Molecular genetics and comparative genomics reveal RNAi is not functional in malaria parasites

Techniques for targeted genetic disruption in Plasmodium, the causative agent of malaria, are currently intractable for those genes that are essential for blood stage development. The ability to use RNA interference (RNAi) to silence gene expression
would provide a powerful means to gain valuable insight into the pathogenic blood stages but its functionality in Plasmodium remains controversial. Here we have used various RNA-based gene silencing approaches to test the utility of RNAi in malaria
parasites and have undertaken an extensive comparative genomics search using profile hidden Markov models to clarify whether RNAi machinery
exists in malaria. These investigative approaches revealed that Plasmodium lacks the enzymology required for RNAi-based ablation of gene expression
and indeed no experimental evidence for RNAi was observed. In its absence, the most likely explanations for previously reported RNAi-mediated knockdown are either the general toxicity of introduced RNA (with global down-regulation of gene expression) or a specific antisense effect mechanistically distinct from RNAi, which will need systematic
analysis if it is to be of use as a molecular genetic tool for malaria parasites.

Molecular genetic studies of the yabby, Cherax destructor Clark

A study of genetic variation in the yabby (Cherax destructor) was undertaken using a number of different molecular techniques. The results have significantly improved our understanding of the genetic structure, evolution and taxonomy of this important freshwater species. The findings also contribute to the sustainable exploitation of yabbies for aquaculture.

Cystic fibrosis in children of the eastern Arabian peninsula : a clinical, spatial and genetic study

Aim: The aim of this thesis is to describe the process by which the inherited disease, cystic fibrosis, (CF) was recognised as an important clinical entity in the United Arab Emirates (UAE) and the Sultanate of Oman (Oman). It examines the clinical presentation of the first patients and assesses their degree of severity. Further, it describes the first studies carried out to determine the underlying CF mutations associated with the disease in the UAE and Oman. An estimate is offered of the birth frequency of the condition. Overall, the cultural, geographical and historical aspect of the societies in which the disease occurs is stressed. Methods: An initial literature search was carried out using Medline of any literature pertaining to the Arab World and CF. this was read and classified into the relevance to Arabs in general, the Middle East and then specifically the Arab (Persian) Gulf societies. Thereafter, a clinic was established at Tawam Hospital, Al Ain, UAE, for children presenting With chronic respiratory disease that could serve as a national referral centre. It was run by the Author as a service of the Paediatric Department of the UAE University Medical School. I sent a letter to every Paediatrician working in the UAE informing them of our clinic and offering our services for the diagnosis and management of chronic respiratory disease in children. This was based on the author's experience as a respiratory paediatrician in Australia and New Zealand and as the Professor of Paediatrics in the UAE. No such service then existed in the UAE. Funding was sought to establish a research programme and develop a molecular genetics laboratory in the UAE Medical School. A series of successful research applications provided the grants to commence the investigations. Once a small number of children had been identified as having CF from those referred to the respiratory clinic, the initial project was to assess and report their clinical presentation. Following this an early start was made on the identification of the mutations responsible. Once these were established an attempt was made to estimate the frequency of the condition at birth. Additional clinical studies revolved around assessing the severity of the condition that was associated with the main mutations that were identified. A clinical comparison was made with those with the mutation AF508 and the other main mutation, despite the obvious limitation of small numbers then available. Radiological assessment was made to evaluate the progression of the disease. The final aspect of the study was to assess patients from Oman and compare their findings and mutations with the neighbouring UAE. Based on information gained hypotheses are proposed regarding the spread of the gene mutation by population drift. Thesis outline: A literature review is presented in the form of a critique on the disease and a resume of the relevant aspects of the genetics of CF. Additionally, facts about the two countries' geography and history are presented. Finally, knowledge about CF mutations and population origins from other areas is presented. The second main section deals with the clinical features of the disorder as it presents in the UAE. Molecular findings are then presented and details of the common mutation found in Bedouin Arabs. Hypotheses are then presented based on the information gathered. Results: CF is not a rare disease in the Arab children of the UAE and Oman. These findings refute previous reports of CF being a rare or non-existent disease in Arabs. The condition presents with a severe clinical picture, with early colonisation of the respiratory tract with staphylococcus, haemophilus and pseudomonas organisms, even with conventional CF management practices in place. The CF mutation S549R is prevalent in Arabs of Bedouin stock, while AF508 is found in those of Baluch origin. The former may be descendants of Arabs who left southern Arabia and travelled to the Trucial Coast at the time of the destruction of the great dam at Marib. The origins of this mutation may lie in the area that corresponds to the modern Republic of Yemen. The latter groups are descendants of those who came originally from Baluchistan. It is hypothesised also that the ancestral home of the AF508 mutation may be in the geographical area now known as Baluchistan, that spans three separate modern political territories. The evidence presented supports the concept that the S549R mutation may be associated with a severe, if not the severest, clinical pattern recognised. It equates with that seen with the homozygous AF508 genotype. The absence of an additional mutation in the promoter region accounts for the different clinical pattern seen in previously described patients. Conclusions: There needs to be a major awareness of the presence of CF as a severe clinical disease in the children of the Gulf States. The clinical presentation and findings support the concept of under recognition of the disease. Climatic conditions put the children at special risk of hyponatraemia and electrolyte imbalance. The absence of surviving adults with the disease suggests premature deaths have occurred, but the high fertility rates have maintained the gene pool for this recessive disorder.

Isolation and characterisation of Arabidopsis RAD30 and ERCC1

This thesis describes the isolation and characterisation of two plant genes, AtERCC1 and AtRAD30. Evidence from protein homology comparisons, and functional complementation, in vitro mutagenesis, or interaction assays suggests the involvement of these genes in the repair or tolerance, respectively, of UV-induced DNA change.

Morphological, bioacoustical, and genetic variation in Miniopterus bats from eastern Madagascar, with the description of a new species

Recent molecular genetic work, combined with morphological comparisons, of Malagasy members of the bat genus Miniopterus (Family Miniopteridae), has revealed several cryptic species. Based on new specimens and associated tissues, we examine patterns of variation in the recently described species M. petersoni, the holotype of which comes from extreme southeastern Madagascar, and for which specimens from more northerly portions of eastern Madagascar were noted to show some morphological divergence from typical M. petersoni. On the basis of morphological and genetic (cytochrome b) characters we described a new species, M. egeri sp. nov. This taxon also shows bioacoustical differences from M. petersoni. Miniopterus egeri is widely distributed in the eastern portion of Madagascar across an elevational range from near sea level to 550 m. The specific status of moderately small Miniopterus from Montagne d'Ambre in the far north remains to be determined.

Description of a new species of the miniopterus aelleni group (chiroptera: miniopteridae) from upland areas of central and northern Madagascar

Recent molecular genetic work, combined with morphological comparisons, on Malagasy members of the bat genus Miniopterus (Family Miniopteridae), has uncovered a number of cryptic species. Based on recently collected specimens and associated tissues, we examine patterns of variation in M. aelleni, the holotype of which comes from Ankarana in northern Madagascar. Using molecular genetic (mitochondrial cytochrome b) and morphological characters we describe a new species, M. ambohitrensis sp. nov. In northern Madagascar, M. ambohitrensis and M. aelleni are allopatric, but occur in relatively close geographical contact (approximately 40 km direct line distance) with M. ambohitrensis found at Montagne d'Ambre in montane humid forest and M. aelleni sensu stricto at Ankarana in dry deciduous forest. Morphologically, this new taxon is differentiated from M. aelleni based on pelage coloration, external measurements, craniodental differences, and tragus shape. Comparisons using 725 bp of cytochrome b found a divergence of 1.1% within M. aelleni sensu stricto, 0.8% within M. ambohitrensis, and 3.3% between these two clades. The two sister species do not demonstrate acoustical differences based on recordings made in a flight cage. Miniopterus ambohitrensis is known from four localities in the northern and central portions of Madagascar, all from montane regions and across an elevational range from about 800 to 1600 m; its calculated "Extent of occurrence" is 15,143 km². It is possible that this species is at least partially migratory.

Inheritance of molecular markers and sex in the Australian freshwater crayfish, Cherax destructor Clark

Inheritance of three kinds of molecular genetic markers (mtDNA, random-amplified polymorphic DNAs (RAPDs) and allozymes) and sex were investigated in crossbreeding experiments between three populations of the Australian freshwater crayfish Cherax destructor. Crossbreeding did not disrupt the ively maternally inherited, and allozyme and RAPD markers were transmitted following expected Mendelian principles for co-dominant and dominant traits respectively. Unlike these three markers, sex ratios were found to be distorted by crossbreeding in some families. Two crossbred families produced only females. The implications of these findings for freshwater crayfish population genetics, taxonomy and aquaculture are discussed.

Manual on application of molecular tools in aquaculture and inland fisheries management. Part I : conceptual basis of population genetic approaches

The aim of this manual is to provide a comprehensive practical tool for the generation and analysis of genetic data for subsequent application in aquatic resources management in relation to genetic stock identification in inland fisheries and aquaculture. The material only covers general background on genetics in relation to aquaculture and fisheries resource management, the techniques and relevant methods of data analysis that are commonly used to address questions relating to genetic resource characterisation and population genetic analyses. No attempt is made to include applications of genetic improvement techniques e.g. selective breeding or producing genetically modified organisms (GMOs). The manual includes two ‘stand-alone’ parts, of which this is the first volume: Part 1 – Conceptual basis of population genetic approaches: will provide a basic foundation on genetics in general, and concepts of population genetics. Issues on the choices of molecular markers and project design are also discussed. Part 2 – Laboratory protocols, data management and analysis: will provide step-by-step protocols of the most commonly used molecular genetic techniques utilised in population genetics and systematic studies. In addition, a brief discussion and explanation of how these data are managed and analysed is also included. This manual is expected to enable NACA member country personnel to be trained to undertake molecular genetic studies in their own institutions, and as such is aimed at middle and higher level technical grades. The manual can also provide useful teaching material for specialised advanced level university courses in the region and postgraduate students. The manual has gone through two development/improvement stages. The initial material was tested at a regional workshop and at the second stage feedback from participants was used to improve the contents.

Manual on application of molecular tools in aquaculture and inland fisheries management. Part I : laboratory protocols, data management and analysis

The aim of this manual is to provide a comprehensive practical tool for the generation and analysis of genetic data for subsequent application in aquatic resources management in relation to genetic stock identification in inland fisheries and aquaculture. The material only covers general background on genetics in relation to aquaculture and fisheries resource management, the techniques and relevant methods of data analysis that are commonly used to address questions relating to genetic resource characterisation and population genetic analyses. No attempt is made to include applications of genetic improvement techniques e.g. selective breeding or producing genetically modified organisms (GMOs). The manual includes two ‘stand-alone’ parts, of which this is the second volume: Part 1 – Conceptual basis of population genetic approaches: will provide a basic foundation on genetics in general, and concepts of population genetics. Issues on the choices of molecular markers and project design are also discussed. Part 2 – Laboratory protocols, data management and analysis: will provide step-by-step protocols of the most commonly used molecular genetic techniques utilised in population genetics and systematic studies. In addition, a brief discussion and explanation of how these data are managed and analysed is also included. This manual is expected to enable NACA member country personnel to be trained to undertake molecular genetic studies in their own institutions, and as such is aimed at middle and higher level technical grades. The manual can also provide useful teaching material for specialised advanced level university courses in the region and postgraduate students. The manual has gone through two development/improvement stages. The initial material was tested at a regional workshop and at the second stage feedback from participants was used to improve the contents.

Development of molecular techniques for the genetic analysis of abalone populations

Three DNA techniques: random amplified polymorphic DNA (RAPD), minisatellite, and microsatellite analyses, were developed for use in abalone population genetic structure studies. The techniques were assessed using sample sets of blacklip and greenlip abalone. The study identifies a potential for the application of these DNA markers in abalone fisheries management, but microsatellites are the recommended method for future studies.

Molecular systematic studies of freshwater crayfish of the the genus Cherax

This study investigated the molecular systematics and zoogeography of freshwater crayfish species (yabbies) of the genus Cherax throughout Australia. Using DNA sequencing of up to five gene regions and phylogenetic methods of analyses, this study provided new insights into the taxonomy and evolution of this important group of Australian freshwater invertebrates.

Molecular genetic markers for abalone aquaculture

Describes the development and mapping of molecular markers in the blacklip and greenlip abalones. By means of a genome scan using a novel selective DNA pooling strategy, markers associated with growth were discovered that could potentially be applied to increase genetic gain in abalone aquaculture, whilst minimising inbreeding.