Packed bed bioreactor for the isolation and expansion of placental-derived Mesenchymal Stromal Cells

Large numbers of Mesenchymal stem/stromal cells (MSCs) are required for clinical relevant doses to treat a number of diseases. To economically manufacture these MSCs, an automated bioreactor system will be required. Herein we describe the development of a scalable closed-system, packed bed bioreactor suitable for large-scale MSCs expansion. The packed bed was formed from fused polystyrene pellets that were air plasma treated to endow them with a surface chemistry similar to traditional tissue culture plastic. The packed bed was encased within a gas permeable shell to decouple the medium nutrient supply and gas exchange. This enabled a significant reduction in medium flow rates, thus reducing shear and even facilitating single pass medium exchange. The system was optimised in a small-scale bioreactor format (160 cm2) with murine-derived green fluorescent protein-expressing MSCs, and then scaled-up to a 2800 cm2 format. We demonstrated that placental derived MSCs could be isolated directly within the bioreactor and subsequently expanded. Our results demonstrate that the closed system large-scale packed bed bioreactor is an effective and scalable tool for large-scale isolation and expansion of MSCs.

Law of the Republic of Uzbekistan "On Protection of Mental Health in the Population of Uzbekistan"

The law provides rights for PWD for easy access of public goods, including education, social security, medical treatment, occupational and social rehabilitation and establishes an extent of responsibility of the government and its bodies for the creation of favourable conditions for the social adaptation of PWDs in market environment conditions.

Inter-seasonal population dynamics and pest status of Bemisia tabaci (Gennadius) biotype B in an Australian cropping system

Bemisia tabaci, biotype B, commonly known as the silverleaf whitefly (SLW) is an alien species that invaded Australia in the mid-90s. This paper reports on the invasion ecology of SLW and the factors that are likely to have contributed to the first outbreak of this major pest in an Australian cotton cropping system, population dynamics of SLW within whitefly-susceptible crop (cotton and cucurbit) and non-crop vegetation (sowthistle, Sonchus spp.) components of the cropping system were investigated over four consecutive growing seasons (September-June) 2001/02-2004/05 in the Emerald Irrigation Area (EIA) of Queensland, Australia. Based on fixed geo-referenced sampling sites, variation in spatial and temporal abundance of SLW within each system component was quantified to provide baseline data for the development of ecologically sustainable pest management strategies. Parasitism of large (3rd and 4th instars) SLW nymphs by native aphelinid wasps was quantified to determine the potential for natural control of SLW populations. Following the initial outbreak in 2001/02, SLW abundance declined and stabilised over the next three seasons. The population dynamics of SLW is characterised by inter-seasonal population cycling between the non-crop (weed) and cotton components of the EIA cropping system. Cotton was the largest sink for and source of SLW during the study period. Over-wintering populations dispersed from weed host plant sources to cotton in spring followed by a reverse dispersal in late summer and autumn to broad-leaved crops and weeds. A basic spatial source-sink analysis showed that SLW adult and nymph densities were higher in cotton fields that were closer to over-wintering weed sources throughout spring than in fields that were further away. Cucurbit fields were not significant sources of SLW and did not appear to contribute significantly to the regional population dynamics of the pest. Substantial parasitism of nymphal stages throughout the study period indicates that native parasitoid species and other natural enemies are important sources of SLW mortality in Australian cotton production systems. Weather conditions and use of broad-spectrum insecticides for pest control are implicated in the initial outbreak and on-going pest status of SLW in the region.

Modelling management options for management of feral camels in central Australia

Camels (Camelus dromedarius) were introduced into Australia from the 1840s to the early 1900s for transport and hauling cargo in arid regions. Feral populations remained small until the 1930s when many were released after they were superseded for transport by trucks and rail. Although camels have a relatively slow population growth (<10% per annum), the population has not reached carrying capacity and therefore, requires control to reduce the increasing impacts on central Australia. The model developed for the Northern Territory suggested that currently there are insufficient numbers being removed. The model also investigated which control options would have greatest impacts and found harvesting to be most important. The extent to which commercial harvesting can feasibly reduce camel populations requires further analysis. Due to the wide dispersal of camels in Australia, fertility control, even if technically feasible, will not target adults, the most important age class of the population. Habitat preferences were also investigated in the model but more validation is required as the population is still under range expansion. Immediate action is suggested to alleviate future costs as camel populations and their impacts rise.

Population dynamics of the European rabbit (Oryctolagus cuniculus) in north eastern Australia: simulated responses to control

Wild European rabbits are a serious problem to agriculture in Australia, with an estimated annual cost of A$ 113 million. Biological control agents (myxomatosis and rabbit haemorrhagic disease virus) have caused large and sustained declines in rabbit populations throughout Australia. A simulation model incorporates these diseases as well as warren destruction as methods of controlling rabbit populations in Queensland, north eastern Australia. These diseases reduced populations by 90-99% and the combination of these and warren destruction led to 100% control in simulations at six sites across southern Queensland. Increasing monthly pasture growth by 15% had little effect on simulated populations whereas a 15% decrease reduced populations by 0-50%. An increase in temperature of 2.5 °C would lead to a 15-60% decrease in populations. These effects suggest that climate change will lead to a decrease in the population of rabbits in Queensland and a retraction in the northern limit of their distribution in Australia.

Stochastic demography and population dynamics in the red kangaroo Macropus rufus

1. Many organisms inhabit strongly fluctuating environments but their demography and population dynamics are often analysed using deterministic models and elasticity analysis, where elasticity is defined as the proportional change in population growth rate caused by a proportional change in a vital rate. Deterministic analyses may not necessarily be informative because large variation in a vital rate with a small deterministic elasticity may affect the population growth rate more than a small change in a less variable vital rate having high deterministic elasticity. 2. We analyse a stochastic environment model of the red kangaroo (Macropus rufus), a species inhabiting an environment characterized by unpredictable and highly variable rainfall, and calculate the elasticity of the stochastic growth rate with respect to the mean and variability in vital rates. 3. Juvenile survival is the most variable vital rate but a proportional change in the mean adult survival rate has a much stronger effect on the stochastic growth rate. 4. Even if changes in average rainfall have a larger impact on population growth rate, increased variability in rainfall may still be important also in long-lived species. The elasticity with respect to the standard deviation of rainfall is comparable to the mean elasticities of all vital rates but the survival in age class 3 because increased variation in rainfall affects both the mean and variability of vital rates. 5. Red kangaroos are harvested and, under the current rainfall pattern, an annual harvest fraction of c. 20% would yield a stochastic growth rate about unity. However, if average rainfall drops by more than c. 10%, any level of harvesting may be unsustainable, emphasizing the need for integrating climate change predictions in population management and increase our understanding of how environmental stochasticity translates into population growth rate.

Demography of feral camels in central Australia and its relevance to population control

Since their release over 100 years ago, camels have spread across central Australia and increased in number. Increasingly, they are being seen as a pest, with observed impacts from overgrazing and damage to infrastructure such as fences. Irregular aerial surveys since 1983 and an interview-based survey in 1966 suggest that camels have been increasing at close to their maximum rate. A comparison of three models of population growth fitted to these, albeit limited, data suggests that the Northern Territory population has indeed been growing at an annual exponential rate of r = 0.074, or 8% per year, with little evidence of a density-dependent brake. A stage-structured model using life history data from a central Australian camel population suggests that this rate approximates the theoretical maximum. Elasticity analysis indicates that adult survival is by far the biggest influence on rate of increase and that a 9% reduction in survival from 96% is needed to stop the population growing. In contrast, at least 70% of mature females need to be sterilised to have a similar effect. In a benign environment, a population of large mammals such as camels is expected to grow exponentially until close to carrying capacity. This will frustrate control programs, because an ever-increasing number of animals will need to be removed for zero growth the longer that culling or harvesting effort is delayed. A population projection for 2008 suggests ~10 500 animals need to be harvested across the Northern Territory. Current harvests are well short of this. The ability of commercial harvesting to control camel populations in central Australia will depend on the value of animals, access to animals and the presence of alternative species to harvest when camels are at low density.

Population structure of South African and Australian Pyrenophora teres isolates.

There are two recognized forms of the disease net blotch of barley: the net form caused by Pyrenophora teres f. teres (PTT) and the spot form caused by P. teres f. maculata (PTM). In this study, amplified fragment length polymorphism analysis was used to investigate the genetic diversity and population structure of 60 PTT and 64 PTM isolates collected across Australia (66 isolates) and in the south-western Cape of South Africa (58 isolates). For comparison, P. tritici-repentis, Exserohilum rostratum and Bipolaris sorokiniana samples were also included in the analyses. Both distance-and model-based cluster analyses separated the PTT and PTM isolates into two strongly divergent genetic groups. Significant variation was observed both among the South African and Australian populations of PTT and PTM and among sampling locations for the PTT samples. Results suggest that sexual reproduction between the two forms is unlikely and that reproduction within the PTT and PTM groups occurs mainly asexually.

Impacts of grazing management options on pasture and animal productivity in a Heteropogon contortus (black speargrass) pasture in central Queensland. 2. Population dynamics of Heteropogon contortus and Stylosanthes scabra cv. Seca.

The dynamics of Heteropogon contortus and Stylosanthes scabra cv. Seca populations were studied in a subset of treatments in an extensive grazing study conducted in central Queensland between 1988 and 2001. These treatments were 4 stocking rates in native pasture and 2 of these stocking rates in legume oversown and supplement/spring burning treatments. For the 1999-2000 summer, population data for H. contortus in 5 of these native pasture and supplement/burning treatments were compared with those for an additional burnt treatment. Seasonal rainfall throughout this study was below the long-term mean and mean annual pasture utilisation ranged from 24 to 61%. Increasing stocking rate from 5 to 2 ha/steer in native pasture reduced H. contortus plant density. Increasing stocking rate reduced seedling recruitment as a result of its effect on soil seedbanks. Seedling recruitment was the major determinant of change in plant density, although some individual H. contortus plants did survive throughout the study. Burning in spring 1999, particularly at light stocking rate, promoted seedling recruitment above that in both unburnt native and legume oversown pasture and resulted in increased H. contortus plant density. In the legume oversown treatments, S. scabra cv. Seca density increased rapidly from 15 plants/m2 in 1988 to 140 plants/m2 in 2001 following a lag phase between 1988 and 1993. This increased S. scabra density was associated with an eventual decline in H. contortus plant density through reduced seedling recruitment. It was concluded that H. contortus population density is sustainable at stocking rates of 4 and 5 ha/steer (30% pasture utilisation) and that spring burning at light stocking rate can promote H. contortus populations. Increasing densities of S. scabra need to be managed to prevent its dominance.

Can we safely deform a plate to fit every bone? Population-based fit assessment and finite element deformation of a distal tibial plate

Anatomically precontoured plates are commonly used to treat periarticular fractures. A well-fitting plate can be used as a tool for anatomical reduction of the fractured bone. Recent studies highlighted that some plates fit poorly for many patients due to considerable shape variations between bones of the same anatomical site. While it is impossible to design one shape that fits all, it is also burdensome for the manufacturers and hospitals to produce, store and manage multiple plate shapes without the certainty of utilization by a patient population. In this study, we investigated the number of shapes required for maximum fit within a given dataset, and if they could be obtained by manually deforming the original plate. A distal medial tibial plate was automatically positioned on 45 individual tibiae, and the optimal deformation was determined iteratively using finite element analysis simulation. Within the studied dataset, we found that: (i) 89% fit could be achieved with four shapes, (ii) 100% fit was impossible through mechanical deformation, and (iii) the deformations required to obtain the four plate shapes were safe for the stainless steel plate for further clinical use. The proposed framework is easily transferable to other orthopaedic plates.

Intrapopulation variation in an Australian population of the North American thrips, Bagnalliella yuccae (Thysanoptera: Phlaeothripidae), a new record from Australia.

The genus Bagnalliella Karny is an endemic North American genus of Phlaeothripidae with 7 species associated with the New World plant genus Yucca; 2 Old World species currently placed in the genus are probably not congeneric. The number of sensoria on antennal segments III and IV has been used to distinguish the Bagnalliella species on Yucca, but an invasive population of Bagnalliella yuccae (Hinds) is reported here from Australia, at Brisbane, Queensland, in which the number of sensoria varied between individuals and even between left and right antennae of single individuals. These observations cast considerable doubt on the validity of some of the North American species of Bagnalliella. The Australian population was damaging young leaves of Yucca elephantipes, and indicates the ease with which thrips can be distributed by the horticultural trade.

Cellular and network mechanisms generating spontaneous population events in the immature rat hippocampus

Distinct endogenous network events, generated independently of sensory input, are a general feature of various structures of the immature central nervous system. In the immature hippocampus, these type of events are seen as "giant depolarizing potentials" (GDPs) in intracellular recordings in vitro. GABA, the major inhibitory neurotransmitter of the adult brain, has a depolarizing action in immature neurons, and GDPs have been proposed to be driven by GABAergic transmission. Moreover, GDPs have been thought to reflect an early pattern that disappears during development in parallel with the maturation of hyperpolarizing GABAergic inhibition. However, the adult hippocampus in vivo also generates endogenous network events known as sharp (positive) waves (SPWs), which reflect synchronous discharges of CA3 pyramidal neurons and are thought to be involved in cognitive functions. In this thesis, mechanisms of GDP generation were studied with intra- and extracellular recordings in the neonatal rat hippocampus in vitro and in vivo. Immature CA3 pyramidal neurons were found to generate intrinsic bursts of spikes and to act as cellular pacemakers for GDP activity whereas depolarizing GABAergic signalling was found to have a temporally non-patterned facilitatory role in the generation of the network events. Furthermore, the data indicate that the intrinsic bursts of neonatal CA3 pyramidal neurons and, consequently, GDPs are driven by a persistent Na+ current and terminated by a slow Ca2+-dependent K+ current. Gramicidin-perforated patch recordings showed that the depolarizing driving force for GABAA receptor-mediated actions is provided by Cl- uptake via the Na-K-C1 cotransporter, NKCC1, in the immature CA3 pyramids. A specific blocker of NKCC1, bumetanide, inhibited SPWs and GDPs in the neonatal rat hippocampus in vivo and in vitro, respectively. Finally, pharmacological blockade of the GABA transporter-1 prolonged the decay of the large GDP-associated GABA transients but not of single postsynaptic GABAA receptor-mediated currents. As a whole the data in this thesis indicate that the mechanism of GDP generation, based on the interconnected network of bursting CA3 pyramidal neurons, is similar to that involved in adult SPW activity. Hence, GDPs do not reflect a network pattern that disappears during development but they are the in vitro counterpart of neonatal SPWs.

Population structure, effective population size and adverse effects of stocking in the endangered Australian eastern freshwater cod Maccullochella ikei

Microsatellite markers were used to examine spatio-temporal genetic variation in the endangered eastern freshwater cod Maccullochella ikei in the Clarence River system, eastern Australia. High levels of population structure were detected. A model-based clustering analysis of multilocus genotypes identified four populations that were highly differentiated by F-statistics (FST = 0· 09 − 0· 49; P < 0· 05), suggesting fragmentation and restricted dispersal particularly among upstream sites. Hatchery breeding programmes were used to re-establish locally extirpated populations and to supplement remnant populations. Bayesian and frequency-based analyses of hatchery fingerling samples provided evidence for population admixture in the hatchery, with the majority of parental stock sourced from distinct upstream sites. Comparison between historical and contemporary wild-caught samples showed a significant loss of heterozygosity (21%) and allelic richness (24%) in the Mann and Nymboida Rivers since the commencement of stocking. Fragmentation may have been a causative factor; however, temporal shifts in allele frequencies suggest swamping with hatchery-produced M. ikei has contributed to the genetic decline in the largest wild population. This study demonstrates the importance of using information on genetic variation and population structure in the management of breeding and stocking programmes, particularly for threatened species.

Negligible evidence for regional genetic population structure for two shark species Rhizoprionodon acutus (Rüppell, 1837) and Sphyrna lewini (Griffith & Smith, 1834) with contrasting biology

Biodiversity of sharks in the tropical Indo-Pacific is high, but species-specific information to assist sustainable resource exploitation is scarce. The null hypothesis of population genetic homogeneity was tested for scalloped hammerhead shark (Sphyrna lewini, n = 237) and the milk shark (Rhizoprionodon acutus, n = 207) from northern and eastern Australia, using nuclear (S. lewini, eight microsatellite loci; R. acutus, six loci) and mitochondrial gene markers (873 base pairs of NADH dehydrogenase subunit 4). We were unable to reject genetic homogeneity for S. lewini, which was as expected based on previous studies of this species. Less expected were similar results for R. acutus, which is more benthic and less vagile than S. lewini. These features are probably driving the genetic break found between Australian and central Indonesian R. acutus (F-statistics; mtDNA, 0.751–0.903, respectively; microsatellite loci, 0.038–0.047 respectively). Our results support the spatially homogeneous monitoring and management plan for shark species in Queensland, Australia.

Evidence for extensive population structure in the white-spotted eagle ray within the Indo-Pacific inferred from mitochondrial gene sequences

The white-spotted eagle ray Aetobatus narinari is a species complex that occurs circumglobally throughout warm-temperate waters. Aetobatus narinari is semi-pelagic and large (up to 300 cm disc width), suggesting high dispersal capabilities and gene flow on a wide spatial scale. Sequence data from two mitochondrial genes, cytochrome b (cytb) and NADH dehydrogenase subunit 4 (ND4), were used to determine the genetic variability within and among 18 sampling locations in the central Indo-Pacific biogeographical region. Populations in the Indo-Pacific were highly genetically structured with c. 70% of the total genetic variation found among three geographical regions (East China Sea, Southeast Asia and Australia). FST was 0.64 for cytb and 0.53 for ND4, with φST values being even larger, that is, 0.78 for cytb and 0.65 for ND4. This high-level genetic partitioning provides strong evidence against extensive gene flow in A. narinari. The degree of genetic population structuring in the Indo-Pacific was similar to that found on a global scale. Global FST was 0.63 for cytb and 0.57 for ND4, and global φST values were 0.94 for cytb and 0.82 for ND4. This suggests that the A. narinari complex may be more speciose than the two or three species proposed to date. Further sampling and genetic analyses are likely to uncover the ‘evolutionarily significant’ and ‘management’ units that are critical to determine the susceptibilities of individual populations to regional fishing pressures and to provide advice on management options. Network analyses showed a close genetic relationship between haplotypes from the central Indo-Pacific and South Africa, providing support for a proposed dispersal pathway from the possible centre of origin of the A. narinari species complex in the Indo-Pacific into the Atlantic Ocean.