Aspects of the biology of the yabby Trypea australiensis (Dana) (Decapoda : Thalassinidea) and the potential of burrow counts as an indirect measure of population density

The thalassinidean shrimp Trypea australiensis (the yabby) commonly occurs on intertidal sandflats and subtidal regions of sheltered embayments and estuaries along the east coast of Australia and is harvested commercially and recreationally for use as bait by anglers. The potential for counts of burrow openings to provide a reliable indirect estimate of the abundance of yabbies was examined on intertidal sandflats on North Stradbroke Island (Queensland, Australia). The relationship between the number of burrow openings and the abundance of yabbies was generally poor and also varied significantly through time, casting doubt on previous estimates of abundance for this species based on unvalidated hole counts. Spatial and temporal variation in population density, the size at maturity and the reproductive period of the yabby were also assessed. Except for an initial peak in abundance as a result of recruitment, the density of yabbies was constant throughout the study but considerably less than that estimated from a previous study in the same area. Ovigerous females were recorded at 3 mm carapace length (CL) which is smaller than previously recorded for this species and thalassinideans in general. Small ovigerous females were found throughout the study, including the summer months, which is unusual for thalassinideans in the intertidal zone. It was hypothesised that in the intertidal zone, small female yabbies may be able to balance the metabolic demands of reproduction and respiration at higher temperatures than can larger females allowing them to reproduce in the warmer months.

Efficacy of Lentivirus-mediated and MUC1 antibody-targeted VP22-TK/GCV suicide gene therapy for ovarian cancer

Transfer of the herpes simplex virus type I thymidine kinase (HSV-TK) gene into tumor cells using virus-based vectors in conjunction with ganciclovir (GCV) exposure provides a potential gene therapy strategy for the treatment of cancer. Effective gene therapy,, depends on the efficient transfer and specific targeting of therapeutic genes and their protein products to target cells. The purpose of this study was to investigate the anti-tumor effect of Lentivirus-mediated and MUC1 antibody-targeted VP22-TK/GCV suicide gene therapy in animal models. Mouse models were generated with intraperitoneal injection of human epithelial ovarian cancer cells 3AO, which are MUC1-positive. HTV-1-based lentiviral vectors carrying VP22-TK or scFv-VP22-TK were prepared. The animals were injected intraperitoneally with lentivirus containing scFv-VP22-TK, VP22-TK followed by GCV treatment. Combined treatment of lentivirus-expressed scFv-VP22-TK or VP22-TK with GCV inhibited the proliferation and prolonged survival times compared with the control vector. The survival time of animals treated with scFv-VP22-TK/GCV was significantly longer than that of animals treated with VP22-TK/GCV (p = 0.006). Conclusion: Our results suggest that MUC1 antibody-targeted VP22-TK/GCV suicide gene therapy can efficiently inhibit ovarian tumor growth and increase survival in a nude mouse model of ovarian carcinoma. These data support the development of this method for human clinical trials.

Variation in the dispersal potential of non-feeding invertebrate larvae: the desperate larva hypothesis and larval size

For many species of marine invertebrates, variability in larval settlement behaviour appears to be the rule rather than the exception. This variability has the potential to affect larval dispersal, because settlement behaviour will influence the length of time larvae are in the plankton. Despite the ubiquity and importance of this variability, relatively few sources of variation in larval settlement behaviour have been identified. One important factor that can affect larval settlement behaviour is the nutritional state of larvae. Non-feeding larvae often become less discriminating in their 'choice' of settlement substrate, i.e. more desperate to settle, when energetic reserves run low. We tested whether variation in larval size (and presumably in nutritional reserves) also affects the settlement behaviour of 3 species of colonial marine invertebrate larvae, the bryozoans Bugula neritina and Watersipora subtorquata and the ascidian Diplosoma listerianum. For all 3 species, larger larvae delayed settlement for longer in the absence of settlement cues, and settlement of Bugula neritina larvae was accelerated by the presence of settlement cues, independently of larval size. In the field, larger W subtorquata larvae also took longer to settle than smaller larvae and were more discriminating towards settlement surfaces. These differences in settlement time are likely to result in differences in the distance that larvae disperse in the field. We suggest that species that produce non-feeding larvae can affect the dispersal potential of their offspring by manipulating larval size and thus larval desperation.

Potential triggers for akinete differentiation in an Australian strain of the cyanobacterium Cylindrospermopsis raciborskii (AWT 205/1)

Understanding the triggers for some cyanobacteria of the Nostocales and Stigonematales orders to produce specialised reproductive cells termed akinetes, is very important to gain further insights into their ecology. By improving our understanding of their life cycle, appropriate management options may be devised to control the formation of these cells, and therefore the potential bloom inoculum which they are thought to provide, may be reduced. This study investigated the effect of chemical (phosphorus limitation), and environmental variables (temperature shock) on akinete differentiation in the freshwater cyanobacterium Cylindrospermopsis raciborskii (AWT 205/1). From the preliminary results, it is suggested that the availability of phosphorus and changes in temperature were a necessary requirement for the formation of akinetes in this particular strain of C. raciborskii. In the four phosphorus treatments investigated (0, 3, 38 and 75 mug l(-1) P), only the two higher treatments produced akinetes (approximately 220 ml(-1)). When the first akinetes were observed in the 38 and 75 mug l(-1) P treatments, filterable reactive phosphorus (FRP) concentrations in the medium were approximately 22 and 52 mug l(-1) P, respectively, indicating that there was no phosphorus limitation. In the temperature shock experiment, akinetes were observed in the 15 and 20degreesC treatments. However, akinetes were degraded (pale yellow colour, limited swelling and shrivelled edges) and in much lower concentrations, which was thought to be a result of the daily temperature shock. We suggest that the formation of akinetes in C. raciborskii (AWT 205/1) can be triggered by an initial temperature shock and that phosphorus is a necessary requirement to allow further growth and full development of akinetes.

Estimating the potential refolding yield of recombinant proteins expressed as inclusion bodies

Recombinant protein production in bacteria is efficient except that insoluble inclusion bodies form when some gene sequences are expressed. Such proteins must undergo renaturation, which is an inefficient process due to protein aggregation on dilution from concentrated denaturant. In this study, the protein-protein interactions of eight distinct inclusion-body proteins are quantified, in different solution conditions, by measurement of protein second virial coefficients (SVCs). Protein solubility is shown to decrease as the SVC is reduced (i.e., as protein interactions become more attractive). Plots of SVC versus denaturant concentration demonstrate two clear groupings of proteins: a more aggregative group and a group having higher SVC and better solubility. A correlation of the measured SVC with protein molecular weight and hydropathicity, that is able to predict which group each of the eight proteins falls into, is presented. The inclusion of additives known to inhibit aggregation during renaturation improves solubility and increases the SVC of both protein groups. Furthermore, an estimate of maximum refolding yield (or solubility) using high-performance liquid chromatography was obtained for each protein tested, under different environmental conditions, enabling a relationship between yield and SVC to be demonstrated. Combined, the results enable an approximate estimation of the maximum refolding yield that is attainable for each of the eight proteins examined, under a selected chemical environment. Although the correlations must be tested with a far larger set of protein sequences, this work represents a significant move beyond empirical approaches for optimizing renaturation conditions. The approach moves toward the ideal of predicting maximum refolding yield using simple bioinformatic metrics that can be estimated from the gene sequence. Such a capability could potentially screen, in silico, those sequences suitable for expression in bacteria from those that must be expressed in more complex hosts. (C) 2004 Wiley Periodicals, Inc.

Bioprocess-centered molecular design (BMD) for the efficient production of an interfacially active peptide

The efficient expression and purification of an interfacially active peptide (mLac21) was achieved by using bioprocess-centered molecular design (BMD), wherein key bioprocess considerations are addressed during the initial molecular biology work. The 21 amino acid mLac21 peptide sequence is derived from the lac repressor protein and is shown to have high affinity for the oil-water interface, causing a substantial reduction in interfacial tension following adsorption. The DNA coding for the peptide sequence was cloned into a modified pET-31(b) vector to permit the expression of mLac21 as a fusion to ketosteroid isomerase (KSI). Rational iterative molecular design, taking into account the need for a scaleable bioprocess flowsheet, led to a simple and efficient bioprocess yielding mLac21 at 86% purity following ion exchange chromatography (and >98% following chromatographic polishing). This case study demonstrates that it is possible to produce acceptably pure peptide for potential commodity applications using common scaleable bioprocess unit operations. Moreover, it is shown that BMD is a powerful strategy that can be deployed to reduce bioseparation complexity. (C) 2004 Wiley Periodicals, Inc.

New gene technologies and their potential value for sugarcane

An efficient system is now in place for improving diverse sugarcane cultivars by genetic transformation, that is, the insertion of useful new genes into single cells followed by the regeneration of genetically modified (transgenic) plants. The method has already been used to introduce genes for resistance to several major diseases, insect pests and a herbicide, Field testing has begun, and research is underway to identify other genes for increased environmental stress resistance, agronomic efficiency and yield of sucrose or other valuable products. Experience in other crops has shown that genetically improved varieties which provide genuine environmental and consumer benefits are welcomed by producers and consumers. Substantial research is still needed, but these new gene technologies will reshape the sugar industry and determine the international competitive efficiency of producers.