The application of next generation sequencing to profile microbe related cheese quality defects

High throughput next generation sequencing, together with advanced molecular methods, has considerably enhanced the field of food microbiology. By overcoming biases associated with culture dependant approaches, it has become possible to achieve novel insights into the nature of food-borne microbial communities. In this thesis, several different sequencing-based approaches were applied with a view to better understanding microbe associated quality defects in cheese. Initially, a literature review provides an overview of microbe-associated cheese quality defects as well as molecular methods for profiling complex microbial communities. Following this, 16S rRNA sequencing revealed temporal and spatial differences in microbial composition due to the time during the production day that specific commercial cheeses were manufactured. A novel Ion PGM sequencing approach, focusing on decarboxylase genes rather than 16S rRNA genes, was then successfully employed to profile the biogenic amine producing cohort of a series of artisanal cheeses. Investigations into the phenomenon of cheese pinking formed the basis of a joint 16S rRNA and whole genome shotgun sequencing approach, leading to the identification of Thermus species and, more specifically, the pathway involved in production of lycopene, a red coloured carotenoid. Finally, using a more traditional approach, the effect of addition of a facultatively heterofermentative Lactobacillus (Lactobacillus casei) to a Swiss-type cheese, in which starter activity was compromised, was investigated from the perspective of its ability to promote gas defects and irregular eye formation. X-ray computed tomography was used to visualise, using a non-destructive method, the consequences of the undesirable gas formation that resulted. Ultimately this thesis has demonstrated that the application of molecular techniques, such as next generation sequencing, can provide a detailed insight into defect-causing microbial populations present and thereby may underpin approaches to optimise the quality and consistency of a wide variety of cheeses.

Semantic markup in discussion fora

Discussion tools in existing LEs have few or no integrated tools to analyse student learning. This paper proposes tools not only for integrating social network analytics, but also why we need to semantically tag and track key concepts within posts in order to make student learning in discussions visible. This paper will argue for the importance of semantic markup in discussion tools using screenshots of existing LEs and UI mockups of semantically aware discussion tools to argue the case for this element of next generation LEs

Design, fabrication and characterization of resonant waveguide grating based optical biosensors

The absence of rapid, low cost and highly sensitive biodetection platform has hindered the implementation of next generation cheap and early stage clinical or home based point-of-care diagnostics. Label-free optical biosensing with high sensitivity, throughput, compactness, and low cost, plays an important role to resolve these diagnostic challenges and pushes the detection limit down to single molecule. Optical nanostructures, specifically the resonant waveguide grating (RWG) and nano-ribbon cavity based biodetection are promising in this context. The main element of this dissertation is design, fabrication and characterization of RWG sensors for different spectral regions (e.g. visible, near infrared) for use in label-free optical biosensing and also to explore different RWG parameters to maximize sensitivity and increase detection accuracy. Design and fabrication of the waveguide embedded resonant nano-cavity are also studied. Multi-parametric analyses were done using customized optical simulator to understand the operational principle of these sensors and more important the relationship between the physical design parameters and sensor sensitivities. Silicon nitride (SixNy) is a useful waveguide material because of its wide transparency across the whole infrared, visible and part of UV spectrum, and comparatively higher refractive index than glass substrate. SixNy based RWGs on glass substrate are designed and fabricated applying both electron beam lithography and low cost nano-imprint lithography techniques. A Chromium hard mask aided nano-fabrication technique is developed for making very high aspect ratio optical nano-structure on glass substrate. An aspect ratio of 10 for very narrow (~60 nm wide) grating lines is achieved which is the highest presented so far. The fabricated RWG sensors are characterized for both bulk (183.3 nm/RIU) and surface sensitivity (0.21nm/nm-layer), and then used for successful detection of Immunoglobulin-G (IgG) antibodies and antigen (~1μg/ml) both in buffer and serum. Widely used optical biosensors like surface plasmon resonance and optical microcavities are limited in the separation of bulk response from the surface binding events which is crucial for ultralow biosensing application with thermal or other perturbations. A RWG based dual resonance approach is proposed and verified by controlled experiments for separating the response of bulk and surface sensitivity. The dual resonance approach gives sensitivity ratio of 9.4 whereas the competitive polarization based approach can offer only 2.5. The improved performance of the dual resonance approach would help reducing probability of false reading in precise bio-assay experiments where thermal variations are probable like portable diagnostics.

Modifying the band gap and optical properties of Germanium nanowires by surface termination

Semiconductor nanowires, based on silicon (Si) or germanium (Ge) are leading candidates for many ICT applications, including next generation transistors, optoelectronics, gas and biosensing and photovoltaics. Key to these applications is the possibility to tune the band gap by changing the diameter of the nanowire. Ge nanowires of different diameter have been studied with H termination, but, using ideas from chemistry, changing the surface terminating group can be used to modulate the band gap. In this paper we apply the generalised gradient approximation of density functional theory (GGA-DFT) and hybrid DFT to study the effect of diameter and surface termination using –H, –NH2 and –OH groups on the band gap of (001), (110) and (111) oriented germanium nanowires. We show that the surface terminating group allows both the magnitude and the nature of the band gap to be changed. We further show that the absorption edge shifts to longer wavelength with the –NH2 and –OH terminations compared to the –H termination and we trace the origin of this effect to valence band modifications upon modifying the nanowire with –NH2 or –OH. These results show that it is possible to tune the band gap of small diameter Ge nanowires over a range of ca. 1.1 eV by simple surface chemistry.