Molecular biology techniques as a tool for detection and characterisation of Mycobacterium avium subsp. paratuberculosis

Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) is the causative agent of paratuberculosis, also known as Johne’s disease, a chronic intestinal infection in cattle and other ruminants. Paratuberculosis is characterised by diarrhea and weight loss that occurs after a period of a few months up to several years without any clinical signs. The considerable economic losses to dairy and beef cattle producers are caused by reduced milk production and poor reproduction performance in subclinically infected animals. Early diagnosis of infected cattle is essential to prevent the spread of the disease. Efforts have been made to eradicate paratuberculosis by using a detection and cull strategy, but eradication is hampered by the lack of suitable and sensitive diagnostic methods. This thesis, based on five scientific investigations, describes the development of different DNA amplification strategies for detection and characterisation of M. paratuberculosis. Various ways to pre-treat bacterial cultures, tissue specimens and fecal samples prior to PCR analysis were investigated. Internal positive PCR control molecules were developed and used in PCR analyses to improve the reliability and to facilitate the interpretation of the results. The sensitivity of the ultimate methods was found to be approximate that of culture and allowed detection of low numbers of M. paratuberculosis expected to be found in subclinically infected animals. Genomic DNA of a Swedish mycobacterial isolate, incorrectly identified by PCR as M. paratuberculosis was characterised. The isolate was closely related to M. cookii and harboured one copy of a DNA segment with 94% similarity to IS900, the target sequence used in diagnostic PCR for detection of M. paratuberculosis. This finding highlighted the urgency of developing or evaluating PCR systems based on genes other than IS900. A PCR-based fingerprinting method using primers targeting the enterobacterial intergenic consensus sequence (ERIC) and the IS900 sequence was developed and successfully used to distinguish M. paratuberculosis from closely related mycobacteria, including the above mentioned mycobacterial isolate. In conclusion, the molecular biology techniques developed in these studies have proved useful for accelerating the diagnostic detection and characterisation of M. paratuberculosis

Defining bacterial species in the genomic era : insights from the genus Acinetobacter

Background: Microbial taxonomy remains a conservative discipline, relying on phenotypic information derived from growth in pure culture and techniques that are time-consuming and difficult to standardize, particularly when compared to the ease of modern high-throughput genome sequencing. Here, drawing on the genus Acinetobacter as a test case, we examine whether bacterial taxonomy could abandon phenotypic approaches and DNA-DNA hybridization and, instead, rely exclusively on analyses of genome sequence data. Results: In pursuit of this goal, we generated a set of thirteen new draft genome sequences, representing ten species, combined them with other publically available genome sequences and analyzed these 38 strains belonging to the genus. We found that analyses based on 16S rRNA gene sequences were not capable of delineating accepted species. However, a core genome phylogenetic tree proved consistent with the currently accepted taxonomy of the genus, while also identifying three misclassifications of strains in collections or databases. Among rapid distance-based methods, we found average-nucleotide identity (ANI) analyses delivered results consistent with traditional and phylogenetic classifications, whereas gene content based approaches appear to be too strongly influenced by the effects of horizontal gene transfer to agree with previously accepted species. Conclusion: We believe a combination of core genome phylogenetic analysis and ANI provides an appropriate method for bacterial species delineation, whereby bacterial species are defined as monophyletic groups of isolates with genomes that exhibit at least 95% pair-wise ANI. The proposed method is backwards compatible; it provides a scalable and uniform approach that works for both culturable and non-culturable species; is faster and cheaper than traditional taxonomic methods; is easily replicable and transferable among research institutions; and lastly, falls in line with Darwin’s vision of classification becoming, as far as is possible, genealogical

OntoMathPROOntoMath^{PRO} Ontology: A Linked Data Hub for Mathematics

In this paper, we present an ontology of mathematical knowledge concepts that covers a wide range of the fields of mathematics and introduces a balanced representation between comprehensive and sensible models. We demonstrate the applications of this representation in information extraction, semantic search, and education. We argue that the ontology can be a core of future integration of math-aware data sets in the Web of Data and, therefore, provide mappings onto relevant datasets, such as DBpedia and ScienceWISE.Comment: 15 pages, 6 images, 1 table, Knowledge Engineering and the Semantic Web - 5th International Conferenc

Cryptic Species: A Mismatch between Genetics and Morphology in Millepora

Millepore morphology is highly variable and shows signs of phenotypic plasticity. Two species of Millepora are present around the islands of the Bahamas: one exhibiting a strong, blade-like structure, Millepora complanata, and the other having a delicate branch-like structure, Millepora alcicornis. The phylogenetic relationship of these corals has been under considerable debate for many years. The existence of a range of intermediate growth forms exhibiting characteristics of both recognized species has led to the re-examination of this species complex. Several methods were employed to examine the taxonomic relationship including ecological abundance surveys, morphological thin-section analysis, and sequencing of rDNA internal transcribed spacer (ITS) regions. Abundance surveys showed a demarcation of growth forms by depth at two sites but an intermingling of growth forms at a third site. Morphometric analysis resulted in discrimination between M. alcicornis, M. complanata and the intermediate growth forms. However, rDNA sequence differences revealed the presence of two distinct clades, each containing members of the two currently recognized species as well as intermediate growth forms. The sequence analysis suggests the presence of two, phenotypically plastic cryptic species. Although limited in scope, our results indicate that caution should be exercised when describing species based on morphology alone and that multiple characters, including genetic information, should be used when describing species relationships

A Taxonomy of Data Grids for Distributed Data Sharing, Management and Processing

Data Grids have been adopted as the platform for scientific communities that need to share, access, transport, process and manage large data collections distributed worldwide. They combine high-end computing technologies with high-performance networking and wide-area storage management techniques. In this paper, we discuss the key concepts behind Data Grids and compare them with other data sharing and distribution paradigms such as content delivery networks, peer-to-peer networks and distributed databases. We then provide comprehensive taxonomies that cover various aspects of architecture, data transportation, data replication and resource allocation and scheduling. Finally, we map the proposed taxonomy to various Data Grid systems not only to validate the taxonomy but also to identify areas for future exploration. Through this taxonomy, we aim to categorise existing systems to better understand their goals and their methodology. This would help evaluate their applicability for solving similar problems. This taxonomy also provides a "gap analysis" of this area through which researchers can potentially identify new issues for investigation. Finally, we hope that the proposed taxonomy and mapping also helps to provide an easy way for new practitioners to understand this complex area of research.Comment: 46 pages, 16 figures, Technical Repor