Masters of Science / A nitrile and amide hydrolysing Nesterenkonia sp. was isolated from Antarctic soil and was characterised as a psychrotolerant, halotolerant and alkaliphilic extremophile. Amidases are widely distributed in both prokaryotic and eukaryotic organisms. These enzymes hydrolyze C-N bonds other than peptide bonds and are particularly interesting for their potential industrial application. This study aimed to identify and characterize amidase genes from this novel psychrotolerant microorganism. Using BLAST analysis, two ORFs with conserved amidase sequences were identified from the complete genome sequence of the organism. Two ORFs, AmiF and AmiS, were assigned to two different gene families, the aceta/formamidase family and amidase signature family, respectively. On the genome, the spatial orientation and intergenic distance (1bp overlap) of the ORF‟s suggested that amiF and amiS could possibly be cotranscribed which was confirmed by reverse transcription PCR. A third ORF with a conserved amidase sequence was found ±500bps downstream from amiS, suggesting the possible presence of a multi amidase operon. The two genes were cloned and expressed as N-terminal 6x His-Tag fusion proteins. AmiS and Ami F were partially purified using Ni-chelation chromatography. Although both proteins were subjected to activity assay, their activities are yet to be established. Homology modeling of the AmiF and AmiS translated sequences showed that the proteins had the significant similarities to the members of their families. Although the sequence identities between the AmiF and AmiS and their templates were very low (24 % and 25% respectively), the evaluation of the models showed that the quality of the models were good. This study reports the genetic and functional characterisation of amidase genes from the cold adapted microorganisms. / South Africa

Flyrock remains a significant risk to the health and safety of the mine’s employees and infrastructure as well as the safety of the neighbouring communities and their property. Losses and damages can result in significant financial and reputation consequences. The lack of fundamental research in recent years and quantifiable data relating to the relationship between blast design parameters and the risk of flyrock motivated this project. A number of authors concluded that major gaps in knowledge relative to flyrock caused by its random nature still remain a weakness in the field. Recent papers published (since 2010) proposed a wide range of potential approaches and techniques to predict or investigate flyrock. However, the majority of these papers concluded that the proposed results were site-specific and could not be applied to other environments. The focus of this project was to develop a concept that is able to quantify the flight path of the flyrock resulting from a blast. The motivation behind the development of this concept was to enable future researchers to quantify the impact of the different blast design parameters on the measured flyrock. Various technologies were considered and investigated during this project. After a comparative analysis of these technologies, it was decided to use photogrammetry as the foundation of the proposed concept tool. The proposed concept consists of three main phases, namely (1) data acquisition, (2) image processing and data analysis and (3) data interpretation. To date, progress has been achieved with phase one and phase two. In phase one, all objectives have been met. However, there are still areas which need refinement, specifically regarding the placement of the cameras in the field. In phase two, success was achieved with the proof of concept exercise in a controlled environment using a clay pigeon as the projectile. The process of calibrating the lenses has been established, however, further optimization is possible. Point-cloud data was successfully generated in the concept test, but converting the image data from subsequent quarry test blasts proved more challenging and is still a work in progress. Once phase two has been satisfactorily resolved, attention will focus on phase three. Results to date have given a positive indication that the concept is viable and that additional work will prove the technology functional. Ultimately, it is envisioned that this tool can be used for one of three purposes, namely: • Mines can generate a database with accurate historical flyrock of their blasting operations. • Research teams can implement this tool to conduct quantitative research and investigations into flyrock and the impact of different blast design parameters on the risk of flyrock. • Point-cloud data combined with ballistics calculations can be used to visualise blasts and flyrock in Virtual Reality for training and education. / Dissertation (MEng)--University of Pretoria, 2019. / AEL Mining Services Chair in Innovative Rock Breaking / Mining Engineering / MEng / Unrestricted

The MSU Loci/BLAST CFD code was used to study blast wave interactions with structures for different urban environments. A series of analyses which included single building structures inside of ERDC's Blast Load Simulator (BLS) with different obliquity orientations to the flow direction, two building structures inside the BLS with varying gap distances between the structures, and open air blast simulations with four structure scenarios at different building spacings and different blast orientations were performed. Unstructured and overset grid techniques were used during the modeling process and were compared for consistency with shock physics and computational performance. Results show Loci/BLAST's capability to accurately model blast wave interactions in urban environments for both unstructured and overset grids.

<p>Bacteriophages are viruses that infect and kill bacteria. They are the most abundant organism on the planet and the largest source of untapped genetic information. Every year, more bacteriophages are isolated from the environment, purified, and sequenced. Once sequenced, their genomes are annotated to determine the location and putative function of each gene expressed by the phage. Phages have been used in the past for genetic engineering and new research is being done into how they can be used for the treatment of disease, water safety, agriculture, and food safety. </p> <p>Despite the influx of sequenced bacteriophages, a majority of the genes annotated are hypothetical proteins, also known as No Known Function (NKF) proteins. They are expressed by the phages, but research has not identified a possible function. Wet lab research into the functions of the hundreds of NKF phages genes would be costly and could take years. Bioinformatics methods could be used to determine putative functions and functional categories for these hypothetical proteins. A new bioinformatics method using algorithms such as Domain Assignments, Hidden Markov Models, Structure Prediction, Sub-Cellular Localization, and iterative algorithms is proposed here. This new method was tested on the bacteriophage genome PotatoSplit and dropped the number of NKF genes from 57 to 40. A total of 17 new functions were found. The functional class was identified for an additional six proteins, though no specific functions were named. Structure Prediction and Simulations were tested with a focus on two NKF proteins within lytic phages and both returned possible functional categories with high confidence.</p> <p>Additionally, this research focuses on the possibility of phage therapy and FDA regulation. A database of phage proteins was built and tested using R Statistical Analysis to determine proteins significant to phage infecting <i>M. tuberculosis</i> and to the lytic cycle of phages. The statistical methods were also tested on both pharmaceutical products recalled by the FDA between 2012 and 2018 to determine ingredients/manufacturing steps that could affect product quality and on the FDA Adverse Event Reporting System (FAERS) data to determine if AERs could be used to judge the quality of a product. Many significant excipients/manufacturing steps were identified and used to score products on their quality. The AERs were evaluated on two case studies with mixed results. </p>