A sensitive and affordable multiplex RT-qPCR assay for SARS-CoV-2 detection.

With the ongoing COVID-19 (Coronavirus Disease 2019) pandemic, caused by the novel coronavirus SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2), there is a need for sensitive, specific, and affordable diagnostic tests to identify infected individuals, not all of whom are symptomatic. The most sensitive test involves the detection of viral RNA using RT-qPCR (quantitative reverse transcription PCR), with many commercial kits now available for this purpose. However, these are expensive, and supply of such kits in sufficient numbers cannot always be guaranteed. We therefore developed a multiplex assay using well-established SARS-CoV-2 targets alongside a human cellular control (RPP30) and a viral spike-in control (Phocine Herpes Virus 1 [PhHV-1]), which monitor sample quality and nucleic acid extraction efficiency, respectively. Here, we establish that this test performs as well as widely used commercial assays, but at substantially reduced cost. Furthermore, we demonstrate >1,000-fold variability in material routinely collected by combined nose and throat swabbing and establish a statistically significant correlation between the detected level of human and SARS-CoV-2 nucleic acids. The inclusion of the human control probe in our assay therefore provides a quantitative measure of sample quality that could help reduce false-negative rates. We demonstrate the feasibility of establishing a robust RT-qPCR assay at approximately 10% of the cost of equivalent commercial assays, which could benefit low-resource environments and make high-volume testing affordable.

The PHF21A neurodevelopmental disorder: an evaluation of clinical data from 13 patients.

Potocki-Shaffer syndrome (PSS) is a rare neurodevelopmental disorder caused by deletions involving the 11p11.2-p12 region, encompassing the plant homeodomain finger protein 21A (PHF21A) gene. PHF21A has an important role in epigenetic regulation and PHF21A variants have previously been associated with a specific disorder that, whilst sharing some features of PSS, has notable differences. This study aims to expand the phenotype, particularly in relation to overgrowth, associated with PHF21A variants. Analysis of phenotypic data was undertaken on 13 individuals with PHF21A constitutional variants including four individuals described in the current series. Of those individuals where data were recorded, postnatal overgrowth was reported in 5/6 (83%). In addition, all had both an intellectual disability and behavioural issues. Frequent associations included postnatal hypotonia (7/11, 64%); and at least one afebrile seizure episode (6/12, 50%). Although a recognizable facial gestalt was not associated, subtle dysmorphic features were shared amongst some individuals and included a tall broad forehead, broad nasal tip, anteverted nares and full cheeks. We provide further insight into the emerging neurodevelopmental syndrome associated with PHF21A disruption. We present some evidence that PHF21A might be considered a new member of the overgrowth-intellectual disability syndrome (OGID) family.

Microcephalic osteodysplastic primordial dwarfism type II is associated with global vascular disease.

BACKGROUND: Microcephalic osteodysplastic primordial dwarfism type II (MOPDII) is the most common form of primordial dwarfism, caused by bialleic mutations in the pericentrin gene (PCNT). Aside from its classic features, there are multiple associated medical complications, including a well-documented risk of neurovascular disease. Over the past several years, it has become apparent that additional vascular issues, as well as systemic hypertension and kidney disease may also be related to MOPDII. However, the frequency and extent of the vasculopathy was unclear. To help address this question, a vascular substudy was initiated within our Primordial Dwarfism Registry. RESULTS: Medical records from 47 individuals, living and deceased, ranging in age from 3 to 41 years of age were interrogated for this purpose. Of the total group, 64% were diagnosed with moyamoya, intracranial aneurysms, or both. In general, the age at diagnosis for moyamoya was younger than aneurysms, but the risk for neurovascular disease was throughout the shortened lifespan. In addition to neurovascular disease, renal, coronary and external carotid artery involvement are documented. 43% of the total group was diagnosed with hypertension, and 17% had myocardial infarctions. A total of 32% of the entire cohort had some form of chronic kidney disease, with 4% of the total group necessitating a kidney transplant. In addition, 38% had diabetes/insulin resistance. Ages of diagnoses, treatment modalities employed, and location of vasculopathies were notated as available and applicable, as well as frequencies of other comorbidities. CONCLUSIONS: It is now clear that vascular disease in MOPDII is global and screening of the cardiac and renal vessels is warranted along with close monitoring of blood pressure. We recommend a blood pressure of 110/70 mmHg as a starting point for an upper limit, especially if the individual has a history of neurovascular disease, chronic kidney disease and/or diabetes. Additionally, providers need to be at high alert for the possibility of myocardial infarctions in young adults with MOPDII, so that appropriate treatment can be initiated promptly in an acute situation.

Soluble Non-Starch Polysaccharides From Plantain (Musa x paradisiaca L.) Diminish Epithelial Impact of Clostridioides difficile.

Clostridioides difficile infection (CDI) is a leading cause of antibiotic-associated diarrhoea. Adhesion of this Gram-positive pathogen to the intestinal epithelium is a crucial step in CDI, with recurrence and relapse of disease dependent on epithelial interaction of its endospores. Close proximity, or adhesion of, hypervirulent strains to the intestinal mucosa are also likely to be necessary for the release of C. difficile toxins, which when internalized, result in intestinal epithelial cell rounding, damage, inflammation, loss of barrier function and diarrhoea. Interrupting these C. difficile-epithelium interactions could therefore represent a promising therapeutic strategy to prevent and treat CDI. Intake of dietary fibre is widely recognised as being beneficial for intestinal health, and we have previously shown that soluble non-starch polysaccharides (NSP) from plantain banana (Musa spp.), can block epithelial adhesion and invasion of a number of gut pathogens, such as E. coli and Salmonellae. Here, we assessed the action of plantain NSP, and a range of alternative soluble plant fibres, for inhibitory action on epithelial interactions of C. difficile clinical isolates, purified endospore preparations and toxins. We found that plantain NSP possessed ability to disrupt epithelial adhesion of C. difficile vegetative cells and spores, with inhibitory activity against C. difficile found within the acidic (pectin-rich) polysaccharide component, through interaction with the intestinal epithelium. Similar activity was found with NSP purified from broccoli and leek, although seen to be less potent than NSP from plantain. Whilst plantain NSP could not block the interaction and intracellular action of purified C. difficile toxins, it significantly diminished the epithelial impact of C. difficile, reducing both bacteria and toxin induced inflammation, activation of caspase 3/7 and cytotoxicity in human intestinal cell-line and murine intestinal organoid cultures. Dietary supplementation with soluble NSP from plantain may therefore confer a protective effect in CDI patients by preventing adhesion of C. difficile to the mucosa, i.e. a "contrabiotic" effect, and diminishing its epithelial impact. This suggests that plantain soluble dietary fibre may be a therapeutically effective nutritional product for use in the prevention or treatment of CDI and antibiotic-associated diarrhoea.

NF-κB2 signalling in enteroids modulates enterocyte responses to secreted factors from bone marrow-derived dendritic cells.

Alternative pathway NF-κB signalling regulates susceptibility towards developing inflammatory bowel disease (IBD), colitis-associated cancer and sepsis-associated intestinal epithelial cell apoptosis and shedding. However, the cell populations responsible for the perturbed alternative pathway NF-κB signalling in intestinal mucosal pathology remain unclear. In order to investigate the contribution of the epithelial compartment, we have tested whether NF-κB2 regulated transcription in intestinal epithelial cells controls the intestinal epithelial response to cytokines that are known to disrupt intestinal barrier permeability. Enteroids were generated from the proximal, middle and distal regions of small intestine (SI) from C57BL/6J wild-type mice and displayed region-specific morphology that was maintained during sub-culture. Enteroids treated with 100 ng/mL TNF were compared with corresponding regions of SI from C57BL/6J mice treated systemically with 0.33 mg/kg TNF for 1.5 h. TNF-induced apoptosis in all regions of the intestine in vitro and in vivo but resulted in Paneth cell degranulation only in proximal tissue-derived SI and enteroids. TNF also resulted in increased enteroid sphericity (quantified as circularity from two-dimensional bright field images). This response was dose and time-dependent and correlated with active caspase-3 immunopositivity. Proximal tissue-derived enteroids generated from Nfκb2-/- mice showed a significantly blunted circularity response following the addition of TNF, IFNγ, lipopolysaccharide (LPS) activated C57BL/6J-derived bone marrow-derived dendritic cells (BMDC) and secreted factors from LPS-activated BMDCs. However, Nfκb1-/- mouse-derived enteroids showed no significant changes in response to these stimuli. In conclusion, the selection of SI region is important when designing enteroid studies as region-specific identity and response to stimuli such as TNF are maintained in culture. Intestinal epithelial cells are at least partially responsible for regulating their own fate by modulating NF-κB2 signalling in response to stimuli known to be involved in multiple intestinal and systemic diseases. Future studies are warranted to investigate the therapeutic potential of intestinal epithelial NF-κB2 inhibition.

A Reassessment of Genes Modulating Aging in Mice Using Demographic Measurements of the Rate of Aging.

Many studies have reported genetic interventions that have an effect on mouse life span; however, it is crucial to discriminate between manipulations of aging and aging-independent causes of life extension. Here, we used the Gompertz equation to determine whether previously reported aging-related mouse genes statistically affect the demographic rate of aging. Of 30 genetic manipulations previously reported to extend life span, for only two we found evidence of retarding demographic aging: Cisd2 and hMTH1 Of 24 genetic manipulations reported to shorten life span and induce premature aging features, we found evidence of five accelerating demographic aging: Casp2, Fn1, IKK-ß, JunD, and Stub1 Overall, our reassessment found that only 15% of the genetic manipulations analyzed significantly affected the demographic rate of aging as predicted, suggesting that a relatively small proportion of interventions affecting longevity do so by regulating the rate of aging. By contrast, genetic manipulations affecting longevity tend to impact on aging-independent mortality. Our meta-analysis of multiple mouse longevity studies also reveals substantial variation in the controls used across experiments, suggesting that a short life span of controls is a potential source of bias. Overall, the present work leads to a reassessment of genes affecting the aging process in mice, with broad implications for our understanding of the genetics of mammalian aging and which genes may be more promising targets for drug discovery.

The development of a Spanish language version of the Worn Out scale of the General Well-Being Questionnaire (GWBQ).

The paper describes the development of a short Spanish-language version of the General Well-Being Questionnaire (GWBQ; Cox and Gotts, 1987), based on the 12 items of its Worn Out scale. Research has shown the English-version Worn Out scale to be sensitive to aspects of the design and management of work. This study aimed to test its cross-cultural consistency in a Spanish-language workplace context. The data were collected from a sample of 229 workers in Valencia (Spain). Confirmatory Factor Analyses showed the factorial validity, reliability, and concurrent validity of the new Spanish version to be adequate. The sensitivity of the new measure to safety behavior and the reporting of accidents was also assessed and shown to be good. The new questionnaire extends the usefulness of the parent questionnaire to occupational health psychology research in the Spanish language by offering a short assessment tool appropriate for workplace studies.

Books on Prescription - community-based health initiative to increase access to mental health treatment: an evaluation.

OBJECTIVE: To evaluate the implementation of a regional Books on Prescription (BoP) programme. METHODS: Seven data collection pathways were used to provide indicators of program implementation, utilisation, acceptability and reach. RESULTS: BoP is seen by prescribers, librarians and consumers as a valuable mental health resource. Key areas for improvement were identified: booklist literacy level and breadth of topic area; confidentiality; program promotion; and prescriber/librarian training. CONCLUSIONS: Recommendations are made regarding the improvement of programme acceptability, accessibility and assessment. IMPLICATIONS: The establishment of a national BoP scheme would facilitate sustainable and consistent methods for BoP promotion and assessment. The authors hope this evaluation is a step towards actualising this goal.

Evidence for a common pattern of peptidergic innervation of cnidocytes.

Tentacles from representatives of all four classes of the phylum Cnidaria were examined using antibodies against the neuropeptides FMRFamide and RFamide to reveal the organization of neurons and nerve nets associated with cnidocytes. The tentacles of all species examined contained FMRFamide- or RFamide-immunoreactive neurons, in varying densities. In representatives from the Scyphozoa, Hydrozoa, and Cubozoa, the FMRFamide-immunoreactive neurons formed plexuses at the base of the cnidocyte assemblages; in anthozoans, the absence of discrete assemblies of cnidocytes precluded visual co-localization of cnidocytes and immunoreactive neurons. In all four classes, immunoreactive sensory cells connected these peptidergic nerve nets to the surface of the tentacle. These findings suggest that members of all four cnidarian classes share a common organizational pattern, and it is proposed that this peptidergic innervation may be involved in the chemosensory regulation of cnidocyte discharge.

A Lilliputian army under the floorboards: persistent delirium with complete though prolonged recovery.

We report the case of an 83-year-old man who presented with a history of fluctuating delirium of insidious onset, secondary to an amoebic liver abscess more than 30 years after acute exposure. We describe a 2-year clinical journey that started with a fall and was additionally complicated by severe weight loss and acute kidney injury (AKI). The likely prognosis for such a combination of comorbidities in an older person is for lasting morbidity, institutionalisation and significant mortality. However, the case demonstrates that with timely assessment and care complete recovery is possible though it may take many months. It reminds us of the catalytic implications of falls for older persons and to maintain a differential diagnostic approach to delirium of insidious onset avoiding misdiagnosis as dementia with which it may be associated. Our case report includes extracts from the patient's own account providing added insight into such experiences.

Right atrial mass in patient undergoing chemotherapy.

A 58-year-old lady, with a Hickman line in situ for chemotherapy for invasive ductal breast carcinoma, presented with fever. Blood cultures grew coagulase negative staphylococci. Transoesophageal echocardiography revealed a mass in the right atrium in the region of the Eustachian apparatus. The Hickman line was removed and the patient was treated for right-sided endocarditis. However, the mass persisted after prolonged intravenous antibiotics and a decision was made to remove it surgically. Histology revealed organised thrombus. The differential investigation of a right atrial mass in this position is discussed.

Patient motion correction for multiplanar, multi-breath-hold cardiac cine MR imaging.

PURPOSE: To correct for spatial misregistration of multi-breath-hold short-axis (SA), two-chamber (2CH), and four-chamber (4CH) cine cardiac MR (CMR) images caused by respiratory and patient motion. MATERIALS AND METHODS: Twenty CMR studies from consecutive patients with separate breath-hold 2CH, 4CH, and SA 20-phase cine images were considered. We automatically registered the 2CH, 4CH, and SA images in three dimensions by minimizing the cost function derived from plane intersections for all cine phases. The automatic alignment was compared with manual alignment by two observers. RESULTS: The processing time for the proposed method was <20 seconds, compared to 14-24 minutes for the manual correction. The initial plane displacement identified by the observers was 2.8 +/- 1.8 mm (maximum = 14 mm). A displacement of >/=5 mm was identified in 15 of 20 studies. The registration accuracy (defined as the difference between the automatic parameters and those obtained by visual registration) was 1.0 +/- 0.9 mm, 1.1 +/- 1.0 mm, 1.1 +/- 1.2 mm, and 2.0 +/- 1.8 mm for 2CH-4CH alignment and SA alignment in the mid, basal, and apical regions, respectively. The algorithm variability was higher in the apex (2.0 +/- 1.9 mm) than in the mid (1.4 +/- 1.4 mm) or basal (1.2 +/- 1.2 mm) regions (ANOVA, P < 0.05). CONCLUSION: An automated preprocessing algorithm can reduce spatial misregistration between multiple CMR images acquired at different breath-holds and plane orientations.

The development of a Spanish language version of the worn out scale of the general well-being questionnaire (GWBQ)

El artículo describe el desarrollo de la versión corta en español del General Well-Being Questionnaire (GWBQ; Cox & Gotts, 1987; en español el «Cuestionario de Bienestar General»), basada en los 12 ítems de su subescala de «Agotamiento». La investigación ha mostrado que la versión en inglés de la escala de Agotamiento es sensible a ciertos aspectos del diseño y gestión del trabajo. El propósito de este estudio fue comprobar la consistencia transcultural en un contexto de trabajo de habla española. Los datos se recogieron de una muestra de 229 trabajadores en Valencia (España). Análisis factoriales confirmatorios demostraron la adecuación de la validez factorial, la fiabilidad, y la validez concurrente de la nueva versión española. También se evaluó la sensibilidad de la nueva medida hacia la conducta de seguridad y distintas medidas de siniestralidad, comprobando asimismo su adecuación. El cuestionario nuevo amplía la utilidad del cuestionario original al ofrecerse como herramienta para los investigadores en psicología de la salud ocupacional en lengua española, ofreciendo un instrumento corto de evaluación apropiado para el lugar de trabajo (AU)

The paper describes the development of a short Spanish-language version of the General Well-Being Questionnaire (GWBQ; Cox & Gotts, 1987), based on the 12 items of its Worn Out scale. Research has shown the English-version Worn Out scale to be sensitive to aspects of the design and management of work. This study aimed to test its cross cultural consistency in a Spanish-language workplace context. The data were collected from a sample of 229 workers in Valencia (Spain). Confirmatory Factor Analyses showed the factorial validity, reliability, and concurrent validity of the new Spanish version to be adequate. The sensitivity of the new measure to safety behavior and the reporting of accidents was also assessed and shown to be good. The new questionnaire extends the usefulness of the parent questionnaire to occupational health psychology research in the Spanish language by offering a short assessment tool appropriate for workplace studies (AU)