Transcriptome analysis of Neotyphodium and Epichloë grass endophytes

Large-scale gene discovery has been performed for the grass fungal endophytes Neotyphodium coenophialum, Neotyphodium lolii, and Epichloë festucae. The resulting sequences have been annotated by comparison with public DNA and protein sequence databases and using intermediate gene ontology annotation tools. Endophyte sequences have also been analysed for the presence of simple sequence repeat and single nucleotide polymorphism molecular genetic markers. Sequences and annotation are maintained within a MySQL database that may be queried using a custom web interface. Two cDNA-based microarrays have been generated from this genome resource. They permit the interrogation of 3806 Neotyphodium genes (NchipTM microarray), and 4195 Neotyphodium and 920 Epichloë genes (EndoChipTM microarray), respectively. These microarrays provide tools for high-throughput transcriptome analysis, including genome-specific gene expression studies, profiling of novel endophyte genes, and investigation of the host grass–symbiont interaction. Comparative transcriptome analysis in Neotyphodium and Epichloë was performed

First record of a smut fungus on Byblidaceae: Yelsemia lowrieana, a new species from Australia.

Yelsemia lowrieana sp . nov. (Ustilaginomycetes) is described and illustrated from Byblis rorida collected in northwestern Western Australia. Infected plants had galls filled with spores on stems and pedicels. The spores were unusual in that each could be separated from a dark outer spore wall. This is the first record of a smut fungus on the dicotyledonous host family Byblidaceae.

Effects of foliar application of potassium nitrate on suppression of Alternaria leaf blight of cotton (Gossypium hirsutum) in northern Australia

Alternaria leaf blight is the most prevalent disease of cotton in northern Australia. A trial was conducted at Katherine Research Station, Northern Territory, Australia, to determine the effects of foliar application of potassium nitrate (KNO3) on the suppression of Alternaria leaf blight of cotton. Disease incidence, severity and leaf shedding were assessed at the bottom (1-7 nodes), middle (8-14 nodes) and the top (15+ nodes) of plants at weekly intervals from 7 July to 22 September 2004. Disease incidence, severity and shedding at the middle canopy level were significantly higher for all treatments than those from bottom and top canopies. Foliar KNO3, applied at 13 kg/ha, significantly (P < 0.05) reduced the mean disease incidence, severity and leaf shedding assessed during the trial period. KNO 3 significantly (P < 0.001) reduced the disease severity and leaf shedding at the middle canopy level. Almost all leaves in the middle canopy became infected in the first week of July in contrast to infection levels of 50-65% at the bottom and top of the canopy. Disease severity and leaf shedding in the middle canopy were significantly (P < 0.05) lower in KNO 3-treated plots than the control plots from the second and third weeks of July to the second and third weeks of August. This study demonstrates that foliar application of KNO3 may be effective in reducing the effect of Alternaria leaf blight of cotton in northern Australia.

Gene expression profiling of cuticular proteins across the moult cycle of the crab Portunus pelagicus

Background: Crustaceans represent an attractive model to study biomineralization and cuticle matrix formation, as these events are precisely timed to occur at certain stages of the moult cycle. Moulting, the process by which crustaceans shed their exoskeleton, involves the partial breakdown of the old exoskeleton and the synthesis of a new cuticle. This cuticle is subdivided into layers, some of which become calcified while others remain uncalcified. The cuticle matrix consists of many different proteins that confer the physical properties, such as pliability, of the exoskeleton. Results: We have used a custom cDNA microarray chip, developed for the blue swimmer crab Portunus pelagicus, to generate expression profiles of genes involved in exoskeletal formation across the moult cycle. A total of 21 distinct moult-cycle related differentially expressed transcripts representing crustacean cuticular proteins were isolated. Of these, 13 contained copies of the cuticle_1 domain previously isolated from calcified regions of the crustacean exoskeleton, four transcripts contained a chitin_bind_4 domain (RR consensus sequence) associated with both the calcified and un-calcified cuticle of crustaceans, and four transcripts contained an unannotated domain (PfamB_109992) previously isolated from C. pagurus. Additionally, cryptocyanin, a hemolymph protein involved in cuticle synthesis and structural integrity, also displays differential expression related to the moult cycle. Moult stage-specific expression analysis of these transcripts revealed that differential gene expression occurs both among transcripts containing the same domain and among transcripts containing different domains. Conclusion: The large variety of genes associated with cuticle formation, and their differential expression across the crustacean moult cycle, point to the complexity of the processes associated with cuticle formation and hardening. This study provides a molecular entry path into the investigation of the gene networks associated with cuticle formation.

Differential expression profiling of components associated with exoskeletal hardening in crustaceans

Background: Exoskeletal hardening in crustaceans can be attributed to mineralization and sclerotization of the organic matrix. Glycoproteins have been implicated in the calcification process of many matrices. Sclerotization, on the other hand, is catalysed by phenoloxidases, which also play a role in melanization and the immunological response in arthropods. Custom cDNA microarrays from Portunus pelagicus were used to identify genes possibly associated with the activation pathways involved in these processes. Results: Two genes potentially involved in the recognition of glycosylation, the C-type lectin receptor and the mannose-binding protein, were found to display molt cycle-related differential expression profiles. C-type lectin receptor up-regulation was found to coincide with periods associated with new uncalcified cuticle formation, while the up-regulation of mannose-binding protein occurred only in the post-molt stage, during which calcification takes place, implicating both in the regulation of calcification. Genes presumed to be involved in the phenoloxidase activation pathway that facilitates sclerotization also displayed molt cycle-related differential expression profiles. Members of the serine protease superfamily, trypsin-like and chymotrypsin-like, were up-regulated in the intermolt stage when compared to post-molt, while trypsin-like was also up-regulated in pre-molt compared to ecdysis. Additionally, up-regulation in pre- and intermolt stages was observed by transcripts encoding other phenoloxidase activators including the putative antibacterial protein carcinin-like, and clotting protein precursor-like. Furthermore, hemocyanin, itself with phenoloxidase activity, displayed an identical expression pattern to that of the phenoloxidase activators, i.e. up-regulation in pre- and intermolt. Conclusion: Cuticle hardening in crustaceans is a complex process that is precisely timed to occur in the post-molt stage of the molt cycle. We have identified differential expression patterns of several genes that are believed to be involved in biomineralization and sclerotization and propose possible regulatory mechanisms for these processes based on their expression profiles, such as the potential involvement of C-type lectin receptors and mannose binding protein in the regulation of calcification.

Heritability estimates for growth in the tropical abalone Haliotis asinina using microsatellites to assign parentage

The tropical abalone Haliotis asinina is a wild-caught and cultured species throughout the Indo-Pacific as well as being an emerging model species for the study of haliotids. H. asinina has the fastest recorded natural growth rate of any abalone and reaches sexual maturity within one year. As such, it is a suitable abalone species for selective breeding for commercially important traits such as rapid growth. Estimating the amount of variation in size that is attributable to heritable genetic differences can assist the development of such a selective breeding program. Here we estimated heritability for growth-related traits at 12 months of age by creating a single cohort of 84 families in a full-factorial mating design consisting of 14 sires and 6 dams. Of 500 progeny sampled, 465 were successfully assigned to their parents based on shared alleles at 5 polymorphic microsatellite loci. Using an animal model, heritability estimates were 0.48 ± 0.15 for shell length, 0.38 ± 0.13 for shell width and 0.36 ± 0.13 for weight. Genetic correlations were > 0.98 between shell parameters and weight, indicating that breeding for weight gains could be successfully achieved by selecting for shell length.

Domestication for Conservation of an Endangered Species: The Case of the Wollemi Pine

A small population of tall slender conifers was discovered in 1994 in a deep rainforest canyon of the Wollemi National Park, New SouthWales, Australia. The living trees closely resembled fossils that were more than 65 million years old, and this ‘living fossil’ was recognised as a third extant genus in the Araucariaceae (Araucaria, Agathis and now Wollemia). The species was named the Wollemi pine (W. nobilis). Extensive searches uncovered very few populations, with the total number of adult trees being less than 100. Ex situ collections were quickly established in Sydney as part of the Wollemi Pine Recovery Plan. The majority of the ex situ population was later transferred to our custom-built facility in Queensland for commercial multiplication. Domestication has relied very heavily on the species’ amenability to vegetative propagation because seed collection from the natural populations is dangerous, expensive, and undesirable for conservation reasons. Early propagation success was poor, with only about 25% of cuttings producing roots. However, small increases in propagation success have a very large impact on a domestication program because plant production can be modelled on an exponential curve where each rooted cutting develops into a mother plant that, in turn, provides more rooted cuttings. An extensive research program elevated rooting percentages to greater than 80% and also provided in vitro methods for plant multiplication. These successes have enabled international release of the Wollemi pine as a new and attractive species for ornamental horticulture.

Gene expression evidence for off-target effects caused by RNA interference-mediated gene silencing of Ubiquitin-63E in the cattle tick Rhipicephalus microplus.

Knowledge of cattle tick (Rhipicephalus (Boophilus) microplus; Acari: Ixodidae) molecular and cellular pathways has been hampered by the lack of an annotated genome. In addition, most of the tick expressed sequence tags (ESTs) available to date consist of similar to 50% unassigned sequences without predicted functions. The most common approach to address this has been the application of RNA interference (RNAi) methods to investigate genes and their pathways. This approach has been widely adopted in tick research despite minimal knowledge of the tick RNAi pathway and double-stranded RNA (dsRNA) uptake mechanisms. A strong knockdown phenotype of adult female ticks had previously been observed using a 594 bp dsRNA targeting the cattle tick homologue for the Drosophila Ubiquitin-63E gene leading to nil or deformed eggs. A NimbleGen cattle tick custom microarray based on the BmiGI.V2 database of R. microplus ESTs was used to evaluate the expression of mRNAs harvested from ticks treated with the tick Ubiquitin-63E 594 bp dsRNA compared with controls. A total of 144 ESTs including TC6372 (Ubiquitin-63E) were down-regulated with 136 ESTs up-regulated following treatment. The results obtained substantiated the knockdown phenotype with ESTs identified as being associated with ubiquitin proteolysis as well as oogenesis, embryogenesis, fatty acid synthesis and stress responses. A bioinformatics analysis was undertaken to predict off-target effects (OTE) resulting from the in silico dicing of the 594 bp Ubiquitin-63E dsRNA which identified 10 down-regulated ESTs (including TC6372) within the list of differentially expressed probes on the microarrays. Subsequent knockdown experiments utilising 196 and 109 bp dsRNAs, and a cocktail of short hairpin RNAs (shRNA) targeting Ubiquitin-63E, demonstrated similar phenotypes for the dsRNAs but nil effect following shRNA treatment. Quantitative reverse transcriptase PCR analysis confirmed differential expression of TC6372 and selected ESTs. Our study demonstrated the minimisation of predicted OTEs in the shorter dsRNA treatments (similar to 100-200 bp) and the usefulness of microarrays to study knockdown phenotypes.

Economic case study of ABCD cane management practices in the Burdekin River Irrigation Area (BRIA) region

A case study was undertaken to determine the economic impact of a change in management class as detailed in the A, B, C and D management class framework. This document focuses on the implications of changing from D to C, C to B and B to A class management in the Burdekin River irrigation area (BRIA) and if the change is worthwhile from an economic perspective. This report provides a guide to the economic impact that may be expected when undertaking a particular change in farming practices and will ultimately lead to more informed decisions being made by key industry stakeholders. It is recognised that these management classes have certain limitations and in many cases the grouping of practices may not be reflective of the real situation. The economic case study is based on the A, B, C and D management class framework for water quality improvement developed in 2007/2008 for the Burdekin natural resource management region. The framework for the Burdekin is currently being updated to clarify some issues and incorporate new knowledge since the earlier version of the framework. However, this updated version is not yet complete and so the Paddock to Reef project has used the most current available version of the framework for the modelling and economics. As part of the project specification, sugarcane crop production data for the BRIA was provided by the APSIM model. The information obtained from the APSIM crop modelling programme included sugarcane yields and legume grain yield (legume grain yield only applies to A class management practice). Because of the complexity involved in the economic calculations, a combination of the FEAT, PiRisk and a custom made spreadsheet was used for the economic analysis. Figures calculated in the FEAT program were transferred to the custom made spreadsheet to develop a discounted cash flow analysis. The marginal cash flow differences for each farming system were simulated over a 5-year and 10-year planning horizon to determine the net present value of changing across different management practices. PiRisk was used to test uncertain parameters in the economic analysis and the potential risk associated with a change in value.

Economic case study of ABCD cane management practices in the Burdekin Delta region.

A case study was undertaken to determine the economic impact of a change in management class as detailed in the A, B, C and D management class framework. This document focuses on the implications of changing from D to C, C to B and B to A class management in the Burdekin Delta region and if the change is worthwhile from an economic perspective. This report provides a guide to the economic impact that may be expected when undertaking a particular change in farming practices and will ultimately lead to more informed decisions being made by key industry stakeholders. It is recognised that these management classes have certain limitations and in many cases the grouping of practices may not be reflective of the real situation. The economic case study is based on the A, B, C and D management class framework for water quality improvement developed in 2007/2008 for the Burdekin natural resource management region. The framework for the Burdekin is currently being updated to clarify some issues and incorporate new knowledge since the earlier version of the framework. However, this updated version is not yet complete and so the Paddock to Reef project has used the most current available version of the framework for the modelling and economics. As part of the project specification, sugarcane crop production data for the Burdekin Delta region was provided by the APSIM model. The information obtained from the APSIM crop modelling programme included sugarcane yields and legume grain yield (legume grain yield only applies to A class management practice). Because of the complexity involved in the economic calculations, a combination of the FEAT, PiRisk and a custom made spreadsheet was used for the economic analysis. Figures calculated in the FEAT program were transferred to the custom made spreadsheet to develop a discounted cash flow analysis. The marginal cash flow differences for each farming system were simulated over a 5-year and 10-year planning horizon to determine the Net Present Value of changing across different management practices. PiRisk was used to test uncertain parameters in the economic analysis and the potential risk associated with a change in value.

Economic case study of ABCD cane management practices in the Tully region

A case study was undertaken to determine the economic impact of a change in management class as detailed in the A, B, C and D management class framework. This document focuses on the implications of changing from D to C, C to B and B to A class management in the Tully region and if the change is worthwhile from an economic perspective. This report provides a guide to the economic impact that may be expected when undertaking a particular change in farming practices and will ultimately lead to more informed decisions being made by key industry stakeholders. It is recognised that these management classes have certain limitations and in many cases the grouping of practices may not be reflective of the real situation. The economic case study is based on the A, B, C and D management class framework for water quality improvement developed in 2007/2008 by the wet tropics natural resource management region. The framework for wet tropics is currently being updated to clarify some issues and incorporate new knowledge since the earlier version of the framework. However, this updated version is not yet complete and so the Paddock to Reef project has used the most current available version of the framework for the modelling and economics. As part of the project specification, sugarcane crop production data for the Tully region was provided by the APSIM model. Because of the complexity involved in the economic calculations, a combination of the FEAT, PiRisk and a custom made spreadsheet was used for the economic analysis. Figures calculated in the FEAT program were transferred to the custom made spreadsheet to develop a discounted cash flow analysis. The marginal cash flow differences for each farming system were simulated over a 5-year and 10-year planning horizon to determine the Net Present Value of changing across different management practices. PiRisk was used to test uncertain parameters in the economic analysis and the potential risk associated with a change in value.

Gene Expression Profiling of the Cephalothorax and Eyestalk in Penaeus Monodon during Ovarian Maturation.

In crustaceans, a range of physiological processes involved in ovarian maturation occurs in organs of the cephalothorax including the hepatopancrease, mandibular and Y-organ. Additionally, reproduction is regulated by neuropeptide hormones and other proteins released from secretory sites within the eyestalk. Reproductive dysfunction in captive-reared prawns, Penaeus monodon, is believed to be due to deficiencies in these factors. In this study, we investigated the expression of gene transcripts in the cephalothorax and eyestalk from wild-caught and captive-reared animals throughout ovarian maturation using custom oligonucleotide microarray screening. We have isolated numerous transcripts that appear to be differentially expressed throughout ovarian maturation and between wild-caught and captive-reared animals. In the cephalothorax, differentially expressed genes included the 1,3-beta-D-glucan-binding high-density lipoprotein, 2/3-oxoacyl-CoA thiolase and vitellogenin. In the eyestalk, these include gene transcripts that encode a protein that modulates G-protein coupled receptor activity and another that encodes an architectural transcription factor. Each may regulate the expression of reproductive neuropeptides, such as the crustacean hyperglycaemic hormone and molt-inhibiting hormone. We could not identify differentially expressed transcripts encoding known reproductive neuropeptides in the eyestalk of either wild-caught or captive-reared prawns at any ovarian maturation stage, however, this result may be attributed to low relative expression levels of these transcripts. In summary, this study provides a foundation for the study of target genes involved in regulating penaeid reproduction.

Fencing artificial waterpoints failed to influence density and distribution of red kangaroos (Macropus rufus)

Provision of artificial waterpoints in Australian rangelands has resulted in an increase in the range and density of kangaroos. At high densities, kangaroos can inhibit vegetation regeneration, particularly in some protected areas where harvesting is prohibited. Fencing off waterpoints has been proposed to limit these impacts. Our aim was to determine whether fencing off waterpoints during a drought (when kangaroos would be especially water-limited) would influence the density and distribution of red kangaroos (Macropus rufus). Two waterpoints were fenced within the first 6 months of the 27-month study and a further two waterpoints were kept unfenced as controls in Idalia National Park, western Queensland. We estimated kangaroo densities around waterpoints from walked line-transect counts, and their grazing distribution from dung-pellet counts. Fencing off waterpoints failed to influence either the density or distribution up to 4 km from the waterpoints. Our results indicate that food availability, rather than the location of waterpoints, determines kangaroo distribution. Few areas in the rangelands are beyond kangaroos' convenient reach from permanent waterpoints. Therefore, fencing off waterpoints without explicitly considering the spatial context in relation to other available water sources will fail to achieve vegetation regeneration.

Mass transfer properties (permeability and mass diffusivity) of four Australian hardwood species

Characterisation of mass transfer properties was achieved in the longitudinal, radial, and tangential directions for four Australian hardwood species: spotted gum, blackbutt, jarrah and messmate. Measurement of mass transfer properties for these species was necessary to complement current vacuum drying modelling research. Water-vapour diffusivity was determined in steady state using a specific vapometer. Permeability values of some species and material directions were extremely low and undetectable by the mass flow meter device. Hence, a custom system based on volume evolution was conceived to determine very low, previously unpublished, wood permeability values. Mass diffusivity and permeability were lowest for spotted gum and highest for messmate. Except for messmate, in the radial direction, the four species measured were less permeable in all directions than the lowest published figures, demonstrating the high impermeability of Australian hardwoods and partly accounting for their relatively slow drying rates. Premeability, water-vapour diffusivity, and associated anisotropic ratio data obtained for messmate were extreme or did not follow typical trends and is consequently the most difficult of the four woods to dry in terms of collapse and checkinng degradation.

Small-scale capture, transport and tank adaptation of live, medium-sized Scombrids using "Tuna Tubes"

The transport of live fish is a crucial step to establish fish culture in captivity, and is especially challenging for species that have not been commonly cultured before, therefore transport and handling methods need to be optimized and tailored. This study describes the use of tuna tubes for small-scale transport of medium-sized pelagic fish from the Scombridae family. Tuna tubes are an array of vertical tubes that hold the fish, while fresh seawater is pumped up the tubes and through the fish mouth and gills, providing oxygen and removing wastes. In this study, 19 fish were captured using rod and line and 42% of the captured fish were transported alive in the custom-designed tuna tubes to an on-shore holding tank: five mackerel tuna (Euthynnus affinis) and three leaping bonito (Cybiosarda elegans). Out of these, just three (15.8% of total fish) acclimatized to the tank's condition. Based on these results, we discuss an improved design of the tuna tubes that has the potential to increase survival rates and enable a simple and low cost method of transporting of live pelagic fish.