advanced liver disease. Our cross-sectional analysis assessed the association of the GM with hepatic adiposity among African American, Japanese American, White, Latino, and Native Hawaiian participants in the Multiethnic Cohort. We used MRI to measure liver fat and determine nonalcoholic fatty liver disease (NAFLD) status (n = 511 cases) in 1,544 participants, aged 60–77 years, with 12–53% overall adiposity (BMI of 17.8–46.2 kg/m2). The GM was measured by 16S rRNA gene sequencing and, on a subset, by metagenomic sequencing. Alpha diversity was lower overall with NAFLD and in certain ethnicities (African Americans, Whites, and Latinos). In models regressing genus on NAFLD status, 62 of 149 genera (40%) exhibited a significant interaction between NAFLD and ethnicity stratified analysis found 69 genera significantly associated with NAFLD in at least one ethnic group. No single genus was significantly associated with NAFLD across all ethnicities. In contrast, the same bacterial metabolic pathways were over-represented in participants with NAFLD regardless of ethnicity. Imputed secondary bile acid and carbohydrate pathways were associated with NAFLD, the latter of which was corroborated by metagenomics, although different genera in different ethnicities were associated with these pathways. Overall, we found that NAFLD was associated with altered bacterial composition and metabolism, and that bacterial endotoxin, assessed by plasma lipopolysaccharide binding protein (LBP), may mediate liver fat-associated systemic inflammation in a manner that seems to vary by ethnicity.

Remdesivir (RDV), a broadly acting nucleoside analogue, is the only FDA approved small molecule antiviral for the treatment of COVID-19 patients. To date, there are no reports identifying SARS-CoV-2 RDV resistance in patients, animal models or in vitro. Here, we selected drug-resistant viral populations by serially passaging SARS-CoV-2 in vitro in the presence of RDV. Using high throughput sequencing, we identified a single mutation in RNA-dependent RNA polymerase (NSP12) at a residue conserved among all coronaviruses in two independently evolved populations displaying decreased RDV sensitivity. Introduction of the NSP12 E802D mutation into our SARS-CoV-2 reverse genetics backbone confirmed its role in decreasing RDV sensitivity in vitro. Substitution of E802 did not affect viral replication or activity of an alternate nucleoside analogue (EIDD2801) but did affect virus fitness in a competition assay. Analysis of the globally circulating SARS-CoV-2 variants (>800,000 sequences) showed no evidence of widespread transmission of RDV-resistant mutants. Surprisingly, we observed an excess of substitutions in spike at corresponding sites identified in the emerging SARS-CoV-2 variants of concern (i.e., H69, E484, N501, H655) indicating that they can arise in vitro in the absence of immune selection. The identification and characterisation of a drug resistant signature within the SARS-CoV-2 genome has implications for clinical management and virus surveillance., Author summary The emergence of SARS-CoV-2 has led to a worldwide pandemic with significant morbidity and mortality. Remdesivir is the only antiviral with FDA approval for treatment. Antivirals use comes at a risk, as viruses may acquire mutations to overcome the inhibition. We identified a mutation in the virus polymerase responsible for decreased sensitivity to Remdesivir. A change at this conserved site was not predicted, and the mutation did not cause a replication advantage or change in sensitivity to another antiviral drug. Importantly, this change occurred at very low frequency globally. Unexpectedly, passage of SARS-CoV-2 led to an accumulation of mutations in spike. A number occurred at the same sites but to different residues as those in emerging variants of concern indicating they arise in the absence of immune pressure. Our data indicates low-level Remdesivir resistance in SARS-CoV-2 is different to other RNA viruses and monitoring changes in vitro provides insight into general virus adaptation of newly emerging viruses.

Everyone who serves in the military (and survives the experience) will eventually become a Veteran, and most will face the challenge of finding a civilian job. This paper investigates how contemporary Veterans experience the transition period between military exit and entrance into civilian life and how their own actions before separation shape their post-transition outcomes. We follow 35 servicemembers through the transition process, interviewing them before and several months after they left the military. These interviews reveal the importance of three factors – the conditions triggering their exit, the strength of their military identity, and their own planning (or lack thereof) for the transition to civilian life – in enhancing or detracting from achievement of a stated post-military goals and objective success in moving into work or school. The strength and depth of an individual’s institutional identity shapes how and when servicemembers plan for military exit and how they adjust to unanticipated military exits. Early planning reflects anticipatory socialization for new civilian roles and is associated with better post-military outcomes. But early planning is often hindered by a strong military identity or facilitated by a weak military identity. These findings have important implications for the military and advocates who serve them with the recent military policy changes to transition assistance and the retirement pension system that encourage early planning for post-military life.

Patients who recover from nosocomial pneumonia oftentimes exhibit long-lasting cognitive impairment comparable with what is observed in Alzheimer’s disease patients. We previously hypothesized that the lung endothelium contributes to infection-related neurocognitive dysfunction, because bacteria-exposed endothelial cells release a form(s) of cytotoxic tau that is sufficient to impair long-term potentiation in the hippocampus. However, the full-length lung and endothelial tau isoform(s) have yet to be resolved and it remains unclear whether the infection-induced endothelial cytotoxic tau triggers neuronal tau aggregation. Here, we demonstrate that lung endothelial cells express a big tau isoform and three additional tau isoforms that are similar to neuronal tau, each containing four microtubule-binding repeat domains, and that tau is expressed in lung capillaries in vivo. To test whether infection elicits endothelial tau capable of causing transmissible tau aggregation, the cells were infected with Pseudomonas aeruginosa. The infection-induced tau released from endothelium into the medium-induced neuronal tau aggregation in reporter cells, including reporter cells that express either the four microtubule-binding repeat domains or the full-length tau. Infection-induced release of pathological tau variant(s) from endothelium, and the ability of the endothelial-derived tau to cause neuronal tau aggregation, was abolished in tau knockout cells. After bacterial lung infection, brain homogenates from WT mice, but not from tau knockout mice, initiated tau aggregation. Thus, we conclude that bacterial pneumonia initiates the release of lung endothelial-derived cytotoxic tau, which is capable of propagating a neuronal tauopathy.

SARS-CoV-2 is notable both for its rapid spread, and for the heterogeneity of its patterns of transmission, with multiple published incidences of superspreading behaviour. Here, we applied a novel network reconstruction algorithm to infer patterns of viral transmission occurring between patients and health care workers (HCWs) in the largest clusters of COVID-19 infection identified during the first wave of the epidemic at Cambridge University Hospitals NHS Foundation Trust, UK. Based upon dates of individuals reporting symptoms, recorded individual locations, and viral genome sequence data, we show an uneven pattern of transmission between individuals, with patients being much more likely to be infected by other patients than by HCWs. Further, the data were consistent with a pattern of superspreading, whereby 21% of individuals caused 80% of transmission events. Our study provides a detailed retrospective analysis of nosocomial SARS-CoV-2 transmission, and sheds light on the need for intensive and pervasive infection control procedures., eLife digest The COVID-19 pandemic, caused by the SARS-CoV-2 virus, presents a global public health challenge. Hospitals have been at the forefront of this battle, treating large numbers of sick patients over several waves of infection. Finding ways to manage the spread of the virus in hospitals is key to protecting vulnerable patients and workers, while keeping hospitals running, but to generate effective infection control, researchers must understand how SARS-CoV-2 spreads. A range of factors make studying the transmission of SARS-CoV-2 in hospitals tricky. For instance, some people do not present any symptoms, and, amongst those who do, it can be difficult to determine whether they caught the virus in the hospital or somewhere else. However, comparing the genetic information of the SARS-CoV-2 virus from different people in a hospital could allow scientists to understand how it spreads. Samples of the genetic material of SARS-CoV-2 can be obtained by swabbing infected individuals. If the genetic sequences of two samples are very different, it is unlikely that the individuals who provided the samples transmitted the virus to one another. Illingworth, Hamilton et al. used this information, along with other data about how SARS-CoV-2 is transmitted, to develop an algorithm that can determine how the virus spreads from person to person in different hospital wards. To build their algorithm, Illingworth, Hamilton et al. collected SARS-CoV-2 genetic data from patients and staff in a hospital, and combined it with information about how SARS-CoV-2 spreads and how these people moved in the hospital . The algorithm showed that, for the most part, patients were infected by other patients (20 out of 22 cases), while staff were infected equally by patients and staff. By further probing these data, Illingworth, Hamilton et al. revealed that 80% of hospital-acquired infections were caused by a group of just 21% of individuals in the study, identifying a ‘superspreader’ pattern. These findings may help to inform SARS-CoV-2 infection control measures to reduce spread within hospitals, and could potentially be used to improve infection control in other contexts.

The purpose of this consensus document was to develop feasible, evidence‐based occupational heat safety recommendations to protect the US workers that experience heat stress. Heat safety recommendations were created to protect worker health and to avoid productivity losses associated with occupational heat stress. Recommendations were tailored to be utilized by safety managers, industrial hygienists, and the employers who bear responsibility for implementing heat safety plans. An interdisciplinary roundtable comprised of 51 experts was assembled to create a narrative review summarizing current data and gaps in knowledge within eight heat safety topics: (a) heat hygiene, (b) hydration, (c) heat acclimatization, (d) environmental monitoring, (e) physiological monitoring, (f) body cooling, (g) textiles and personal protective gear, and (h) emergency action plan implementation. The consensus‐based recommendations for each topic were created using the Delphi method and evaluated based on scientific evidence, feasibility, and clarity. The current document presents 40 occupational heat safety recommendations across all eight topics. Establishing these recommendations will help organizations and employers create effective heat safety plans for their workplaces, address factors that limit the implementation of heat safety best‐practices and protect worker health and productivity., Key Points This document presents feasible, evidenced‐based occupational heat safety recommendations to protect workers from the dangers of heatA roundtable of 51 experts created 40 heat safety recommendations within eight heat safety topics: heat hygiene, hydration, heat acclimatization, environmental monitoring, physiological monitoring, body cooling, textiles and personal protective gear, and emergency action plan implementationImplementing feasible and effective heat safety plans in the workplace will protect worker health and mitigate productivity losses

Monitoring the spread of SARS-CoV-2 and reconstructing transmission chains has become a major public health focus for many governments around the world. The modest mutation rate and rapid transmission of SARS-CoV-2 prevents the reconstruction of transmission chains from consensus genome sequences, but within-host genetic diversity could theoretically help identify close contacts. Here we describe the patterns of within-host diversity in 1181 SARS-CoV-2 samples sequenced to high depth in duplicate. 95.1% of samples show within-host mutations at detectable allele frequencies. Analyses of the mutational spectra revealed strong strand asymmetries suggestive of damage or RNA editing of the plus strand, rather than replication errors, dominating the accumulation of mutations during the SARS-CoV-2 pandemic. Within- and between-host diversity show strong purifying selection, particularly against nonsense mutations. Recurrent within-host mutations, many of which coincide with known phylogenetic homoplasies, display a spectrum and patterns of purifying selection more suggestive of mutational hotspots than recombination or convergent evolution. While allele frequencies suggest that most samples result from infection by a single lineage, we identify multiple putative examples of co-infection. Integrating these results into an epidemiological inference framework, we find that while sharing of within-host variants between samples could help the reconstruction of transmission chains, mutational hotspots and rare cases of superinfection can confound these analyses., eLife digest The COVID-19 pandemic has had major health impacts across the globe. The scientific community has focused much attention on finding ways to monitor how the virus responsible for the pandemic, SARS-CoV-2, spreads. One option is to perform genetic tests, known as sequencing, on SARS-CoV-2 samples to determine the genetic code of the virus and to find any differences or mutations in the genes between the viral samples. Viruses mutate within their hosts and can develop into variants that are able to more easily transmit between hosts. Genetic sequencing can reveal how genetically similar two SARS-CoV-2 samples are. But tracking how SARS-CoV-2 moves from one person to the next through sequencing can be tricky. Even a sample of SARS-CoV-2 viruses from the same individual can display differences in their genetic material or within-host variants. Could genetic testing of within-host variants shed light on factors driving SARS-CoV-2 to evolve in humans? To get to the bottom of this, Tonkin-Hill, Martincorena et al. probed the genetics of SARS-CoV-2 within-host variants using 1,181 samples. The analyses revealed that 95.1% of samples contained within-host variants. A number of variants occurred frequently in many samples, which were consistent with mutational hotspots in the SARS-CoV-2 genome. In addition, within-host variants displayed mutation patterns that were similar to patterns found between infected individuals. The shared within-host variants between samples can help to reconstruct transmission chains. However, the observed mutational hotspots and the detection of multiple strains within an individual can make this challenging. These findings could be used to help predict how SARS-CoV-2 evolves in response to interventions such as vaccines. They also suggest that caution is needed when using information on within-host variants to determine transmission between individuals.

U2 snRNP is an essential component of the spliceosome. It is responsible for branch point recognition in the spliceosome A-complex via base-pairing of U2 snRNA with an intron to form the branch helix. Small molecule inhibitors target the SF3B component of the U2 snRNP and interfere with A-complex formation during spliceosome assembly. We previously found that the first SF3B inhibited-complex is less stable than A-complex and hypothesized that SF3B inhibitors interfere with U2 snRNA secondary structure changes required to form the branch helix. Using RNA chemical modifiers, we probed U2 snRNA structure in A-complex and SF3B inhibited splicing complexes. The reactivity pattern for U2 snRNA in the SF3B inhibited-complex is indistinguishable from that of A-complex, suggesting that they have the same secondary structure conformation, including the branch helix. This observation suggests SF3B inhibited-complex instability does not stem from an alternate RNA conformation and instead points to the inhibitors interfering with protein component interactions that normally stabilize U2 snRNP’s association with an intron. In addition, we probed U2 snRNA in the free U2 snRNP in the presence of SF3B inhibitor and again saw no differences. However, increased protection of nucleotides upstream of Stem I in the absence of SF3A and SF3B proteins suggests a change of secondary structure at the very 5′ end of U2 snRNA. Chemical probing of synthetic U2 snRNA in the absence of proteins results in similar protections and predicts a previously uncharacterized extension of Stem I. Because this stem must be disrupted for SF3A and SF3B proteins to stably join the snRNP, the structure has the potential to influence snRNP assembly and recycling after spliceosome disassembly.

Adaptation of group A Streptococcus (GAS) to its human host is mediated by two-component systems that transduce external stimuli to regulate bacterial physiology. Among such systems, CsrRS (also known as CovRS) is the most extensively characterized for its role in regulating ∼10% of the GAS genome, including several virulence genes. Here, we show that extracellular magnesium and the human antimicrobial peptide LL-37 have opposing effects on the phosphorylation of the response regulator CsrR by the receptor kinase CsrS. Genetic inactivation of CsrS phosphatase or kinase activity, respectively, had similar but more pronounced effects on CsrR phosphorylation compared to growth in magnesium or LL-37. These changes in CsrR phosphorylation were correlated with the repression or activation of CsrR-regulated genes as assessed by NanoString analysis. Chromatin immunoprecipitation and DNA sequencing (ChIP-seq) revealed CsrR occupancy at CsrRS-regulated promoters and lower-affinity associations at many other locations on the GAS chromosome. Because ChIP-seq did not detect CsrR occupancy at promoters associated with some CsrR-regulated genes, we investigated whether these genes might be controlled indirectly by intermediate regulators whose expression is modulated by CsrR. Transcriptional profiling of mutant strains deficient in the expression of either of two previously uncharacterized transcription regulators in the CsrR regulon indicated that one or both proteins participated in the regulation of 22 of the 42 CsrR-regulated promoters for which no CsrR association was detected by ChIP-seq. Taken together, these results illuminate CsrRS-mediated regulation of GAS gene expression through modulation of CsrR phosphorylation, CsrR association with regulated promoters, and the control of intermediate transcription regulators. IMPORTANCE Group A Streptococcus (GAS) is an important public health threat as a cause of sore throat, skin infections, life-threatening invasive infections, and the postinfectious complications of acute rheumatic fever, a leading cause of acquired heart disease. This work characterizes CsrRS, a GAS system for the detection of environmental signals that enables adaptation of the bacteria for survival in the human throat by regulating the production of products that allow the bacteria to resist clearance by the human immune system. CsrRS consists of two proteins: CsrS, which is on the bacterial surface to detect specific stimuli, and CsrR, which receives signals from CsrS and, in response, represses or activates the expression of genes coding for proteins that enhance bacterial survival. Some of the genes regulated by CsrR encode proteins that are themselves regulators of gene expression, thereby creating a regulatory cascade.

Apolipoprotein B-containing lipoproteins (B-lps) are essential for the transport of hydrophobic dietary and endogenous lipids through the circulation in vertebrates. Zebrafish embryos produce large numbers of B-lps in the yolk syncytial layer (YSL) to move lipids from yolk to growing tissues. Disruptions in B-lp production perturb yolk morphology, readily allowing for visual identification of mutants with altered B-lp metabolism. Here we report the discovery of a missense mutation in microsomal triglyceride transfer protein (Mtp), a protein that is essential for B-lp production. This mutation of a conserved glycine residue to valine (zebrafish G863V, human G865V) reduces B-lp production and results in yolk opacity due to aberrant accumulation of cytoplasmic lipid droplets in the YSL. However, this phenotype is milder than that of the previously reported L475P stalactite (stl) mutation. MTP transfers lipids, including triglycerides and phospholipids, to apolipoprotein B in the ER for B-lp assembly. In vitro lipid transfer assays reveal that while both MTP mutations eliminate triglyceride transfer activity, the G863V mutant protein unexpectedly retains ~80% of phospholipid transfer activity. This residual phospholipid transfer activity of the G863V mttp mutant protein is sufficient to support the secretion of small B-lps, which prevents intestinal fat malabsorption and growth defects observed in the mttpstl/stl mutant zebrafish. Modeling based on the recent crystal structure of the heterodimeric human MTP complex suggests the G865V mutation may block triglyceride entry into the lipid-binding cavity. Together, these data argue that selective inhibition of MTP triglyceride transfer activity may be a feasible therapeutic approach to treat dyslipidemia and provide structural insight for drug design. These data also highlight the power of yolk transport studies to identify proteins critical for B-lp biology., Author summary Cardiovascular disease affects about a third of the world’s population and is mediated by the buildup of lipids (primarily derived from plasma lipoproteins) in the wall of blood vessels. An essential protein required for the synthesis of the lipoproteins, from insects to humans, is microsomal triglyceride transfer protein (MTP). This protein transfers lipids, including triglycerides and phospholipids, to apolipoprotein B (APOB) for the assembly of ApoB-containing lipoproteins. Here we report a mutation in MTP that blocks the transfer of triglycerides but not phospholipids. Modeling based on the recent crystal structure of the MTP complex suggests the G865V mutation may block access to the region of the protein that binds triglyceride. In zebrafish, the residual phospholipid transfer activity of the mutant protein is sufficient to support secretion of small ApoB-containing lipoproteins and prevent a number of serious health conditions observed in humans harboring null MTP mutations (e.g., intestinal fat malabsorption, growth retardation). These results suggest that selective inhibitors of MTP that can mimic this mutation may be a feasible therapeutic approach to treat dyslipidemias in humans.

Maintaining the essential functions of mitochondria requires mechanisms to recognize and remove misfolded proteins. However, quality control (QC) pathways for misfolded mitochondrial proteins remain poorly defined. Here, we establish temperature-sensitive (ts-) peripheral mitochondrial outer membrane (MOM) proteins as novel model QC substrates in Saccharomyces cerevisiae. The ts- proteins sen2-1HAts and sam35-2HAts are degraded from the MOM by the ubiquitin-proteasome system. Ubiquitination of sen2-1HAts is mediated by the ubiquitin ligase (E3) Ubr1, while sam35-2HAts is ubiquitinated primarily by San1. Mitochondria-associated degradation (MAD) of both substrates requires the SSA family of Hsp70s and the Hsp40 Sis1, providing the first evidence for chaperone involvement in MAD. In addition to a role for the Cdc48-Npl4-Ufd1 AAA-ATPase complex, Doa1 and a mitochondrial pool of the transmembrane Cdc48 adaptor, Ubx2, are implicated in their degradation. This study reveals a unique QC pathway comprised of a combination of cytosolic and mitochondrial factors that distinguish it from other cellular QC pathways., eLife digest Proteins are molecules that need to fold into the right shape to do their job. If proteins lose that shape, not only do they stop working but they risk clumping together and becoming toxic, potentially leading to disease. Fortunately, the cell has quality control systems that normally detect and remove misfolded proteins before they can cause damage to the cell. First, sets of proteins known as chaperones recognize the misfolded proteins, and then another class of proteins attaches a molecular tag, known as ubiquitin, to the misshapen proteins. When several ubiquitin tags are attached to a protein, forming chains of ubiquitin, it is transported to a large molecular machine within the cell called the proteasome. The proteasome unravels the protein and breaks it down into its constituent building blocks, which can then be used to create new proteins. Proteins are found throughout the different compartments of the cell and quality control processes have been well-studied in some parts of the cell but not others. Metzger et al. have now revealed how the process works on the surface of mitochondria, the compartment that provides the cell with most of its energy. To do this, they used baker’s yeast, a model laboratory organism that shares many fundamental properties with animal cells, but which is easier to manipulate genetically. The quality control process was studied using two mitochondrial proteins that had been mutated to make them sensitive to changes in temperature. This meant that, when the temperature increased from 25°C to 37°C, these proteins would begin to unravel and trigger the clean-up operation. This approach has been used previously to understand the quality control processes in other parts of the cell. By removing different quality control machinery in turn from the yeast cells, Metzger et al. could detect which were necessary for the process on mitochondria. This showed that there were many similarities with how this process happen in other parts of the cell but that the precise combination of chaperones and enzymes involved was distinct. Furthermore, when the proteasome was not working, the misfolded proteins remained on the mitochondria, showing that they are not transported to other parts of the cell to be broken down. In the future, understanding this process could help to find potential drug targets for mitochondrial diseases. The next steps will be to see how well these findings apply to human and other mammalian cells.

Antibiotic-resistant Staphylococcus aureus remains a leading cause of antibiotic resistance-associated mortality in the United States. Given the reality of multi-drug resistant infections, it is imperative that we establish and maintain a pipeline of new compounds to replace or supplement our current antibiotics. A first step towards this goal is to prioritize targets by identifying the genes most consistently required for survival across the S. aureus phylogeny. Here we report the first direct comparison of multiple strains of S. aureus via transposon sequencing. We show that mutant fitness varies by strain in key pathways, underscoring the importance of using more than one strain to differentiate between core and strain-dependent essential genes. We treated the libraries with daptomycin to assess whether the strain-dependent differences impact pathways important for survival. Despite baseline differences in gene importance, several pathways, including the lipoteichoic acid pathway, consistently promote survival under daptomycin exposure, suggesting core vulnerabilities that can be exploited to resensitize daptomycin-nonsusceptible isolates. We also demonstrate the merit of using transposons with outward-facing promoters capable of overexpressing nearby genes for identifying clinically-relevant gain-of-function resistance mechanisms. Together, the daptomycin vulnerabilities and resistance mechanisms support a mode of action with wide-ranging effects on the cell envelope and cell division. This work adds to a growing body of literature demonstrating the nuanced insights gained by comparing Tn-Seq results across multiple bacterial strains., Author summary Antibiotic-resistant Staphylococcus aureus kills thousands of people every year in the United States alone. To stay ahead of the looming threat of multidrug-resistant infections, we must continue to develop new antibiotics and find ways to make our current repertoire of antibiotics more effective, including by finding pairs of compounds that perform best when administered together. In the age of next-generation sequencing, we can now use transposon sequencing to find potential targets for new antibiotics on a genome-wide scale, identified as either essential genes or genes that positively influence survival in the presence of an antibiotic. In this work, we created a compendium of genes that are essential across a range of S. aureus strains, as well as those that are important for growth in the presence of the antibiotic daptomycin. The results will be a resource for researchers working to develop the next generation of antibiotic therapies.

Mammalian circadian (24 h) rhythms are timed by the pattern of spontaneous action potential firing in the suprachiasmatic nucleus (SCN). This oscillation in firing is produced through circadian regulation of several membrane currents, including large-conductance Ca(2+)- and voltage-activated K(+) (BK) and L-type Ca(2+) channel (LTCC) currents. During the day steady-state BK currents depend mostly on LTCCs for activation, whereas at night they depend predominantly on ryanodine receptors (RyRs). However, the contribution of these Ca(2+) channels to BK channel activation during action potential firing has not been thoroughly investigated. In this study, we used a pharmacological approach to determine that both LTCCs and RyRs contribute to the baseline membrane potential of SCN action potential waveforms, as well as action potential-evoked BK current, during the day and night, respectively. Since the baseline membrane potential is a major determinant of circadian firing rate, we focused on the LTCCs contributing to low voltage activation of BK channels during the subthreshold phase. For these experiments, two LTCC subtypes found in SCN (Ca(V)1.2 and Ca(V)1.3) were coexpressed with BK channels in heterologous cells, where their differential contributions could be separately measured. Ca(V)1.3 channels produced currents that were shifted to more hyperpolarized potentials compared with Ca(V)1.2, resulting in increased subthreshold Ca(2+) and BK currents during an action potential command. These results show that although multiple Ca(2+) sources in SCN can contribute to the activation of BK current during an action potential, specific BK-Ca(V)1.3 partnerships may optimize the subthreshold BK current activation that is critical for firing rate regulation. NEW & NOTEWORTHY BK K(+) channels are important regulators of firing. Although Ca(2+) channels are required for their activation in excitable cells, it is not well understood how BK channels activate using these Ca(2+) sources during an action potential. This study demonstrates the differences in BK current activated by Ca(V)1.2 and Ca(V)1.3 channels in clock neurons and heterologous cells. The results define how specific ion channel partnerships can be engaged during distinct phases of the action potential.