An investigation into the molecular determinants of salmon louse (Lepeophtheirus salmonis (Krøyer, 1837)) susceptibility to the antiparasitic drug emamectin benzoate.

Caligid copepods, also called sea lice, are ectoparasites of marine fish, with Lepeophtheirus salmonis (Krøyer, 1837) emerging as a problem for mariculture of Atlantic salmon (Salmo salar Linnaeus, 1758) in the northern hemisphere. Annual costs of sea lice to global salmon farming was estimated to be in excess of €300 million in 2006, with the majority of this accounted for through expenses accrued from chemical treatments. Only a limited range of anti-sea louse drugs are available and licensed for the treatment of fish, and the continued use of only a few compounds creates a situation potentially favouring the development of drug resistance. Emamectin benzoate (EMB) is currently used as a salmon delousing agent, being employed as a 0.2 % in-feed pre-mix (SLICE®). Atlantic salmon farmers have reported increased incidence of reduced L. salmonis sensitivity to SLICE®, which has highlighted the requirement for further research into the molecular mechanisms controlling salmon louse resistance to EMB. Genomic and transcriptomic research concerning L. salmonis drug resistance mechanisms has not often been reported, with previous transcriptomic studies using candidate gene approaches and genetic studies focussing on population genetics. Drug resistance in ecdysozoan invertebrates is associated with a variety of molecular mechanisms including target site mutations and changes in the expression of components in drug detoxification pathways. The research reported in this thesis was aimed at the exploration of mechanisms employed by L. salmonis to reduce the toxicity of EMB exposure, following a transcriptomic approach that utilised custom oligonucleotide (oligo) microarrays and a genetic approach that utilised Restriction-site associated DNA sequencing (RAD-seq) to identify Single Nucleotide Polymorphism (SNP) markers. An EMB-resistant (PT) and drug-susceptible (S) L. salmonis laboratory-maintained strain were to be used as a model for this research, as these two strains differ in EMB susceptibility (~ 7-fold) and show stable susceptibility profiles through multiple generations, suggesting that this drug resistance phenotype may be a heritable trait. Sequence resources available for salmon lice are limited as an annotated L. salmonis genome is currently under construction. Therefore, a significant amount of this study involved creating new resources to facilitate the analysis of EMB susceptibility. Suppression subtractive hybridisation (SSH) was used to enrich for transcripts that were differentially expressed between strains PT and S, which provided sufficient target sequence for the development of 15K oligo microarrays when combined with sequences assembled from existing L. salmonis ESTs. Additionally, transcripts were generated through sequencing a pooled sample representing key developmental stages of the L. salmonis life cycle, which were later used in the construction of a 44K oligo microarray. The toxicity of EMB and other avermectins (AVMs) against ecdysozoan invertebrates is reported to be based mainly on their interaction with ligand-gated ion channels (LGIC), specifically glutamate-gated chloride channels (GluCl). However, -aminobutyric acid (GABA)-gated chloride channels (GABA-Cls) are also believed to be targeted by AVMs and neuronal acetylcholine receptors (nAChRs) can be allosterically modulated by the AVM compound ivermectin. Transcriptional responses in PT and S salmon lice were investigated using custom 15K L. salmonis oligo microarrays. In the absence of EMB exposure, 359 targets differed in transcript abundance between the two strains. GABA-Cl and nAChR subunits showed significantly lower transcript levels in PT compared to S lice, which was estimated at ~1.4-fold for GABA-Cl and ~2.8-fold for nAChR using RT-qPCR, suggesting their involvement in AVM toxicity in caligids. Although, salmon lice from the PT strain showed few transcriptional responses following acute exposure (1 or 3 h) to 200 µg L-1 of EMB, a drug concentration tolerated by PT lice, but toxic for S lice. RAD-seq analysis of both genders from L. salmonis strains S and PT identified 15 RAD-markers that show complete association with salmon louse strain, although these preliminary results will need further analysis to confirm marker association with reduced EMB susceptibility. Additionally, RAD marker Lsa101901 showed complete association with sex for all individuals analysed, being heterozygous in females and homozygous in males. Using an allele-specific PCR assay, this SNP association pattern was further confirmed for three unrelated salmon louse strains. Marker Lsa101901 was located in the coding region of the prohibitin-2 gene, which showed a sex-dependent differential expression, with mRNA levels determined by RT-qPCR about 1.8-fold higher in adult female than adult male salmon lice. In conclusion, the identification of decreased transcript abundances for LGIC subunits in EMB-resistant salmon lice, and polymorphic SNP markers showing complete association with L. salmonis strains S or PT, provides suitable candidates for further investigation into their association with reduced EMB susceptibility. Further analysis will also be required to confirm whether EMB-induced mechanisms are not associated with reduced EMB susceptibility in L. salmonis. Additionally, the identification of sex-linked SNP Lsa101901 suggests that sex determination in the salmon louse is genetic and follows a female heterozygous system, with marker Lsa101901 providing a tool to determine the genetic sex of salmon lice. Improved knowledge of L. salmonis biology and the mechanisms potentially involved in EMB resistance, obtained during this study, may provide molecular markers that contribute to successful monitoring and management of this commercially important parasite of Atlantic salmon

Developing the Surface Chemistry of Transparent Butyl Rubber for Impermeable Stretchable Electronics

Transparent butyl rubber is a new elastomer that has the potential to revolutionize stretchable electronics due to its intrinsically low gas permeability. Encapsulating organic electronic materials and devices with transparent butyl rubber protects them from problematic degradation due to oxygen and moisture, preventing premature device failure and enabling the fabrication of stretchable organic electronic devices with practical lifetimes. Here, we report a methodology to alter the surface chemistry of transparent butyl rubber to advance this material from acting as a simple device encapsulant to functioning as a substrate primed for direct device fabrication on its surface. We demonstrate a combination of plasma and chemical treatment to deposit a hydrophilic silicate layer on the transparent butyl rubber surface to create a new layered composite that combines Si-OH surface chemistry with the favorable gas-barrier properties of bulk transparent butyl rubber. We demonstrate that these surface Si-OH groups react with organosilanes to form self-assembled monolayers necessary for the deposition of electronic materials, and furthermore demonstrate the fabrication of stretchable gold wires using nanotransfer printing of gold films onto transparent butyl rubber modified with a thiol-terminated self-assembled monolayer. The surface modification of transparent butyl rubber establishes this material as an important new elastomer for stretchable electronics and opens the way to robust, stretchable devices

Metal-Interface-Elastomer (MINE) Structures for Stretchable Electronics

The future of soft, conformable, and robust wearable electronics will require elastomers to provide mechanical stabilization, a soft surface to interact with human wearers, and a crucial physical barrier to protect stretchable devices from the environment. It is a difficult challenge, however, for a single elastomer to fulfill each of these needs. Here, we present a new approach that fuses a membrane of poly(dimethylsiloxane) (PDMS) onto the surface of a transparent butyl rubber (T-IIR) substrate using an organosilane-based molecular glue. The resulting membrane-interface-elastomer (MINE) structures uniquely combine the surface chemistry of PDMS with the intrinsically low gas permeability of T-IIR for the fabrication of robust stretchable devices. Our most intriguing finding, however, is that the T-IIR-PDMS interface, buried microns below the PDMS surface, exerts a remarkable influence on metal films deposited on the PDMS membrane surface from below, improving stretching and conductance performance by orders of magnitude

Stretchable Ultrasheer Fabrics as Semitransparent Electrodes for Wearable Light-Emitting e-Textiles with Changeable Display Patterns

Despite the development throughout human history of a wealth of textile materials and structures, the porous structures and non-planar surfaces of textiles are often viewed as problematic for the fabrication of wearable e-textiles and smart clothing. Here, we demonstrate a new textile-centric design paradigm in which we use the textile structure as an integral part of wearable device design. We coat the open framework structure of an ultrasheer knitted textile with a conformal gold film using solution-based metallization to form gold-coated ultrasheer electrodes that are highly conductive (3.6 ± 0.9 Ω/sq) and retain conductivity to 200% strain with R/R0 \u3c 2. The ultrasheer electrodes produce wearable, highly stretchable light-emitting e-textiles that function to 200% strain. Stencil printing a wax resist provides patterned electrodes for patterned light emission; furthermore, incorporating soft-contact lamination produces light-emitting textiles that exhibit, for the first time, readily changeable patterns of illumination

Avian adrenal medulla : cytomorphology and function

The purpose of this review is to explore the world literature on the avian adrenal medulla from the last 20 years. Unlike the mammalian adrenal medulla, the adrenal gland in birds has chromaffin cells mixed with cortical cells. Studies have investigated the ultrastructure (both transmission and scanning electron microscopy), biochemistry, and physiology (partic-ularly interactions with other endocrine glands) of the avian adrenal medulla. Although progress has been made, it is apparent that research on the avian adrenal medulla still lags behind work on the mammalian organ

Avian adrenal medulla: cytomorphology and function

The purpose of this review is to explore the world literature on the avian adrenal medulla from the last 20 years. Unlike the mammalian adrenal medulla, the adrenal gland in birds has chromaffin cells mixed with cortical cells. Studies have investigated the ultrastructure (both transmission and scanning electron microscopy), biochemistry, and physiology (particularly interactions with other endocrine glands) of the avian adrenal medulla. Although progress has been made, it is apparent that research on the avian adrenal medulla still lags behind work on the mammalian organ

The Bristol Health and Planning Protocol - First year evaluation

The report documents the evaluation of the first year of ‘the Bristol Protocol’. This is an agreement between NHS Bristol and Bristol City Council to receive public health consultations for selected planning applications and preapplication processes. The purpose was to support the consideration of the impact on health outcomes in the assessment and determination of planning applications. Based on audit of planning applications referred to NHS Bristol, surveys of Bristol City Council’s planners, interviews and documentary analysis, this evaluation assesses how and to what extent public health colleagues inBristol have been able to influence decisions on planning applications. The analysis also helps to determine whether there have been changes in knowledge and attitudes of development management planners about the links between health and planning. The research covered the period 17 May 2011 to 31 August 2012

Transparent, Stretchable, and Conductive SWNT Films Using Supramolecular Functionalization and Layer-by-Layer Self-Assembly

We demonstrate films of single-walled carbon nanotubes (SWNTs) on the elastomer polydimethylsiloxane (PDMS) that are stretchable, conductive, and transparent. Our fabrication method uses the supramolecular functionalization of SWNTs with conjugated polyelectrolytes to generate aqueous dispersions of positively- and negatively-charged SWNTs, followed by layer-by-layer self-assembly onto a PDMS substrate. Adding bilayers of positively- and negatively-charged SWNTs to the surface causes the sheet resistance and the % transmittance of the film to both progressively decrease. The sheet resistance decreases sharply in the first five bilayers as the layer-by-layer process efficiently establishes the percolation network, whereas the % transmittance declines more gradually. Films with 25 bilayers are transparent (75% at 550 nm) and conductive (560 ± 90 ohms/sq). The combination of electrostatic and pi-stacking forces very effectively bind the SWNTs within the film, producing smooth film surfaces (root-mean-square roughness of 18 nm) and enabling the films to remain conductive up to 80% elongation. We demonstrate the use of the SWNT films as transparent conductive electrodes in light-emitting devices and as soft strain sensors that are both wearable and transparent