Association between early and current gastro-intestinal symptoms and co-morbidities in children and adolescents with Angelman syndrome.

BACKGROUND: Angelman syndrome (AS) is a neurogenetic disorder that causes severe intellectual disability, expressive language deficits, motor impairment, ataxia, sleep problems, epileptic seizures and a happy disposition. People with AS frequently experience gastrointestinal (GI) symptoms. METHOD: This study used data from the Global Angelman Syndrome Registry to explore the relationship between early and current GI symptoms and co-morbidity in children and adolescents with AS (n = 173). Two groups that experienced a high (n = 91) and a low (n = 82) frequency of GI symptoms were examined in relation to feeding and GI history in infancy, sleep and toileting problems, levels of language and communication and challenging behaviours. Predictors of GI symptoms were then investigated using a series of logistic regressions. RESULTS: This analysis found that constipation and gastroesophageal reflux affected 84% and 64%, of the sample, respectively. The high frequency of GI symptoms were significantly associated with: 'refusal to nurse', 'vomiting', 'arching', 'difficulty gaining weight', gastroesophageal reflux, 'solid food transition', frequency of night-time urinary continence and sleep hyperhidrosis during infancy. GI symptoms were not significantly associated with sleep, toileting, language or challenging behaviours. Significant predictors of high frequency GI symptoms were gastroesophageal reflux and sleep hyperhidrosis. CONCLUSIONS: Future research needs to investigate the association between AS and GI co-morbidity in adults with AS.

Research protocol: The initiation, design and establishment of the Global Angelman Syndrome Registry.

BACKGROUND: Angelman syndrome (AS) is a rare neurodevelopmental disorder affecting between 1 in 15 000 and 1 in 24 000 individuals. The condition results in severe developmental and expressive language delays, motor impairments and a unique behavioural phenotype consisting of excessive laughter, smiling and sociability. While many studies have contributed knowledge about the causes and natural history of the syndrome, large scale longitudinal studies are required to advance research and therapeutics for this rare syndrome. METHOD: This article describes the protocol for the Global Angelman Syndrome Registry, and some initial findings. Due to the rarity of AS and the variability in symptom presentation, the registry team will strive for complete case ascertainment. Parents and caregivers will submit data to the registry via a secure internet connection. The registry consists of 10 modules that cover patient demographics; developmental, diagnostic, medical and surgical history, behaviour and development, epilepsy, medications and interventions and sleep. RESULTS: Since its launch at https://angelmanregistry.info in September 2016, almost 470 individuals with AS have been signed up to the registry worldwide: 59% are from North and South America, 23% are from Europe, 17% are from the Asia Pacific region and 1% are from the Middle East or Africa. The majority of registrants are children, with only 16% aged over 20 years. Most participants indicated a chromosome deletion (76%), with fewer participants indicating a mutation, uniparental disomy or imprinting defect (20%). CONCLUSION: Findings indicate a need to consider recruitment strategies that target caregivers of older children and adults, and parents and caregivers from non-English speaking backgrounds.

Advances in biotechnology and informatics to link variation in the genome to phenotypes in plants and animals.

Advances in our understanding of genome structure provide consistent evidence for the existence of a core genome representing species classically defined by phenotype, as well as conditionally dispensable components of the genome that shows extensive variation between individuals of a given species. Generally, conservation of phenotypic features between species reflects conserved features of the genome; however, this is evidently not necessarily always the case as demonstrated by the analysis of the tunicate chordate Oikopleura dioica. In both plants and animals, the methylation activity of DNA and histones continues to present new variables for modifying (eventually) the phenotype of an organism and provides for structural variation that builds on the point mutations, rearrangements, indels, and amplification of retrotransposable elements traditionally considered. The translation of the advances in the structure/function analysis of the genome to industry is facilitated through the capture of research outputs in "toolboxes" that remain accessible in the public domain.

Advances in biotechnology and linking outputs to variation in complex traits: Plant and Animal Genome meeting January 2012.

The Plant and Animal Genome (PAG, held annually) meeting in January 2012 provided insights into the advances in plant, animal, and microbe genome studies particularly as they impact on our understanding of complex biological systems. The diverse areas of biology covered included the advances in technologies, variation in complex traits, genome change in evolution, and targeting phenotypic changes, across the broad spectrum of life forms. This overview aims to summarize the major advances in research areas presented in the plenary lectures and does not attempt to summarize the diverse research activities covered throughout the PAG in workshops, posters, presentations, and displays by suppliers of cutting-edge technologies.

Genome studies at the PAG 2011 conference.

The contents of the plenary lectures presented at the Plant and Animal Genome (PAG) meeting in January 2011 are summarized in order to provide some insights into the advances in plant, animal and microbe genome studies as they impact on our understanding of complex biological systems. The areas of biology covered include the dynamics of genome change, biological recognition processes and the new processes that underpin investment in science. This overview does not attempt to summarize the diversity of activities that are covered during the PAG through workshops, posters and the suppliers of cutting-edge technologies, but reviews major advances in specific research areas.

Advances in genome studies: The PAG 2010 conference.

An overview is provided of the advances in plant, animal and human genome studies by summarizing the contents of seven plenary lectures presented at the Plant and Animal Genome (PAG) meeting in January 2010. The area of biology covered was wide and reflected the nature of this fast moving science.

Identification of genes involved with tick infestation in Bos taurus and Bos indicus.

Tick resistant cattle could provide a potentially sustainable and environmentally sound method of controlling cattle ticks. Advances in genomics and the availability of the bovine genome sequence open up opportunities to identify useful and selectable genes controlling cattle tick resistance. Using quantitative real-time PCR and theAffymetrix bovine array platform, differences in gene expression of skin biopsies from tick resistant Bos indicus (Brahman) and tick susceptible Bos taurus (Holstein-Friesian) cattle following tick challenge were examined. We identified 138 significant differentially-expressed genes, including several immunologicallhost defence genes, extracellularmatrix proteins, and transcription factors as well as genes involved in lipid metabolism. Three key pathways, represented by genes differentially expressed in resistant Brahmans, were identified; the development of the cell-mediated immune response, structural integrity of the dermis and intracellular Ca2+ levels. Ca2+, which is implicated in host responses to microbial stimuli, may be required for the enhancement or fine-tuning of transcriptional activation of Ca2+ -dependant host defence signalling pathways.

Translational development of splice-modifying antisense oligomers.

INTRODUCTION: Antisense nucleic acid analogues can interact with pre-mRNA motifs and influence exon or splice site selection and thereby alter gene expression. Design of antisense molecules to target specific motifs can result in either exon exclusion or exon inclusion during splicing. Novel drugs exploiting the antisense concept are targeting rare, life-limiting diseases; however, the potential exists to treat a wide range of conditions by antisense-mediated splice intervention. Areas covered: In this review, the authors discuss the clinical translation of novel molecular therapeutics to address the fatal neuromuscular disorders Duchenne muscular dystrophy and spinal muscular atrophy. The review also highlights difficulties posed by issues pertaining to restricted participant numbers, variable phenotype and disease progression, and the identification and validation of study endpoints. Expert opinion: Translation of novel therapeutics for Duchenne muscular dystrophy and spinal muscular atrophy has been greatly advanced by multidisciplinary research, academic-industry partnerships and in particular, the engagement and support of the patient community. Sponsors, supporters and regulators are cooperating to deliver new drugs and identify and define meaningful outcome measures. Non-conventional and adaptive trial design could be particularly suited to clinical evaluation of novel therapeutics and strategies to treat serious, rare diseases that may be problematic to study using more conventional clinical trial structures.

Significant differences between the G+C content of synonymous codons in orthologous genes and the genomic G+C content.

The relationship between the overall G+C content of the genome (GC) and the GC content at the third codon positions (GC3) of genes, which we refer to as a GC3-plot, was examined using 15 currently available complete genome sequences. A remarkably linear relationship was found between these two quantities, confirming previous observations of a strong positive correlation in the GC3-plot. In order to conduct a more detailed analysis of the GC3-plot, we examined the GC3 content by separating orthologous codons into three categories: synonymously different codons (namely identical amino acids, IA), different amino acids (DA), and identical codons (IC), for a pairwise comparison of two closely related species. When we took pairwise species comparisons between Mycoplasma genitalium (Mg) and Mycoplasma pneumoniae (Mp) and between Mycobacterium tuberculosis (Mt) and Mycobacterium leprae (Ml) as examples, we found that for Mp and Ml, the GC3 for IA deviated the most from the linear expectation in the GC3-plot, whereas for Mg and Mt the deviation was minimal. These findings suggest that the major changes of GC content took place in Mp and Ml, but not in Mg and Mt. This analysis also enables us to predict the future direction of the evolutionary changes of the genomic GC content.

Identification of a ribonuclease H gene in both Mycoplasma genitalium and Mycoplasma pneumoniae by a new method for exhaustive identification of ORFs in the complete genome sequences.

Exhaustive identification of open reading frames in complete genome sequences is a difficult task. It is possible that important genes are missed. In our efforts to reanalyze the intergenic regions of Mycoplasma genitalium and Mycoplasma pneumoniae, we have newly identified a number of new open reading frames (ORFs) in both M. genitalium and M. pneumoniae. The most significant identification was that of a ribonuclease H enzyme in both species which until now has not been identified or assumed absent and interpreted as such. In this paper we discuss the biological importance of RNase H and its evolutionary implication. We also stress the usefulness of our method for identifying new ORFs by reanalyzing intergenic regions of existing ORFs in complete genome sequences.

MHC haplotype analysis by artificial neural networks.

Conventional matching is based on numbers of alleles shared between donor and recipient. This approach, however, ignores the degree of relationship between alleles and haplotypes, and therefore the actual degree of difference. To address this problem, we have compared family members using a block matching technique which reflects differences in genomic sequences. All parents and siblings had been genotyped using conventional MHC typing so that haplotypes could be assigned and relatives could be classified as sharing 0, 1 or 2 haplotypes. We trained an Artificial Neural Network (ANN) with subjects from 6 families (85 comparisons) to distinguish between relatives. Using the outputs of the ANN, we developed a score, the Histocompatibility Index (HI), as a measure of the degree of difference. Subjects from a further 3 families (106 profile comparisons) were tested. The HI score for each comparison was plotted. We show that the HI score is trimodal allowing the definition of three populations corresponding to approximately 0, 1 or 2 haplotype sharing. The means and standard deviations of the three populations were found. As expected, comparisons between family members sharing 2 haplotypes resulted in high HI scores with one exception. More interestingly, this approach distinguishes between the 1 and 0 haplotype groups, with some informative exceptions. This distinction was considered too difficult to attempt visually. The approach provides promise in the quantification of degrees of histocompatibility.

Dynamic evolution of genomes and the concept of genome space.

A new era in the elucidation of genome evolution has been heralded with the availability of numerous genome sequences. With these data, it has been possible to study evolutionary processes at a greater level of detail in order to characterize features such as gene shuffling, genome rearrangements, base bias composition, and horizontal gene transfer. In this paper, we discuss the evolutionary implications of significant rearrangements within genomes as well as characteristic genomic regions that have been conserved across genomes. This is based on our analysis of orthologous and paralogous genes. We argue that genome plasticity has most likely contributed substantially to the dynamic evolution of genomes. We also describe the characteristic mosaic features of an archaea genome that is comprised of both bacterial and eukaryal elements. Here we investigate base compositional differences as well as the similarity of this species' genes to either bacteria or eukarya. We conclude that these features can be largely explained by the mechanism of horizontal gene transfer. Finally, we introduce the concept of genome space which is defined as the entire set of genomes of all living organisms. We explain its usefulness to describe as well as to gain deeper insight into the general features of the dynamic genomic evolutionary process.

Bioinformatics issues for automating the annotation of genomic sequences.

The rapid explosion in the amount of biological data being generated worldwide is surpassing efforts to manage analysis of the data. As part of an ongoing project to automate and manage bioinformatics analysis, the authors have designed and implemented a simple automated annotation system, which is described in this paper. The system is applied to existing GenBank/DDBJ/EMBL entries and compared with existing annotations to illustrate not only potential errors but also that they are generally not up-to-date, as a result of new versions of analysis tools and updates of genomic repositories. We highlight the important Bioinformatics issues of storage and management of information to ensure data and results are kept up-to-date in light of new information becoming available. Surprisingly, from just four database entries, a significant number of new features were found. We describe the results as well as identify important issues that need to be addressed in order to automate the re-analysis/re-annotation of genomic sequences within a reasonable timeframe.

Structure and polymorphism of two stress-activated protein kinase genes centromeric of the MHC: SAPK2a and SAPK4.

As MHC genes are potent determinants of susceptibility to immunopathological diseases, the mapping of SAPK2a (CSBP) and SAPK4 to chromosome 6p 21.2-21.3 suggested that these genes may mediate the effects of the MHC on disease. Here we describe the genomic structure and localisation of both genes approximately 2.3Mb centromeric of HLA-DP. Examination of the complete coding region and selected intronic regions of SAPK2a and SAPK4 from 22 human EBV-transformed B-cell lines of different MHC haplotypes and racial background revealed complete sequence conservation. There were no notable differences in levels of expression of SAPK2a and SAPK4 mRNA in cell lines of different MHC haplotypes or racial origin. Examination of the SAPK2a and SAPK4 sequences from two chimpanzees revealed 3 nucleotide differences between human and chimpanzee in each gene resulting in only one amino acid change in SAPK4, and 6 nucleotide substitutions plus 2 deletions in 600bp of intronic sequence from SAPK4. This highlights the selective pressure placed on these genes to maintain their protein sequence, but does not favour a role in genetic regulation of disease or provide evidence of linkage disequilibrium with the MHC.

Gene expression evidence for off-target effects caused by RNA interference-mediated gene silencing of Ubiquitin-63E in the cattle tick Rhipicephalus microplus.

Knowledge of cattle tick (Rhipicephalus (Boophilus) microplus; Acari: Ixodidae) molecular and cellular pathways has been hampered by the lack of an annotated genome. In addition, most of the tick expressed sequence tags (ESTs) available to date consist of ∼50% unassigned sequences without predicted functions. The most common approach to address this has been the application of RNA interference (RNAi) methods to investigate genes and their pathways. This approach has been widely adopted in tick research despite minimal knowledge of the tick RNAi pathway and double-stranded RNA (dsRNA) uptake mechanisms. A strong knockdown phenotype of adult female ticks had previously been observed using a 594 bp dsRNA targeting the cattle tick homologue for the Drosophila Ubiquitin-63E gene leading to nil or deformed eggs. A NimbleGen cattle tick custom microarray based on the BmiGI.V2 database of R. microplus ESTs was used to evaluate the expression of mRNAs harvested from ticks treated with the tick Ubiquitin-63E 594 bp dsRNA compared with controls. A total of 144 ESTs including TC6372 (Ubiquitin-63E) were down-regulated with 136 ESTs up-regulated following treatment. The results obtained substantiated the knockdown phenotype with ESTs identified as being associated with ubiquitin proteolysis as well as oogenesis, embryogenesis, fatty acid synthesis and stress responses. A bioinformatics analysis was undertaken to predict off-target effects (OTE) resulting from the in silico dicing of the 594 bp Ubiquitin-63E dsRNA which identified 10 down-regulated ESTs (including TC6372) within the list of differentially expressed probes on the microarrays. Subsequent knockdown experiments utilising 196 and 109 bp dsRNAs, and a cocktail of short hairpin RNAs (shRNA) targeting Ubiquitin-63E, demonstrated similar phenotypes for the dsRNAs but nil effect following shRNA treatment. Quantitative reverse transcriptase PCR analysis confirmed differential expression of TC6372 and selected ESTs. Our study demonstrated the minimisation of predicted OTEs in the shorter dsRNA treatments (∼100-200 bp) and the usefulness of microarrays to study knockdown phenotypes.

A descriptive profile of ß-thalassaemia mutations in India, Pakistan and Sri Lanka.

Thalassaemia is a common and debilitating autosomal recessive disorder affecting many populations in South Asia. To date, efforts to create a regional profile of ß-thalassaemia mutations have largely concentrated on the populations of India. The present study updates and expands an earlier profile of ß-thalassaemia mutations in India, and incorporates comparable data from Pakistan and Sri Lanka. Despite limited data availability, clear patterns of historical and cultural population movements were observed relating to major ß-thalassaemia mutations. The current regional mutation profiles of ß-thalassaemia have been influenced by historical migrations into and from the Indian sub-continent, by the development and effects of Hindu, Buddhist, Muslim and Sikh religious traditions, and by the major mid-twentieth century population translocations that followed the Partition of India in 1947. Given the resultant genetic complexity revealed by the populations of India, Pakistan and Sri Lanka, to ensure optimum diagnostic efficiency and the delivery of appropriate care, it is important that screening and counselling programmes for ß-thalassaemia mutations recognise the underlying patterns of population sub-division throughout the region.

Profiling ß-thalassaemia mutations in India at state and regional levels: implications for genetic education, screening and counselling programmes.

UNLABELLED: Thalassaemia and sickle cell disease have been recognized by the World Health Organization as important inherited disorders principally impacting on the populations of low income countries. To create a national and regional profile of ß-thalassaemia mutations in the population of India, a meta-analysis was conducted on 17 selected studies comprising 8,505 alleles and offering near-national coverage for the disease. At the national level 52 mutations accounted for 97.5% of all ß-thalassaemia alleles, with IVSI-5(G>C) the most common disease allele (54.7%). Population stratification was apparent in the mutation profiles at regional level with, for example, the prevalence of IVSI-5(G>C) varying from 44.8% in the North to 71.4% in the East. A number of major mutations, such as Poly A(T>C), were apparently restricted to a particular region of the country, although these findings may in part reflect the variant test protocols adopted by different centres. Given the size and genetic complexity of the Indian population, and with specific mutations for ß-thalassaemia known to be strongly associated with individual communities, comprehensive disease registries need to be compiled at state, district and community levels to ensure the efficacy of genetic education, screening and counselling programmes. At the same, time appropriately designed community-based studies are required as a health priority to correct earlier sampling inequities which resulted in the under-representation of many communities, in particular rural and socioeconomically under-privileged groups. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11568-010-9132-3) contains supplementary material, which is available to authorized users.

LegumeDB1 bioinformatics resource: comparative genomic analysis and novel cross-genera marker identification in lupin and pasture legume species.

The identification of markers in legume pasture crops, which can be associated with traits such as protein and lipid production, disease resistance, and reduced pod shattering, is generally accepted as an important strategy for improving the agronomic performance of these crops. It has been demonstrated that many quantitative trait loci (QTLs) identified in one species can be found in other plant species. Detailed legume comparative genomic analyses can characterize the genome organization between model legume species (e.g., Medicago truncatula, Lotus japonicus) and economically important crops such as soybean (Glycine max), pea (Pisum sativum), chickpea (Cicer arietinum), and lupin (Lupinus angustifolius), thereby identifying candidate gene markers that can be used to track QTLs in lupin and pasture legume breeding. LegumeDB is a Web-based bioinformatics resource for legume researchers. LegumeDB analysis of Medicago truncatula expressed sequence tags (ESTs) has identified novel simple sequence repeat (SSR) markers (16 tested), some of which have been putatively linked to symbiosome membrane proteins in root nodules and cell-wall proteins important in plant-pathogen defence mechanisms. These novel markers by preliminary PCR assays have been detected in Medicago truncatula and detected in at least one other legume species, Lotus japonicus, Glycine max, Cicer arietinum, and (or) Lupinus angustifolius (15/16 tested). Ongoing research has validated some of these markers to map them in a range of legume species that can then be used to compile composite genetic and physical maps. In this paper, we outline the features and capabilities of LegumeDB as an interactive application that provides legume genetic and physical comparative maps, and the efficient feature identification and annotation of the vast tracks of model legume sequences for convenient data integration and visualization. LegumeDB has been used to identify potential novel cross-genera polymorphic legume markers that map to agronomic traits, supporting the accelerated identification of molecular genetic factors underpinning important agronomic attributes in lupin.