CDKN2A/p16 is inactivated in most melanoma cell lines

The CDKN2A gene maps to chromosome 9p21-22 and is responsible for melanoma susceptibility in some families. Its product, p16, binds specifically to CDK4 and CDK6 in vitro and in vivo, inhibiting their kinase activity. CDKN2A is homozygously deleted or mutated in a large proportion of tumor cell lines and some primary tumors, including melanomas. The aim of this study was to investigate the involvement of CDKN2A and elucidate the mechanisms of p16 inactivation in a panel of 60 cell lines derived from sporadic melanomas. Twenty-six (43%) of the melanoma lines were homozygously deleted for CDKN2A, and an additional 15 (25%) lines carried missense, nonsense, or frameshift mutations. All but one of the latter group were shown by microsatellite analysis to be hemizygous for the region of 9p surrounding CDKN2A. p16 was detected by Western blotting in only five of the cell lines carrying mutations. Immunoprecipitation of p16 in these lines, followed by Western blotting to detect the coprecipitation of CDK4 and CDK6, revealed that p16 was functionally compromised in all cell lines but the one that carried a heterozygous CDKN2A mutation. In the remaining 19 lines that carried wild-type CDKN2A alleles, Western blot analysis and immunoprecipitation indicated that 11 cell lines expressed a wild-type protein. Northern blotting was performed on the remaining eight cell lines and revealed that one cell line carried an aberrantly sized RNA transcript, and two other cell lines failed to express RNA. The promoter was found to be methylated in five cell lines that expressed CDKN2A transcript but not p16. Presumably, the message seen by Northern blotting in these cell lines is the result of cross-hybridization of the total cDNA probe with the exon 1beta transcript. Microsatellite analysis revealed that the majority of these cell lines were hemi/homozygous for the region surrounding CDKN2A, indicating that the wild-type allele had been lost. In the 11 cell lines that expressed functional p16, microsatellite analysis revealed loss of heterozygosity at the markers immediately surrounding CDKN2A in five cases, and the previously characterized R24C mutation of CDK4 was identified in one of the remaining 6 lines. These data indicate that 55 of 60 (92%) melanoma cell lines demonstrated some aberration of CDKN2A or CDK4, thus suggesting that this pathway is a primary genetic target in melanoma development.

From margins to mainstream : how screen and creative industries developed in the Northern Rivers region of NSW : 2000–2010

This thesis explores the proposition that growth and development in the screen and creative industries is not confined to the major capital cities. Lifestyle considerations, combined with advances in digital technology, convergence and greater access to broadband are altering requirements for geographic location, and creative workers are being drawn away from the big metropolises to certain regional areas. Regional screen industry enclaves are emerging outside of London, in the Highlands and Islands of Scotland, in Nova Scotia in Canada and in New Zealand. In the Australian context, the proposition is tested in an area regarded as a ‘special case’ in creative industry expansion: the Northern Rivers region of NSW. A key feature of the ‘specialness’ of this region is the large number of experienced, credited producers who live and operate their businesses within the region. The development of screen and creative industries in the Northern Rivers over the decade 2000 – 2010 has implications for regional regeneration and offers new insights into the rapidly changing screen industry landscape. This development also has implications for creative industry discourse, especially the dominance of the urban in creative industries thought. The research is pioneering in a number of ways. Building on the work conducted for my Masters thesis in 2000, a second study was conducted during the research phase, adapting creative industries theory and mapping methods, which have been largely city and nation-centric, and applying them to a regional context. The study adopted an action research approach as an industry development strategy for screen industries, while at the same time developing fine-grained ground up methods for collecting primary quantitative data on the size and scope of the creative industries. In accordance with the action research framework, the researcher also acted in the dual roles of industry activist and screen industry producer in the region. The central focus of the research has been both to document and contribute to the growth and development of screen and creative industries over the past decade in the Northern Rivers region. These interventions, along with policy developments at both a local and national level, and broader global shifts, have had the effect of repositioning the sector from a marginal one to a priority area considered integral to the future economic and cultural life of the region. The research includes a detailed mapping study undertaken in 2005 with comparisons to an earlier 2000 study and to ABS data for 2001 and 2006 to reveal growth trends. It also includes two case studies of projects that developed from idea to production and completion in the region during the decade in question. The studies reveal the drivers, impediments and policy implications for sustaining the development of screen industries in a regional area. A major finding of the research was the large and increasing number of experienced producers who operate within the region and the leadership role they play in driving the development of the emerging local industry. The two case studies demonstrate the impact of policy decisions on local screen industry producers and their enterprises. A brief overview of research in other regional areas is presented, including two international examples, and what they reveal about regional regeneration. Implications are drawn for creative industries discourse and regional development policy challenges for the future.

The flammability of metallic materials in normal and reduced gravity

Metallic materials exposed to oxygen-enriched atmospheres – as commonly used in the medical, aerospace, aviation and numerous chemical processing industries – represent a significant fire hazard which must be addressed during design, maintenance and operation. Hence, accurate knowledge of metallic materials flammability is required. Reduced gravity (i.e. space-based) operations present additional unique concerns, where the absence of gravity must also be taken into account. The flammability of metallic materials has historically been quantified using three standardised test methods developed by NASA, ASTM and ISO. These tests typically involve the forceful (promoted) ignition of a test sample (typically a 3.2 mm diameter cylindrical rod) in pressurised oxygen. A test sample is defined as flammable when it undergoes burning that is independent of the ignition process utilised. In the standardised tests, this is indicated by the propagation of burning further than a defined amount, or „burn criterion.. The burn criterion in use at the onset of this project was arbitrarily selected, and did not accurately reflect the length a sample must burn in order to be burning independent of the ignition event and, in some cases, required complete consumption of the test sample for a metallic material to be considered flammable. It has been demonstrated that a) a metallic material.s propensity to support burning is altered by any increase in test sample temperature greater than ~250-300 oC and b) promoted ignition causes an increase in temperature of the test sample in the region closest to the igniter, a region referred to as the Heat Affected Zone (HAZ). If a test sample continues to burn past the HAZ (where the HAZ is defined as the region of the test sample above the igniter that undergoes an increase in temperature of greater than or equal to 250 oC by the end of the ignition event), it is burning independent of the igniter, and should be considered flammable. The extent of the HAZ, therefore, can be used to justify the selection of the burn criterion. A two dimensional mathematical model was developed in order to predict the extent of the HAZ created in a standard test sample by a typical igniter. The model was validated against previous theoretical and experimental work performed in collaboration with NASA, and then used to predict the extent of the HAZ for different metallic materials in several configurations. The extent of HAZ predicted varied significantly, ranging from ~2-27 mm depending on the test sample thermal properties and test conditions (i.e. pressure). The magnitude of the HAZ was found to increase with increasing thermal diffusivity, and decreasing pressure (due to slower ignition times). Based upon the findings of this work, a new burn criterion requiring 30 mm of the test sample to be consumed (from the top of the ignition promoter) was recommended and validated. This new burn criterion was subsequently included in the latest revision of the ASTM G124 and NASA 6001B international test standards that are used to evaluate metallic material flammability in oxygen. These revisions also have the added benefit of enabling the conduct of reduced gravity metallic material flammability testing in strict accordance with the ASTM G124 standard, allowing measurement and comparison of the relative flammability (i.e. Lowest Burn Pressure (LBP), Highest No-Burn Pressure (HNBP) and average Regression Rate of the Melting Interface(RRMI)) of metallic materials in normal and reduced gravity, as well as determination of the applicability of normal gravity test results to reduced gravity use environments. This is important, as currently most space-based applications will typically use normal gravity information in order to qualify systems and/or components for reduced gravity use. This is shown here to be non-conservative for metallic materials which are more flammable in reduced gravity. The flammability of two metallic materials, Inconel® 718 and 316 stainless steel (both commonly used to manufacture components for oxygen service in both terrestrial and space-based systems) was evaluated in normal and reduced gravity using the new ASTM G124-10 test standard. This allowed direct comparison of the flammability of the two metallic materials in normal gravity and reduced gravity respectively. The results of this work clearly show, for the first time, that metallic materials are more flammable in reduced gravity than in normal gravity when testing is conducted as described in the ASTM G124-10 test standard. This was shown to be the case in terms of both higher regression rates (i.e. faster consumption of the test sample – fuel), and burning at lower pressures in reduced gravity. Specifically, it was found that the LBP for 3.2 mm diameter Inconel® 718 and 316 stainless steel test samples decreased by 50% from 3.45 MPa (500 psia) in normal gravity to 1.72 MPa (250 psia) in reduced gravity for the Inconel® 718, and 25% from 3.45 MPa (500 psia) in normal gravity to 2.76 MPa (400 psia) in reduced gravity for the 316 stainless steel. The average RRMI increased by factors of 2.2 (27.2 mm/s in 2.24 MPa (325 psia) oxygen in reduced gravity compared to 12.8 mm/s in 4.48 MPa (650 psia) oxygen in normal gravity) for the Inconel® 718 and 1.6 (15.0 mm/s in 2.76 MPa (400 psia) oxygen in reduced gravity compared to 9.5 mm/s in 5.17 MPa (750 psia) oxygen in normal gravity) for the 316 stainless steel. Reasons for the increased flammability of metallic materials in reduced gravity compared to normal gravity are discussed, based upon the observations made during reduced gravity testing and previous work. Finally, the implications (for fire safety and engineering applications) of these results are presented and discussed, in particular, examining methods for mitigating the risk of a fire in reduced gravity.

The validation of the rating for subdivision neighbourhood sustainability design (RSNSD) In Bangkok Metropolitan Region (BMR), Thailand

In recent years, the problems resulting from unsustainable subdivision development have become significant problems in the Bangkok Metropolitan Region (BMR), Thailand. Numbers of government departments and agencies have tried to eliminate the problems by introducing the rating tools to encourage the higher sustainability levels of subdivision development in BMR, such as the Environmental Impact Assessment Monitoring Award (EIA-MA) and the Thai’s Rating for Energy and Environmental Sustainability of New construction and major renovation (TREES-NC). However, the EIA-MA has included the neighbourhood designs in the assessment criteria, but this requirement applies to large projects only. Meanwhile, TREES-NC has focused only on large scale buildings such as condominiums, office buildings, and is not specific for subdivision neighbourhood designs. Recently, the new rating tool named “Rating for Subdivision Neighbourhood Sustainability Design (RSNSD)” has been developed. Therefore, the validation process of RSNSD is still required. This paper aims to validate the new rating tool for subdivision neighbourhood design in BMR. The RSNSD has been validated by applying the rating tool to eight case study subdivisions. The result of RSNSD by data generated through surveying subdivisions will be compared to the existing results from the EIA-MA. The selected cases include of one “Excellent Award”, two “Very Good Award”, and five non-rated subdivision developments. This paper expects to prove the credibility of RSNSD before introducing to the real subdivision development practises. The RSNSD could be useful to encourage higher sustainability subdivision design level, and then protect the problems from further subdivision development in BMR.

Evaluation of novel Streptococcus pyogenes vaccine candidates incorporating multiple conserved sequences from the C-repeat region of the M-protein

A major challenge for Streptococcus pyogenes vaccine development is the identification of epitopes that confer protection from infection by multiple S. pyogenes M-types. Here we have identified and characterised the distribution of common variant sequences from individual repeat units of the C-repeat region (CRR) of M-proteins representing 77 different M-types. Three polyvalent fusion vaccine candidates (SV1, SV2 and SV3) incorporating the most common variants were subsequently expressed and purified, and demonstrated to be alpha-helical by Circular Dichroism (CD), a secondary conformational characteristic of the CRR in the M-protein. Antibodies raised against each of these constructs recognise M-proteins that vary in their CRR, and bind to the surface of multiple S. pyogenes isolates. Antibodies raised against SV1, containing five variant sequences, also kill heterologous S. pyogenes isolates in in vitro bactericidal assays. Further structural characterisation of this construct demonstrated the conformation of SV1 was stable at different pHs, and thermal unfolding of SV1 a reversible process. Our findings demonstrate that linkage of multiple variant sequences into a single recombinant construct overcomes the need to embed the variant sequences in foreign helix promoting flanking sequences for conformational stability, and demonstrates the viability of the polyvalent candidates as global S. pyogenes vaccine candidates.

Defining the role of phytoene synthase in carotenoid accumulation of high provitamin A bananas

Vitamin A deficiency (VAD) is a serious problem in developing countries, affecting approximately 127 million children of preschool age and 7.2 million pregnant women each year. However, this deficiency is readily treated and prevented through adequate nutrition. This can potentially be achieved through genetically engineered biofortification of staple food crops to enhance provitamin A (pVA) carotenoid content. Bananas are the fourth most important food crop with an annual production of 100 million tonnes and are widely consumed in areas affected by VAD. However, the fruit pVA content of most widely consumed banana cultivars is low (~ 0.2 to 0.5 ìg/g dry weight). This includes cultivars such as the East African highland banana (EAHB), the staple crop in countries such as Uganda, where annual banana consumption is approximately 250 kg per person. This fact, in addition to the agronomic properties of staple banana cultivars such as vegetative reproduction and continuous cropping, make bananas an ideal target for pVA enhancement through genetic engineering. Interestingly, there are banana varieties known with high fruit pVA content (up to 27.8 ìg/g dry weight), although they are not widely consumed due to factors such as cultural preference and availability. The genes involved in carotenoid accumulation during banana fruit ripening have not been well studied and an understanding of the molecular basis for the differential capacity of bananas to accumulate carotenoids may impact on the effective production of genetically engineered high pVA bananas. The production of phytoene by the enzyme phytoene synthase (PSY) has been shown to be an important rate limiting determinant of pVA accumulation in crop systems such as maize and rice. Manipulation of this gene in rice has been used successfully to produce Golden Rice, which exhibits higher seed endosperm pVA levels than wild type plants. Therefore, it was hypothesised that differences between high and low pVA accumulating bananas could be due either to differences in PSY enzyme activity or factors regulating the expression of the psy gene. Therefore, the aim of this thesis was to investigate the role of PSY in accumulation of pVA in banana fruit of representative high (Asupina) and low (Cavendish) pVA banana cultivars by comparing the nucleic acid and encoded amino acid sequences of the banana psy genes, in vivo enzyme activity of PSY in rice callus and expression of PSY through analysis of promoter activity and mRNA levels. Initially, partial sequences of the psy coding region from five banana cultivars were obtained using reverse transcriptase (RT)-PCR with degenerate primers designed to conserved amino acids in the coding region of available psy sequences from other plants. Based on phylogenetic analysis and comparison to maize psy sequences, it was found that in banana, psy occurs as a gene family of at least three members (psy1, psy2a and psy2b). Subsequent analysis of the complete coding regions of these genes from Asupina and Cavendish suggested that they were all capable of producing functional proteins due to high conservation in the catalytic domain. However, inability to obtain the complete mRNA sequences of Cavendish psy2a, and isolation of two non-functional Cavendish psy2a coding region variants, suggested that psy2a expression may be impaired in Cavendish. Sequence analysis indicated that these Cavendish psy2a coding region variants may have resulted from alternate splicing. Evidence of alternate splicing was also observed in one Asupina psy1 coding region variant, which was predicted to produce a functional PSY1 isoform. The complete mRNA sequence of the psy2b coding regions could not be isolated from either cultivar. Interestingly, psy1 was cloned predominantly from leaf while psy2 was obtained preferentially from fruit, suggesting some level of tissue-specific expression. The Asupina and Cavendish psy1 and psy2a coding regions were subsequently expressed in rice callus and the activity of the enzymes compared in vivo through visual observation and quantitative measurement of carotenoid accumulation. The maize B73 psy1 coding region was included as a positive control. After several weeks on selection, regenerating calli showed a range of colours from white to dark orange representing various levels of carotenoid accumulation. These results confirmed that the banana psy coding regions were all capable of producing functional enzymes. No statistically significant differences in levels of activity were observed between banana PSYs, suggesting that differences in PSY activity were not responsible for differences in the fruit pVA content of Asupina and Cavendish. The psy1 and psy2a promoter sequences were isolated from Asupina and Cavendish gDNA using a PCR-based genome walking strategy. Interestingly, three Cavendish psy2a promoter clones of different sizes, representing possible allelic variants, were identified while only single promoter sequences were obtained for the other Asupina and Cavendish psy genes. Bioinformatic analysis of these sequences identified motifs that were previously characterised in the Arabidopsis psy promoter. Notably, an ATCTA motif associated with basal expression in Arabidopsis was identified in all promoters with the exception of two of the Cavendish psy2a promoter clones (Cpsy2apr2 and Cpsy2apr3). G1 and G2 motifs, linked to light-regulated responses in Arabidopsis, appeared to be differentially distributed between psy1 and psy2a promoters. In the untranscribed regulatory regions, the G1 motifs were found only in psy1 promoters, while the G2 motifs were found only in psy2a. Interestingly, both ATCTA and G2 motifs were identified in the 5’ UTRs of Asupina and Cavendish psy1. Consistent with other monocot promoters, introns were present in the Asupina and Cavendish psy1 5’ UTRs, while none were observed in the psy2a 5’ UTRs. Promoters were cloned into expression constructs, driving the â-glucuronidase (GUS) reporter gene. Transient expression of the Asupina and Cavendish psy1 and psy2a promoters in both Cavendish embryogenic cells and Cavendish fruit demonstrated that all promoters were active, except Cpsy2apr2 and Cpsy2apr3. The functional Cavendish psy2a promoter (Cpsy2apr1) appeared to have activity similar to the Asupina psy2a promoter. The activities of the Asupina and Cavendish psy1 promoters were similar to each other, and comparable to those of the functional psy2a promoters. Semi-quantitative PCR analysis of Asupina and Cavendish psy1 and psy2a transcripts showed that psy2a levels were high in green fruit and decreased during ripening, reinforcing the hypothesis that fruit pVA levels were largely dependent on levels of psy2a expression. Additionally, semi-quantitative PCR using intron-spanning primers indicated that high levels of unprocessed psy2a and psy2b mRNA were present in the ripe fruit of Cavendish but not in Asupina. This raised the possibility that differences in intron processing may influence pVA accumulation in Asupina and Cavendish. In this study the role of PSY in banana pVA accumulation was analysed at a number of different levels. Both mRNA accumulation and promoter activity of psy genes studied were very similar between Asupina and Cavendish. However, in several experiments there was evidence of cryptic or alternate splicing that differed in Cavendish compared to Asupina, although these differences were not conclusively linked to the differences in fruit pVA accumulation between Asupina and Cavendish. Therefore, other carotenoid biosynthetic genes or regulatory mechanisms may be involved in determining pVA levels in these cultivars. This study has contributed to an increased understanding of the role of PSY in the production of pVA carotenoids in banana fruit, corroborating the importance of this enzyme in regulating carotenoid production. Ultimately, this work may serve to inform future research into pVA accumulation in important crop varieties such as the EAHB and the discovery of avenues to improve such crops through genetic modification.

Measuring bitrate and quality trade-off in a fast region-of-interest based video coding

Prevailing video adaptation solutions change the quality of the video uniformly throughout the whole frame in the bitrate adjustment process; while region-of-interest (ROI)-based solutions selectively retains the quality in the areas of the frame where the viewers are more likely to pay more attention to. ROI-based coding can improve perceptual quality and viewer satisfaction while trading off some bandwidth. However, there has been no comprehensive study to measure the bitrate vs. perceptual quality trade-off so far. The paper proposes an ROI detection scheme for videos, which is characterized with low computational complexity and robustness, and measures the bitrate vs. quality trade-off for ROI-based encoding using a state-of-the-art H.264/AVC encoder to justify the viability of this type of encoding method. The results from the subjective quality test reveal that ROI-based encoding achieves a significant perceptual quality improvement over the encoding with uniform quality at the cost of slightly more bits. Based on the bitrate measurements and subjective quality assessments, the bitrate and the perceptual quality estimation models for non-scalable ROI-based video coding (AVC) are developed, which are found to be similar to the models for scalable video coding (SVC).

Study of the effectiveness of outrigger system for high-rise composite buildings for cyclonic region

The demands of taller structures are becoming imperative almost everywhere in the world in addition to the challenges of material and labor cost, project time line etc. This paper conducted a study keeping in view the challenging nature of high-rise construction with no generic rules for deflection minimizations and frequency control. The effects of cyclonic wind and provision of outriggers on 28-storey, 42-storey and 57-storey are examined in this paper and certain conclusions are made which would pave way for researchers to conduct further study in this particular area of civil engineering. The results show that plan dimensions have vital impacts on structural heights. Increase of height while keeping the plan dimensions same, leads to the reduction in the lateral rigidity. To achieve required stiffness increase of bracings sizes as well as introduction of additional lateral resisting system such as belt truss and outriggers is required.

Neoproterozoic glacial deposits of the Kimberly Region and northwestern Northern Territory, Australia

Neoproterozoic glacigenic formations are preserved in the Kimberley region and northwestern Northern Territory of northern Australia. They are distributed in the west Kimberley adjacent to the northern margins of the King Leopold Orogen, the Mt Ramsay area at the junction of the King Leopold and Halls Creek Orogens, and the east Kimberley, adjacent to the eastern margin of the Halls Creek Orogen. Small outlier glacigenic deposits are preserved in the Litchfield Province, Northern Territory (Uniya Formation) and Georgina Basin, western Queensland (Little Burke Formation). Glacigenic strata comprise diamictite, conglomerate, sandstone and pebbly mudstone and characterize the Walsh, Landrigan and Fargoo/Moonlight Valley formations. Thin units of laminated dolomite sit conformably at the top of the Walsh, Landrigan and Moonlight Valley formations. Glacigenic units are also interbedded with the carbonate platform deposits of the Egan Formation and Boonall Dolomite. δ13C data are available for all carbonate units. There is no direct chronological constraint on these successions. Dispute over regional correlation of the Neoproterozoic succession has been largely resolved through biostratigraphic, chemostratigraphic and lithostratigraphic analysis. However, palaeomagnetic results from the Walsh Formation are inconsistent with sedimentologically based correlations. Two stratigraphically defined glaciations are preserved in northwestern Australia: the ‘Landrigan Glaciation’, characterized by southwest-directed continental ice-sheet movement and correlated with late Cryogenian glaciation elsewhere in Australia and the world; and, the ‘Egan Glaciation’, a more localized glaciation of the Ediacaran Period. Future research focus should include chronology, palaeomagnetic constraint and tectonostratigraphic controls on deposition.

Expression Pattern of the Alpha-Kafirin Promoter Coupled with a Signal Peptide from Sorghum bicolor L. Moench

Regulatory sequences with endosperm specificity are essential for foreign gene expression in the desired tissue for both grain quality improvement and molecular pharming. In this study, promoters of seed storage α-kafirin genes coupled with signal sequence (ss) were isolated from Sorghum bicolor L. Moench genomic DNA by PCR. The α-kafirin promoter (α-kaf) contains endosperm specificity-determining motifs, prolamin-box, the O2-box 1, CATC, and TATA boxes required for α-kafirin gene expression in sorghum seeds. The constructs pMB-Ubi-gfp and pMB-kaf-gfp were microprojectile bombarded into various sorghum and sweet corn explants. GFP expression was detected on all explants using the Ubi promoter but only in seeds for the α-kaf promoter. This shows that the α-kaf promoter isolated was functional and demonstrated seed-specific GFP expression. The constructs pMB-Ubi-ss-gfp and pMB-kaf-ss-gfp were also bombarded into the same explants. Detection of GFP expression showed that the signal peptide (SP)::GFP fusion can assemble and fold properly, preserving the fluorescent properties of GFP.

Limestone and speleothem trace element geochemistry as tools for palaeoclimatic reconstruction, Mount Etna region, central-coastal Queensland

This study investigated potential palaeoclimate proxies provided by rare earth element (REE) geochemistry in speleothems and in clay mineralogy of cave sediments. Speleothem and sediment samples were collected from a series of cave fill deposits that occurred with rich vertebrate fossil assemblages in and around Mount Etna National Park, Rockhampton (central coastal Queensland). The fossil deposits range from Plio- Pleistocene to Holocene in age (based on uranium/thorium dating) and appear to represent depositional environments ranging from enclosed rainforest to semi-arid grasslands. Therefore, the Mount Etna cave deposits offer the perfect opportunity to test new palaeoclimate tools as they include deposits that span a known significant climate shift on the basis of independent faunal data. The first section of this study investigates the REE distribution of the host limestone to provide baseline geochemistry for subsequent speleothem investigations. The Devonian Mount Etna Beds were found to be more complex than previous literature had documented. The studied limestone massif is overturned, highly recrystallised in parts and consists of numerous allochthonous blocks with different spatial orientations. Despite the complex geologic history of the Mount Etna Beds, Devonian seawater-like REE patterns were recovered in some parts of the limestone and baseline geochemistry was determined for the bulk limestone for comparison with speleothem REE patterns. The second part of the study focused on REE distribution in the karst system and the palaeoclimatic implications of such records. It was found that REEs have a high affinity for calcite surfaces and that REE distributions in speleothems vary between growth bands much more than along growth bands, thus providing a temporal record that may relate to environmental changes. The morphology of different speleothems (i.e., stalactites, stalagmites, and flowstones) has little bearing on REE distributions provided they are not contaminated with particulate fines. Thus, baseline knowledge developed in the study suggested that speleothems were basically comparable for assessing palaeoclimatically controlled variations in REE distributions. Speleothems from rainforest and semi-arid phases were compared and it was found that there are definable differences in REE distribution that can be attributed to climate. In particular during semiarid phases, total REE concentration decreased, LREE became more depleted, Y/Ho increased, La anomalies were more positive and Ce anomalies were more negative. This may reflect more soil development during rainforest phases and more organic particles and colloids, which are known to transport REEs, in karst waters. However, on a finer temporal scale (i.e. growth bands) within speleothems from the same climate regime, no difference was seen. It is suggested that this may be due to inadequate time for soil development changes on the time frames represented by differences in growth band density. The third part of the study was a reconnaissance investigation focused on mineralogy of clay cave sediments, illite/kaolinite ratios in particular, and the potential palaeoclimatic implications of such records. Although the sample distribution was not optimal, the preliminary results suggest that the illite/kaolinite ratio increased during cold and dry intervals, consistent with decreased chemical weathering during those times. The study provides a basic framework for future studies at differing latitudes to further constrain the parameters of the proxy. The identification of such a proxy recorded in cave sediment has broad implications as clay ratios could potentially provide a basic local climate proxy in the absence of fossil faunas and speleothem material. This study suggests that REEs distributed in speleothems may provide information about water throughput and soil formation, thus providing a potential palaeoclimate proxy. It highlights the importance of understanding the host limestone geochemistry and broadens the distribution and potential number of cave field sites as palaeoclimate information no longer relies solely on the presence of fossil faunas and or speleothems. However, additional research is required to better understand the temporal scales required for the proxies to be recognised.

A 2-h diagnostic protocol to assess patients with chest pain symptoms in the Asia-Pacific region (ASPECT): a prospective observational validation study

Background: Patients with chest pain contribute substantially to emergency department attendances, lengthy hospital stay, and inpatient admissions. A reliable, reproducible, and fast process to identify patients presenting with chest pain who have a low short-term risk of a major adverse cardiac event is needed to facilitate early discharge. We aimed to prospectively validate the safety of a predefined 2-h accelerated diagnostic protocol (ADP) to assess patients presenting to the emergency department with chest pain symptoms suggestive of acute coronary syndrome. Methods: This observational study was undertaken in 14 emergency departments in nine countries in the Asia-Pacific region, in patients aged 18 years and older with at least 5 min of chest pain. The ADP included use of a structured pre-test probability scoring method (Thrombolysis in Myocardial Infarction [TIMI] score), electrocardiograph, and point-of-care biomarker panel of troponin, creatine kinase MB, and myoglobin. The primary endpoint was major adverse cardiac events within 30 days after initial presentation (including initial hospital attendance). This trial is registered with the Australia-New Zealand Clinical Trials Registry, number ACTRN12609000283279. Findings: 3582 consecutive patients were recruited and completed 30-day follow-up. 421 (11•8%) patients had a major adverse cardiac event. The ADP classified 352 (9•8%) patients as low risk and potentially suitable for early discharge. A major adverse cardiac event occurred in three (0•9%) of these patients, giving the ADP a sensitivity of 99•3% (95% CI 97•9–99•8), a negative predictive value of 99•1% (97•3–99•8), and a specificity of 11•0% (10•0–12•2). Interpretation: This novel ADP identifies patients at very low risk of a short-term major adverse cardiac event who might be suitable for early discharge. Such an approach could be used to decrease the overall observation periods and admissions for chest pain. The components needed for the implementation of this strategy are widely available. The ADP has the potential to affect health-service delivery worldwide.