Standard-volume polyethylene glycol (PEG) gut lavage solutions are safe and effective, but they require the consumption of large volumes of fluid. A new lower-volume solution of PEG plus ascorbic acid has been used recently as a preparation for colonoscopy.A meta-analysis was performed to compare the performance of low-volume PEG plus ascorbic acid with standard-volume PEG as bowel preparation for colonoscopy.Electronic and manual searches were performed to identify randomized controlled trials (RCTs) that compared the performance of low-volume PEG plus ascorbic acid with standard-volume PEG as bowel preparation for colonoscopy. After a methodological quality assessment and data extraction, the pooled estimates of bowel preparation efficacy during bowel cleansing, compliance with preparation, willingness to repeat the same preparation, and the side effects were calculated. We calculated pooled estimates of odds ratios (OR) by fixed- and/or random-effects models. We also assessed heterogeneity among studies and the publication bias.Eleven RCTs were identified for analysis. The pooled OR for preparation efficacy during bowel cleansing and for compliance with preparation for low-volume PEG plus ascorbic acid were 1.08  and 2.23 , respectively, compared with those for standard-volume PEG. The side effects of vomiting and nausea for low-volume PEG plus ascorbic acid were reduced relative to standard-volume PEG. There was no significant publication bias, according to a funnel plot.Low-volume PEG plus ascorbic acid gut lavage achieved non-inferior efficacy for bowel cleansing, is more acceptable to patients, and has fewer side effects than standard-volume PEG as a bowel preparation method for colonoscopy. Colorectal cancer, a major cause of cancer-associated morbidity and mortality, is one of the most common cancers Thus, the level of bowel cleansing, to a large extent, determines the success of colonoscopy The quality of bowel preparation is dependent on patient compliance, the type of bowel preparation, and the timing of ingestion Sodium phosphate solution (SPS) has been used since the 1990s An alternative preparation of polyethylene glycol (PEG)-based gut lavage, which was introduced in 1980 Recently, combining ascorbic acid, a laxative, with PEG showed potential to reduce the volume necessary for effective colonic cleansing, while possibly improving tolerance. The absorption of ascorbic acid reaches saturation at high doses In the present study, we performed a systematic review and meta-analysis to qualitatively and quantitatively summarize previous high-quality RCTs that compared low-volume PEG plus ascorbic acid with standard-volume PEG preparation in terms of bowel preparation quality. We also sought to statistically summarize secondary outcomes, such as compliance with the preparation, the willingness of the patient to repeat the same bowel preparation, and side effects.surgical endoscopy and other interventional techniques, gastrointestinal endoscopy, endoscopy, digestive endoscopy and diseases of the colon and rectum. The abstracts submitted to the Digestive Disease Week and the ACG national meeting, were also manually searched for accuracy and completeness of data retrieval. The literature search was performed using the following terms: 2L or low-volume polyethylene glycol plus ascorbic acid and 4 L or standard-volume polyethylene glycol and colonoscopy.A computerized search was performed by two independent investigators (W-L.W. and Q-S.X.) in PubMed/Medline, EMBASE, the Cochrane Library, and Google Scholar to identify relevant articles published between 2000 and 2013. We scanned publishers’ databases and conducted manual searches among Two reviewers (W-L.W. and Q-S.X.) read the titles and abstracts of original articles that compared the performance of low-volume PEG plus ascorbic acid with standard-volume PEG as the preparation method for colonoscopy to select potentially relevant articles. All of the selected articles were collected and reviewed independently by the same reviewers to determine their eligibility for detailed analysis. The inclusion criteria were: (i) randomized controlled trials (RCTs), (ii) adult patients undergoing elective colonoscopy, and (iii) using 2 L PEG plus ascorbic acid and 4 L PEG preparations. Exclusion criteria were duplicate publications (based on the same primary study) and a lack of categorical data on preparation quality or adherence. Review articles, editorials, letters to the editor, and articles enrolling patients younger than 18 years old were also excluded. Disagreements between the two reviewers regarding study selection were resolved by consensus after a face-to-face discussion. If data were missing from a study, the investigator was contacted to provide the missing data if possible. Each study was evaluated by a Jada score The primary outcome was bowel preparation efficacy. This was prespecified as an Ottawa score less than 5, or an excellent or good bowel preparation designation on the Aronchik scale or other non-validated 3-, 4-, or 5-point scales . As an assessment of bowel preparation tolerability and side effects, a patient’s subjective evaluation of their level of satisfaction and acceptability of the bowel preparation was recorded by studies that administered a periprocedural non-standardized questionnaire to the patient. Data for the secondary outcomes were extracted from the results of these questionnaires. Compliance with bowel preparation was defined as adherence to the bowel preparation prescribed or consumption of at least 75% of the prescribed bowel preparation. The additional secondary outcomes of willingness to repeat the same bowel preparation and side effects, including abdominal cramping/pain, abdominal bloating, vomiting, and nausea, represented affirmative responses to the relevant question from the questionnaires.I2 measure of inconsistency, which was considered significant if I2>50%. The RevMan 5.2 software from the Cochrane Collaboration was used for the statistical analysis.A meta-analysis was performed comparing low-volume PEG plus ascorbic acid solution with standard-volume PEG solution as bowel preparation for colonoscopy by calculating pooled estimates of the quality of the bowel preparation, compliance with the preparation, willingness to repeat the same preparation, and side effects using the odds ratio (OR) with fixed- or random-effects models. Publication bias was assessed using funnel plots. Heterogeneity among studies was assessed by calculating the I2 = 42%), showing no statistically significant difference, indicating that low-volume PEG plus ascorbic acid solution achieved equal bowel preparation efficacy compared with standard-volume PEG.The forest plot in I2 = 30%), compared with standard-volume PEG , 0.82 , 1.10 , 1.00 , 0.74 , and 0.80 , compared with standard-volume PEG, respectively  has similar bowel preparation efficacy to and better tolerability than that of PEG Because the ideal bowel cleanser is effective, safe for all patient groups, and acceptable to patients, the combined use of low-volume (2 L) PEG and ascorbic acid was recently introduced into clinical practice. However, several investigators have argued that low-volume PEG provides inadequate cleansing of the upper colon and significantly worse bowel preparation than either standard-volume PEG or SP I2 = 42%). The two studies comparing low-volume PEG plus ascorbic acid with standard-volume PEG plus simethicone also reported similar bowel preparation quality. This non-inferior efficacy in bowel cleansing benefited substantially from the synergistic osmotic effect achieved with low-volume PEG Compared to Godfrey’s study, in the present study, we included 2 more RCTs and confirmed by meta-analysis that low-volume PEG plus ascorbic acid solution achieved the same bowel preparation efficacy as did standard-volume PEG . The individual studies Colonoscopy is increasingly being used to screen healthy patients (or those with relatively minor symptoms) for bowel cancer In the present study, we also compared overall adverse events, abdominal cramping/pain and bloating, vomiting, and nausea between the low-volume PEG plus ascorbic acid and the standard-volume PEG groups. We found that there was no significant difference in abdominal cramping/pain or bloating between the groups. However, we did see that the low-volume group had significantly fewer overall adverse events and less vomiting and nausea than did the standard-volume group. This significant difference may result from the safety of ascorbic acid, even at high doses In summary, optimal bowel cleansing should be not only effective but also safe for all patients. With non-inferior bowel preparation efficacy and the advantages of fewer adverse events and better compliance, the low-volume PEG plus ascorbic acid solution was clearly more applicable to bowel preparation for colonoscopy.Checklist S1PRISMA checklist.(DOC)Click here for additional data file.


Implementation of user-friendly, real-time, electronic medical records for patient management may lead to improved adherence to clinical guidelines and improved quality of patient care. We detail the systematic, iterative process that implementation partners, Lighthouse clinic and Baobab Health Trust, employed to develop and implement a point-of-care electronic medical records system in an integrated, public clinic in Malawi that serves HIV-infected and tuberculosis (TB) patients.Baobab Health Trust, the system developers, conducted a series of technical and clinical meetings with Lighthouse and Ministry of Health to determine specifications. Multiple pre-testing sessions assessed patient flow, question clarity, information sequencing, and verified compliance to national guidelines. Final components of the TB/HIV electronic medical records system include: patient demographics; anthropometric measurements; laboratory samples and results; HIV testing; WHO clinical staging; TB diagnosis; family planning; clinical review; and drug dispensing.Our experience suggests that an electronic medical records system can improve patient management, enhance integration of TB/HIV services, and improve provider decision-making. However, despite sufficient funding and motivation, several challenges delayed system launch including: expansion of system components to include of HIV testing and counseling services; changes in the national antiretroviral treatment guidelines that required system revision; and low confidence to use the system among new healthcare workers. To ensure a more robust and agile system that met all stakeholder and user needs, our electronic medical records launch was delayed more than a year. Open communication with stakeholders, careful consideration of ongoing provider input, and a well-functioning, backup, paper-based TB registry helped ensure successful implementation and sustainability of the system. Additional, on-site, technical support provided reassurance and swift problem-solving during the extended launch period.Even when system users are closely involved in the design and development of an electronic medical record system, it is critical to allow sufficient time for software development, solicitation of detailed feedback from both users and stakeholders, and iterative system revisions to successfully transition from paper to point-of-care electronic medical records. For those in low-resource settings, electronic medical records for integrated care is a possible and positive innovation.The online version of this article (doi:10.1186/s13104-016-1943-4) contains supplementary material, which is available to authorized users. As the numbers of patients co-infected with human immunodeficiency virus (HIV) and tuberculosis (TB) increase in sub-Saharan Africa, reliance on paper-based registers may result in poor record management and, ultimately, suboptimal clinical care. One innovative approach to improve data quality and strengthen the continuum of care is the implementation of user-friendly, real-time electronic medical record (EMR) system. EMR systems may be well received by clinicians , improveIn 2005, Lighthouse Trust, the largest public provider of antiretroviral therapy (ART) and TB services in Malawi’s central region, in partnership with Baobab Health Trust, a local non-governmental organisation that leads the development of EMR system in the country, developed and launched an ART-focused EMR system. By 2006, both of Lighthouse Trust’s clinics, Lighthouse and Martin Preuss centre (MPC), successfully employed the real-time, touchscreen-based EMR system to manage ART patients. Data management for more than 36,000 ART patients at both clinics is now entirely based on the EMR system resulting in near completeness of patient level data. Due to the EMR system success in improving both patient management and reliable record keeping, described previously [Although the current ART EMR system in Malawi is successful, one significant limitation is its singular programmatic focus. Nationally, almost 60 % of TB patients are co-infected with HIV . At MPC In 2010, Lighthouse Trust, with its partners Baobab Health Trust, the Malawi National TB Program (NTP), and the IeDEA Collaboration at the Institute of Social and Preventive Medicine at University of Bern in Switzerland, received a one-year innovation development grant from US President’s Emergency Plan for AIDS Relief (PEPFAR) to develop and implement a real-time, touchscreen-based EMR system for TB and TB/HIV integrated care. The TB/HIV EMR system is expected to significantly increase service quality and facilitate program monitoring at MPC. If proven beneficial, the system will expand from MPC to other clinics.This paper shares insights and lessons learned from our three-phased approach to planning, developing, and implementing the point-of-care TB/HIV EMR system for TB and TB/HIV co-infected patient management implemented at MPC clinic. We first detail the steps taken in conjunction with developers, system users and local stakeholders during system planning. Then, we note the system components designed to aid patient care followed by our experiences in system deployment. Then, we discuss the delays and unexpected challenges we overcame during implementation. These experiences and lessons may assist providers, clinics and organizations in other resource limited settings that are considering developing and deploying TB/HIV EMR systems.The Malawi national Health Science Research Committee (NHSRC) provided a general blanket approval for the use of routine programmatic data for monitoring and evaluation every year; and as such, a separate formal ethics approval was not required from NHSRC.https://github.com/baobabhealthtrust/TB-ART.The TB/HIV EMR system and its components are Open Source. The code and system components are available for non-commercial, scientific use at While the technical processes were being discussed with management and technical staff at Lighthouse and Baobab, a series of concurrent meetings were conducted by Lighthouse staff with Baobab system developers, DHO TB staff, NTP, and MoH’s HIV department. Due to the rapid nature of the award and expected results, these initial, highly-participative meetings were held over the course of several weeks, helping ensure buy-in, create ownership, and discuss perceived barriers to TB/HIV EMR implementation , pharmacy, and exit. Unlike the simpler flow for ART patients who largely follow the same visit flow, most TB suspects and patients fell under one of five, distinct, patient-provider encounter types depending on HIV status, TB suspects, TB patients , follow-up TB patients already in care with unknown HIV status or not yet on ART, follow-up patients on ART at MPC, and transfer-in TB patients  are moved to archive areas; the process of identifying the longest dormant file is automated in the TB/HIV EMR system. As patient numbers continue to increase, filing space will run out, leaving the potential for missing or misplaced files. To address this challenge, the TB/HIV EMR system reduces the need for complete paper files; however, the system does retain capacity for generating printed summary files for patient who transfer. As more ART facilities adopt TB/HIV EMR technology for ART and TB/HIV services, we expect the use of paper-based records to stop.Compliance with guidelines can present challenges especially with complicated patients. The TB/HIV EMR system is designed to enforce compliance to national TB and ART/PMTCT guidelines by requiring clinicians to complete key tasks such as ascertainment of HIV status and ART status, as needed. The user interface software was designed to reduce system complexity by presenting only one question and possible answers on one screen page, helping to reduce user error and ensure data completeness. The questions are simple and concise. Each question requires an answer to progress. The system prompts healthcare providers to consider additional prevention and treatment tasks during reviews, including ordering lab samples , consider drug contraindications, and counsel on drug adherence.Drug dispensing with multiple regimens, including those for TB and HIV can become difficult and riddled with errors in stock management. In the TB/HIV EMR system, the drug dispensing process uses the same barcode scanning technology employed for patient IDs. Unique standard barcodes for different drug regimens and quantities are pre-printed on labels and placed in the pharmacy. For each patient receiving TB and/or ART drugs, pharmacy technicians scan the corresponding barcode for the prescribed drugs and quantities Fig. . This teDetermination of next appointment date requires consideration of each patient’s drug requirements, adherence, and clinical review schedule—a process that is hard to do correctly by hand. In the TB/HIV EMR system, the system automatically calculates and presents the next appointment date in a calendar-format for provider confirmation based on the drug quantities dispensed and adherence profile, allowing the provider to change the date based on clinical or patient need Fig. . At the The primary TB/HIV EMR system users are TB clinical officers, nurses, clinic clerks and health surveillance assistants. Access rights for specific tasks are assigned according to cadre and system management level. For instance, the clinic clerk cadre has access to registration and lab information modules only and cannot access tasks assigned to clinicians. A TB clinician trained in ART has access to all tasks through TB/HIV patient clinical review and prescription while a pharmacy technician is only allowed to dispense drugs. Super-users  and system developers have access to edit patient data including amending current and previous patients’ visit records. System usage logs record usernames of all users who enter or change patient data records for quality assurance and supervision purposes.As components of the system were being developed, a series of provider-focused pre-testing sessions were organized with mixed groups of up to 10, total, clinicians, clerks, and data staff to assess clarity and order of questions, and to verify compliance to national guidelines. We also tested various patient flow scenarios to ensure skip patterns were followed, e.g. family planning questions were not asked to young children or assessment of ART eligibility in confirmed HIV-negative patients. Also, pre-test sessions ensured that system alerts designed to remind clinicians to conduct HIV tests, order follow-up lab tests, consider drug contraindications, etc., worked as expected. Early pre-testing of TB/HIV EMR question flow and content helped system users understand the questions, identify missing or misplaced content, and avoid errors in misinterpretation.After successfully pre-testing the system off-site, an intensive, two-day practical training was provided to approximately 20 healthcare providers and support staff at MPC—those with and without previous EMR experience. This training session was held over a weekend so that all staff could attend without affecting patients. Service providers first practiced specific tasks  according to their cadre  using touchscreen computers. Subsequent TB/HIV EMR practice activities mimicked diverse patient visit types requiring system users to “move” patients from registration through various clinic review services to drug dispensing. As system developers also attended the training, many issues in patient management and flow identified by users on Saturday could be immediately addressed by code fixes for further review and testing on Sunday. More complicated coding issues required additional days to correct and follow-up test with a smaller group of staff. Subsequently, through the initial on-site testing processes, system users identified various additional bottlenecks and system inconsistencies that required the developers to further refine system before a full clinic-based launch. After each system revision, additional, brief hands-on trainings were held on site with users to gain confidence and provide further feedback. When the system was completed, another weekend training was held with all clinic staff to demonstrate the final system and allow users to gain confidence in the system.As an indication of the volume of data that can now be more easily accessed for monitoring and evaluation, in only our first 4 months of full system implementation, from April and July 2013, 787 individuals were registered in the TB/HIV EMR system: 369 (45 %) TB suspects and 418 (55 %) TB patients  system intranet and terminals were tested and (2) a system dry-run from clinic registration to drug dispensing was conducted using diverse imaginary patients to identify and rectify additional issues before live deployment. Still, we encountered problems that required software developers to provide additional on-site assistance to avoid challenges in data quality while providers gained TB/HIV EMR skills and confidence.EMR systems require strict data security protocols to ensure both patient confidentiality and data privacy. The TB/HIV EMR system has several built-in security features: (1) each user  has a unique username and password for logging in; (2) each user or cadre is restricted to their specific domain (e.g. service area); (3) only select “super users” may edit or revise patient entries; (4) patient information accessed by providers is restricted to the current patient (through entering patient’s unique ID or scanning its barcode); (5) data are stored in a password-protected, database and are accessible only by highly-trained internal staff; (6) the TB/HIV EMR system keeps an audit trail by tracking users, locations and timestamps for all data changes; and (7) data is encrypted during off-site data back up using secure encryption protocols.Securing start-up and sustainable funding are critical for system initiation and maintenance. Financial requirements were categorized into two groups; start-up and ongoing costs. Start-up (one-time) costs included purchasing hardware and developing software (Additional fileAlthough implementation of large TB/HIV EMR system is complex, our three-phase approach to planning, design, and implementation allowed us to successfully launch the TB/HIV system in MPC clinic due to several key factors. First, an extensive, highly-participatory planning period with stakeholders ranging from service providers to MoH officials before and during the system development process, resulting in generating provider buy-in and ownership by the NTP who plan to adapt the system for national use. Second, alerts to order lab samples encourage high quality clinical practices while maintaining user-end simplicity, ensuing that providers were reminded to include all components of patient care even when faced with complicated patient management. Features such as prompts to order lab sample have the potential to decrease disparities in patient care because system-decision support is given to all providers. Third, working to ensure a consistent power-supply with a mix of electrical power and reliable backup system helped avoid unnecessary system failures and lost data. Fourth, a well-functioning paper-based TB registry greatly aided adoption of the TB/HIV EMR system because the clinic and all service providers already understood the importance of accurate and quality data in patient management. Lastly, adequate time should be allocated to mapping patient flows, assessing clarity and order of questions, development of software development and pre-test sessions to avoid unplanned delays and maintain user confidence.Overall, the TB/HIV EMR system demonstrates the potential to enhance integration of TB/HIV services and facilitate improved decision-making among providers. We plan to further evaluate the data collected to inform service integration, strengthen monitoring and evaluation, and, ultimately, improve patient treatment outcomes. In the meantime, the system will be scaled up to other high-burden TB/HIV centers in Malawi for additional testing and evaluation at the national level. We believe the additional benefits of the TB/HIV EMR system towards improving patient care will become apparent as the system is expanded to other clinics and clinicians over time. Despite the personnel, financial, and technical investments required for success, other clinics, organizations, and public healthcare service providers should consider TB/HIV EMR systems as a beneficial investment in patient care.


N = 1332, aged 16–69 years). Participants completed online, self-report measures assessing self-harm, momentary affect, experiential avoidance and coping in response to a recent stressor. Participants who had self-harmed reported significantly higher levels of experiential avoidance and avoidance coping, as well as lower levels of approach, reappraisal and emotional regulation coping, than those with no self-harm history. Moreover, more recent self-harm was associated with lower endorsement of approach, reappraisal and emotion regulation coping, and also higher levels of both avoidance coping and experiential avoidance. Higher experiential avoidance and avoidance coping also predicted increased lifetime frequency of self-harm. Conversely, increased approach and reappraisal coping were associated with a decreased likelihood of high frequency self-harm. Although some of the effects were small, particularly in relation to lifetime frequency of self-harm, overall our results suggest that experiential avoidance tendency may be an important psychological factor underpinning self-harm, regardless of suicidal intent , which is not accounted for in existing models of self-harm.This study investigated the relationship between experiential avoidance, coping and the recency and frequency of self-harm, in a community sample ( Self-harm, defined as “self-injury or self-poisoning irrespective of the apparent purpose of the act” , is a maSelf-harm is highly prevalent, with onset typically occurring in adolescence ,6. GivenIn the broadest sense, the term experiential avoidance (EA) encompasses all forms of avoidance and escape behaviour that are utilised as methods to alter the form, intensity and/or function of experience; EA is defined by the drive to avoid distress . The terWhile some have outlined EA as a discrete strategy of emotion regulation , in its Experiential avoidance has been outlined as a central component in the development of increased perceived stress due to its relationship with coping dynamics; emotional avoidance demonstrates a propensity to engender coping responses that diminish resources, thus reducing an individual’s capacity to deal with day-to-day activities and tasks . As highTherefore, while experiential avoidance may be an effective way of terminating unwanted emotional states in the short-term, pervasive reliance has detrimental consequences and can lead to a severely constricted life . ExperieAlthough it is argued that self-harm serves multiple functions simultaneously, the reasons most consistently endorsed in both clinical and theoretical literature relate to the avoidance and elimination of, or escape from, aversive internal experience –18. PrelThe experiential avoidance model (EAM)  providesResearch has begun to delineate factors which may be associated with an increased propensity to adopt experientially avoidant strategies. Avoidant response tendencies are especially likely in individuals experiencing heightened emotional intensity, those with deficits in arousal regulation or those who have poor distress tolerance . AdditioThese factors may be important in developing an understanding of the continuation/ exacerbation of self-harm, as well as an ability to discern these individuals from those who stop, or who have engaged in self-harm less recently or frequently. This has implications for intervention planning.Both coping responses and coping strategies refer to intentional behavioural and cognition coping efforts, employed in response to a specific stressor . Trait mFunctional accounts of coping are distinguishable from coping style accounts by a shift in emphasis from what the individual does, to what these behavioural and/or cognitive efforts do  for that individual ,25. ThatDivergent transactional accounts differ in terms of number of functional dimensions conceptualised. The most parsimonious accounts consider all coping functions subsumed under a single uni-dimensional construct, approach- avoidance . More coResearch indicates that coping and experiential avoidance are overlapping, but discrete, constructs . AvoidanThe present study focuses on experiential avoidance and the implementation of coping strategies, specifically the perceived functional dynamics of elected coping responses. While both structural and functional accounts are paramount to building a comprehensive understanding of coping, there is a lack of research addressing functional coping dynamics in relation to self-harm. Indeed—despite an increased understanding that functionality is paramount in understand the complex and dynamic nature of self-harm —to the bThe EAM  proposesGiven that the EAM  is a behExperiential avoidance tendency is associated with the expression of both negative affect  and selfIn its original conceptualisation, the EAM focuses exclusively on ‘self-injurious behavior that occurs in the absence of any intent to die’ , arguingRecent research evidence indicates that experiential avoidance may be a key psychological variable in suicidality  and, moreover, that addressing experiential avoidance therapeutically may contribute to a reduction in ideation in suicidal individuals, independently of decreased hopelessness and depressive symptomology . Moreoven = 42 ‘prefer not to say’; 11.4%, n = 152 missing), 151 for age (11.3%). Ethnicity demographics were not captured; however country of completion was assessed. Over forty percent (41.7%) of participants were based in North America, .3% South America, .8% Asia, 6.3% Australasia, .1% Africa and 50.8% Europe (data were missing for one participant).One thousand, three hundred and thirty two (1332) adult participants took part in the study. Participants varied in age between 16 and 69 Years . The majority of the sample was female (75.2%). However it is important to note the omission of demographic information for 194 participants for gender  via e-mail listings  and the School of Psychology, Research Participation Scheme. Where applicable, first year undergraduate students received partial course credit for their participation. In line with ethical considerations regarding informed consent, the exclusion criteria prohibited those aged under 16 years from participation. Recruitment was not topic blind; the study was advertised as, ‘part of an on-going project investigating coping function and self-harmful behaviours’. All advertising materials highlighted that we were ‘recruiting participants who have never self-harmed, as well as those who have.’The work received ethical approval from the University of Nottingham, School of Psychology Ethics Committee [Ref. 335R]. All participants provided written informed consent.Age, gender and continent of residency demographics were collected.Acceptance and Action Questionnaire-II (AAQII)  is a 7-iFunctional Dimensions of Coping scale (FDC)  is a thrThe measure pertains to four dimensions of coping: emotional regulation , approach functions , reappraisal  and avoidance functions .It is important to note that the FDC measure deliberately does not restrict focus exclusively to a single coping response. The multi-dimensional nature of the scale allows for easy accommodation of the simultaneous, multiple functions typical of self-harm .The measPositive And Negative Affect Scale (PANAS)  is a 20-Section one of the Inventory of Statements about Self-Injury (ISAS) ,51 was ath of that month .The Inventory of Statements about Self-Injury (ISAS) ,51 captuData were analysed using SPSS V21 for Windows. Experiential avoidance, approach, avoidance, emotion regulation and reappraisal were normally distributed. Recency and lifetime frequency demonstrated significant skew. It is also important to note the markedly unequal group sizes when considering lifetime dichotomised engagement with self-harm. Therefore, in all analyses non-parametric tests were performed. In all instances of missing data, cases were excluded pairwise.A series of Spearman Rho correlations were conducted to test whether coping functions and experiential avoidance were associated with age and affect. As significant relationships were observed, age and affect were included in subsequent analyses and multivariate analyses were adjusted for their effect.In order to compare the endorsement of experiential avoidance and coping functions between those with and without a lifetime history of self-harm a series of binary logistic regressions were conducted. Experiential avoidance and coping functions were then entered into multivariate binary logistic regressions, to explore effects when adjusting for age and affect.p is the number of known model parameters).A series of univariate negative binomial regressions were conducted to explore whether there is a relationship between experiential avoidance, coping function and the recency of self-harm. The influence of age and affect on self-harm recency were also explored. Experiential avoidance and coping functions were then entered into multivariate negative binomial regression, to explore effects when adjusting for age and affect. Finally, the effect of experiential avoidance, coping function, age and affect on lifetime frequency of self-harm was assessed by a series of univariate negative binomial regressions before these variables were entered into multivariate analysis. Negative binomial regressions were selected as, in addition to being suited to count data, this analytic approach models over-dispersion. Negative binomial regression models therefore provided a better fit to the data than scale weighted Poisson regressions  reported lifetime self-harm. Of those who had ever self-harmed, 115 (10.1%) participants had self-harmed on the day of the study, 45.3% (cumulative percentage) had within the last week, 54.2% the last fortnight, 65.6% in the last month (31 days). The number of days since last episode of self-harm ranged from 0 to 12,775 days (35 years).The majority of the sample who reported self-harm histories reported high frequencies of behaviour engagement. Nearly one third (29.8%) of participants reported more than 500 self-harm episodes during their lifetime , with a further 44.5% of participants reporting having self-harmed 101–500 times. Around twenty-five percent (25.8%) of participants reported engaging 100 times or less . Self-cutting was the most commonly reported method of self-harm see .Thirteen participants (<1%) reported extremely high frequency estimates . On examination of the ISAS responses it appeared that these participants had entered predominantly large, rounded figures on all indicated behaviours. The figures were therefore taken to indicate frequent engagement  rather than a literal count and lifetime frequency was capped at 50,000 for subsequent analyses. This did not affect the sample median (= 198) or IQR (= 475).The relationships between experiential avoidance, coping functions and demographic/ contextual factors across both conditions were explored see . Spearman = 1173) showed significantly higher levels of experiential avoidance than those with no self-harm history (n = 159)  = 403.713, p <.001). Further, differences in coping function were also apparent; those with a lifetime history of self-harm were significantly more likely to endorse avoidance  = 32.010, p <.001) coping than those who have never self-harmed. Those with a lifetime history of self-harm were also significantly less likely than those who have never self-harmed to endorse approach  = 155.462, p <.001), reappraisal  = 141.386, p <.001) and emotion regulation coping  = 12.806, p <.001) in response to a stressor.Results of a series of univariate binary logistic regressions indicate that those who have ever engaged in self-harm  = 54.094, p< .001). Therefore EA related to increased risk of higher frequency self-harm. Coping functions were also predictive of self-harm. Increased avoidance  = 17.377, p< .001) was also associated with higher frequency self-harm engagement. The incident rate for number of lifetime episode of self-harm increases by 2.8% for every unit increase in functionally avoidant coping. Conversely, increases in both approach  = 45.947, p< .001) and reappraisal coping  = 34.674, p< .001) were associated with a decreased likelihood of high frequency self-harm. For every unit increase in approach coping, the incident rate for number of lifetime episodes of self-harm decreased by 6.0%; for reappraisal coping the decrease was 4.2%. Emotion regulation coping was not predictive of lifetime frequency of self-harm  = 1.394, p = .238).A series of univariate negative binomial regressions were conducted to explore whether, in those with a history of self-harm, experiential avoidance and coping functions predicted lifetime frequency of engagement. The effects of age and affect were also explored. Experiential avoidance significantly predicted the frequency of self-harm engagement; the incident rate for lifetime frequency of self-harm increases by 4.6% for every unit increase in experiential avoidance tendency  = 4.530, p = .033) and positive affect  = 1.327, p = .249) were not predictive of lifetime frequency of self-harm.Age  = 47.041, p< .001). Therefore negative affect related to increased risk of higher frequency self-harm.Negative affect significantly predicted the frequency of self-harm engagement; the incident rate for lifetime frequency of self-harm increases by 3.6% for every unit increase in negative affect  directly confront problems faced (approach), and 2) readdress the problems faced and reinterpret the meaning of a situation   may be pIn the current study, approach and reappraisal coping functions were highly correlated. This has implications for the conceptualisation of coping. While Cox and Ferguson ,25 outliAs hypothesised, increased endorsement of experiential avoidance and avoidance coping functions were associated with increased negative affect. Reduced approach, reappraisal and contrary to expectations, emotional regulation coping functions were associated with increased negative affect. A pattern of relationships with converse directionality is observed in positive affect. In line with the hypothesis, older age was related to increased approach and reappraisal coping as well as decreased experiential avoidance tendency and functionally avoidance coping. Older participants also reported significantly higher levels of emotion regulation coping.While mixed intent, or varying degrees of ambivalence are commonly reported in self-harm ,52, it iWhen interpreting these findings it is important to note the nature of the sample. A key advantage of the study is the large, non-clinical sample. The majority of empirical support for the EAM  and the A large majority of participants had a lifetime history of self-harm (88.1%) which is markedly higher than rates typically reported for self-harm ,57 and NDespite efforts to recruit a diverse sample, the sample achieved contained a large proportion of recent, high frequency self-harm; 65.3% had self-harmed in the last month. Therefore, while considering ‘lifetime’ engagement, for many this was very recent, or in fact current, engagement. This is important when considering ‘recovery’ and the dynamics that may underpin it: while the results suggest that coping dynamics and levels of emotional avoidance differ dependant on whether an individual has ever engaged in self-harm, caution is advised in the strength given to these claims. It is less clear if, and how, the relationships between variables may differ when considering more long-term resolution of self-harm engagement. Future work should seek to recruit a more varied sample in terms of the volume and recency of behaviour engagement.The primary limitation of the work is the cross-sectional nature of the study, precluding the ability to draw conclusions regarding causality. While the study offers a preliminary consideration of coping function and experiential avoidance in relation to the recency and frequency of self-harm, longitudinal research is optimally placed to explore these dynamics more fully. Such research offers the potential to address the current lack of clarity regarding how functional coping dynamics and non-acceptance of internal negative experiences may map to behaviour change. Research exploring coping which an individual believes will 1) allow them to deal directly with a stressor, and/or 2) allow them to reinterpret or construct the problem faced, may be particularly important. Moreover, in addition to insight into temporal dynamics, such investigations would permit the exploration of interactions with proposed moderators, such as distress tolerance . ImportaThe current study considers the recency and frequency of self-harm independently. While this offers novel insight, future research may benefit from the exploration of the combined effects of recency and frequency. For example, equivalent overall lifetime frequency may be reported by an individual who previously self-harmed, yet has not done so for some years and by an individual who currently self-harms often. Arguably, it could be expected that the dynamics underlying these situations may be divergent. The duration of time over which an individual has engaged in self-harm  and any effect this may have on the associations between self-harm, experiential avoidance and coping functions, also warrants future attention.Although the study recruited a community sample this sample was not representative. A disproportionate number of participants were under 25 years of age (88.9%) and female (74.9%). It would be interesting to test the replicability of findings in an older, or more diverse sample.Finally, the study is limited by its reliance on mono-method, self-report assessment. The ability to accurately report on internal experiences may be confounded by an individual’s aptitude for accessing states, and willingness to engage with aversive experience. A reluctance, or difficulty in doing so, is likely to be marked in those with heightened tendencies for experiential avoidance. Thus, multimodal assessment may be optimal, with the inclusion of behavioural and/or experimental paradigms adding additional strength to protocols investigating experiential avoidance tendencies .Notwithstanding these limitations, the study offers novel insight into the relationship between self-harm, experiential avoidance and coping function, expanding on the existing literature to consider the recency and frequency of self-harm engagement. Additionally, and for the first time, the study explores these relationships in both non-suicidal self-injury and self-harm irrespective of  intent, comparing findings between groups.


AbstractOver the past 50 years, Southeast Asia has suffered the greatest losses of biodiversity of any tropical region in the world. Malaysia is a biodiversity hotspot in the heart of Southeast Asia with roughly the same number of mammal species, three times the number of butterfly species, but only 4% of the land area of Australia. Consequently, in Malaysia, there is an urgent need for biodiversity monitoring and also public engagement with wildlife to raise awareness of biodiversity loss. Citizen science is “on the rise” globally and can make valuable contributions to long-term biodiversity monitoring, but perhaps more importantly, involving the general public in science projects can raise public awareness and promote engagement. Butterflies are often the focus of citizen science projects due to their charisma and familiarity and are particularly valuable “ambassadors” of biodiversity conservation for public outreach.Appiasolferna, followed by Junoniaorithya and Zizinaotis. Twenty-two species were only sampled once, five were only sampled twice, and four were only sampled three times. Three DNA barcodes could not be assigned species names. The sampled butterflies revealed that widely distributed, cosmopolitan species, often those recently arrived to the peninsula or with documented "invasive" potential, dominated the habitat types sampled by the participants. Data from this first Butterfly Count helps establish a baseline from which we can monitor the patterns and changes in butterfly communities in Peninsular Malaysia.Here we present the data from our citizen science project, the first “Peninsular Malaysia Butterfly Count”. Participants were asked to go outdoors on June 6, 2015, and  sample butterfly legs for species identification through DNA barcoding. Fifty-seven citizens responded to our adverts and registered to take part in the butterfly count with many registering on behalf of groups. Collectively the participants sampled 220 butterfly legs from 26 mostly urban and suburban sampling localities. These included our university campus, a highschool, several public parks and private residences. On the basis of 192 usable DNA barcodes, 43 species were sampled by the participants. The most sampled species was  Over the past 50 years, Southeast Asia has suffered the greatest losses of biodiversity of any tropical region in the world . Malaysihttp://www.bigbutterflycount.org; Table Citizen science is “on the rise” globally and can make valuable contributions to long-term biodiversity monitoring , althougAgainst the backdrop of these challenges, here we present data and insights from our citizen science project, the first “Peninsular Malaysia Butterfly Count”.http://scistarter.com/blog/2012/07/summer-is-busy-season-for-butterflies-and-citizen-scientists; http://www.pierisproject.org) due to their charisma and familiarity and are particularly valuable “ambassadors” of biodiversity conservation for public outreach (http://www.thestar.com.my/News/Education/2014/10/26/The-butterfly-effect/). Butterflies are thought to react rapidly to environmental changes due to their short generation time and high mobility . By coincidence, in Malaysia, the first Saturday of June is the Yang di-Pertuan Agong's (King's) official birthday, a public holiday. On such occasions families often like to go out for picnics in local parks, so it seemed ideal to have our first Butterfly Count on BEAD - June 6, 2015. We restricted our project to Peninsular Malaysia  to avoid complications due to the shipping overseas and because the peninsula and East Malaysia (part of the island of Borneo) are governed by a different set of wildlife laws.This project builds on our experience with another ongoing citizen science project, "The School Butterfly Project", which is reported elsewhere . When wehttp://www.facebook.com/butterflycount). We created an advertisement for the project in facebook and purchased “boost post” across Malaysia to encourage maximum participation across the peninsula. According to facebook statistics the advert "reached" 27,392 people at a cost of RM73.00. We also contacted national newspapers to request coverage to encourage registration. This request was taken up by two national English-language newspapers  and one national Chinese-language newspaper  linked to the facebook page, or by phone. Registration was closed on May 31, 2015.Registered citizens were sent (via Pos Laju) a Butterfly Count Pack containing:i) Butterfly Count Guide  Ten 1.5ml microcentifuge tubes.iii) Pair of tweezers.iv) Butterfly net.v) Prepaid addressed envelope (Pos Laju).vi) Souvenir button badges.https://www.facebook.com/butterflycount/photos_stream).Following guidelines in the Butterfly Count Guide . The NParks program involved a butterfly identification training workshop and assigned participants to a specified count location, requiring a significant commitment (time) and investment (travel costs) by the participants. In contrast, for the Peninsular Malaysia Butterfly Count, in order to reduce costs and encourage participation, we allowed the participants to choose their own count location, and did not provide identification training . The participants were asked to collect non-lethal tissue samples (butterfly legs) to enable accurate species identification through DNA barcoding , also conducted an inaugural butterfly count in neighbouring Singapore .Of the 220 legs received, 192 (87%) generated 'usable' DNA barcodes of varying quality and length. The DNA barcodes and associated collection data are available on BOLD in the publicly accessible datatset - Peninsular Malaysia Butterfly Count [PMBC] (Appiasolferna (BOLD:AAZ4640), followed by Junoniaorithya (BOLD:ABZ6191)  were assigned to the genus Ypthima  using the strict tree-based criterion for DNA barcode-based higher-taxon assignment  Fig. . Twenty-signment .Of the 192 usable DNA barcodes, the participants had attempted a family-level identification for 108 56%) of these butterflies. Based on the DNA barcode assigments, 60% of these family-level identifications were correct . We need to review how to attract participation from the east coast of Peninsular Malaysia and rural areas, as it was clear we failed to reach out to those communities. Based on the registered names of participants it was also clear we were more successful in attracting participants from Malaysia's ethnic Chinese community than the other ethnic groups represented in Peninsular Malaysia. We were unable to obtain any coverage in Malay language newspapers, which may partially account for this trend, but it also likely reflects the rural-urban divide.All of those who registered to take part in the Butterfly Count live on the west coast of Peninsular Malaysia. Furthermore, most of the participants live in the Klang Valley, the large urban agglomeration surrounding Kuala Lumpur  generated 'usable' DNA barcodes  of varying quality and length. There are several explanations for the relatively low success rate of PCR and sequencing. As noted above, it took some butterfly legs quite a while to reach us after the count day. Depending on the storage conditions, this could have boosted DNA degradation, which is particularly a problem in hot and humid Malaysia . SecondlA comparison of the DNA barcode identifications and the family-level identifications provided by the participants revealed that the participants were able to correctly identify the family of butterfly specimens 60% of the time. The relatively low success rate suggests that our butterfly family identification guide could undergo some improvement. Preparation of an identification guide represents a trade-off between being "user-friendly" and technical, and it is important not to discourage participants from attempting identifications by providing overly-complex guidelines. The relatively low success of family identifications suggests that asking participants for species identifications would not be very useful, either as a learning experience, or for contributing data on species occurrences. This finding further validates the continued use of the non-lethal DNA barcoding model used for this project.Although the primary purpose of the Peninsular Malaysia Butterfly Count was to promote awareness and engage the public with biodiversity, the Butterfly Count did produce some ecologically interesting findings.Appiasolferna, commonly known as the Striped Albatross. Although most often treated as a distinct species (http://www.nic.funet.fi/pub/sci/bio/life/insecta/lepidoptera/ditrysia/papilionoidea/pieridae/pierinae/appias/index.html), Appiasolferna is sometimes considered a subspecies of Appiaslibythea and DNA barcodes currently named as A.libythea and A.olferna share the same BIN in BOLD (BOLD:AAZ4640). A.olferna shows extreme sexual dimorphism with the male being predominantly white, but the female having broad black stripes on the upperside of the wings. This could potentially explain some of the family-level misidentifications by the participants, as our guide relied heavily on wing colour. According to Corbet and Pendlebury . C.rutidosperma is a common weed found growing in disturbed habitats such as roadsides, gardens, and abandoned land. C.rutidosperma is native to Africa and is an invasive species in Peninsular Malaysia as well as other parts of Asia, Australia and the Domican Republic (http://www.cabi.org/isc/datasheet/14044#20087204071).The most sampled butterfly species was ndlebury , A.olfeoadsides . In SingJunoniaorithya, the Blue Pansy (BOLD:ABZ6191), which has a distribution covering Central Asia, India, Southeast Asia, southern China, Taiwan, Australia, and Africa , at positions four and five amongst the most collected species are, together with Euremahecabe (BOLD:AAA6082) at position nine, known to be paticularly common in gardens and along roadsides , fell within the Ypthima group of DNA barcodes on BOLD but without species-level matches. This suggests new species records for Peninsular Malaysia. At position six-equal amongst the most sampled species was Acraeaviolae, the Tawny Coster . The species, native to India, was first recorded in Peninsular Malaysia in 1992. The recent range expansion, reaching Australia in 2012, has been reviewed by Chiladespandava , the tenth-equal collected species, is of concern in Taiwan and China as an "invasive" species with a history of populations "outbreaks"  The level of participation in the first Peninsular Malaysia Butterfly Count was encouraging, but reaching and engaging rural communities remains a challenge.2) The non-lethal DNA barcoding approach for species identification worked effectively, however, protocols could be improved to limit the number of returned samples which could not be identified. The family-level identification guide could use some improvement but provides an important educational tool for the participants.3) The sampled butterflies revealed that widely distributed, cosmopolitan species, often recently arrived to the peninsula or with documented "invasive" potential, dominate the habitats sampled by the participants. Data from the first Butterfly Count helps establish a baseline from which we can monitor changes in butterfly communities in Peninsular Malaysia.Supplementary material 1Butterfly Count GuideData type: Butterfly Count GuideFile: oo_48731.pdfShi-Wei Jisming-See, Guo-Jie Brandon-Mong, Kong-Wah Sing, John-James WilsonSupplementary material 2Bulletin of the Museum of Zoology (3)(4)Data type: NewsletterFile: oo_60225.pdfJohn-James Wilson, Shi-Wei Jisming-See, Guo-Jie Brandon-Mong, Aik-Hean Lim, Voon-Ching Lim, Ping-Shin Lee, Kong-Wah SingSupplementary material 3Butterfly Count Raw DataData type: Data spreadsheetFile: oo_63256.xlsxJohn-James Wilson, Shi-Wei Jisming-See, Guo-Jie Brandon-Mong, Aik-Hean Lim, Voon-Ching Lim, Ping-Shin Lee, Kong-Wah Sing


Panthera pardus), puma (Puma concolor) snow leopard (Panthera uncia), African lions (Panthera leo), cheetahs (Acinonyx jubatus), and tigers (Panthera tigris)) for the purpose of conservation. Sensory analyses of scent-markings could address knowledge gaps in ethochemistry. The objective of this review is to summarize the current state-of-the art of both the chemical and sensory analyses of scent-markings in wild mammals. Specific focus is placed on sampling and sample preparation, chemical analysis, sensory analysis, and simultaneous chemical and sensory analyses. Constituents of exocrine and endocrine secretions have been most commonly studied with chromatography-based analytical separations. Odor analysis of scent-markings provides an insight into the animal's sensory perception. A limited number of articles have been published in the area of sensory characterization of scent marks. Simultaneous chemical and sensory analyses with chromatography-olfactometry hyphenation could potentially aid conservation efforts by linking perceived odor, compounds responsible for odor, and resulting behavior.In conjoining the disciplines of “ethology” and “chemistry” the field of “Ethochemistry” has been instituted. Ethochemistry is an effective tool in conservation efforts of endangered species and the understanding of behavioral patterns across all species. Chemical constituents of scent-markings have an important, yet poorly understood function in territoriality, reproduction, dominance, and impact on evolutionary biology, especially in large mammals. Particular attention has recently been focused on scent-marking analysis of great cats (Kalahari leopards ( This developing field is limited in the scope of information available about chemosensory analysis of wild animal markings. The use of scent- markings as a method for aiding conservation has been reviewed , but lacThe objectives of this large mammal and great cat scent-marking review are to: (1) classify different sample preparation techniques for their analysis of scent-markings; (2) summarize existing information on the use of advanced analytical methods on these scent-markings; (3) identify different sensory techniques used to characterize odors of these scent-markings; and (4) classify different sample preparation techniques for the analysis of these scent-markings.i.e., leopards, snow leopard, lions, cheetahs, and tigers), scent-markings. Development in the area of sampling and analysis of semiochemicals aids in understanding animal behavior that can be used, for example, toward efforts such as conservation of great cats.This review provides an overall perspective of literature on the subject of chemical and sensory analysis of large wild mammals, particularly great cats  to the receptive animal (receiver) .Scent-markings require accuracy of olfactory detection to send and receive the correct signal. Scent-markings contain a complex mixture of chemical compounds at varying concentrations based on its chemical message . If an a1.3.Chemicals that act between organisms are called semiochemicals ,8. In a In group living species, for example, it is essential that an individual can recognize members of its social group as individuals and distinguish them from non-group members. . LimitedSemiochemicals can be classified as kairomones or pheromones ,12. WhenAntilocapra americana), cheek rubbing and interdigital scrapping in domestic cats, interdigital scrapping in white-tailed deer (Odocoileus virginianus), and head rubbing in goats [Pheromones are extensively used in territory marking by mammals. Although pheromones are often thought of as odorants (volatile organic compounds), they can be odorless (nonvolatile organic compounds) . Often tin goats –18.Pheromones are classified into two categories: (1) primers, which prolong a shift in the physiology of the recipient and (2) releasers, which trigger a rapid behavioral response . Primer Alarm substances communicate that there is a possibility of danger. Recruitment pheromones are commonly found in social insects. They are generally employed by worker castes of social insects to guide their nest mates to a food source . ReproduIn many vertebrates mother-young recognition is contingent on chemical cues . Territo1.4.Scent-marking is described as the most ubiquitous form of chemical signaling in mammals . ChemicaScent-marks are placed on objects in the environment, frequently in the absence of the receiver, and may only be detected later, in the absence of the signaler . SendersTo counteract degradation, male mammals generally will remark active scent-markings. Compounds in scent-markings that have longevity under environmental conditions tend to have high molecular weights and low vapor pressures. Some examples of compounds that are found ubiquitously in scent-markings are: squalene, cholesterol, and long-chained carboxylic acids. These compounds are primarily in the secretions/excretions of mammals .The most common form of marking is for resource defense territories. Scent-marking by resource holders presents an opportunity for competitor assessment . Scent-mDetection of scent-marks is dependent upon the sensory neurons for olfaction within the vomeronasal organ (VNO) and the main olfactory epithelium (MOE) ,21. UnivAccording to the stereochemical theory of olfaction, mammals bind odorants to specific OR sites based on the size and shape of the molecule , which rThe persistence time of the mark is the interval between deposition and the time when the mark can no longer be sensed ,30. The 1.5.Sample preparation serves an important role in the efficient extraction of components of interest from the sample matrix. The results of this extraction process are later used with analytical instrumentation for target analyte: separation and isolation into constituents, identification, and quantitation . Some biRecent advancements in sample preparation and analysis of biological samples can aid in addressing needs and knowledge gaps when applied to scent-markings. Reduced sampling, sample preparation time, and faster, more sensitive and precise analytical procedures have the potential to help scientists working in the field of scent-marking analysis .1.5.1.There are two main approaches to sample preparation techniques; solventless and solvent-based.Sample preparation methods are categorized by the compound's class, polarity, molecular weight (MW), volatility in which it can be extracted, the physical state , and the analytical instrument used for chemical characterization ,37,38. Sin vivo extractions from biota [Modern day sample preparation has advanced dramatically in the area of solvent-free extraction processes ,44–49. Som biota –53. Refeom biota  reviewed1.5.2.Analytical methods are designed to separate, isolate, identify, and quantify analytes of interest within a sample. There are various techniques and reviews on the separation of these components, specifically in mammals ,54. WithIn GC, the most widely used analytical tool, a mixture of volatile organic compounds (VOCs) is separated into individual VOCs and semi-VOCs, which are eluted out of the GC column at different times . This al1.6.Odor detection is a critical constituent in animal interpretation of scent-markings. Inferences into the actual chemicals and odors sensed by animals have been sought through the use of chemical and sensory analytical instrumentation and the use of animals. Rodents have been commonly used to measure the efficacy of the longevity of scent-marks –66. ConsHomo sapiens) olfactory system's measurement of volatiles [In recent years, the introduction of application-specific sensor array systems, otherwise known as “electronic noses”, were developed and combined with GC, MS, and infrared spectroscopy to mimic the sensitivity of the human  within the marking and to detect their presence in the wild. Often this identification is performed via conspecific confirmation. Mice have been the primary models of olfactory detection and interpretation of markings, such as in deciphering the age and reproductive messages in urine ,64,72,73Dogs have also been used in the estimation of wild animal populations based on individual scent-mark recognition ,74. The Damilscus dorcas dorcas) interdigital gland secretions [Sus scrofa) [The human nose has been an olfactory detection system in various studies of animal pheromones. When m-cresol, 2-heptylpyridine, hexanal, (Z)-6-dodecen-4-olide, and α-terpineol were present in high concentrations, they were identified by human nasal detection as the compounds responsible for the pleasant herbal smell of bontebok (cretions . The sen scrofa) ,76. Huma scrofa) .Simple human nasal detection was performed for the determination of the characteristic odor of tiger marking fluid ,33. They1.6.2.The implementation of simultaneous chemical and sensory analyses is the modern approach to investigating the odors, tastes, and visual appearance of chemical compounds in biological samples. Based on their detection mechanisms, these systems can be classified into several categories, including chemical sensors, biosensors, GC-based systems, MS-based detectors, and hybrid GC/chemical sensors. Specifically, ‘electronic noses’ (‘e-noses’), multidimensional gas chromatography-mass spectrometry-olfactometry (md-gc-ms-o), ‘electronic tongues’, and visual analyzers are a few types of biosensory technologies available for the characterization of biological compounds. The reaction between odor molecules and the target sensing materials on the sensor surface triggers changes in mass, volume, or other physical properties. This reaction is then converted to an electronic signal by a transducer.Widely used types of transducers include optical, electrochemical, heat-sensitive, and mass-sensitive. Some common chemical sensors are: surface acoustic wave sensor, quartz crystal microbalance sensor, metal oxide semiconductor sensor, and polymer composite-based sensor. An ‘e-nose’ is an instrument that is designed to mimic the function of the natural nose. By definition, it uses a sensor array to not only detect but also discriminate among complex odors ,78,79.−7 to 10−11 M in humans) [The ideal example for the detection of odors is the mammalian nose because of its ability to evaluate with both high sensitivity and specificity. Olfactory receptors make these properties possible, as they support combinatorial detection of odors at trace levels  ,81. Exha humans) –85.The detection mechanism of these biosensors is based on the specific interaction between olfactory receptors and odorant molecules. Biosensors have been known to demonstrate better detection selectivity than chemical sensors. The ‘bio-sniffer’ is another example of a type of biosensor developed for VOC detection that is based on biochemical reactions between a biomolecule and a VOC, or a chemical reaction catalyzed by biomolecules ,87.md-gc-ms-o is capable of removing the interference effect from non-target components. This system allows the users to separate components of interest, identify character defining compounds, and identify those components using modern mass spectral techniques ,88–94. MA few examples of research that have been performed using MD-GC-MS-O and simultaneous chemical and odor identification are: identification of compounds responsible for the characteristic odor of livestock and poultry manure and rumen of beef cattle; association of a specific odor with a volatile compound; the role of particulate matter as a carrier of odor; characterization of kairomones and characteristic odorants released by insects; and quantification of nutraceuticals in wine ,89–98.This analytical tool is a state-of-the-art technology that is particularly suited for identification of chemical-odor association. This instrument can be used to explain the association between VOCs and their odors in wild mammal secretions and excretions. MD-GC-MS-O is capable of determining the concentrations of these compounds and evaluating the intensity and aroma of the odors of the entire scent-mark. Identification of compounds responsible for specific odors and signaling could aid wild mammal conservation, and it would serve in giving some insight into how and why animals are detecting these scents.2.Articles were obtained through searches on Science Direct, Academic Search Premier (EBSCO), and Google Scholar article databases. Keywords and phrases that were used in the searches included: “conservation”, “GC-MS”, “GC-MS-O”, “gas chromatography”, “chromatography”, “endangered species”, “odor”, “chemosensory”, “simultaneous chemical and odor analysis”, “panthera”, “elephas”, “odocoileus”, “TAARs”, “olfactory receptors”, “scent-marks”, “urine”, “feces”, “mammals”, “scent-marking”, “conservation”, “animals”, “volatile organic compounds”, “sample preparation”, “analytical techniques”, “large mammals”, “pheromones”, and “marking fluid.” Articles selected for this review focused on the use of modern analytical techniques to identify and/or quantify chemical compounds detected in scent-markings of large wild mammals and great cats for the purpose of sensory and chemical identification, conservation, behavioral understanding, and evaluation of sampling and sample preparation effectiveness.Citations from the initial search were downloaded into EndNote, a reference management database. Duplicate citations were removed. Assessment of the identified studies for relevance was based on a standardized criterion developed by all co-authors: (1) the focal animal reported was a large wild mammal; (2) analytical techniques were utilized for chemical identification of scent marks; (3) sample preparation was defined; (4) the articles were peer-reviewed; (5) if sensory analysis was performed the method needed to be clearly defined; and (6) the co-authors had no objections, such as quality or topic focus of the articles.If any of the five criteria were not met, the reference was omitted. For articles that remained in the review after applicability and quality selection, data were summarized and reported. Data extraction from these articles was completed by one reviewer and when uncertain this reviewer consulted with the other authors. Data extracted from the research articles included: (1) sample preparation technique; (2) analytical methods; (3) animal species; (4) sensory analysis approach; (5) relationship to conservation; and (6) scent-markings being collected. Conclusions were based on a summary of the data.3.3.1.3.1.1.This section summarizes sampling and sample preparation methods performed for the analysis of scent-markings of large mammals. It discusses solvent-free and solvent-based extraction methods and the advantages and disadvantages of these methods. The sampling and sample preparation section also explains the similarities and differences between the uses of various techniques for the identification of chemical constituents in scent-markings.2 being released simultaneously and driving the volatile material into a needle at the end of the cylinder [Chrysocyn brachurus), when SPME was ineffective [Solvent-free extraction methods often reduce sample preparation time and eliminate multiple step procedures for the extraction of a component from a sample. Conventional solvent-free extraction methods implemented for wild mammal scent-marking characterization included: headspace extraction, direct injection, precolumn heaters, solid phase extraction (SPE), stir bar absorptive extraction (SBSE), and solid phase microextraction (SPME). Headspace extraction is the process of transferring a substance from a solid or liquid matrix to the vapor phase by heating, and removing analytes from the headspace in a carrier gas . Direct cylinder ,102. SPEcylinder . SPDE haffective . SPME isffective . In someffective ,92.Elephas maximus) blood volatiles in 35 min in comparison to other lengthier procedures.Headspace extraction results in the emissions of volatile compounds to the headspace, and thus provides some information about the fate of semiochemicals based on their physicochemical properties. This is particularly important when providing evidence of an animal's ability to identify compounds in the air from extreme distances. These volatile compounds are essential to our comprehension of animal communication. Headspace autosampling extraction of gases emitted from urine can provide information on compounds potentially detected by passing animals, specifically lions . HeadspaPhascolarctos cinereus) were analyzed using a solvent-free technique [VOCs in sternal secretions from koalas (echnique . The steechnique .Rangifer tarandus) [Odocoileus hemionus columbianus) and reindeer. This scent is recognized through tugging and licking the tarsal gland and is used to identify individuals by the scent associated with them [The PH technique allowed for the identification of compounds in the interdigital glands of reindeer (arandus) ,102 and ith them . The cheStrepsirrhini urine for characterization. This urine characterization led to the phylogenetic construction of the Strepsirrhini suborder [Solid phase dynamic extraction (SPDE) is an extraction process that can be utilized at ambient room temperature to extract semi-VOCs. When coupled with an automated sampling system that can regulate temperature, a higher number of volatile compounds can be extracted. Using a SPDE needle internally coated with a modified activated charcoal-polydimethylsiloxane (AC-PDMS) allowed for a small sample size of 0.5 mL of suborder . Utilizisuborder .Stir bar absorptive extraction (SBSE) techniques have been advantageous in measuring small sample sizes and diluted media . VolatilThe polydimethylsiloxane (PDMS) coating on the stir bar and constant stirring agitation allows for a more precise and reliable extraction, and decent analytical precision . In SBSEin vivo extractions of volatiles. SPME is a solventless extraction technology that incorporates fibers of assorted coatings and a fiber holder (Solid phase microextraction (SPME) is particularly suited for characterization of volatiles from biota. SPME can be used for r holder  and 3 thr holder  can be uThus, the sampling results in high preconcentration and enrichment of compounds that did not require use of solvents and additional steps. Specific SPME coatings can be used for optimization of extraction processes favoring certain groups of compounds varying by MW, polarity, and functional groups. Often fibers with Carboxen polydimethylsiloxane (Car-PDMS) coating are used for the detection of VOCs with low MW. Divinylbenzene/Carboxen/PDMS coating is used on a broad range of analytes, specifically volatile and/or semi-volatile compounds. SPME combines sampling and sample preparation to minimize the sample preparation step with a process that is simple, reusable and efficient.There are relatively few publications that report the use of SPME for characterization of scent- markings of large wild mammals ,90, HoweLoxodonta africana) urine analyzed with SPME used a chiral column to detect the pheromone, frontalin [Automating headspace extraction with SPME was useful and a non-invasive method for monitoring reproductive status via the urine in elephants and other species . Africanrontalin . When SProntalin .Ailuropoda melanoleuca), ultrasound was used for 15 min to separate anogenital gland secretions from tampons [The use of ultrasound as a tool for compound separation has proven to be less effective than SPME. In the case of giant panda  urine used dichloromethane as the solvent and liquid N2 to condense the extracted sample. This resulted in a total sample preparation time that was less than 1 h [Methanol extraction of koala sternal gland secretions required upwards of 8 hours . The extthan 1 h . Solventthan 1 h .Summary of sampling and sample preparation techniques with references used for the chemical and sensory characterization of scent-markings in wild mammals is presented in . To dateIt appears that in the last decade there has been a rise in the implementation of SPME for the sample preparation and sampling of scent-marks . This in3.2.Research in chemical signaling plays an important role in the conservation of many endangered large animals. This section summarizes analytical methods performed for the analysis of scent- markings of large mammals. The use of various GC- and high performance liquid chromatography (HPLC)-based techniques with an assortment of detectors is summarized with the advantages and disadvantages of each method.3.2.1.Canis lupus) urine and feces volatiles [Gas chromatography (GC) is a very useful analytical technique for the analysis of mammal scent- markings . The useolatiles . Anotherolatiles .GC combined with a detector allows for the identification of compounds within the sample. The most commonly used detectors were: MS, FID, and FT-IR. MS was the most widely used because of its capability to perform a spectral search and match for over 200,000 compounds within its spectral library. Also, MS detection was preferred with GC analysis because of its compound identification abilities and sensitivity ,122. TheAiluropoda melanoleuca) anogenital gland secretions, urine, feces, and blood serum [While GC-MS is a well-established and often preferred technology for detecting volatile compounds with MW below 300, it is not ideal for the detection of higher MW compounds ,118. Theod serum ; all of od serum .gray wolves was deemed efficient [(Lycaon pictus).In the case of urine from gray wolves, notable peaks from the GC were identified through matching GC retention times and MS spectral patterns . The usefficient . SPME-GCOut of all of the 11 species-specific compounds, 8 were confirmed. The confirmed compounds were: 1,3-propandiol, N,N-dimethylacetamide, 1-methyl-2,4-imidazolidinedione, 1-methylimidazole-5-carbox-aldehyde, and quinazoline. The aforementioned compounds were at three times the level in urine than feces . This an(Saguinus fusciollis), compounds with concentration levels of 0.01% were omitted from analysis, possibly excluding the incorporation of specific pheromone or semiochemicals that are essential in animal communication but present in very low abundance [Although GC is the modern system for separations and chemical composition determination, the use of variable detectors, in conjunction with the GC, may impact the ability to quantify or qualitatively define scent-markings. While GC-MS analysis allowed for quantification of the compounds in the scent-markings of brown-mantled tamarin bundance . The usebundance  resultedbundance .Lynx rufus), MS and retention time identification allowed for first time confirmation of compounds in urine [In addition, nondistillable compounds in the tarsal gland were also not identified through GC-MS detection . In the in urine . Neverthi.e., dehydration of the oximes occurred during the desorption of the swab in the GC injection port. In the identification of castoreum composition in the American beaver (Castor canadensis), GC analysis may have impacted the analysis of highly volatile phenol constituents [GC-based analyses had some additional drawbacks such as sample dehydration/alteration. Dehydration was observed when characterizing koala sternal gland secretions , i.e., dtituents . Previoutituents . GC-FID tituents . The inttituents ,125,126.Elephants have been a major focal animal in the area of scent-marking and its role in reproduction and socialization. They have been used to understand how scent-marking impacts mating and interaction of males and females of various ages and social levels within herds ,146,147.gray wolf' urine and feces volatiles [For several chemical component identifications, a combination of capillary GC with Fourier-transformed infrared spectroscopy FTIR was essential for accurate identification of olatiles . MALDI holatiles .3.3.A three step process is needed to fully comprehend the role of cues in scent-markings in animal behavior. First, an understanding of which chemical constituents constitute the marking must be determined. Next, an odor characterization of these specific compounds must be performed. Lastly, a behavioral analysis of how the animal reacts to these specific odorous compounds to determine the relationship between behavior and scent must be completed. Without the input of sensory analysis, the interpretation of cues in scent-markings can be limited. The use of the human nose for sensory analyses, as opposed to the use of animal olfactory sensing further complicates this process. This section summarizes the limited information available on the use of chemical and sensory analysis for the characterization of large mammal scent-markings .3.3.1.(Crocuta crocuta) as being responsible for eliciting behavioral responses without detecting specific odorous compounds [GC-MS were able to generalize all compounds in spotted hyena ompounds . This stAn ‘electronic-nose’ (E-nose) indicated that VOCs emitted from the body vary with age, diet, sex, physiological status and genetics . The mai3.3.2.Animals are frequently the objects of sensory evaluation . Gray woOdor detection thresholds for humans are different for each chemical . The same principle is thought to apply in wild mammals. In complex mixtures of scent-markings reside distinct odorous compounds responsible for the longevity of its scent availability. An example of a compound that constitutes a large mammal scent-marking is cyclohexanone. Cyclohexanone elicits flehmen responses from sub-dominant females, but in males there is no response . ElephanOreotragus oreotragus) have scent marks that remain active for as long as 7 days [Helogale parvula) after 10 days and in hamsters (Mesocricetus auratus), for 100 days. Even humans, however, can detect scent from anal gland marks of hyenids after 1 to 6 months [Panthera uncia) territories and marking behaviors by differentiating the age of different urine and scat markings over a period of months. Frequency of marking coincided with the winter/early spring mating season. This marking rate potentially serves to maintain awareness of conspecific presence and also distance between snow leopards [Persistence of scent-markings in the environment has been recorded at a wide variety of lengths. In the case of dominant male mice, urine has been avoided by other males for up to 72 h. Klipspringer antelope (s 7 days . Scent m6 months . Humans leopards .3.4.3.4.1.GC×GC) (in this instance the whole sample is eluted from the first column to the second column in some specific time frame); and (c) by using practices, where only a small part of the sample elutes to the second column either via backflash, foreflash, and heart-cut [Multi-dimensional-gas chromatography (MDGC) has previously been defined as, “the process of selecting a (limited) region or zone of eluted compounds from the end of one GC column, subjecting the zone to a further GC displacement” . Two-dimeart-cut . Backflaeart-cut . Foreflaeart-cut . Heart-ceart-cut .A series of detectors can be used for two-dimensional GC: flame ionization detector (FID), electron capture detector (ECD), atomic emission detector (AED), nitrogen-phosphorus detector (NPD), and olfactory detector and mass spectrometer (MS) ,158. MDGtrans-resveratrol in red wine [The characteristic or overall aroma of a sample is an intricate combination of various odorants. Simultaneous analyses can potentially identify links between certain scents and the exact chemical compounds causing them. Simultaneous chemical and sensory analyses have the potential of linking both chemical and sensory analyses that are often analyzed independently. MD-GC-MS-O can be described as a two-way split detection system. In this arrangement, compounds are quantitatively trapped in a capillary column loop, which isolates them online from preceding and following peaks, and splits the target region into the second column for effective resolution from interfering matrix compounds ; this alred wine –91,96.Simultaneous chemical and sensory analysis is very rarely performed in the area of wild large mammal scent-markings. The only instances of sensory analysis were the use of conspecifics after chemical identification ,131,161.It has been reported that olfactory receptors in biosensors are more sensitive detectors of ligands than GC-MS and chemical “noses” . An E-noElectro-olfactograms (EOG) are “electrical potentials of the olfactory epithelium that occur in response to olfactory stimulation” . ‘EOGs a3.5.Great cat markings have been studied to aid in conservation, specifically focusing on territoriality, dominance, and reproduction  31,33,4,163–165.3.5.1.Behavioral studies of free-ranging tigers have determined that marking functions to establish and maintain territorial boundaries and advertise female reproductive status  Table 3Table 3. The focus of previous studies has been on identifying total compound composition, neglecting the study of olfaction's relationship to scent-mark identification by animals. Application of MD-GC-MS-O has the potential to measure the influence of odor in scent-marking detection in species that use chemical cues as their communication method., African cheetahs, Indian leopards (Panthera pardus fusca), and African lions  in the wild . Bengal Panthera tigris altaica) and the Amur leopard (Panthera pardus) are needed to restore their populations. Previous felid research has led to their species and sex identification from fecal and hair samples [Genetic characterization and definition of Siberian tigers ( samples . Referen samples  used sce samples . This in samples . Scent-mPanthera leo persica); Sumatran tigers (Panthera tigirs sumatrae); Persian leopards (Panthera pardus saxicolor); jaguar (Panthera onca); and clouded leopard (Neofelis nebulosa) because of its similar mass to urinary serum albumin [The volatile constituents of lion urine have been reported . The use albumin .To date, there is no published research on domestic or wild cats linking a chemical with specific odors associated with their scent marks. Thus, there is clearly a need to define characteristic odors by identifying key chemical constituents responsible for odor in a more reliable approach using analytical tools. Several studies have established the importance of odor in scent mark detection and signalling in domestic cats ,206–208.4.Chemical and sensory analyses of semiochemicals can potentially aid wildlife conservation. These volatile compounds are essential to the comprehension of animal communication. Large mammal scent-markings are of particular interest because they have not been studied in as much depth as insects and small mammals . Great cats, specifically, are facing complete eradication and could benefit from alternative and improved conservation approaches. Scent-marking sample and analytical techniques have their pitfalls and advantages, but have evolved in efficiency over the last decade. The most frequently implemented analytical techniques for characterizing scent marks of wildlife are: GC , GC-MS ,56–59, GUnderstanding of scent-marking constituency aids in the identification of key chemical markers responsible for behavior associated with mating, territoriality, and resource management. Without the input of sensory analysis, the last two steps in the understanding of ethochemistry cannot be executed. The use of animals, human olfaction, and simple GC analysis in the determination of odor composition is limiting at best. The implementation of MD-GC-MS-O, E-noses, and EOGs can help to bridge the knowledge gap about total odor composition of scent marks. This new found information can lead to wildlife management improvement and protection of large mammals and other groups of endangered species.


Alphacoronavirus, the Porcine epidemic diarrhea virus (PEDV), was first described in Europe in the 1970s and since then has spread over many Asian and American countries, where it recently led to devastating effects on swine health and pork industry. While the disease was seldom reported in Europe within the last few decades, a few recent reports re-emergence of PED in German pig farms. The hitherto isolated German strain seems to be closely related to a low pathogenic PEDV variant from the USA. This case report describes the first detection of PEDV in Austria.Porcine epidemic diarrhea (PED) is a syndrome that is characterized by rapidly spreading watery diarrhea affecting pigs of all ages, but with major effects on suckling piglets. The disease, as well as the causative Reduced feed uptake and occasional diarrhea were observed in December 2014 in a group of fattening pigs, kept on an Austrian swine farm. The concerned pigs had been recently purchased from Germany. Within a few weeks, diarrhea became apparent also in pigs of Austrian origin, which were kept in a different stable on the same farm. Gastrointestinal symptoms among fattening pigs were generally mild, quickly resolving and did not lead to death. PEDV RNA was identified by RT-qPCR in pooled feces and serum and PEDV antibodies were detectable in serum in both groups of pigs. Phylogenetic analysis of the nearly complete PEDV spike gene shows that the Austrian PEDV strain is highly similar to other strains involved in recent outbreaks in Western and Central Europe.This is the first report demonstrating the presence of PEDV in Austria. The virus was probably introduced by purchasing piglets from a German source, which underlines the significance of trans-boundary animal trade for the distribution of highly contagious diseases, such as PED.The online version of this article (doi:10.1186/s12917-015-0624-1) contains supplementary material, which is available to authorized users. Coronaviridae family, genus Alphacoronavirus, was first identified as the cause of porcine epidemic diarrhea (PED) in European pigs in 1978 [Porcine epidemic diarrhea virus (PEDV), a member of the  in 1978 , it beca in 1978 . Variant in 1978 . In 2013 in 1978 . Severe  in 1978 . The dis in 1978 , 11. Mea in 1978 –14. The  in 1978 , 16. Cli in 1978 . PEDV wa in 1978 , 19. Ful in 1978 , 17–19.Here, we report the first detection of PEDV in pigs on an Austrian fattening farm and provide information about disease symptoms, diagnostic test results, possible route of PEDV entry and genetic relatedness of the newly detected Austrian PEDV to published sequences.Actinobacillus pleuropneumoniae (APP) infection by pathomorphological examination and bacterial isolation, performed in a private diagnostic laboratory. Thus, antibiotic treatment of the German-origin group was initiated: pigs were treated via feed with 16 mg/kg/day Tilmicosin  for 8 days. In addition to the pigs of German origin, about 1000 fatteners of Austrian origin  were housed in a different stable on the same farm. The pigs of Austrian origin were clinically inconspicuous at that time. Change of clothing between different stables was regularly practiced. Although gastrointestinal symptoms in the German-origin group were mild, the veterinary practitioner submitted a pooled feces sample for PCR testing for porcine coronaviruses, Brachyspira spp. and Lawsonia intracellularis as well as for bacterial culture and parasitological examination. Commercial reverse transcriptase quantitative PCR (RT-qPCR) kits were used for porcine coronavirus testing . RT-qPCR testing revealed a high PEDV load (quantification cycle (Cq)-value of 17) in pooled feces, while tests for Transmissible gastroenteritis virus and Porcine deltacoronavirus were negative. In addition, Brachyspira intermedia/innocens was identified by conventional gel-based PCR , while the sample tested negative by Lawsonia intracellularis real-time PCR [Salmonella enterica serovar Typhimurium was isolated following pre-enrichment in buffered peptone water and typed by standard methods (not shown). No endoparasites were detected in pooled feces.In mid-December 2014, reduced feed uptake and mild diarrhea were observed in a pig farm located in Upper Austria in about 10–20 % of 550 fattening pigs (about 11–12 weeks of age) that had been purchased from a German source a few days before. Two pigs had died and were diagnosed with acute Brachyspira spp. was identified by PCR in a fraction of samples from both groups. Oesophagostomum sp. was demonstrated in a single pooled feces sample from the Austrian-origin pigs, while Salmonella Typhimurium was again identified in feces from the German-origin pigs, but not from those of Austrian origin.By early January 2015, four weeks after the introduction of the German-origin pigs, the clinical situation in the German-origin pigs had improved and feed uptake and daily weight gain were back to normal levels. However, gastrointestinal symptoms were now apparent in >80 % of the Austrian-origin pigs, typically shown by strongly reduced feed uptake and diarrhea that lasted for 2–3 days. No vomiting was observed and no deaths occurred. Pooled feces collected from both German- and Austrian-origin animals (each ten pen-wise pooled samples) tested positive for PEDV in RT-qPCR , with lowest Cq-values (indicative of highest PEDV-load) in samples originating from pigs that had been introduced just a few days ago. These animals were diarrheic at the time of sampling. The mean PEDV Cq-value was lower in feces from the Austrian-origin pigs than from the German-origin pigs (23 Cq vs. 31 Cq). PEDV was further demonstrated by RT-qPCR in 40–50 % of sera collected from both German- and Austrian-origin pigs (each ten samples), respectively. PEDV Cq-values in sera were relatively high > Cq 31) in both groups. PEDV-specific antibodies (PEDV-Ab) were detectable by commercial ELISA  in all sera from the German-origin group and five from the Austrian-origin group; the Austrian-origin pigs that showed diarrhea (pens A7 - A10) were PEDV-Ab negative , clinical symptoms were no longer evident in any of the groups. Nevertheless, 70–91 % of pooled feces samples from both groups still tested positive by PEDV RT-qPCR. In comparison to the previous sampling time-point (early January), mean Cq-values in feces had decreased in both groups, ranging now from Cq 32 - Cq 34, which indicates a drop in PEDV fecal shedding. Furthermore, PEDV-Ab were now detectable in sera collected from those pens that had shown PEDV-Ab negative results 11 days before. Sera from two pens were still positive for PEDV RNA at low level , but absent or low (<1 %) mortality. All affected farms were specialized on finishing pigs. Permission for taking samples was granted by all farm owners.To determine the genetic relationship of PEDV detected on Austrian and Slovenian pig farms to hitherto published strains, sequencing of the PEDV S-gene from selected RT-qPCR positive samples was conducted. Briefly, the complete S-gene was amplified by RT-PCR, utilizing primers S-F1 and S-R1 , followeBrachyspira spp., Oesophagostomum sp. and Salmonella Typhimurium was demonstrated in pooled feces. The chronological appearance of clinical symptoms in the two different groups of pigs was in accordance with semi-quantitative RT-qPCR results  in pen-wise pooled feces and with the order of occurrence of PEDV-Ab in serum. PEDV RNA was detected in sera from both diarrheic and convalescent pigs, although in the latter at very low level. Presence of PEDV RNA in serum is consistent with the acute and later phases of infection, as already described by others [In this report, the first detection of PEDV on an Austrian pig fattening farm is described. Clinical symptoms, such as decreased feed uptake, reduced weight gain and diarrhea initially affected pigs that had been introduced from Germany a few days before. Within a few weeks, this condition had already spread to pigs of Austrian origin that were kept in a different stable on the same farm. PEDV was demonstrated by RT-qPCR in both feces and serum of affected pigs. RT-qPCR results in feces were further confirmed by conventional PCR and sequencing , 23. Fury others .A similar onset of disease following introduction of feeder pigs and rapid spread within the farms was reported from several recent outbreaks in German fattening farms , 17. In High morbidity, but low or no mortality among fattening pigs was also observed during a PED epidemic in 2005–2006 in Italy , 18. WhiThis case report describes the first detection and characterization of PEDV on an Austrian pig farm. PEDV was introduced most likely by purchasing piglets from a German source. The infection spread quickly between different stables of the same farm and led to mild disease in fattening pigs without any deaths related to enteric disease. PEDV RNA was detected in pooled feces for an extended period of time after cessation of clinical symptoms and in the absence of new infections. So far, all available sequences point to a single PEDV S-INDEL strain as the cause of recent PED outbreaks in Western and Central Europe that are, nevertheless, characterized by varying disease severity.All diagnostic and therapeutic procedures involving live animals were performed by an approved veterinarian in the course of routine veterinary health management.Supporting data (Additional file