Genomic deletions and mutations resulting in the loss of eight genes reduce the in vivo replication capacity of Meleagrid herpesvirus 1

Meleagrid herpesvirus 1 (MeHV-1 or turkey herpesvirus) has been widely used as a vaccine in commercial poultry. Initially, these vaccine applications were for the prevention of Marek’s disease resulting from Gallid herpesvirus 2 infections, while more recently MeHV-1 has been used as recombinant vector for other poultry infections. The construction of herpesvirus infectious clones that permit propagation and manipulation of the viral genome in bacterial hosts has advanced the studies of herpesviral genetics. The current study reports the construction of five MeHV-1 infectious clones. The in vitro properties of viruses recovered from these clones were indistinguishable from the parental MeHV-1. In contrast, the rescued MeHV-1 viruses were significantly attenuated when used in vivo. Complete sequencing of the infectious clones identified the absence of two regions of the MeHV-1 genome compared to the MeHV-1 reference sequence. These analyses determined the rescued viruses have seven genes, UL43, UL44, UL45, UL56, HVT071, sorf3 and US2 either partially or completely deleted. In addition, single nucleotide polymorphisms were identified in all clones compared with the MeHV-1 reference sequence. As a consequence of one of the polymorphisms identified in the UL13 gene, four of the rescued viruses were predicted to encode a serine/threonine protein kinase lacking two of three domains required for activity. Thus four of the recovered viruses have a total of eight missing or defective genes. The implications of these findings in the context of herpesvirus biology and infectious clone construction are discussed.

Post-head-emergence frost in wheat and barley: defining the problem, assessing the damage, and identifying resistance

Radiant frost is a significant production constraint to wheat (Triticum aestivum) and barley (Hordeum vulgare), particularly in regions where spring-habit cereals are grown through winter, maturing in spring. However, damage to winter-habit cereals in reproductive stages is also reported. Crops are particularly susceptible to frost once awns or spikes emerge from the protection of the flag leaf sheath. Post-head-emergence frost (PHEF) is a problem distinct from other cold-mediated production constraints. To date, useful increased PHEF resistance in cereals has not been identified. Given the renewed interest in reproductive frost damage in cereals, it is timely to review the problem. Here we update the extent and impacts of PHEF and document current management options to combat this challenge. We clarify terminology useful for discussing PHEF in relation to chilling and other freezing stresses. We discuss problems characterizing radiant frost, the environmental conditions leading to PHEF damage, and the effects of frost at different growth stages. PHEF resistant cultivars would be highly desirable, to both reduce the incidence of direct frost damage and to allow the timing of crop maturity to be managed to maximize yield potential. A framework of potential adaptation mechanisms is outlined. Clarification of these critical issues will sharpen research focus, improving opportunities to identify genetic sources for improved PHEF resistance.

Post-head-emergence frost in wheat and barley: defining the problem, assessing the damage, and identifying resistance

Radiant frost is a significant production constraint to wheat (Triticum aestivum) and barley (Hordeum vulgare), particularly in regions where spring-habit cereals are grown through winter, maturing in spring. However, damage to winter-habit cereals in reproductive stages is also reported. Crops are particularly susceptible to frost once awns or spikes emerge from the protection of the flag leaf sheath. Post-head-emergence frost (PHEF) is a problem distinct from other cold-mediated production constraints. To date, useful increased PHEF resistance in cereals has not been identified. Given the renewed interest in reproductive frost damage in cereals, it is timely to review the problem. Here we update the extent and impacts of PHEF and document current management options to combat this challenge. We clarify terminology useful for discussing PHEF in relation to chilling and other freezing stresses. We discuss problems characterizing radiant frost, the environmental conditions leading to PHEF damage, and the effects of frost at different growth stages. PHEF resistant cultivars would be highly desirable, to both reduce the incidence of direct frost damage and to allow the timing of crop maturity to be managed to maximize yield potential. A framework of potential adaptation mechanisms is outlined. Clarification of these critical issues will sharpen research focus, improving opportunities to identify genetic sources for improved PHEF resistance.

Genomic deletions and mutations resulting in the loss of eight genes reduce the in vivo replication capacity of Meleagrid herpesvirus 1

Meleagrid herpesvirus 1 (MeHV-1 or turkey herpesvirus) has been widely used as a vaccine in commercial poultry. Initially, these vaccine applications were for the prevention of Marek’s disease resulting from Gallid herpesvirus 2 infections, while more recently MeHV-1 has been used as recombinant vector for other poultry infections. The construction of herpesvirus infectious clones that permit propagation and manipulation of the viral genome in bacterial hosts has advanced the studies of herpesviral genetics. The current study reports the construction of five MeHV-1 infectious clones. The in vitro properties of viruses recovered from these clones were indistinguishable from the parental MeHV-1. In contrast, the rescued MeHV-1 viruses were significantly attenuated when used in vivo. Complete sequencing of the infectious clones identified the absence of two regions of the MeHV-1 genome compared to the MeHV-1 reference sequence. These analyses determined the rescued viruses have seven genes, UL43, UL44, UL45, UL56, HVT071, sorf3 and US2 either partially or completely deleted. In addition, single nucleotide polymorphisms were identified in all clones compared with the MeHV-1 reference sequence. As a consequence of one of the polymorphisms identified in the UL13 gene, four of the rescued viruses were predicted to encode a serine/threonine protein kinase lacking two of three domains required for activity. Thus four of the recovered viruses have a total of eight missing or defective genes. The implications of these findings in the context of herpesvirus biology and infectious clone construction are discussed.

Understanding local processes: Contemplating franchisation

Increasing, there is growing acknowledgement of the importance of franchising within all modern global economies. Despite this, little is understood with regards the actual impact of franchising on local economies. This research aims to reframe the contribution of franchising by considering the process of franchisation. This study employed a mixed-method approach, utilizing critical realism to facilitate an outcomes-based explanation of firm survival. The focus of the study was upon generative mechanisms that were assumed to give rise to particular events from which (pizza) firm survival was enhanced vis-à-vis all other community members. A database of 2440 firms (or in excess of 21,000 company years) combined with archival records, interviews and the researcher’s observations provided the researcher with access to the nature of interaction occurring between firms. It was found that the survival of local firms was influenced positively by the day-to-day actions of franchise operators. However, it is argued that to understand how any such advantage my fall to local independent firms, we need too better appreciate the multitude of local processes related to such industries. This research re-examines several ecological concepts with the view of enabling a clearer investigation of underlying local processes. It also represents an authentic autecological approach to the study of firms.

Computational Techniques for Haplotype Inference and for Local Alignment Significance

This thesis which consists of an introduction and four peer-reviewed original publications studies the problems of haplotype inference (haplotyping) and local alignment significance. The problems studied here belong to the broad area of bioinformatics and computational biology. The presented solutions are computationally fast and accurate, which makes them practical in high-throughput sequence data analysis. Haplotype inference is a computational problem where the goal is to estimate haplotypes from a sample of genotypes as accurately as possible. This problem is important as the direct measurement of haplotypes is difficult, whereas the genotypes are easier to quantify. Haplotypes are the key-players when studying for example the genetic causes of diseases. In this thesis, three methods are presented for the haplotype inference problem referred to as HaploParser, HIT, and BACH. HaploParser is based on a combinatorial mosaic model and hierarchical parsing that together mimic recombinations and point-mutations in a biologically plausible way. In this mosaic model, the current population is assumed to be evolved from a small founder population. Thus, the haplotypes of the current population are recombinations of the (implicit) founder haplotypes with some point--mutations. HIT (Haplotype Inference Technique) uses a hidden Markov model for haplotypes and efficient algorithms are presented to learn this model from genotype data. The model structure of HIT is analogous to the mosaic model of HaploParser with founder haplotypes. Therefore, it can be seen as a probabilistic model of recombinations and point-mutations. BACH (Bayesian Context-based Haplotyping) utilizes a context tree weighting algorithm to efficiently sum over all variable-length Markov chains to evaluate the posterior probability of a haplotype configuration. Algorithms are presented that find haplotype configurations with high posterior probability. BACH is the most accurate method presented in this thesis and has comparable performance to the best available software for haplotype inference. Local alignment significance is a computational problem where one is interested in whether the local similarities in two sequences are due to the fact that the sequences are related or just by chance. Similarity of sequences is measured by their best local alignment score and from that, a p-value is computed. This p-value is the probability of picking two sequences from the null model that have as good or better best local alignment score. Local alignment significance is used routinely for example in homology searches. In this thesis, a general framework is sketched that allows one to compute a tight upper bound for the p-value of a local pairwise alignment score. Unlike the previous methods, the presented framework is not affeced by so-called edge-effects and can handle gaps (deletions and insertions) without troublesome sampling and curve fitting.

Optimisation Problems in Wireless Sensor Networks : Local Algorithms and Local Graphs

This thesis studies optimisation problems related to modern large-scale distributed systems, such as wireless sensor networks and wireless ad-hoc networks. The concrete tasks that we use as motivating examples are the following: (i) maximising the lifetime of a battery-powered wireless sensor network, (ii) maximising the capacity of a wireless communication network, and (iii) minimising the number of sensors in a surveillance application. A sensor node consumes energy both when it is transmitting or forwarding data, and when it is performing measurements. Hence task (i), lifetime maximisation, can be approached from two different perspectives. First, we can seek for optimal data flows that make the most out of the energy resources available in the network; such optimisation problems are examples of so-called max-min linear programs. Second, we can conserve energy by putting redundant sensors into sleep mode; we arrive at the sleep scheduling problem, in which the objective is to find an optimal schedule that determines when each sensor node is asleep and when it is awake. In a wireless network simultaneous radio transmissions may interfere with each other. Task (ii), capacity maximisation, therefore gives rise to another scheduling problem, the activity scheduling problem, in which the objective is to find a minimum-length conflict-free schedule that satisfies the data transmission requirements of all wireless communication links. Task (iii), minimising the number of sensors, is related to the classical graph problem of finding a minimum dominating set. However, if we are not only interested in detecting an intruder but also locating the intruder, it is not sufficient to solve the dominating set problem; formulations such as minimum-size identifying codes and locating dominating codes are more appropriate. This thesis presents approximation algorithms for each of these optimisation problems, i.e., for max-min linear programs, sleep scheduling, activity scheduling, identifying codes, and locating dominating codes. Two complementary approaches are taken. The main focus is on local algorithms, which are constant-time distributed algorithms. The contributions include local approximation algorithms for max-min linear programs, sleep scheduling, and activity scheduling. In the case of max-min linear programs, tight upper and lower bounds are proved for the best possible approximation ratio that can be achieved by any local algorithm. The second approach is the study of centralised polynomial-time algorithms in local graphs these are geometric graphs whose structure exhibits spatial locality. Among other contributions, it is shown that while identifying codes and locating dominating codes are hard to approximate in general graphs, they admit a polynomial-time approximation scheme in local graphs.

Managing reforestation to sequester carbon, increase biodiversity potential and minimize loss of agricultural land

Reforestation will have important consequences for the global challenges of mitigating climate change, arresting habitat decline and ensuring food security. We examined field-scale trade-offs between carbon sequestration of tree plantings and biodiversity potential and loss of agricultural land. Extensive surveys of reforestation across temperate and tropical Australia (N = 1491 plantings) were used to determine how planting width and species mix affect carbon sequestration during early development (< 15 year). Carbon accumulation per area increased significantly with decreasing planting width and with increasing proportion of eucalypts (the predominant over-storey genus). Highest biodiversity potential was achieved through block plantings (width > 40 m) with about 25% of planted individuals being eucalypts. Carbon and biodiversity goals were balanced in mixed-species plantings by establishing narrow belts (width < 20 m) with a high proportion (>75%) of eucalypts, and in monocultures of mallee eucalypt plantings by using the widest belts (ca. 6–20 m). Impacts on agriculture were minimized by planting narrow belts (ca. 4 m) of mallee eucalypt monocultures, which had the highest carbon sequestering efficiency. A plausible scenario where only 5% of highly-cleared areas (<30% native vegetation cover remaining) of temperate Australia are reforested showed substantial mitigation potential. Total carbon sequestration after 15 years was up to 25 Mt CO2-e year−1 when carbon and biodiversity goals were balanced and 13 Mt CO2-e year−1 if block plantings of highest biodiversity potential were established. Even when reforestation was restricted to marginal agricultural land (<$2000 ha−1 land value, 28% of the land under agriculture in Australia), total mitigation potential after 15 years was 17–26 Mt CO2-e year−1 using narrow belts of mallee plantings. This work provides guidance on land use to governments and planners. We show that the multiple benefits of young tree plantings can be balanced by manipulating planting width and species choice at establishment. In highly-cleared areas, such plantings can sequester substantial biomass carbon while improving biodiversity and causing negligible loss of agricultural land.

New yellow head virus genotype (YHV7) in giant tiger shrimp Penaeus monodon indigenous to northern Australia

ABSTRACT: In 2012, giant tiger shrimp Penaeus monodon originally sourced from Joseph Bonaparte Gulf in northern Australia were examined in an attempt to identify the cause of elevated mortalities among broodstock at a Queensland hatchery. Nucleic acid extracted from ethanol-fixed gills of 3 individual shrimp tested positive using the OIE YHV Protocol 2 RT-PCR designed to differentiate yellow head virus (YHV1) from gill-associated virus (GAV, synonymous with YHV2) and the OIE YHV Protocol 3 RT-nested PCR designed for consensus detection of YHV genotypes. Sequence analysis of the 794 bp (Protocol 2) and 359 bp (Protocol 3) amplicons from 2 distinct regions of ORF1b showed that the yellow-head-complex virus detected was novel when compared with Genotypes 1 to 6. Nucleotide identity on the Protocol 2 and Protocol 3 ORF1b sequences was highest with the highly pathogenic YHV1 genotype (81 and 87%, respectively) that emerged in P. monodon in Thailand and lower with GAV (78 and 82%, respectively) that is enzootic to P. monodon inhabiting eastern Australia. Comparison of a longer (725 bp) ORF1b sequence, spanning the Protocol 3 region and amplified using a modified YH30/31 RT-nPCR, provided further phylogenetic evidence for the virus being distinct from the 6 described YHV genotypes. The virus represents a unique seventh YHV genotype (YHV7). Despite the mortalities observed, the role of YHV7 remains unknown.

Head motion and Inattention/Hyperactivity share common genetic influences: Implications for fMRI studies of ADHD

Head motion (HM) is a well known confound in analyses of functional MRI (fMRI) data. Neuroimaging researchers therefore typically treat HM as a nuisance covariate in their analyses. Even so, it is possible that HM shares a common genetic influence with the trait of interest. Here we investigate the extent to which this relationship is due to shared genetic factors, using HM extracted from resting-state fMRI and maternal and self report measures of Inattention and Hyperactivity-Impulsivity from the Strengths and Weaknesses of ADHD Symptoms and Normal Behaviour (SWAN) scales. Our sample consisted of healthy young adult twins (N = 627 (63% females) including 95 MZ and 144 DZ twin pairs, mean age 22, who had mother-reported SWAN; N = 725 (58% females) including 101 MZ and 156 DZ pairs, mean age 25, with self reported SWAN). This design enabled us to distinguish genetic from environmental factors in the association between head movement and ADHD scales. HM was moderately correlated with maternal reports of Inattention (r = 0.17, p-value = 7.4E-5) and Hyperactivity-Impulsivity (r = 0.16, p-value = 2.9E-4), and these associations were mainly due to pleiotropic genetic factors with genetic correlations [95% CIs] of rg = 0.24 [0.02, 0.43] and rg = 0.23 [0.07, 0.39]. Correlations between self-reports and HM were not significant, due largely to increased measurement error. These results indicate that treating HM as a nuisance covariate in neuroimaging studies of ADHD will likely reduce power to detect between-group effects, as the implicit assumption of independence between HM and Inattention or Hyperactivity-Impulsivity is not warranted. The implications of this finding are problematic for fMRI studies of ADHD, as failing to apply HM correction is known to increase the likelihood of false positives. We discuss two ways to circumvent this problem: censoring the motion contaminated frames of the RS-fMRI scan or explicitly modeling the relationship between HM and Inattention or Hyperactivity-Impulsivity

Development of a transient gradient enhanced non local continuum damage mechanics model for masonry

Due to the advent of varied types of masonry systems a comprehensive failure mechanism of masonry essential for the understanding of its behaviour is impossible to be determined from experimental testing. As masonry is predominantly used in wall structures a biaxial stress state dominates its failure mechanism. Biaxial testing will therefore be necessary for each type of masonry, which is expensive and time consuming. A computational method would be advantageous; however masonry is complex to model which requires advanced computational modelling methods. This thesis has formulated a damage mechanics inspired modelling method and has shown that the method effectively determines the failure mechanisms and deformation characteristics of masonry under biaxial states of loading.

Inhibition of Human Two-Pore Domain K+ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

TWIK-related K+ channel TREK1, a background leak K+ channel, has been strongly implicated as the target of several general and local anesthetics. Here, using the whole-cell and single-channel patch-clamp technique, we investigated the effect of lidocaine, a local anesthetic, on the human (h) TREK1 channel heterologously expressed in human embryonic kidney 293 cells by an adenoviral-mediated expression system. Lidocaine, at clinical concentrations, produced reversible, concentration-dependent inhibition of hTREK1 current, with IC50 value of 180 mu M, by reducing the single-channel open probability and stabilizing the closed state. We have identified a strategically placed unique aromatic couplet (Tyr352 and Phe355) in the vicinity of the protein kinase A phosphorylation site, Ser348, in the C-terminal domain (CTD) of hTREK1, that is critical for the action of lidocaine. Furthermore, the phosphorylation state of Ser348 was found to have a regulatory role in lidocaine-mediated inhibition of hTREK1. It is interesting that we observed strong intersubunit negative cooperativity (Hill coefficient = 0.49) and half-of-sites saturation binding stoichiometry (half-reaction order) for the binding of lidocaine to hTREK1. Studies with the heterodimer of wild-type (wt)-hTREK1 and Delta 119 C-terminal deletion mutant (hTREK1(wt)-Delta 119) revealed that single CTD of hTREK1 was capable of mediating partial inhibition by lidocaine, but complete inhibition necessitates the cooperative interaction between both the CTDs upon binding of lidocaine. Based on our observations, we propose a model that explains the unique kinetics and provides a plausible paradigm for the inhibitory action of lidocaine on hTREK1.

Chapter 1: Precarious creativity: Global media, local labor

In most parts of the world, screen media workers—actors, directors, gaffers, and makeup artists—consider Hollywood to be glamorous and aspirational. If given the opportunity to work on a major studio lot, many would make the move, believing the standards of professionalism are high and the history of accomplishment is renowned. Moreover, as a global leader, Hollywood offers the chance to rub shoulders with talented counterparts and network with an elite labor force that earns top-tier pay and benefits. Yet despite this reputation, veterans say the view from inside isn’t so rosy, that working conditions have been deteriorating since the 1990s if not earlier. This grim outlook is supported by industry statistics that show the number of good jobs has been shrinking as studios outsource production to Atlanta, London, and Budapest, among others...