The dual orexin/hypocretin receptor antagonist, almorexant, in the ventral tegmental area attenuates ethanol self-administration

Recent studies have implicated the hypocretin/orexinergic system in reward-seeking behavior. Almorexant, a dual orexin/hypocretin R1 and R2 receptor antagonist, has proven effective in preclinical studies in promoting sleep in animal models and was in Phase III clinical trials for sleep disorders. The present study combines behavioral assays with in vitro biochemical and electrophysiological techniques to elucidate the role of almorexant in ethanol and sucrose intake. Using an operant self-administration paradigm, we demonstrate that systemic administration of almorexant decreased operant selfadministration of both 20% ethanol and 5% sucrose. We further demonstrate that intraventral tegmental area (VTA) infusions, but not intra substantia nigra infusions, of almorexant reduced ethanol self-administration. Extracellular recordings performed in VTA neurons revealed that orexin-A increased firing and this enhancement of firing was blocked by almorexant. The results demonstrate that orexin/hypocretin receptors in distinct brain regions regulate ethanol and sucrose mediated behaviors.

A review of the pharmacobiotic regulation of gastrointestinal inflammation by probiotics, commensal bacteria and prebiotics.

The idea that microbes induce disease has steered medical research toward the discovery of antibacterial products for the prevention and treatment of microbial infections. The twentieth century saw increasing dependency on antimicrobials as mainline therapy accentuating the notion that bacterial interactions with humans were to be avoided or desirably controlled. The last two decades, though, have seen a refocusing of thinking and research effort directed towards elucidating the critical inter-relationships between the gut microbiome and its host that control health/wellness or disease. This research has redefined the interactions between gut microbes and vertebrates, now recognizing that the microbial active cohort and its mammalian host have shared co-evolutionary metabolic interactions that span millennia. Microbial interactions in the gastrointestinal tract provide the necessary cues for the development of regulated pro- and anti-inflammatory signals that promotes immunological tolerance, metabolic regulation and other factors which may then control local and extra-intestinal inflammation. Pharmacobiotics, using nutritional and functional food additives to regulate the gut microbiome, will be an exciting growth area of therapeutics, developing alongside an increased scientific understanding of gut-microbiome symbiosis in health and disease.

Dual-frequency decoupling for two distinct antennas

Constant development of new wireless standards increases the demand for more radiating elements in compact end-user platforms. A decrease in antenna separation gives rise to increased antenna coupling, resulting in a reduction of the signal-to-interference-plus-noise-ratio (SINR) between transmitter and receiver. This paper proposes a decoupling network which provides dual band port isolation for a pair of distinct antennas. A prototype has been fabricated to verify the theory.

The relationship between drivers’ perceptions toward police speed enforcement and self-reported speeding behaviour

Exceeding the speed limit and driving too fast for the conditions are regularly cited as significant contributing factors in traffic crashes, particularly fatal and serious injury crashes. Despite an extensive body of research highlighting the relationship between increased vehicle speeds and crash risk and severity, speeding remains a pervasive behaviour on Australian roads. The development of effective countermeasures designed to reduce the prevalence of speeding behaviour requires that this behaviour is well understood. The primary aim of this program of research was to develop a better understanding of the influence of drivers’ perceptions and attitudes toward police speed enforcement on speeding behaviour. Study 1 employed focus group discussions with 39 licensed drivers to explore the influence of perceptions relating to specific characteristics of speed enforcement policies and practices on drivers’ attitudes towards speed enforcement. Three primary factors were identified as being most influential: site selection; visibility; and automaticity (i.e., whether the enforcement approach is automated/camera-based or manually operated). Perceptions regarding these enforcement characteristics were found to influence attitudes regarding the perceived legitimacy and transparency of speed enforcement. Moreover, misperceptions regarding speed enforcement policies and practices appeared to also have a substantial impact on attitudes toward speed enforcement, typically in a negative direction. These findings have important implications for road safety given that prior research has suggested that the effectiveness of speed enforcement approaches may be reduced if efforts are perceived by drivers as being illegitimate, such that they do little to encourage voluntary compliance. Study 1 also examined the impact of speed enforcement approaches varying in the degree of visibility and automaticity on self-reported willingness to comply with speed limits. These discussions suggested that all of the examined speed enforcement approaches (see Section 1.5 for more details) generally showed potential to reduce vehicle speeds and encourage compliance with posted speed limits. Nonetheless, participant responses suggested a greater willingness to comply with approaches operated in a highly visible manner, irrespective of the corresponding level of automaticity of the approach. While less visible approaches were typically associated with poorer rates of driver acceptance (e.g., perceived as “sneaky” and “unfair”), participants reported that such approaches would likely encourage long-term and network-wide impacts on their own speeding behaviour, as a function of the increased unpredictability of operations and increased direct (specific deterrence) and vicarious (general deterrence) experiences with punishment. Participants in Study 1 suggested that automated approaches, particularly when operated in a highly visible manner, do little to encourage compliance with speed limits except in the immediate vicinity of the enforcement location. While speed cameras have been criticised on such grounds in the past, such approaches can still have substantial road safety benefits if implemented in high-risk settings. Moreover, site-learning effects associated with automated approaches can also be argued to be a beneficial by-product of enforcement, such that behavioural modifications are achieved even in the absence of actual enforcement. Conversely, manually operated approaches were reported to be associated with more network-wide impacts on behaviour. In addition, the reported acceptance of such methods was high, due to the increased swiftness of punishment, ability for additional illegal driving behaviours to be policed and the salutary influence associated with increased face-to-face contact with authority. Study 2 involved a quantitative survey conducted with 718 licensed Queensland drivers from metropolitan and regional areas. The survey sought to further examine the influence of the visibility and automaticity of operations on self-reported likelihood and duration of compliance. Overall, the results from Study 2 corroborated those of Study 1. All examined approaches were again found to encourage compliance with speed limits, such that all approaches could be considered to be “effective”. Nonetheless, significantly greater self-reported likelihood and duration of compliance was associated with visibly operated approaches, irrespective of the corresponding automaticity of the approach. In addition, the impact of automaticity was influenced by visibility; such that significantly greater self-reported likelihood of compliance was associated with manually operated approaches, but only when they are operated in a less visible fashion. Conversely, manually operated approaches were associated with significantly greater durations of self-reported compliance, but only when they are operated in a highly visible manner. Taken together, the findings from Studies 1 and 2 suggest that enforcement efforts, irrespective of their visibility or automaticity, generally encourage compliance with speed limits. However, the duration of these effects on behaviour upon removal of the enforcement efforts remains questionable and represents an area where current speed enforcement practices could possibly be improved. Overall, it appears that identifying the optimal mix of enforcement operations, implementing them at a sufficient intensity and increasing the unpredictability of enforcement efforts (e.g., greater use of less visible approaches, random scheduling) are critical elements of success. Hierarchical multiple regression analyses were also performed in Study 2 to investigate the punishment-related and attitudinal constructs that influence self-reported frequency of speeding behaviour. The research was based on the theoretical framework of expanded deterrence theory, augmented with three particular attitudinal constructs. Specifically, previous research examining the influence of attitudes on speeding behaviour has typically focussed on attitudes toward speeding behaviour in general only. This research sought to more comprehensively explore the influence of attitudes by also individually measuring and analysing attitudes toward speed enforcement and attitudes toward the appropriateness of speed limits on speeding behaviour. Consistent with previous research, a number of classical and expanded deterrence theory variables were found to significantly predict self-reported frequency of speeding behaviour. Significantly greater speeding behaviour was typically reported by those participants who perceived punishment associated with speeding to be less certain, who reported more frequent use of punishment avoidance strategies and who reported greater direct experiences with punishment. A number of interesting differences in the significant predictors among males and females, as well as younger and older drivers, were reported. Specifically, classical deterrence theory variables appeared most influential on the speeding behaviour of males and younger drivers, while expanded deterrence theory constructs appeared more influential for females. These findings have important implications for the development and implementation of speeding countermeasures. Of the attitudinal factors, significantly greater self-reported frequency of speeding behaviour was reported among participants who held more favourable attitudes toward speeding and who perceived speed limits to be set inappropriately low. Disappointingly, attitudes toward speed enforcement were found to have little influence on reported speeding behaviour, over and above the other deterrence theory and attitudinal constructs. Indeed, the relationship between attitudes toward speed enforcement and self-reported speeding behaviour was completely accounted for by attitudes toward speeding. Nonetheless, the complexity of attitudes toward speed enforcement are not yet fully understood and future research should more comprehensively explore the measurement of this construct. Finally, given the wealth of evidence (both in general and emerging from this program of research) highlighting the association between punishment avoidance and speeding behaviour, Study 2 also sought to investigate the factors that influence the self-reported propensity to use punishment avoidance strategies. A standard multiple regression analysis was conducted for exploratory purposes only. The results revealed that punishment-related and attitudinal factors significantly predicted approximately one fifth of the variance in the dependent variable. The perceived ability to avoid punishment, vicarious punishment experience, vicarious punishment avoidance and attitudes toward speeding were all significant predictors. Future research should examine these relationships more thoroughly and identify additional influential factors. In summary, the current program of research has a number of implications for road safety and speed enforcement policy and practice decision-making. The research highlights a number of potential avenues for the improvement of public education regarding enforcement efforts and provides a number of insights into punishment avoidance behaviours. In addition, the research adds strength to the argument that enforcement approaches should not only demonstrate effectiveness in achieving key road safety objectives, such as reduced vehicle speeds and associated crashes, but also strive to be transparent and legitimate, such that voluntary compliance is encouraged. A number of potential strategies are discussed (e.g., point-to-point speed cameras, intelligent speed adaptation. The correct mix and intensity of enforcement approaches appears critical for achieving optimum effectiveness from enforcement efforts, as well as enhancements in the unpredictability of operations and swiftness of punishment. Achievement of these goals should increase both the general and specific deterrent effects associated with enforcement through an increased perceived risk of detection and a more balanced exposure to punishment and punishment avoidance experiences.

Development of a food frequency questionnaire for Sri Lankan adults

Abstract (provisional) Background Food Frequency Questionnaires (FFQs) are commonly used in epidemiologic studies to assess long-term nutritional exposure. Because of wide variations in dietary habits in different countries, a FFQ must be developed to suit the specific population. Sri Lanka is undergoing nutritional transition and diet-related chronic diseases are emerging as an important health problem. Currently, no FFQ has been developed for Sri Lankan adults. In this study, we developed a FFQ to assess the regular dietary intake of Sri Lankan adults. Methods A nationally representative sample of 600 adults was selected by a multi-stage random cluster sampling technique and dietary intake was assessed by random 24-h dietary recall. Nutrient analysis of the FFQ required the selection of foods, development of recipes and application of these to cooked foods to develop a nutrient database. We constructed a comprehensive food list with the units of measurement. A stepwise regression method was used to identify foods contributing to a cumulative 90% of variance to total energy and macronutrients. In addition, a series of photographs were included. Results We obtained dietary data from 482 participants and 312 different food items were recorded. Nutritionists grouped similar food items which resulted in a total of 178 items. After performing step-wise multiple regression, 93 foods explained 90% of the variance for total energy intake, carbohydrates, protein, total fat and dietary fibre. Finally, 90 food items and 12 photographs were selected. Conclusion We developed a FFQ and the related nutrient composition database for Sri Lankan adults. Culturally specific dietary tools are central to capturing the role of diet in risk for chronic disease in Sri Lanka. The next step will involve the verification of FFQ reproducibility and validity.

The impact of fatigue caused by sprint running on the median power frequency of the hamstring muscle group

Hamstring strain injuries (HSI) are the predominant non-contact injury in many sports. Intermittent running has been shown to result in preferential reductions in eccentric hamstring strength, which increase the risk of sustaining a HSI. The eccentric specific nature of this decline in hamstring function implicates central mechanisms, as peripheral fatigue mechanisms tend to impact upon both concentric and eccentric contractions modes. However, neural function of the hamstrings, such as the median power frequency (MPF) of the surface electromyography signal has yet to be examined in the fatigued hamstring following intermittent sprint running. AIM: To determine the impact of fatigue induced by intermittent sprinting on the MPF of the medial and lateral hamstring muscles. METHODS: Fifteen recreationally active males completed 18 × 20m overground sprints. Maximal strength (concentric and eccentric knee flexor and concentric knee extensor) was determined isokinetically at the velocities of ±180.s-1 and ±60.s- while hamstring muscle activation was assessed using surface electromyography, before and 15 minutes after the running protocol. RESULTS: Overground intermittent running caused a significant reduction in eccentric knee flexor strength (27.2 Nm; 95% CI = 11.2 to 43.3; p=0.0001) but not concentric strength (9.3 Nm; 95% CI = -6.7 to 25.3; P=0.6361). Following the overground running, MPF of the lateral hamstrings showed a significant decline eccentrically (0.86; 95% CI = 0.59 to 1.54; P=0.038) and concentrically (0.76; 95%CI = 0.66 to 0.83; P=0.039). Similar declines in MPF were also noted in the medial hamstrings eccentrically (1.54; 95% CI = 0.59 to 7.9; P=0.005) and concentrically (1.18; 95% CI = 0.44 to 6.8; P=0.040). CONCLUSION: Whilst sprint running induced fatigue led to a eccentric specific reduction in knee flexor torque, MPF was suppressed across both contraction modes. This would indicate that factors associated with the decline in MPF do not appear to explain the contraction mode-specific loss of strength after intermittent sprints. This would implicate other central mechanisms, such as declines in voluntary activation, in explaining the eccentric specific decline in strength seen following sprint running.

A three-nucleotide mutation altering the Maize streak virus Rep pRBR-interaction motif reduces symptom severity in maize and partially reverts at high frequency without restoring pRBR-Rep binding

Geminivirus infectivity is thought to depend on interactions between the virus replication-associated proteins Rep or RepA and host retinoblastoma-related proteins (pRBR), which control cell-cycle progression. It was determined that the substitution of two amino acids in the Maize streak virus (MSV) RepA pRBR-interaction motif (LLCNE to LLCLK) abolished detectable RepA-pRBR interaction in yeast without abolishing infectivity in maize. Although the mutant virus was infectious in maize, it induced less severe symptoms than the wild-type virus. Sequence analysis of progeny viral DNA isolated from infected maize enabled detection of a high-frequency single-nucleotide reversion of C(601)A in the 3 nt mutated sequence of the Rep gene. Although it did not restore RepA-pRBR interaction in yeast, sequence-specific PCR showed that, in five out of eight plants, the C(601)A reversion appeared by day 10 post-inoculation. In all plants, the C(601)A revertant eventually completely replaced the original mutant population, indicating a high selection pressure for the single-nucleotide reversion. Apart from potentially revealing an alternative or possibly additional function for the stretch of DNA that encodes the apparently non-essential pRBR-interaction motif of MSV Rep, the consistent emergence and eventual dominance of the C(601)A revertant population might provide a useful tool for investigating aspects of MSV biology, such as replication, mutation and evolution rates, and complex population phenomena, such as competition between quasispecies and population turnover. © 2005 SGM.

The governance of inter-organisational decision-making : understanding the impacts of informal networks on formal decision-making regimes in the Brisbane Airport region

Airports and cities inevitably recognise the value that each brings the other; however, the separation in decision-making authority for what to build, where, when and how provides a conundrum for both parties. Airports often want a say in what is developed outside of the airport fence, and cities often want a say in what is developed inside the airport fence. Defining how much of a say airports and cities have in decisions beyond their jurisdictional control is likely to be a topic that continues so long as airports and cities maintain separate formal decision-making processes for what to build, where, when and how. However, the recent Green and White Papers for a new National Aviation Policy have made early inroads to formalising relationships between Australia’s major airports and their host cities. At present, no clear indication (within practice or literature) is evident to the appropriateness of different governance arrangements for decisions to develop in situations that bring together the opposing strategic interests of airports and cities; thus leaving decisions for infrastructure development as complex decision-making spaces that hold airport and city/regional interests at stake. The line of enquiry is motivated by a lack of empirical research on networked decision-making domains outside of the realm of institutional theorists (Agranoff & McGuire, 2001; Provan, Fish & Sydow, 2007). That is, governance literature has remained focused towards abstract conceptualisations of organisation, without focusing on the minutia of how organisation influences action in real-world applications. A recent study by Black (2008) has provided an initial foothold for governance researchers into networked decision-making domains. This study builds upon Black’s (2008) work by aiming to explore and understand the problem space of making decisions subjected to complex jurisdictional and relational interdependencies. That is, the research examines the formal and informal structures, relationships, and forums that operationalise debates and interactions between decision-making actors as they vie for influence over deciding what to build, where, when and how in airport-proximal development projects. The research mobilises a mixture of qualitative and quantitative methods to examine three embedded cases of airport-proximal development from a network governance perspective. Findings from the research provide a new understanding to the ways in which informal actor networks underpin and combine with formal decision-making networks to create new (or realigned) governance spaces that facilitate decision-making during complex phases of development planning. The research is timely, and responds well to Isett, Mergel, LeRoux, Mischen and Rethemeyer’s (2011) recent critique of limitations within current network governance literature, specifically to their noted absence of empirical studies that acknowledge and interrogate the simultaneity of formal and informal network structures within network governance arrangements (Isett et al., 2011, pp. 162-166). The combination of social network analysis (SNA) techniques and thematic enquiry has enabled findings to document and interpret the ways in which decision-making actors organise to overcome complex problems for planning infrastructure. An innovative approach to using association networks has been used to provide insights to the importance of the different ways actors interact with one another, thus providing a simple yet valuable addition to the increasingly popular discipline of SNA. The research also identifies when and how different types of networks (i.e. formal and informal) are able to overcome currently known limitations to network governance (see McGuire & Agranoff, 2011), thus adding depth to the emerging body of network governance literature surrounding limitations to network ways of working (i.e. Rhodes, 1997a; Keast & Brown, 2002; Rethemeyer & Hatmaker, 2008; McGuire & Agranoff, 2011). Contributions are made to practice via the provision of a timely understanding of how horizontal fora between airports and their regions are used, particularly in the context of how they reframe the governance of decision-making for airport-proximal infrastructure development. This new understanding will enable government and industry actors to better understand the structural impacts of governance arrangements before they design or adopt them, particularly for factors such as efficiency of information, oversight, and responsiveness to change.

Variability, regularity and coupling measures distinguish PD tremor from voluntary 5Hz tremor

A characteristic of Parkinson's disease (PD) is the development of tremor within the 4–6 Hz range. One method used to better understand pathological tremor is to compare the responses to tremor-type actions generated intentionally in healthy adults. This study was designed to investigate the similarities and differences between voluntarily generated 4–6 Hz tremor and PD tremor in regards to their amplitude, frequency and coupling characteristics. Tremor responses for 8 PD individuals (on- and off-medication) and 12 healthy adults were assessed under postural and resting conditions. Results showed that the voluntary and PD tremor were essentially identical with regards to the amplitude and peak frequency. However, differences between the groups were found for the variability (SD of peak frequency, proportional power) and regularity (Approximate Entropy, ApEn) of the tremor signal. Additionally, coherence analysis revealed strong inter-limb coupling during voluntary conditions while no bilateral coupling was seen for the PD persons. Overall, healthy participants were able to produce a 5 Hz tremulous motion indistinguishable to that of PD patients in terms of peak frequency and amplitude. However, differences in the structure of variability and level of inter-limb coupling were found for the tremor responses of the PD and healthy adults. These differences were preserved irrespective of the medication state of the PD persons. The results illustrate the importance of assessing the pattern of signal structure/variability to discriminate between different tremor forms, especially where no differences emerge in standard measures of mean amplitude as traditionally defined.

Comparing the availability, price, variety and quality of fruits and vegetables across retail outlets and by area level socioeconomic position

Abstract Objective: To explore whether area-level socioeconomic position or the form of retail stream (conventional versus farmers’ market) are associated with differences in the price, availability, variety and quality of a range of fresh fruit and vegetables. Design: A multi-site cross-sectional pilot study of farmers’ markets, supermarkets and independent fruit and vegetable retailers. Each was surveyed to assess the price, availability, variety and quality of 15 fruit and 18 vegetable items. Setting: Retail outlets were located in South-East Queensland. Subjects: Fifteen retail outlets were surveyed (five of each retail stream). Results: Average basket prices were not significantly different across the socioeconomic spectrum however prices in low socioeconomic areas were cheapest. Availability, variety, and quality did not differ across levels of socioeconomic position however the areas with the most socioeconomic disadvantage scored poorest for quality and variety. Supermarkets had significantly better fruit and vegetable availability than farmers’ markets however price, variety and quality scores were not different across retail streams. Results demonstrate a trend to fruit and vegetable prices being more expensive at farmers’ markets, with the price of the Fruit basket being significantly greater at the organic farmer’s market compared with the non-organic farmers’ markets. Conclusions: Neither area-level socioeconomic position nor the form of retail stream was significantly associated with differences in the availability, price, variety and quality of fruit and vegetables, except for availability which was higher in supermarkets than farmers’ markets. Further research is needed to determine what role farmers’ markets can play in affecting fruit and vegetable intake.

An ingenuous Decision Support System (iDSS) approach for remote area pregnant women

This paper addresses development of an ingenious decision support system (iDSS) based on the methodology of survey instruments and identification of significant variables to be used in iDSS using statistical analysis. A survey was undertaken with pregnant women and factorial experimental design was chosen to acquire sample size. Variables with good reliability in any one of the statistical techniques such as Chi-square, Cronbach’s α and Classification Tree were incorporated in the iDSS. The ingenious decision support system was implemented with Visual Basic as front end and Microsoft SQL server management as backend. Outcome of the ingenious decision support system include advice on Symptoms, Diet and Exercise to pregnant women.

Within-subject intra- and inter-method consistency of two experimental risk attitude elicitation methods

We compare the consistency of choices in two methods to used elicit risk preferences on an aggregate as well as on an individual level. We asked subjects to choose twice from a list of nine decision between two lotteries, as introduced by Holt and Laury (2002, 2005) alternating with nine decisions using the budget approach introduced by Andreoni and Harbaugh (2009). We find that while on an aggregate(subject pool) level the results are (roughly) consistent, on an individual(within-subject) level,behavior is far from consistent. Within each method as well as across methods we observe low correlations. This again questions the reliability of experimental risk elicitation measures and the ability to use results from such methods to control for the risk aversion of subjects when explaining e�ects in other experimental games.

Transnational higher education programs for facilitating inter-university knowledge transfer : University of Indonesia’s experience

This paper explores the impacts and extent of knowledge transfer (KT) in an undergraduate engineering transnational program with an Australian university partner at the University of Indonesia (UI) using an inter-university KT conceptual framework (Sutrisno, Lisana, & Pillay 2012). For the purpose of this paper, the opportunity for KT in curriculum design is examined. Given the explicit nature of curriculum knowledge, assessing each partner’s curriculum was pivotal in allowing UI to enrich its own curriculum. The KT mechanism of face-to-face contact between Indonesian and Australian academics led to not only transfer of knowledge related to the curriculum of the undergraduate program but also to other cooperation beyond the transnational program in the form of joint research and joint supervision of post-graduate theses. Positive inter-university dynamics, such as trust and willingness to work together between the partners were underpinned by the presence of key actors from both sides at the earlier stages of the partnership. Retrospectively exploring the KT process in the UI’s transnational programs with its Australian partner suggests that there have been both structured and unstructured mechanisms, highlighting the ubiquitous and unbounded nature of KT between universities. While initially successful in facilitating KT, due to rapid succession of persons in charge of the program and the increasing focus on revenue generation, the useful lessons and practices unfortunately are being lost. Although the intention to use the transnational program for KT was always implied, it gradually was overlooked by newer staff members. Based on UI’s experience as the first provider of transnational program in Indonesia and other similar cases in China, seemingly transnational programs driven by short-term immediate financial return are unsuccessful in facilitating KT due to sensitivities to unfavourable economic situation. Those that remain operational and contribute to knowledge exchange between the partners apparently have genuine long-term engagement objective.

Linking three-dimensional geological modelling and multivariate statistical analysis to define groundwater chemistry baseline and identify inter-aquifer connectivity within the Clarence-Moreton Basin, Southeast Queensland, Australia

The Clarence-Moreton Basin (CMB) covers approximately 26000 km2 and is the only sub-basin of the Great Artesian Basin (GAB) in which there is flow to both the south-west and the east, although flow to the south-west is predominant. In many parts of the basin, including catchments of the Bremer, Logan and upper Condamine Rivers in southeast Queensland, the Walloon Coal Measures are under exploration for Coal Seam Gas (CSG). In order to assess spatial variations in groundwater flow and hydrochemistry at a basin-wide scale, a 3D hydrogeological model of the Queensland section of the CMB has been developed using GoCAD modelling software. Prior to any large-scale CSG extraction, it is essential to understand the existing hydrochemical character of the different aquifers and to establish any potential linkage. To effectively use the large amount of water chemistry data existing for assessment of hydrochemical evolution within the different lithostratigraphic units, multivariate statistical techniques were employed.