An innovative microplate assay to facilitate the detection of antimicrobial activity in plant extracts.

A microplate assay was modified for the detection of antimicrobial activity in plant extracts. The aim was to develop an in vitro assay that could rapidly screen plant extracts to provide quantitative data on inhibition of microbial growth. A spectrophotometric assay using a microplate with serial dilutions of the plant extract and the bacteria was developed. Two bacteria, Staphylococcus aureus and Escherichia coli, were used for this study. Essential oils, oregano (Origanum vulgare) and lemon myrtle (Backhousia citriodora), and three active components carvacrol, thymol and citral were evaluated. The reproducibility of the assay was high, with correlation coefficients (r aureus and E. coli between 0.9321 and 0.9816. Similarly, r and 0.9814. This assay could also be used to measure antimicrobial activity in plant extracts which vary in pH and color.

A Newton-Euler formulation for the inverse dynamics of the Stewart platform manipulator

This paper presents an inverse dynamic formulation by the Newton–Euler approach for the Stewart platform manipulator of the most general architecture and models all the dynamic and gravity effects as well as the viscous friction at the joints. It is shown that a proper elimination procedure results in a remarkably economical and fast algorithm for the solution of actuator forces, which makes the method quite suitable for on-line control purposes. In addition, the parallelism inherent in the manipulator and in the modelling makes the algorithm quite efficient in a parallel computing environment, where it can be made as fast as the corresponding formulation for the 6-dof serial manipulator. The formulation has been implemented in a program and has been used for a few trajectories planned for a test manipulator. Results of simulation presented in the paper reveal the nature of the variation of actuator forces in the Stewart platform and justify the dynamic modelling for control.

Early growth and adult health: focus on blood pressure, glucose tolerance status and body composition

Theory of developmental origins of adult health and disease proposes that experiences during critical periods of early development may have consequences on health throughout a lifespan. Thesis studies aimed to characterize associations between early growth and some components of the metabolic syndrome cluster. Participants belong to two epidemiological cohorts with data on birth measurements and, for the younger cohort, on serial recordings of weight and height during childhood. They were born as singletons between 1924-33 and 1934-44 in the Helsinki University Central Hospital, and 500 and 2003 of them, respectively, attended clinical studies at the age of 65-75 and 56-70 years, respectively. In the 65-75 year old men and women, the well-known inverse relationship between birth weight and systolic blood pressure (SBP) was confined to people who had established hypertension. Among them a 1-kg increase in birth weight was associated with a 6.4-mmHg (95% CI: 1.0 to 11.9) decrease in SBP. This relationship was further confined to people with the prevailing Pro12Pro polymorphism of the peroxisome proliferator-activated receptor-γ2 (PPARγ2) gene. People with low birth weight were more likely to receive angiotensin-converting enzyme inhibitors/angiotensin-receptor blockers (ACEI/ARB, p=0.03), and, again, this relationship was confined to the carriers of the Pro12Pro (p=0.01 for interaction). These results suggest that the inverse association between birth weight and systolic BP becomes focused in hypertensive people because pathological features of BP regulation, associated with slow fetal growth, become self-perpetuating in adult life. Insulin resistance of the Pro12Pro carriers with low birth weight may interact with the renin-angiotensin system leading to raised BP levels. Habitual physical activity protected men and women who were small at birth, and thus at increased risk for the development of type 2 diabetes, against glucose intolerance more strongly. Among subjects with birth weight ≤3000 g, the odds ratio (OR) for glucose intolerance was 5.2 (95% CI: 2.1 to 13) in those who exercised less than 3 times per week compared to regular exercisers; in those who scored their exercise light compared with moderate exercisers (defined as comparable to brisk walking) the OR was 3.5 (1.5 to 8.2). In the 56-70 year old men a 1 kg increase in birth weight corresponded to a 4.1 kg (95% CI: 3.1 to 5.1) and in women to a 2.9 kg (2.1 to 3.6) increase in adult lean mass. Rapid gain in body mass index (BMI), i.e. crossing from an original BMI percentile to a higher one, before the age of 2 years increased adult lean mass index (LMI, lean mass/height squared) without excess fat accumulation whereas rapid gain in BMI during later childhood, despite the concurrent rise in LMI, resulted in a relatively higher increase in adult body fat mass. These findings illustrate how genes, the environment and their interactions, early growth patterns, and adult lifestyle modify adult health risks which originate from early life.

Polymorphic microsatellite loci for primitively eusocial wasp Ropalidia marginata

We report here development and characterization of 48 novel microsatellite markers for Ropalidia marginata, a tropical, primitively eusocial polistine wasp from peninsular India. Thirty-two microsatellites showed polymorphism in a wild population of R. marginata (N = 38) collected from Bangalore, India. These markers will facilitate answering some interesting questions in ecology and evolutionary biology of this wasp, such as population structure, serial polygyny, intra-colony genetic relatedness and the pattern of queen succession.

Determinants of farmer retirement and farm succession in Finland

In the past decade, the Finnish agricultural sector has undergone rapid structural changes. The number of farms has decreased and the average farm size has increased when the number of farms transferred to new entrants has decreased. Part of the structural change in agriculture is manifested in early retirement programmes. In studying farmers exit behaviour in different countries, institutional differences, incentive programmes and constraints are found to matter. In Finland, farmers early retirement programmes were first introduced in 1974 and, during the last ten years, they have been carried out within the European Union framework for these programmes. The early retirement benefits are farmer specific and de-pend on the level of pension insurance the farmer has paid over his active farming years. In order to predict the future development of the agricultural sector, farmers have been frequently asked about their future plans and their plans for succession. However, the plans the farmers made for succession have been found to be time inconsistent. This study estimates the value of farmers stated succession plans in predicting revealed succession decisions. A stated succession plan exists when a farmer answers in a survey questionnaire that the farm is going to be transferred to a new entrant within a five-year period. The succession is revealed when the farm is transferred to a suc-cessor. Stated and revealed behaviour was estimated as a recursive Binomial Probit Model, which accounts for the censoring of the decision variables and controls for a potential correlation between the two equations. The results suggest that the succession plans, as stated by elderly farmers in the questionnaires, do not provide information that is significant and valuable in predicting true, com-pleted successions. Therefore, farmer exit should be analysed based on observed behaviour rather than on stated plans and intentions. As farm retirement plays a crucial role in determining the characteristics of structural change in agriculture, it is important to establish the factors which determine an exit from farming among eld-erly farmers and how off-farm income and income losses affect their exit choices. In this study, the observed choice of pension scheme by elderly farmers was analysed by a bivariate probit model. Despite some variations in significance and the effects of each factor, the ages of the farmer and spouse, the age and number of potential successors, farm size, income loss when retiring and the location of the farm together with the production line were found to be the most important determi-nants of early retirement and the transfer or closure of farms. Recently, the labour status of the spouse has been found to contribute significantly to individual retirement decisions. In this study, the effect of spousal retirement and economic incentives related to the timing of a farming couple s early retirement decision were analysed with a duration model. The results suggest that an expected pension in particular advances farm transfers. It was found that on farms operated by a couple, both early retirement and farm succession took place more often than on farms operated by a single person. However, the existence of a spouse delayed the timing of early retirement. Farming couples were found to co-ordinate their early retirement decisions when they both exit through agricultural retirement programmes, but such a co-ordination did not exist when one of the spouses retired under other pension schemes. Besides changes in the agricultural structure, the share and amount of off-farm income of a farm family s total income has also increased. In the study, the effect of off-farm income on farmers retirement decisions, in addition to other financial factors, was analysed. The unknown parameters were first estimated by a switching-type multivariate probit model and then by the simulated maxi-mum likelihood (SML) method, controlling for farmer specific fixed effects and serial correlation of the errors. The results suggest that elderly farmers off-farm income is a significant determinant in a farmer s choice to exit and close down the farm. However, off-farm income only has a short term effect on structural changes in agriculture since it does not significantly contribute to the timing of farm successions.

Infection of mungbean seed by Macrophomina phaseolina is more likely to result from localized pod infection than from systemic plant infection

The ubiquitous fungal pathogen Macrophomina phaseolina is best known as causing charcoal rot and premature death when host plants are subject to post-flowering stress. Overseas reports of M.phaseolina causing a rapid rot during the sprouting of Australian mungbean seed resulted in an investigation of the possible modes of infection of seed. Isolations from serial portions of 10 mungbean plants naturally infected with the pathogen revealed that on most plants there were discrete portions of infected tissue separated by apparently healthy tissue. The results from these studies, together with molecular analysis of isolates collected from infected tissue on two of the plants, suggested that aerial infection of aboveground parts by different isolates is common. Inoculations of roots and aboveground parts of mungbean plants at nine temperaturexsoil moisture incubation combinations and of detached green pods strongly supported the concept that seed infection results from infection of pods by microsclerotia, rather than from hyphae growing systemically through the plant after root or stem infection. This proposal is reinforced by anecdotal evidence that high levels of seed infection are common when rainfall occurs during pod fill, and by the isolation of M.phaseolina from soil peds collected on pods of mungbean plants in the field. However, other experiments showed that when inoculum was placed within 130mm of a green developing pod and a herbicide containing paraquat and diquat was sprayed on the inoculated plants, M.phaseolina was capable of some systemic growth from vegetative tissue into the pods and seeds.

Experimental networking of microcomputers and a minicomputer

The objective of this paper is to discuss some hardware and software features of an experimental network of 8080 and 8085 microcomputers named Micronet. The interprocessor communication in the ring network is established using ring interfaces consisting of universal synchronous-asynchronous receivers-transmitters (USARTs). Another aspect considered is the interfacing of an 8080 microcomputer to a PDP-11/35 minicomputer and the development of the software for the microcomputer-minicomputer link which has been established over a serial line using the USART interface of the microcomputer and the DZ11 module of the minicomputer. This is useful in developing a host-satellite configuration of microcomputers and the minicomputer.

DNP3 network scanning and reconnaissance for critical infrastructure

The Distributed Network Protocol v3.0 (DNP3) is one of the most widely used protocols to control national infrastructure. The move from point-to-point serial connections to Ethernet-based network architectures, allowing for large and complex critical infrastructure networks. However, networks and con- figurations change, thus auditing tools are needed to aid in critical infrastructure network discovery. In this paper we present a series of intrusive techniques used for reconnaissance on DNP3 critical infrastructure. Our algorithms will discover DNP3 outstation slaves along with their DNP3 addresses, their corresponding master, and class object configurations. To validate our presented DNP3 reconnaissance algorithms and demonstrate it’s practicality, we present an implementation of a software tool using a DNP3 plug-in for Scapy. Our implementation validates the utility of our DNP3 reconnaissance technique. Our presented techniques will be useful for penetration testing, vulnerability assessments and DNP3 network discovery.

Change to costs and lengths of stay in the emergency department and the Brisbane protocol: An observational study

- Objective To compare health service cost and length of stay between a traditional and an accelerated diagnostic approach to assess acute coronary syndromes (ACS) among patients who presented to the emergency department (ED) of a large tertiary hospital in Australia. - Design, setting and participants This historically controlled study analysed data collected from two independent patient cohorts presenting to the ED with potential ACS. The first cohort of 938 patients was recruited in 2008–2010, and these patients were assessed using the traditional diagnostic approach detailed in the national guideline. The second cohort of 921 patients was recruited in 2011–2013 and was assessed with the accelerated diagnostic approach named the Brisbane protocol. The Brisbane protocol applied early serial troponin testing for patients at 0 and 2 h after presentation to ED, in comparison with 0 and 6 h testing in traditional assessment process. The Brisbane protocol also defined a low-risk group of patients in whom no objective testing was performed. A decision tree model was used to compare the expected cost and length of stay in hospital between two approaches. Probabilistic sensitivity analysis was used to account for model uncertainty. - Results Compared with the traditional diagnostic approach, the Brisbane protocol was associated with reduced expected cost of $1229 (95% CI −$1266 to $5122) and reduced expected length of stay of 26 h (95% CI −14 to 136 h). The Brisbane protocol allowed physicians to discharge a higher proportion of low-risk and intermediate-risk patients from ED within 4 h (72% vs 51%). Results from sensitivity analysis suggested the Brisbane protocol had a high chance of being cost-saving and time-saving. - Conclusions This study provides some evidence of cost savings from a decision to adopt the Brisbane protocol. Benefits would arise for the hospital and for patients and their families.

Open source computer-vision based guidance system for UAVs on-board decision making

The use of UAVs for remote sensing tasks; e.g. agriculture, search and rescue is increasing. The ability for UAVs to autonomously find a target and perform on-board decision making, such as descending to a new altitude or landing next to a target is a desired capability. Computer-vision functionality allows the Unmanned Aerial Vehicle (UAV) to follow a designated flight plan, detect an object of interest, and change its planned path. In this paper we describe a low cost and an open source system where all image processing is achieved on-board the UAV using a Raspberry Pi 2 microprocessor interfaced with a camera. The Raspberry Pi and the autopilot are physically connected through serial and communicate via MAVProxy. The Raspberry Pi continuously monitors the flight path in real time through USB camera module. The algorithm checks whether the target is captured or not. If the target is detected, the position of the object in frame is represented in Cartesian coordinates and converted into estimate GPS coordinates. In parallel, the autopilot receives the target location approximate GPS and makes a decision to guide the UAV to a new location. This system also has potential uses in the field of Precision Agriculture, plant pest detection and disease outbreaks which cause detrimental financial damage to crop yields if not detected early on. Results show the algorithm is accurate to detect 99% of object of interest and the UAV is capable of navigation and doing on-board decision making.

Phylogeography and Population Genetics of the Moor frog (Rana arvalis Nilsson) in Northern Europe

The ongoing climate change along with increasing levels of pollutants, diseases, habitat loss and fragmentation constitute global threats to the persistence of many populations, species and ecosystems. However, for the long-term persistence of local populations, one of the biggest threats is the intrinsic loss of genetic variation. In order to adapt to changes in the environment, organisms must have a sufficient supply of heritable variation in traits important for their fitness. With a loss of genetic variation, the risk of extinction will increase. For conservational practices, one should therefore understand the processes that shape the genetic population structure and also the broader (historical) phylogenetic patterning of the species in focus. In this thesis, microsatellite markers were applied to study genetic diversity and population differentiation of the protected moor frog (Rana arvalis) in Fennoscandia from both historical (evolutionary) and applied (conservation) perspectives. The results demonstrate that R. arvalis populations are highly structured over rather short geographic distances. Moreover, the results suggest that R. arvalis recolonized Fennoscandia from two directions after the last ice age. This has had implications for the genetic structuring and population differentiation, especially in the northernmost parts where the two lineages have met. Compared to more southern populations, the genetic variation decreases and the interpopulation differentiation increases dramatically towards north. This could be an outcome of serial population bottlenecking along the recolonization route. Also, current isolation and small population sizes increase the effect of drift, thus reinforcing the observed pattern. The same pattern can also be seen in island populations. However, though R. arvalis on the island of Gotland has lost most of its neutral genetic variability, our results indicate that the levels of additive genetic variation have remained high. This conforms to the conjecture that though neutral markers are widely used in conservation purposes, they may be quite uninformative about the levels of genetic variation in ecologically important traits. Finally, the evolutionary impact of the typical amphibian mating behaviour on genetic diversity was investigated. Given the short time available for larval development, it is important that mating takes place as early as possible. The genetic data and earlier capture-recapture data suggest that R. arvalis gather at mating grounds they are familiar with. However, by forming leks in random to relatedness, and having multiple paternities in single clutches, the risk of inbreeding may be minimized in this otherwise highly philopatric species.

Spatial variation of sequestered calcium in the multicellular stage of Dictyostelium discoideum as assayed by chlortetracycline fluorescence

Abstract. We have used chlortetracycline (CTC) as a fluorescent probe to detect the distribution of sequestered calcium in multicellular stages of Dictyostelium discoideum. Tips of late aggregates, slugs and early culminating masses fluoresce very strongly. Most of the fluorescence is intracellular in origin and emanates from a small number of intense punctate sources. The sources correspond in part to autophagic vacuoles viz. neutral-red staining, acidic digestive vesicles, and may also include intracellular organelles; cytoplasmic fluorescence is much weaker in comparison. The level of fluorescence drops in the middle portion of slugs and rises again in the posteriormost region, though not to as high a level as in the tip. This holds good irrespective of whether CTC is applied only in the neighbourhood of the aggregate centre, only in the aggregate periphery, or to the whole aggregate. We infer that there must be a good deal of mixing in the stages leading from aggregation to slug formation; thus the serial order in which cells enter an aggregate does not bear any relation to their ultimate fates. The other implication of our study is that calcium sequestration is much more extensive in prestalk and anterior-like cells than in prespore cells. These findings are discussed with regard to possible implications for pattern formation.

Mechanisms and molecular regulation of mammalian tooth replacement

In most non-mammalian vertebrates, such as fish and reptiles, teeth are replaced continuously. However, tooth replacement in most mammals, including human, takes place only once and further renewal is apparently inhibited. It is not known how tooth replacement is genetically regulated, and little is known on the physiological mechanism and evolutionary reduction of tooth replacement in mammals. In this study I have attempted to address these questions. In a rare human condition cleidocranial dysplasia, caused by a mutation in a Runt domain transcription factor Runx2, tooth replacement is continued. Runx2 mutant mice were used to investigate the molecular mechanisms of Runx2 function. Microarray analysis from dissected embryonic day 14 Runx2 mutant and wild type dental mesenchymes revealed many downstream targets of Runx2, which were validated using in situ hybridization and tissue culture methods. Wnt signaling inhibitor Dkk1 was identified as a candidate target, and in tissue culture conditions it was shown that Dkk1 is induced by FGF4 and this induction is Runx2 dependent. These experiments demonstrated a connection between Runx2, FGF and Wnt signaling in tooth development and possibly also in tooth replacement. The role of Wnt signaling in tooth replacement was further investigated by using a transgenic mouse model where Wnt signaling mediator β-catenin is continuously stabilized in dental epithelium. This stabilization led to activated Wnt signaling and to the formation of multiple enamel knots. In vitro and transplantation experiments were performed to examine the process of extra tooth formation. We showed that new teeth were continuously generated and that new teeth form from pre-existing teeth. A morphodynamic activator-inhibitor model was used to simulate enamel knot formation. By increasing the intrinsic production rate of the activator (β-catenin), the multiple enamel knot phenotype was reproduced by computer simulations. It was thus concluded that β-catenin acts as an upstream activator of enamel knots, closely linking Wnt signaling to the regulation of tooth renewal. As mice do not normally replace teeth, we used other model animals to investigate the physiological and genetic mechanisms of tooth replacement. Sorex araneus, the common shrew was earlier reported to have non-functional tooth replacement in all antemolar tooth positions. We showed by histological and gene expression studies that there is tooth replacement only in one position, the premolar 4 and that the deciduous tooth is diminished in size and disappears during embryogenesis without becoming functional. The growth rates of deciduous and permanent premolar 4 were measured and it was shown by competence inference that the early initiation of the replacement tooth in relation to the developmental stage of the deciduous tooth led to the inhibition of deciduous tooth morphogenesis. It was concluded that the evolutionary loss of deciduous teeth may involve the early activation of replacement teeth, which in turn suppress their predecessors. Mustela putorius furo, the ferret, has a dentition that resembles that of the human as ferrets have teeth that belong to all four tooth families, and all the antemolar teeth are replaced once. To investigate the replacement mechanism, histological serial sections from different embryonic stages were analyzed. It was noticed that tooth replacement is a process which involves the growth and detachment of the dental lamina from the lingual cervical loop of the deciduous tooth. Detachment of the deciduous tooth leads to a free successional dental lamina, which grows deeper into the mesenchyme, and later buds the replacement tooth. A careful 3D analysis of serial histological sections was performed and it was shown that replacement teeth are initiated from the successional dental lamina and not from the epithelium of the deciduous tooth. The molecular regulation of tooth replacement was studied and it was shown by examination of expression patterns of candidate regulatory genes that BMP/Wnt inhibitor Sostdc1 was strongly expressed in the buccal aspect of the dental lamina, and in the intersection between the detaching deciduous tooth and the successional dental lamina, suggesting a role for Sostdc1 in the process of detachment. Shh was expressed in the enamel knot and in the inner enamel epithelium in both generations of teeth supporting the view that the morphogenesis of both generations of teeth is regulated by similar mechanisms. In summary, histological and molecular studies on different model animals and transgenic mouse models were used to investigate tooth replacement. This thesis work has significantly contributed to the knowledge on the physiological mechanisms and molecular regulation of tooth replacement and its evolutionary suppression in mammals.

Ferrous iron oxidation by foam immobilized Acidithiobacillus ferrooxidans: Experiments and modeling

Ferrous iron bio-oxidation by Acidithiobacillus ferrooxidans immobilized on polyurethane foam was investigated. Cells were immobilized on foams by placing them in a growth environment and fully bacterially activated polyurethane foams (BAPUFs) were prepared by serial subculturing in batches with partially bacterially activated foam (pBAPUFs). The dependence of foam density on cell immobilization process, the effect of pH and BAPUF loading on ferrous oxidation were studied to choose operating parameters for continuous operations. With an objective to have high cell densities both in foam and the liquid phase, pretreated foams of density 50 kg/m3 as cell support and ferrous oxidation at pH 1.5 to moderate the ferric precipitation were preferred. A novel basket-type bioreactor for continuous ferrous iron oxidation, which features a multiple effect of stirred tank in combination with recirculation, was designed and operated. The results were compared with that of a free cell and a sheet-type foam immobilized reactors. A fivefold increase in ferric iron productivity at 33.02 g/h/L of free volume in foam was achieved using basket-type bioreactor when compared to a free cell continuous system. A mathematical model for ferrous iron oxidation by Acidithiobacillus ferrooxidans cells immobilized on polyurethane foam was developed with cell growth in foam accounted by an effectiveness factor. The basic parameters of simulation were estimated using the experimental data on free cell growth as well as from cell attachment to foam under nongrowing conditions. The model predicted the phase of both oxidation of ferrous in shake flasks by pBAPUFs as well as by fully activated BAPUFs for different cell loadings in foam. Model for stirred tank basket bioreactor predicted within 5% both transient and steady state of the experiments closely for the simulated dilution rates. Bio-oxidation at high Fe2+ concentrations were simulated with experiments when substrate and product inhibition coefficients were factored into cell growth kinetics.

Inferring neuronal network connectivity from spike data: A temporal data mining approach

Understanding the functioning of a neural system in terms of its underlying circuitry is an important problem in neuroscience. Recent d evelopments in electrophysiology and imaging allow one to simultaneously record activities of hundreds of neurons. Inferring the underlying neuronal connectivity patterns from such multi-neuronal spike train data streams is a challenging statistical and computational problem. This task involves finding significant temporal patterns from vast amounts of symbolic time series data. In this paper we show that the frequent episode mining methods from the field of temporal data mining can be very useful in this context. In the frequent episode discovery framework, the data is viewed as a sequence of events, each of which is characterized by an event type and its time of occurrence and episodes are certain types of temporal patterns in such data. Here we show that, using the set of discovered frequent episodes from multi-neuronal data, one can infer different types of connectivity patterns in the neural system that generated it. For this purpose, we introduce the notion of mining for frequent episodes under certain temporal constraints; the structure of these temporal constraints is motivated by the application. We present algorithms for discovering serial and parallel episodes under these temporal constraints. Through extensive simulation studies we demonstrate that these methods are useful for unearthing patterns of neuronal network connectivity.