Visualisation of quadratic discriminant analysis and its application in exploration of microbial interactions

Background: When comparing diseased and non-diseased patients in order to discriminate between the aspects associated with the specific disease, it is often observed that the diseased patients have more variability than the non-diseased patients. In such cases Quadratic discriminant analysis is required which is based on the estimation of different covariance structures for the different groups. Having different covariance matrices means the Canonical variate transformation cannot be used to obtain a visual representation of the discrimination and group separation. Results: In this paper an alternative method is proposed: combining the different transformations for the different groups into a single representation of the sample points with classification regions. In order to associate the differences in variables with group discrimination, a biplot is produced which include information on the variables, samples and their relationship

Optimizing 16S rRNA gene profile analysis from low biomass nasopharyngeal and induced sputum specimens

Careful consideration of experimental artefacts is required in order to successfully apply high-throughput 16S ribosomal ribonucleic acid (rRNA) gene sequencing technology. Here we introduce experimental design, quality control and “denoising” approaches for sequencing low biomass specimens. Results We found that bacterial biomass is a key driver of 16S rRNA gene sequencing profiles generated from bacterial mock communities and that the use of different deoxyribonucleic acid (DNA) extraction methods [DSP Virus/Pathogen Mini Kit® (Kit-QS) and ZymoBIOMICS DNA Miniprep Kit (Kit-ZB)] and storage buffers [PrimeStore® Molecular Transport medium (Primestore) and Skim-milk, Tryptone, Glucose and Glycerol (STGG)] further influence these profiles. Kit-QS better represented hard-to-lyse bacteria from bacterial mock communities compared to Kit-ZB. Primestore storage buffer yielded lower levels of background operational taxonomic units (OTUs) from low biomass bacterial mock community controls compared to STGG. In addition to bacterial mock community controls, we used technical repeats (nasopharyngeal and induced sputum processed in duplicate, triplicate or quadruplicate) to further evaluate the effect of specimen biomass and participant age at specimen collection on resultant sequencing profiles. We observed a positive correlation (r = 0.16) between specimen biomass and participant age at specimen collection: low biomass technical repeats (represented by < 500 16S rRNA gene copies/μl) were primarily collected at < 14 days of age. We found that low biomass technical repeats also produced higher alpha diversities (r = − 0.28); 16S rRNA gene profiles similar to no template controls (Primestore); and reduced sequencing reproducibility. Finally, we show that the use of statistical tools for in silico contaminant identification, as implemented through the decontam package in R, provides better representations of indigenous bacteria following decontamination. Conclusions We provide insight into experimental design, quality control steps and “denoising” approaches for 16S rRNA gene high-throughput sequencing of low biomass specimens. We highlight the need for careful assessment of DNA extraction methods and storage buffers; sequence quality and reproducibility; and in silico identification of contaminant profiles in order to avoid spurious results

International consensus recommendations for the identification and treatment of tuberous sclerosis complex-associated neuropsychiatric disorders (TAND).

BACKGROUND: Tuberous sclerosis complex (TSC) is associated with a wide range of physical manifestations for which international clinical recommendations for diagnosis and management have been established. TSC is, however, also associated with a wide range of TSC-Associated Neuropsychiatric Disorders (TAND) that are typically under-identified and under-treated yet associated with a profound burden of disease. The contemporary evidence base for the identification and treatment of TAND is much more limited and, to date, consensus recommendations for the diagnosis and management of TAND have also been limited and non-specific. METHODS: The TANDem project was launched with an international, interdisciplinary, and participatory consortium of 24 individuals, including TSC family representatives, from all World Health Organization (WHO) regions but one. One of the aims of the TANDem project was to generate consensus recommendations for the identification and treatment of TAND. At the time of this project, no internationally adopted standard methodology and methodological checklists existed for the generation of clinical practice recommendations. We therefore developed our own systematic procedure for evidence review and consensus-building to generate evidence-informed consensus recommendations of relevance to the global TSC community. RESULTS: At the heart of the consensus recommendations are ten core principles surrounded by cluster-specific recommendations for each of the seven natural TAND clusters identified in the literature (autism-like, dysregulated behavior, eat/sleep, mood/anxiety, neuropsychological, overactive/impulsive, and scholastic) and a set of wraparound psychosocial cluster recommendations. The overarching recommendation is to "screen" for TAND at least annually, to "act" using appropriate next steps for evaluation and treatment, and to "repeat" the process to ensure early identification and early intervention with the most appropriate biological, psychological, and social evidence-informed approaches to support individuals with TSC and their families. CONCLUSIONS: The consensus recommendations should provide a systematic framework to approach the identification and treatment of TAND for health, educational, social care teams and families who live with TSC. To ensure global dissemination and implementation of these recommendations, partnerships with the international TSC community will be important. One of these steps will include the generation of a "TAND toolkit" of "what to seek" and "what to do" when difficulties are identified in TAND clusters

The research landscape of tuberous sclerosis complex-associated neuropsychiatric disorders (TAND)-a comprehensive scoping review.

BACKGROUND: Tuberous sclerosis complex (TSC)-associated neuropsychiatric disorders (TAND) is an umbrella term for the behavioural, psychiatric, intellectual, academic, neuropsychological and psychosocial manifestations of TSC. Although TAND affects 90% of individuals with TSC during their lifetime, these manifestations are relatively under-assessed, under-treated and under-researched. We performed a comprehensive scoping review of all TAND research to date (a) to describe the existing TAND research landscape and (b) to identify knowledge gaps to guide future TAND research. METHODS: The study was conducted in accordance with stages outlined within the Arksey and O'Malley scoping review framework. Ten research questions relating to study characteristics, research design and research content of TAND levels and clusters were examined. RESULTS: Of the 2841 returned searches, 230 articles published between 1987 and 2020 were included (animal studies = 30, case studies = 47, cohort studies = 153), with more than half published since the term TAND was coined in 2012 (118/230; 51%). Cohort studies largely involved children and/or adolescents (63%) as opposed to older adults (16%). Studies were represented across 341 individual research sites from 45 countries, the majority from the USA (89/341; 26%) and the UK (50/341; 15%). Only 48 research sites (14%) were within low-middle income countries (LMICs). Animal studies and case studies were of relatively high/high quality, but cohort studies showed significant variability. Of the 153 cohort studies, only 16 (10%) included interventions. None of these were non-pharmacological, and only 13 employed remote methodologies (e.g. telephone interviews, online surveys). Of all TAND clusters, the autism spectrum disorder-like cluster was the most widely researched (138/230; 60%) and the scholastic cluster the least (53/200; 27%). CONCLUSIONS: Despite the recent increase in TAND research, studies that represent participants across the lifespan, LMIC research sites and non-pharmacological interventions were identified as future priorities. The quality of cohort studies requires improvement, to which the use of standardised direct behavioural assessments may contribute. In human studies, the academic level in particular warrants further investigation. Remote technologies could help to address many of the TAND knowledge gaps identified

Development and Feasibility of the Self-Report Quantified Tuberous Sclerosis Complex-Associated Neuropsychiatric Disorders Checklist (TAND-SQ).

BACKGROUND: Tuberous sclerosis complex-associated neuropsychiatric disorders (TAND) are often present but underidentified and undertreated in individuals with tuberous sclerosis complex (TSC). The clinician-completed TAND-Lifetime Checklist (TAND-L) was developed to address this identification and treatment gap. Stakeholder engagement identified the need for a TAND Checklist that can (1) be completed by caregivers or individuals with TSC and (2) quantify TAND difficulties. The aim of this study was to develop a self-report quantified TAND Checklist (TAND-SQ) and conduct feasibility and acceptability testing. METHODS: This aim was addressed in three phases: (1) development of the TAND-SQ Checklist, (2) feasibility and acceptability testing of the "near-final" TAND-SQ Checklist, and (3) preparation of the final TAND-SQ Checklist. Participants included 23 technical experts from the TAND consortium in all phases and 58 lived experts (caregivers and individuals with TSC) in phase 2. All participants completed a TAND-SQ Checklist and a checklist feedback form. RESULTS: Phase 1 additions to the TAND-SQ, when compared with the TAND-L, included four new items and a quantification rating. Phase 2 showed high ratings for the "near-final" TAND-SQ Checklist on comprehensiveness, clarity, ease of use, and overall acceptability. In phase 3, questions on strengths, strategies, and a TAND Cluster Profile were added. CONCLUSION: The TAND-SQ Checklist is presented here for use by individuals with TSC and their caregivers. The next steps as part of the TANDem project include internal and external validation of the checklist and linking of TAND Cluster Profiles generated from the checklist to evidence-informed consensus recommendations within a smartphone application

Discovery and functional prioritization of Parkinson's disease candidate genes from large-scale whole exome sequencing.

BACKGROUND: Whole-exome sequencing (WES) has been successful in identifying genes that cause familial Parkinson's disease (PD). However, until now this approach has not been deployed to study large cohorts of unrelated participants. To discover rare PD susceptibility variants, we performed WES in 1148 unrelated cases and 503 control participants. Candidate genes were subsequently validated for functions relevant to PD based on parallel RNA-interference (RNAi) screens in human cell culture and Drosophila and C. elegans models. RESULTS: Assuming autosomal recessive inheritance, we identify 27 genes that have homozygous or compound heterozygous loss-of-function variants in PD cases. Definitive replication and confirmation of these findings were hindered by potential heterogeneity and by the rarity of the implicated alleles. We therefore looked for potential genetic interactions with established PD mechanisms. Following RNAi-mediated knockdown, 15 of the genes modulated mitochondrial dynamics in human neuronal cultures and four candidates enhanced α-synuclein-induced neurodegeneration in Drosophila. Based on complementary analyses in independent human datasets, five functionally validated genes-GPATCH2L, UHRF1BP1L, PTPRH, ARSB, and VPS13C-also showed evidence consistent with genetic replication. CONCLUSIONS: By integrating human genetic and functional evidence, we identify several PD susceptibility gene candidates for further investigation. Our approach highlights a powerful experimental strategy with broad applicability for future studies of disorders with complex genetic etiologies

Clinical educators' self-reported personal and professional development after completing a short course in undergraduate clinical supervision at Stellenbosch University

Background. In 2007, a Supervision Course in Undergraduate Clinical Supervision was developed at the Faculty of Medicine and Health Sciencesat Stellenbosch University in South Africa. The target group was inter-professional clinical educators that are involved in student education on the clinical platform. Although the course participants were professionals and specialists in their own fields, the majority of clinical educators have very little or no knowledge of adult education. The Supervision Course aims to develop clinical supervision skills of clinical educators by exposing these supervisors to basic principles of education and specifically clinical teaching, resulting in quality education for undergraduate students. The aim of this study was to determine the impact of this short course on the personal and professional growth of the clinical educator.Methods. A qualitative study was performed, including an open-ended questionnaire that provided opportunity for the clinical educators to elaborate freely on their strengths, weaknesses and areas of desired improvement before and after the Supervision Course, and a semi-structured individual interview after the Supervision Course. The questionnaire data were categorised according to strengths, weaknesses and areas of desired improvement. An inductive approach was used to analyse the qualitative data. Key themes that emerged from the interviews were identified and grouped together in categories.Results. The results are summarised in table format to identify themes with supporting quotes.Conclusion. Although a small sample, this study demonstrates the personal and professional growth reported by attendees of a clinical supervision short course