SARS-CoV-2 reservoir in post-acute sequelae of COVID-19 (PASC).

Millions of people are suffering from Long COVID or post-acute sequelae of COVID-19 (PASC). Several biological factors have emerged as potential drivers of PASC pathology. Some individuals with PASC may not fully clear the coronavirus SARS-CoV-2 after acute infection. Instead, replicating virus and/or viral RNA-potentially capable of being translated to produce viral proteins-persist in tissue as a 'reservoir'. This reservoir could modulate host immune responses or release viral proteins into the circulation. Here we review studies that have identified SARS-CoV-2 RNA/protein or immune responses indicative of a SARS-CoV-2 reservoir in PASC samples. Mechanisms by which a SARS-CoV-2 reservoir may contribute to PASC pathology, including coagulation, microbiome and neuroimmune abnormalities, are delineated. We identify research priorities to guide the further study of a SARS-CoV-2 reservoir in PASC, with the goal that clinical trials of antivirals or other therapeutics with potential to clear a SARS-CoV-2 reservoir are accelerated.

Universal Patterns of Selection in Cancer and Somatic Tissues.

Cancer develops as a result of somatic mutation and clonal selection, but quantitative measures of selection in cancer evolution are lacking. We adapted methods from molecular evolution and applied them to 7,664 tumors across 29 cancer types. Unlike species evolution, positive selection outweighs negative selection during cancer development. On average, <1 coding base substitution/tumor is lost through negative selection, with purifying selection almost absent outside homozygous loss of essential genes. This allows exome-wide enumeration of all driver coding mutations, including outside known cancer genes. On average, tumors carry ∼4 coding substitutions under positive selection, ranging from <1/tumor in thyroid and testicular cancers to >10/tumor in endometrial and colorectal cancers. Half of driver substitutions occur in yet-to-be-discovered cancer genes. With increasing mutation burden, numbers of driver mutations increase, but not linearly. We systematically catalog cancer genes and show that genes vary extensively in what proportion of mutations are drivers versus passengers.

The life history of 21 breast cancers.

Cancer evolves dynamically as clonal expansions supersede one another driven by shifting selective pressures, mutational processes, and disrupted cancer genes. These processes mark the genome, such that a cancer's life history is encrypted in the somatic mutations present. We developed algorithms to decipher this narrative and applied them to 21 breast cancers. Mutational processes evolve across a cancer's lifespan, with many emerging late but contributing extensive genetic variation. Subclonal diversification is prominent, and most mutations are found in just a fraction of tumor cells. Every tumor has a dominant subclonal lineage, representing more than 50% of tumor cells. Minimal expansion of these subclones occurs until many hundreds to thousands of mutations have accumulated, implying the existence of long-lived, quiescent cell lineages capable of substantial proliferation upon acquisition of enabling genomic changes. Expansion of the dominant subclone to an appreciable mass may therefore represent the final rate-limiting step in a breast cancer's development, triggering diagnosis.

Mutational processes molding the genomes of 21 breast cancers.

All cancers carry somatic mutations. The patterns of mutation in cancer genomes reflect the DNA damage and repair processes to which cancer cells and their precursors have been exposed. To explore these mechanisms further, we generated catalogs of somatic mutation from 21 breast cancers and applied mathematical methods to extract mutational signatures of the underlying processes. Multiple distinct single- and double-nucleotide substitution signatures were discernible. Cancers with BRCA1 or BRCA2 mutations exhibited a characteristic combination of substitution mutation signatures and a distinctive profile of deletions. Complex relationships between somatic mutation prevalence and transcription were detected. A remarkable phenomenon of localized hypermutation, termed "kataegis," was observed. Regions of kataegis differed between cancers but usually colocalized with somatic rearrangements. Base substitutions in these regions were almost exclusively of cytosine at TpC dinucleotides. The mechanisms underlying most of these mutational signatures are unknown. However, a role for the APOBEC family of cytidine deaminases is proposed.

Author Correction: Landscape of somatic mutations in 560 breast cancer whole-genome sequences.

In the Methods section of this Article, 'greater than' should have been 'less than' in the sentence 'Putative regions of clustered rearrangements were identified as having an average inter-rearrangement distance that was at least 10 times greater than the whole-genome average for the individual sample. '. The Article has not been corrected.

Prevalence of orthostatic intolerance in long covid clinic patients and healthy volunteers: A multicenter study.

Orthostatic intolerance (OI), including postural orthostatic tachycardia syndrome (PoTS) and orthostatic hypotension (OH), are often reported in long covid, but published studies are small with inconsistent results. We sought to estimate the prevalence of objective OI in patients attending long covid clinics and healthy volunteers and associations with OI symptoms and comorbidities. Participants with a diagnosis of long covid were recruited from eight UK long covid clinics, and healthy volunteers from general population. All undertook standardized National Aeronautics and Space Administration Lean Test (NLT). Participants' history of typical OI symptoms (e.g., dizziness, palpitations) before and during the NLT were recorded. Two hundred seventy-seven long covid patients and 50 frequency-matched healthy volunteers were tested. Healthy volunteers had no history of OI symptoms or symptoms during NLT or PoTS, 10% had asymptomatic OH. One hundred thirty (47%) long covid patients had previous history of OI symptoms and 144 (52%) developed symptoms during the NLT. Forty-one (15%) had an abnormal NLT, 20 (7%) met criteria for PoTS, and 21 (8%) had OH. Of patients with an abnormal NLT, 45% had no prior symptoms of OI. Relaxing the diagnostic thresholds for PoTS from two consecutive abnormal readings to one abnormal reading during the NLT, resulted in 11% of long covid participants (an additional 4%) meeting criteria for PoTS, but not in healthy volunteers. More than half of long covid patients experienced OI symptoms during NLT and more than one in 10 patients met the criteria for either PoTS or OH, half of whom did not report previous typical OI symptoms. We therefore recommend all patients attending long covid clinics are offered an NLT and appropriate management commenced.

Non-BRCA1/BRCA2 high-risk familial breast cancers are not associated with a high prevalence of BRCAness.

BACKGROUND: Familial breast cancer is in most cases unexplained due to the lack of identifiable pathogenic variants in the BRCA1 and BRCA2 genes. The somatic mutational landscape and in particular the extent of BRCA-like tumour features (BRCAness) in these familial breast cancers where germline BRCA1 or BRCA2 mutations have not been identified is to a large extent unknown. METHODS: We performed whole-genome sequencing on matched tumour and normal samples from high-risk non-BRCA1/BRCA2 breast cancer families to understand the germline and somatic mutational landscape and mutational signatures. We measured BRCAness using HRDetect. As a comparator, we also analysed samples from BRCA1 and BRCA2 germline mutation carriers. RESULTS: We noted for non-BRCA1/BRCA2 tumours, only a small proportion displayed high HRDetect scores and were characterized by concomitant promoter hypermethylation or in one case a RAD51D splice variant previously reported as having unknown significance to potentially explain their BRCAness. Another small proportion showed no features of BRCAness but had mutationally active tumours. The remaining tumours lacked features of BRCAness and were mutationally quiescent. CONCLUSIONS: A limited fraction of high-risk familial non-BRCA1/BRCA2 breast cancer patients is expected to benefit from treatment strategies against homologue repair deficient cancer cells.

Somatic mutations reveal asymmetric cellular dynamics in the early human embryo.

Somatic cells acquire mutations throughout the course of an individual's life. Mutations occurring early in embryogenesis are often present in a substantial proportion of, but not all, cells in postnatal humans and thus have particular characteristics and effects. Depending on their location in the genome and the proportion of cells they are present in, these mosaic mutations can cause a wide range of genetic disease syndromes and predispose carriers to cancer. They have a high chance of being transmitted to offspring as de novo germline mutations and, in principle, can provide insights into early human embryonic cell lineages and their contributions to adult tissues. Although it is known that gross chromosomal abnormalities are remarkably common in early human embryos, our understanding of early embryonic somatic mutations is very limited. Here we use whole-genome sequences of normal blood from 241 adults to identify 163 early embryonic mutations. We estimate that approximately three base substitution mutations occur per cell per cell-doubling event in early human embryogenesis and these are mainly attributable to two known mutational signatures. We used the mutations to reconstruct developmental lineages of adult cells and demonstrate that the two daughter cells of many early embryonic cell-doubling events contribute asymmetrically to adult blood at an approximately 2:1 ratio. This study therefore provides insights into the mutation rates, mutational processes and developmental outcomes of cell dynamics that operate during early human embryogenesis.

Indwelling Pleural Catheter: Management of Complications.

Multiple randomized clinical trials have established the advantages of indwelling pleural catheter (IPC) in the management of malignant pleural effusions, resulting in its widespread adoption in clinical practice. Complications can occur with IPC use and must be recognized and managed effectively. This review provides a comprehensive overview of IPC complications and their best care. Pain postinsertion or during drainage of IPC is easily manageable and must be distinguished from tumor-related chest wall pain. IPC-related infections require systemic antibiotics and often intrapleural fibrinolytic/deoxyribonuclease therapy. The removal of IPC for infection is usually unnecessary. Symptomatic loculation usually responds to fibrinolytics but may recur. Catheter tract metastases are common in mesothelioma patients and usually respond to radiotherapy without inducing damages to the IPC. Less common complications include dislodgement, irreversible blockage, and fractures (upon removal) of the catheter. Recommendations on the management of IPC complications by recent consensus statement/guideline are discussed. Expert opinions on management approaches are included in areas where evidence is lacking to guide care.

Reducing sickness absence and stigma due to mental health difficulties: a randomised control treatment trial (RCT) of a low intensity psychological intervention and stigma reduction programme for common mental disorder (Prevail).

BACKGROUND: Common mental disorders are the leading cause of workplace absences. The Prevail intervention programme aims to reduce stigma and to educate staff and managers about evidence-based low intensity psychological interventions for common mental disorders (depression, anxiety, stress, and distress). Prevail is innovative in taking a public health approach. It is designed to be given to all employees irrespective of their past or current mental health. Prevail was evaluated in three studies examining: (1) the acceptability of the intervention and perceived usefulness; (2) whether the intervention altered stigmatic attitudes and motivation to seek help; and (3) whether the intervention reduced sickness absence, both overall and due to mental health problems. METHODS: A two-armed cluster randomised control trial (RCT) evaluated the effectiveness of Prevail. Employees (N = 1051) at a large UK government institution were randomised to an active intervention or control arm in teams identified by their managers (n = 67). Employees in the active arm received the Prevail Staff Intervention. The managers in the active arm also received the Prevail Managers Intervention. Participants' satisfaction and analysis of the Prevail Intervention were gathered by a bespoke questionnaire. Questionnaire measures of attitudes to mental health and mental health stigma were taken 1-2 weeks prior to the intervention and approximately 4 weeks post-intervention. Data relating to sickness absence were gathered via the official records in the time period 3-month post-intervention and for the same period 12 months earlier. RESULTS: Prevail was evaluated highly favourably by both the staff and their managers. Prevail produced significant reductions in self-stigma and anticipated stigma due to mental health difficulties. Crucially, sickness absence was significantly reduced by the Prevail Intervention. DISCUSSION: Prevail achieved its goals of producing a palatable and engaging intervention that altered staff's attitudes and stigmatic beliefs related to mental health and, crucially, produced a strong reduction in work-pace absenteeism. As the Prevail programme is aimed at common mental health problems and was not specialised to this particular workforce, the study provides the evidence-base for a mental health intervention programme that could be used by many organisations across the world. TRIAL REGISTRATION: ISRCTN12040087. Registered 04/05/2020. https://doi.org/10.1186/ISRCTN12040087 . A full protocol for the randomised control trial was published: Gray NS, Davies H, Snowden RJ: Reducing stigma and increasing workplace productivity due to mental health difficulties in a large government organization in the UK: a protocol for a randomised control treatment trial (RCT) of a low intensity psychological intervention and stigma reduction programme for common mental disorder (Prevail). BMC Public Health 2020, 20(1):1-9.

The circular RNome of primary breast cancer.

Circular RNAs (circRNAs) are a class of RNAs that is under increasing scrutiny, although their functional roles are debated. We analyzed RNA-seq data of 348 primary breast cancers and developed a method to identify circRNAs that does not rely on unmapped reads or known splice junctions. We identified 95,843 circRNAs, of which 20,441 were found recurrently. Of the circRNAs that match exon boundaries of the same gene, 668 showed a poor or even negative (R < 0.2) correlation with the expression level of the linear gene. In silico analysis showed only a minority (8.5%) of circRNAs could be explained by known splicing events. Both these observations suggest that specific regulatory processes for circRNAs exist. We confirmed the presence of circRNAs of CNOT2, CREBBP, and RERE in an independent pool of primary breast cancers. We identified circRNA profiles associated with subgroups of breast cancers and with biological and clinical features, such as amount of tumor lymphocytic infiltrate and proliferation index. siRNA-mediated knockdown of circCNOT2 was shown to significantly reduce viability of the breast cancer cell lines MCF-7 and BT-474, further underlining the biological relevance of circRNAs. Furthermore, we found that circular, and not linear, CNOT2 levels are predictive for progression-free survival time to aromatase inhibitor (AI) therapy in advanced breast cancer patients, and found that circCNOT2 is detectable in cell-free RNA from plasma. We showed that circRNAs are abundantly present, show characteristics of being specifically regulated, are associated with clinical and biological properties, and thus are relevant in breast cancer.

Landscape of somatic mutations in 560 breast cancer whole-genome sequences.

We analysed whole-genome sequences of 560 breast cancers to advance understanding of the driver mutations conferring clonal advantage and the mutational processes generating somatic mutations. We found that 93 protein-coding cancer genes carried probable driver mutations. Some non-coding regions exhibited high mutation frequencies, but most have distinctive structural features probably causing elevated mutation rates and do not contain driver mutations. Mutational signature analysis was extended to genome rearrangements and revealed twelve base substitution and six rearrangement signatures. Three rearrangement signatures, characterized by tandem duplications or deletions, appear associated with defective homologous-recombination-based DNA repair: one with deficient BRCA1 function, another with deficient BRCA1 or BRCA2 function, the cause of the third is unknown. This analysis of all classes of somatic mutation across exons, introns and intergenic regions highlights the repertoire of cancer genes and mutational processes operating, and progresses towards a comprehensive account of the somatic genetic basis of breast cancer.

How Artificial Intelligence and Machine Learning Is Assisting Us to Extract Meaning from Data on Bone Mechanics?

Dramatic advancements in interdisciplinary research with the fourth paradigm of science, especially the implementation of computer science, nourish the potential for artificial intelligence (AI), machine learning (ML), and artificial neural network (ANN) algorithms to be applied to studies concerning mechanics of bones. Despite recent enormous advancement in techniques, gaining deep knowledge to find correlations between bone shape, material, mechanical, and physical responses as well as properties is a daunting task. This is due to both complexity of the material itself and the convoluted shapes that this complex material forms. Moreover, many uncertainties and ambiguities exist concerning the use of traditional computational techniques that hinders gaining a full comprehension of this advanced biological material. This book chapter offers a review of literature on the use of AI, ML, and ANN in the study of bone mechanics research. A main question as to why to implement AI and ML in the mechanics of bones is fully addressed and explained. This chapter also introduces AI and ML and elaborates on the main features of ML algorithms such as learning paradigms, subtypes, main ideas with examples, performance metrics, training algorithms, and training datasets. As a frequently employed ML algorithm in bone mechanics, feedforward ANNs are discussed to make their taxonomy and working principles more readily comprehensible to researchers. A summary as well as detailed review of papers that employed ANNs to learn from collected data on bone mechanics are presented. Reviewing literature on the use of these data-driven tools is essential since their wider application has the potential to: improve clinical assessments enabling real-time simulations; avoid and/or minimize injuries; and, encourage early detection of such injuries in the first place.

Early implant placement in ridge preserved extraction sockets: A pre-clinical in vivo study.

OBJECTIVES: The aim was to analyse the outcomes of early implant placement after 6 and 12 weeks of healing in ridge preserved sites in a canine model. MATERIALS AND METHODS: Implants were placed in second maxillary incisors sites in 9 dogs 6 weeks after grafting of the sockets with 90% deproteinized bovine bone mineral in 10% collagen matrix (DBBMC) and closure with resorbable type I/III porcine collagen matrix (PCM). The implants were randomly assigned to 6 (T6) and 12 (T12) weeks of healing. RESULTS: The percentage of bone-to-implant contact (%BIC), old bone, new bone and residual DBBMC was similar between T6 and T12. In relation to the implant shoulder (IS), the original bone crest (IS-ROB) was more apical on the buccal than the palatal side. The regenerated bone crest (IS-C) and IS-ROB were similar between groups. However, the distance from IS to first bone-to implant contact (IS-fBIC) was significantly less in T12 compared with T6 (p = .022; Wilcoxon signed-rank test). The bucco-palatal ridge dimensions between T6 and T12 were similar. CONCLUSIONS: This study confirms that implants can successfully be placed early in ridge preserved maxillary second incisor sites and are osseointegrated by 6 weeks. There were significantly lower IS-fBIC values at 12 weeks than at 6 weeks on the buccal aspect. The original buccal bone crest underwent greater corono-apical resorption than the palatal crest. The %BIC, relative proportions of mineralized tissues and dimensions of the alveolar ridge demonstrated stability between 6 and 12 weeks of healing.

COVID-19 and pneumothorax: a multicentre retrospective case series.

INTRODUCTION: Pneumothorax and pneumomediastinum have both been noted to complicate cases of coronavirus disease 2019 (COVID-19) requiring hospital admission. We report the largest case series yet described of patients with both these pathologies (including nonventilated patients). METHODS: Cases were collected retrospectively from UK hospitals with inclusion criteria limited to a diagnosis of COVID-19 and the presence of either pneumothorax or pneumomediastinum. Patients included in the study presented between March and June 2020. Details obtained from the medical record included demographics, radiology, laboratory investigations, clinical management and survival. RESULTS: 71 patients from 16 centres were included in the study, of whom 60 had pneumothoraces (six with pneumomediastinum in addition) and 11 had pneumomediastinum alone. Two of these patients had two distinct episodes of pneumothorax, occurring bilaterally in sequential fashion, bringing the total number of pneumothoraces included to 62. Clinical scenarios included patients who had presented to hospital with pneumothorax, patients who had developed pneumothorax or pneumomediastinum during their inpatient admission with COVID-19 and patients who developed their complication while intubated and ventilated, either with or without concurrent extracorporeal membrane oxygenation. Survival at 28 days was not significantly different following pneumothorax (63.1±6.5%) or isolated pneumomediastinum (53.0±18.7%; p=0.854). The incidence of pneumothorax was higher in males. 28-day survival was not different between the sexes (males 62.5±7.7% versus females 68.4±10.7%; p=0.619). Patients aged ≥70 years had a significantly lower 28-day survival than younger individuals (≥70 years 41.7±13.5% survival versus <70 years 70.9±6.8% survival; p=0.018 log-rank). CONCLUSION: These cases suggest that pneumothorax is a complication of COVID-19. Pneumothorax does not seem to be an independent marker of poor prognosis and we encourage continuation of active treatment where clinically possible.

Prospective validation of the RAPID clinical risk prediction score in adult patients with pleural infection: the PILOT study.

BACKGROUND: Over 30% of adult patients with pleural infection either die and/or require surgery. There is no robust means of predicting at baseline presentation which patients will suffer a poor clinical outcome. A validated risk prediction score would allow early identification of high-risk patients, potentially directing more aggressive treatment thereafter. OBJECTIVES: To prospectively assess a previously described risk score (the RAPID (Renal (urea), Age, fluid Purulence, Infection source, Dietary (albumin)) score) in adults with pleural infection. METHODS: Prospective observational cohort study that recruited patients undergoing treatment for pleural infection. RAPID score and risk category were calculated at baseline presentation. The primary outcome was mortality at 3 months; secondary outcomes were mortality at 12 months, length of hospital stay, need for thoracic surgery, failure of medical treatment and lung function at 3 months. RESULTS: Mortality data were available in 542 out of 546 patients recruited (99.3%). Overall mortality was 10% at 3 months (54 out of 542) and 19% at 12 months (102 out of 542). The RAPID risk category predicted mortality at 3 months. Low-risk mortality (RAPID score 0-2): five out of 222 (2.3%, 95% CI 0.9 to 5.7%); medium-risk mortality (RAPID score 3-4): 21 out of 228 (9.2%, 95% CI 6.0 to 13.7%); and high-risk mortality (RAPID score 5-7): 27 out of 92 (29.3%, 95% CI 21.0 to 39.2%). C-statistics for the scores at 3 months and 12 months were 0.78 (95% CI 0.71-0.83) and 0.77 (95% CI 0.72-0.82), respectively. CONCLUSIONS: The RAPID score stratifies adults with pleural infection according to increasing risk of mortality and should inform future research directed at improving outcomes in this patient population.

Primary stability and healing outcomes of apically tapered and straight implants placed into fresh extraction sockets. A pre-clinical in vivo study.

OBJECTIVES: To compare the stability of apically tapered and straight (non-tapered cylindrical) implants at the time of immediate placement and to histologically evaluate the healing outcomes after 6 weeks. MATERIALS AND METHODS: The second maxillary incisors were extracted bilaterally in nine dogs. After randomization, apically tapered and straight implants with a 3.3 mm shoulder diameter were inserted into the extraction sockets. The implant stability quotient (ISQ) of the implants was recorded after placement. Peri-implant defects on the buccal aspect were filled with deproteinized bovine bone mineral and covered with resorbable type I/III porcine collagen matrix. After 6 weeks of healing, sections were prepared for histological and morphometric analysis. RESULTS: All implant sites healed uneventfully. The apically tapered implants had significantly higher ISQ values compared to straight implants at placement (p = .009). The histomorphometric outcomes 6 weeks following implant placement in both experimental groups were similar, except in the apico-palatal region. Apically tapered implants demonstrated significantly less percentage bone-to-implant contact (p = .035) in the apico-palatal region. At both implant types, substantial corono-apical resorption of the buccal bone wall was noted in the coronal 2 mm of the implant. CONCLUSION: Apically tapered implants had significantly higher ISQ values at immediate placement compared to straight implants. The healing outcomes and remodelling of the buccal bone wall were similar for both implant designs. In the apico-palatal region, there was less %BIC at the implant surface at apically tapered implants compared to straight implants.

Reducing stigma and increasing workplace productivity due to mental health difficulties in a large government organization in the UK: a protocol for a randomised control treatment trial (RCT) of a low intensity psychological intervention and stigma reduction programme for common mental disorder (Prevail).

BACKGROUND: Common mental disorders are the leading cause of workplace absences. While the reasons for this are multifarious, there is little doubt that stigma related to common mental disorder plays a large role in sickness absence and in poor help-seeking. Frequently both managers and staff are unsure of how to approach and intervene with mental health related problems. We have therefore devised a mental health intervention programme (Prevail) that aims to reduce stigma and to educate staff about evidence-based low intensity psychological interventions. These can be used by the individual, as well as in collaboration with managers via co-production of problem-focussed solutions, with the aim of improving mental health, reducing sickness absence, and increasing workplace productivity. METHODS: This two-armed cluster randomised control trial (RCT) will evaluate the effectiveness of Prevail. Eighty managers at a large UK government institution (the DVLA) and their teams (approximately 960 employees) will be randomised into the active intervention group or control (employment as usual) arms of the study. All participants will be invited to complete a series of questionnaires related to mental health stigma, their current and past mental health, and their recent workplace productivity (absenteeism and presenteeism). All employees in the active arm will receive the Prevail Staff intervention, which covers stigma reduction and includes psychoeducation about evidence-based low intensity psychological interventions for common mental disorder. The managers in the active arm will also receive the Prevail Managers programme which covers communication skills, problem formulation, and problem-solving skills. The questionnaire battery will then be given to both groups again 4 weeks post training, and 12 months post-training. Official records of absenteeism from Human Resources will also be gathered from both active and control groups at 12 months post-training. DISCUSSION: The treatment trial aims to evaluate if Prevail reduces mental health related stigma (of a number of forms), increases help-seeking behaviours, and increases workplace productivity (via decreased absenteeism and presenteeism). TRIAL REGISTRATION: ISRCTN12040087. Retrospectively registered 04/05/2020.