Assessing the risk of ocean acidification for scleractinian corals on the Great Barrier Reef

University of Technology, Sydney. Faculty of Science.Ocean acidification will impact the photo-physiology of reef-building corals as it can lead to dysfunction of the symbiosis and loss of productivity. The major objective of this thesis was to provide insight into the mechanism of CO₂₋induced bleaching and productivity loss across multiple life-history stages and interpret these findings in an ecological context. Chapter 1 provides a review of the literature investigating the photo-physiological impact of ocean acidification, emphasizing the experimental conditions in studies that observed Symbiodinium dysfunction and productivity loss. Chapter 2 presents a working hypothesis to describe the fundamental physiological aspects of coral bleaching under ocean acidification. This research investigates the response of Acropora aspera using pulse amplitude modulation (PAM) fluorometry and oxygen respirometry under increased pCO₂ with concomitant high light conditions. The dinoflagellate density and HPLC pigment analysis are utilised to characterise the CO₂₋induced bleaching response. We present a conceptual model linking photorespiration to CO₂₋induced bleaching and productivity loss. The impact of ocean acidification on coral reef ecosystems is likely to deviate from oceanic climate models due to diel modification of carbonate chemistry by community metabolism. Chapter 3 characterises the diurnal variation in carbonate chemistry at sites around Lizard Island and links this to the ocean acidification response of Acropora millepora collected from these sites. Furthermore, we utilise permutational multivariate statistical analyses to partition the variation in carbonate chemistry attributable to community composition at these sites. It was hypothesized that greater diurnal variation in carbonate chemistry may improve resilience of scleractinian corals to future ocean acidification conditions. This chapter highlights that site-specific physiological trade-offs may influence the response of reef-building corals to future ocean acidification scenarios. Chapter 4 reports a visual bleaching response in A. millepora juveniles under future ocean acidification conditions. The effect of ocean acidification on coral juveniles is hypothesised to impact Symbiodinium uptake and photochemical efficiency. We utilised the iPAM to align the photochemistry in the juveniles with their visual bleaching response and Symbiodnium type, as assessed by denaturing gradient gel electrophoresis (DGGE) of the internal transcribed spacer region 1 (ITS1) of the ribosomal genes. This study links the bleaching response with recruits containing a dominant population of Symbiodinium type D1 or D1-4, with potential implications for post-settlement survivorship and population dynamics. Lastly, in Chapter 5 the key findings of this thesis are discussed in light of the ecological implications for the Great Barrier Reef. The synopsis outlines the effect of ocean acidification on the photo-physiology, productivity, calcification, reproduction and symbiont acquisition of reef-building corals. Future avenues for research are suggested based on new research gaps revealed by this thesis with the aim to continue to provide up-to-date scientific information to policy makers and reef managers

Extensive horizontal gene transfer during Staphylococcus aureus co-colonization in vivo.

Staphylococcus aureus is a commensal and major pathogen of humans and animals. Comparative genomics of S. aureus populations suggests that colonization of different host species is associated with carriage of mobile genetic elements (MGE), particularly bacteriophages and plasmids capable of encoding virulence, resistance, and immune evasion pathways. Antimicrobial-resistant S. aureus of livestock are a potential zoonotic threat to human health if they adapt to colonize humans efficiently. We utilized the technique of experimental evolution and co-colonized gnotobiotic piglets with both human- and pig-associated variants of the lineage clonal complex 398, and investigated growth and genetic changes over 16 days using whole genome sequencing. The human isolate survived co-colonization on piglets more efficiently than in vitro. During co-colonization, transfer of MGE from the pig to the human isolate was detected within 4 h. Extensive and repeated transfer of two bacteriophages and three plasmids resulted in colonization with isolates carrying a wide variety of mobilomes. Whole genome sequencing of progeny bacteria revealed no acquisition of core genome polymorphisms, highlighting the importance of MGE. Staphylococcus aureus bacteriophage recombination and integration into novel sites was detected experimentally for the first time. During colonization, clones coexisted and diversified rather than a single variant dominating. Unexpectedly, each piglet carried unique populations of bacterial variants, suggesting limited transmission of bacteria between piglets once colonized. Our data show that horizontal gene transfer occurs at very high frequency in vivo and significantly higher than that detectable in vitro

Measuring Coverage of Prolog Programs Using Mutation Testing

Testing is an important aspect in professional software development, both to avoid and identify bugs as well as to increase maintainability. However, increasing the number of tests beyond a reasonable amount hinders development progress. To decide on the completeness of a test suite, many approaches to assert test coverage have been suggested. Yet, frameworks for logic programs remain scarce. In this paper, we introduce a framework for Prolog programs measuring test coverage using mutations. We elaborate the main ideas of mutation testing and transfer them to logic programs. To do so, we discuss the usefulness of different mutations in the context of Prolog and empirically evaluate them in a new mutation testing framework on different examples.Comment: 16 pages, Accepted for presentation in WFLP 201

What is the nature of peer interactions in children with language disorders? A qualitative study of parent and practitioner views

Background and aims: Children with Language Disorders (LDs) can exhibit increased levels of social withdrawal, aggression and problems managing social conflicts. The reasons underlying this pattern of social interaction profiles remain unclear. This qualitative study aimed to document the nature of social interactions between children with LDs and their peers, and to evaluate explanations for their social behaviour, as understood by parents and practitioners. Methods: This study focused on children with LDs who spend school hours with other children with LDs. Three parent focus groups (n ¼ 8) and three practitioner focus groups (n ¼ 10) were conducted with parents of children aged 4–12 attending specialist language schools and practitioners working at these schools. This was a mixed clinical sample. All children of participating parents had LD as their primary area of need, which was the reason they required specialist schooling. Focus groups were conducted across two specialist schools in the UK between March and June 2018. Results: An inductive reflective thematic analysis of the data identified three themes; social knowledge, coping strategies, and emotional competence. Parents and school staff reported that children with LDs experience difficulties managing peer interactions due to a combination of challenges including difficulties with understanding and regulating emotions, and difficulties understanding social situations. Some of the children with LDs were described as having developed strategies to cope with their challenges, for example imposing structure on their social interactions to manage uncertainty, which has implications for their social interactions with peers. Conclusions: Children with LDs have difficulties understanding emotions, difficulties understanding their peer’s intentions and difficulties resolving conflict situations independently according to their parents and practitioners working with these children. Participants proposed a novel explanation that social withdrawal may be used adaptively by children with LDs to process information. This study demonstrates the complexity of the relationship between Language Disorders and peer interaction profiles. Implications: Suggestions are offered regarding future research directions, such as investigating the specific contribution language skills make to children’s emotion understanding, to better understand the reasons for peer interaction difficulties in children with Language Disorders

Transcriptome Analysis of Targeted Mouse Mutations Reveals the Topography of Local Changes in Gene Expression.

The unintended consequences of gene targeting in mouse models have not been thoroughly studied and a more systematic analysis is needed to understand the frequency and characteristics of off-target effects. Using RNA-seq, we evaluated targeted and neighboring gene expression in tissues from 44 homozygous mutants compared with C57BL/6N control mice. Two allele types were evaluated: 15 targeted trap mutations (TRAP); and 29 deletion alleles (DEL), usually a deletion between the translational start and the 3' UTR. Both targeting strategies insert a bacterial beta-galactosidase reporter (LacZ) and a neomycin resistance selection cassette. Evaluating transcription of genes in +/- 500 kb of flanking DNA around the targeted gene, we found up-regulated genes more frequently around DEL compared with TRAP alleles, however the frequency of alleles with local down-regulated genes flanking DEL and TRAP targets was similar. Down-regulated genes around both DEL and TRAP targets were found at a higher frequency than expected from a genome-wide survey. However, only around DEL targets were up-regulated genes found with a significantly higher frequency compared with genome-wide sampling. Transcriptome analysis confirms targeting in 97% of DEL alleles, but in only 47% of TRAP alleles probably due to non-functional splice variants, and some splicing around the gene trap. Local effects on gene expression are likely due to a number of factors including compensatory regulation, loss or disruption of intragenic regulatory elements, the exogenous promoter in the neo selection cassette, removal of insulating DNA in the DEL mutants, and local silencing due to disruption of normal chromatin organization or presence of exogenous DNA. An understanding of local position effects is important for understanding and interpreting any phenotype attributed to targeted gene mutations, or to spontaneous indels

Chlorpyrifos- and chlorpyrifos oxon-induced neurite retraction in pre-differentiated N2a cells is associated with transient hyperphosphorylation of neurofilament heavy chain and ERK 1/2

Chlorpyrifos (CPF) and CPF-oxon (CPO) are known to inhibit neurite outgrowth but little is known about their ability to induce neurite retraction in differentiating neuronal cells. The aims of this study were to determine the ability of these compounds to destabilize neurites and to identify the key molecular events involved. N2a cells were induced to differentiate for 20 h before exposure to CPF or CPO for 2-8 h. Fixed cell monolayers labeled with carboxyfluorescein succinimidyl ester or immunofluorescently stained with antibodies to tubulin (B512) or phosphorylated neurofilament heavy chain (Ta51) showed time- and concentration-dependent reductions in numbers and length of axon-like processes compared to the control, respectively, retraction of neurites being observed within 2 h of exposure by live cell imaging. Neurofilament disruption was also observed in treated cells stained by indirect immunofluorescence with anti- phosphorylated neurofilament heavy chain (NFH) monoclonal antibody SMI34, while the microtubule network was unaffected. Western blotting analysis revealed transiently increased levels of reactivity of Ta51 after 2 h exposure and reduced levels of reactivity of the same antibody following 8 h treatment with both compounds, whereas reactivity with antibodies to anti-total NFH or anti-tubulin was not affected. The alteration in NFH phosphorylation at 2 h exposure was associated with increased activation of extracellular signal-regulated protein kinase ERK 1/2. However, increased levels of phosphatase activity were observed following 8 h exposure. These findings suggest for the first time that organophosphorothionate pesticide-induced neurite retraction in N2a cells is associated with transient increases in NFH phosphorylation and ERK1/2 activation

Muscle size explains low passive skeletal muscle force in heart failure patients.

BACKGROUND: Alterations in skeletal muscle function and architecture have been linked to the compromised exercise capacity characterizing chronic heart failure (CHF). However, how passive skeletal muscle force is affected in CHF is not clear. Understanding passive force characteristics in CHF can help further elucidate the extent to which altered contractile properties and/or architecture might affect muscle and locomotor function. Therefore, the aim of this study was to investigate passive force in a single muscle for which non-invasive measures of muscle size and estimates of fiber force are possible, the soleus (SOL), both in CHF patients and age- and physical activity-matched control participants. METHODS: Passive SOL muscle force and size were obtained by means of a novel approach combining experimental data (dynamometry, electromyography, ultrasound imaging) with a musculoskeletal model. RESULTS: We found reduced passive SOL forces (∼30%) (at the same relative levels of muscle stretch) in CHF vs. healthy individuals. This difference was eliminated when force was normalized by physiological cross sectional area, indicating that reduced force output may be most strongly associated with muscle size. Nevertheless, passive force was significantly higher in CHF at a given absolute muscle length (non length-normalized) and likely explained by the shorter muscle slack lengths and optimal muscle lengths measured in CHF compared to the control participants. This later factor may lead to altered performance of the SOL in functional tasks such gait. DISCUSSION: These findings suggest introducing exercise rehabilitation targeting muscle hypertrophy and, specifically for the calf muscles, exercise that promotes muscle lengthening

A critical role for IRF5 in regulating allergic airway inflammation

Interferon regulatory factor 5 (IRF5) is a key transcription factor involved in the control of the expression of pro-inflammatory cytokine and responses to infection, however its role in regulating pulmonary immune responses to allergen is unknown. We used genetic ablation, adenoviral vector-driven overexpression and adoptive transfer approaches to interrogate the role of IRF5 in pulmonary immunity and during challenge with the aero-allergen, house dust mite. Global IRF5 deficiency resulted in impaired lung function and extracellular matrix (ECM) deposition. IRF5 was also essential for effective responses to inhaled allergen, controlling airway hyper- responsiveness, mucus secretion and eosinophilic inflammation. Adoptive transfer of IRF5- deficient alveolar macrophages into the WT pulmonary milieu was sufficient to drive airway hyper-reactivity, at baseline or following antigen challenge. These data identify IRF5-expressing macrophages as a key component of the immune defence of the airways. Manipulation of IRF5 activity in the lung could therefore be a viable strategy for the redirection of pulmonary immune responses and thus, the treatment of lung disorders

New hope or old futures in disguise? Neoliberalism, the Covid-19 pandemic and the possibility for social change

Purpose – The purpose of this paper is to consider the implications of both the Covid-19 pandemic and UK lockdown for the social, political and economic future of the UK. Drawing on primary data obtained during the lockdown and the theoretical concepts of transcendental materialism and the ‘event’, the paper discusses the strength of participants’ attachment to the ‘old normal’ and their dreams of a ‘new normal’. Design – This paper utilises a semi-structured online survey (n=305) with UK residents and Facebook forum debates collected during the lockdown period in the UK. Findings – The findings in this paper suggest that while the lockdown suspended daily routines and provoked participants to reflect upon their consumption habits and the possibility of an alternative future, many of our respondents remained strongly attached to elements of prelockdown normality. Furthermore, the individual impetus for change was not matched by the structures and mechanisms holding up neoliberalism as governments and commercial enterprises merely encouraged people to get back to the shops to spend. Originality – The original contribution of this paper is the strength and depth of empirical data into the Covid-19 pandemic, specifically the lockdown. Additionally, the synthesis of empirical data with the novel theoretical framework of transcendental materialism presents an original and unique perspective on Covid-19