Reconstruction of the insulin-like signalling pathway of Haemonchus contortus

Background: In the present study, we reconstructed the insulin/insulin-like growth factor 1 signalling (IIS) pathway for Haemonchus contortus, which is one of the most important eukaryotic pathogens of livestock worldwide and is related to the free-living nematode Caenorhabditis elegans. Methods: We curated full-length open-reading frames from assembled transcripts, defined the complement of genes that encode proteins involved in this pathway and then investigated the transcription profiles of these genes for all key developmental stages of H. contortus. Results: The core components of the IIS pathway are similar to their respective homologs in C. elegans. However, there is considerable variation in the numbers of isoforms between H. contortus and C. elegans and an absence of AKT-2 and DDL-2 homologs from H. contortus. Interestingly, DAF-16 has a single isoform in H. contortus compared with 12 in C. elegans, suggesting novel functional roles in the parasitic nematode. Some IIS proteins, such as DAF-18 and SGK-1, vary in their functional domains, indicating distinct roles from their homologs in C. elegans. Conclusions: This study paves the way for the further characterization of key signalling pathways in other socioeconomically important parasites and should help understand the complex mechanisms involved in developmental processes

The mitochondrial genome of Angiostrongylus mackerrasae as a basis for molecular, epidemiological and population genetic studies

BACKGROUND: Angiostrongylus mackerrasae is a metastrongyloid nematode endemic to Australia, where it infects the native bush rat, Rattus fuscipes. This lungworm has an identical life cycle to that of Angiostrongylus cantonensis, a leading cause of eosinophilic meningitis in humans. The ability of A. mackerrasae to infect non-rodent hosts, specifically the black flying fox, raises concerns as to its zoonotic potential. To date, data on the taxonomy, epidemiology and population genetics of A. mackerrasae are unknown. Here, we describe the mitochondrial (mt) genome of A. mackerrasae with the aim of starting to address these knowledge gaps. METHODS: The complete mitochondrial (mt) genome of A. mackerrasae was amplified from a single morphologically identified adult worm, by long-PCR in two overlapping amplicons (8 kb and 10 kb). The amplicons were sequenced using the MiSeq Illumina platform and annotated using an in-house pipeline. Amino acid sequences inferred from individual protein coding genes of the mt genomes were concatenated and then subjected to phylogenetic analysis using Bayesian inference. RESULTS: The mt genome of A. mackerrasae is 13,640 bp in size and contains 12 protein coding genes (cox1-3, nad1-6, nad4L, atp6 and cob), and two ribosomal RNA (rRNA) and 22 transfer RNA (tRNA) genes. CONCLUSIONS: The mt genome of A. mackerrasae has similar characteristics to those of other Angiostrongylus species. Sequence comparisons reveal that A. mackerrasae is closely related to A. cantonensis and the two sibling species may have recently diverged compared with all other species in the genus with a highly specific host selection. This mt genome will provide a source of genetic markers for explorations of the epidemiology, biology and population genetics of A. mackerrasae

Uncovering the effects of gender affirming hormone therapy on skeletal muscle and epigenetics: protocol for a prospective matched cohort study in transgender individuals (the GAME study)

INTRODUCTION: Gender affirming hormone therapy (GAHT) is increasingly used by transgender individuals and leads to shifts in sex hormone levels. Skeletal muscle is highly responsive to hormone activity, with limited data on the effects of GAHT on different human tissues. Here, we present the protocol for the GAME study (the effects of Gender Affirming hormone therapy on skeletal Muscle training and Epigenetics), which aims to uncover the effects of GAHT on skeletal muscle 'omic' profiles (methylomics, transcriptomics, proteomics, metabolomics) and markers of skeletal muscle health and fitness. METHODS AND ANALYSIS: This study is a prospective age-matched cohort study in transgender adults commencing GAHT (n=80) and age-matched individuals not commencing GAHT (n=80), conducted at Austin Health and Victoria University in Victoria, Australia. Assessments will take place prior to beginning GAHT and 6 and 12 months into therapies in adults commencing GAHT. Age-matched individuals will be assessed at the same time points. Assessments will be divided over three examination days, involving (1) aerobic fitness tests, (2) muscle strength assessments and (3) collection of blood and muscle samples, as well as body composition measurements. Standardised diets, fitness watches and questionnaires will be used to control for key confounders in analyses. Primary outcomes are changes in aerobic fitness and muscle strength, as well as changes in skeletal muscle DNA methylation and gene expression profiles. Secondary outcomes include changes in skeletal muscle characteristics, proteomics, body composition and blood markers. Linear mixed models will be used to assess changes in outcomes, while accounting for repeated measures within participants and adjusting for known confounders. ETHICS AND DISSEMINATION: The Austin Health Human Research Ethics Committee (HREC) and Victoria University HREC granted approval for this study (HREC/77146/Austin-2021). Findings from this project will be published in open-access, peer-reviewed journals and presented to scientific and public audiences. TRIAL REGISTRATION NUMBER: ACTRN12621001415897; Pre-results

An epigenomic and omics approach to neurodevelopmental disorders

© 2020 Namitha MohandasNeurodevelopmental disorders such as cerebral palsy (CP) and epilepsy are some of the most prevalent childhood neurological disorders caused by damage to the growth and development of the brain. Early life environments predispose children to later health outcomes evidenced by the developmental origins of health and disease (DOHaD) phenomenon. Epigenetics, which refers to modifications of DNA without change in DNA sequence, is one way by which environmental exposures may contribute to development of disease. DNA methylation, arguably the most highly studied epigenetic mark, has been correlated with early life environmental exposures and have implications in both disease mechanisms as well as clinical biomarkers of neurodevelopmental diseases. These modifications most likely originate in utero, in line with the DOHaD hypothesis. The study of monozygotic (MZ) twins, in which genetics, age, sex, parental factors and shared environment are controlled for, helps in distinguishing the extent of effect of genetics and environment. Discordance for neurodevelopmental disorders has been recorded in MZ twins indicating a potential role of non-shared factors in disease risk. The aim of this PhD was to utilise the discordant MZ twin model to understand epigenetic changes associated with neurodevelopmental disorders. Genome-wide DNA methylation was measured within MZ twin cohorts discordant for CP or epilepsy using Illumina’s Infinium HumanMethylation450 and EPIC arrays. Statistical and bioinformatics pipelines were applied to evaluate the association of DNA methylation data to disease phenotypes. As detailed in Chapter Three of this thesis, DNA methylation analysis of CP-discordant twin pairs provides the first evidence that environmentally mediated differential methylation in genes involved in known processes such as hypoxia and inflammation, and processes such as cell adhesion, may contribute to the development of CP. As detailed in Chapter Four, an epigenome-wide analysis of epilepsy discordant MZ twin pairs revealed distinct patterns of DNA methylation within subtypes of epilepsies of unknown cause. Differentially methylated genes within epilepsy subtypes included those with a role in metabolic pathways, voltage-gated channel signalling and neurotransmitter processes. This research paves the way for future larger studies, as understanding DNA methylation profiles associated with neurodevelopmental disorders, may facilitate biomarkers for earlier diagnosis. This could lead to possible intervention strategies for patients suffering from a broad spectrum of disorders. Analysing epigenetic data from disease discordant twins provides an elegant study design and has the power to explore non-shared environmental factors that further refine models of disease mechanisms and biomarkers. The findings of this thesis suggest that epigenetic factors may play a role regulating biological pathways that underlie neurodevelopmental disorders, some of which arise as early as the prenatal period. Replication in other larger and similar cohorts of discordant twin pairs may provide novel targets for biomarker development, thereby allowing for early interventions and helping the health of children

Deciphering the role of epigenetics in self-limited epilepsy with centrotemporal spikes

Objective: The aetiology of self-limited epilepsy with centro-temporal spikes (SECTS) remains controversial and a strong genetic basis has long been presumed. The discordant monozygotic twin (MZ) model controls for shared genetic and environmental factors, enabling focus on the potential role of the non-shared environment. Methods: DNA methylation data was acquired from DNA extracted from three discordant MZ twin pairs, from both new born blood spots before epilepsy onset, and blood samples taken after epilepsy onset. An epigenome-wide analysis was performed, using the Illumina Infinium EPIC array. Differentially methylated regions (DMR) were identified using the bumphunter package in R. Comparative analyses were undertaken at the two different time points as well as a combined analysis independent of time. Results: Many of the top DMR-associated genes have previously been described in neurodevelopmental disorders. The LYPD8 gene was associated with a top-ranked DMR both at birth and across the two time points. Conclusion: We have demonstrated the novel utility of the longitudinal, discordant MZ twin model, to facilitate a deeper appreciation of the complex neurobiology of SECTS. The genetic architecture of SECTS is complex and is likely to involve an interplay between genes and environment, in part mediated by epigenetics

Epigenetic studies of neurodevelopment in twins

Epigenetic studies of neurodevelopment in twin