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The main purpose of the research was to determine if three weekly samplings would produce consistent results for low strongyle egg shedders, generally associated with older horses. Strongyle egg counts per gram of feces (EPG) were done for 661 Thoroughbred mares on 14 farms in 2012-2013 once a week for three consecutive weeks. None of the mares had been treated with ivermectin or moxidectin for more than 90 days and with fenbendazole, oxibendazole, or pyrantel pamoate for more than 60 days. Strongyle EPG count data indicated that 38.3% of the mares were positive at first and second sampling, and 37.8%--at third sampling. There were 5.6-63.3% of horses positive per farm. Correlations between EPG count data in all three weeks are quite high (Rs = 0.84, p<0.001). Level of horse infection was found to be dependent on horse age, type of farm and individual characters of horse. This study documented that strongyle egg counts (EPGs) from individual horses did not differ significantly between weeks. Thus, a single strongyle EPG count can be interpreted as indicative of the level of egg shedding for a given horse. These data support the current trend of not deworming all adult horses on a farm, so that high shedders can be identified and treated. This saves horse owners money and reduces development of potential drug resistance of parasites because of less frequent usage of antiparasitic compounds. Information from this study helps to conclude recommendations for parasite control in horses globally., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The majority of Halicephalobus gingivalis-infections in horses have been fatal and are usually not diagnosed before necropsy. Therefore, knowledge about the nematode and the pathogenesis of infection in horses is limited. This has resulted in an on-going discussion about the port of entry and subsequent dissemination of H. gingivalis within the host. The present case of H. gingivalis-infection in a horse was diagnosed ante mortem. Post mortem findings, the distribution pattern of H. gingivalis nematodes in the brain, a high prevalence of inflammation in close relation to blood vessels, and the presence of the nematode in multiple organs with a disseminated pattern of distribution strongly suggested a haematogenous spread of the nematode in the horse.

Strongylid infections are ubiquitous in grazing horse populations. Infections with cyathostomin (small strongyle) and strongylin (large strongyle) nematodes have long been associated with clinical disease in horses, but little is known about their subclinical impact. A masked, randomized, controlled study was conducted to evaluate the effects of daily administration of pyrantel tartrate on body condition scores, weight gain, fecal egg counts, and total worm counts of young horses repeatedly inoculated with strongylid larvae. Twenty eight immature horses were treated with larvicidal anthelmintic regimens and randomly allocated to two groups. Group 1 horses were given a pelleted placebo product once daily, and those in Group 2 received pyrantel tartrate once daily at ∼ 2.64 mg/kg body weight. On five days during each week, ∼ 5000 infective cyathostomin larvae were administered to each horse. In addition, horses received ∼ 25 infective Strongylus vulgaris larvae once weekly. Horses were maintained on pasture for 154 days and had ad libitum access to grass hay throughout. At approximate, 14-day intervals, body weights were measured, body condition scores were assigned, fecal samples were collected for egg counts, and blood samples were collected for measurement of S. vulgaris antibodies and various physiologic parameters. After 22 weeks at pasture and 14-17 days in confinement, horses were euthanatized and necropsied. Nematodes were recovered and counted from aliquots of organ contents, representative samples of large intestinal mucosa, and the root of the cranial mesenteric artery. Daily treatment with pyrantel tartrate at the recommended dosage significantly reduced numbers of adult cyathostomins in the gut lumen and early third-stage larvae in the cecal mucosa, increased the proportions of fourth-stage larvae in the gut contents, and was accompanied by significant improvements in body condition scores. Fecal egg counts of horses receiving daily pyrantel tartrate were significantly reduced, with percentages of efficacy ranging from 84.4% to 98.9%, but egg counts of both groups increased significantly over the course of the study. Treatment also significantly reduced the numbers of S. vulgaris larvae in the cranial mesenteric artery by 99.2%. Serum antibodies to S. vulgaris apparently persisted from pre-enrollment infections, but ELISA values gradually declined over the course of the study. This study has provided useful insights into the effects of daily pyrantel tartrate on the dynamics of cyathostomin infection, and into some subclinical effects of strongylid parasitism in horses., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Divergent selection in mice for heat loss was conducted in 3 independent replicates creating a high maintenance, high heat loss (MH) and low maintenance, low heat loss (ML) line and unselected control (MC). Improvement in feed efficiency was observed in ML mice due to a reduced maintenance energy requirement but there was also a slight decline in reproductive performance, survivability, and lean content, particularly when compared to MC animals. The objective of this study was to model a life cycle scenario similar to a livestock production system and calculate total inputs and outputs to estimate overall biological efficiency of these lines and determine if reduced feed intake resulted in improved life cycle efficiency. Feed intake, reproductive performance, growth, and body composition were recorded on 21 mating pairs from each line × replicate combination, cohabitated at 7 wk of age and maintained for up to 1 yr unless culled. Proportion of animals at each parity was calculated from survival rates estimated from previous research when enforcing a maximum of 4, 8, or 12 allowed parities. This parity distribution was then combined with values from previous studies to calculate inputs and outputs of mating pairs and offspring produced in a single cycle at equilibrium. Offspring output was defined as kilograms of lean output of offspring at 49 d. Offspring input was defined as megacalories of energy intake for growing offspring from 21 to 49 d. Parent output was defined as kilograms of lean output of culled parents. Parent input was defined as megacalories of energy intake for mating pairs from weaning of one parity to weaning of the next. Offspring output was greatest in MC mice due to superior BW and numbers weaned, while output was lowest in ML mice due to smaller litter sizes and lean content. Parent output did not differ substantially between lines but was greatest in MH mice due to poorer survival rates resulting in more culled animals. Input was greatest in MH and lowest for ML mice for both offspring and parent pairs, consistent with previous results in these lines. Life cycle efficiency was similar in MC and ML mice, while MH mice were least efficient. Ultimately, superior output in MC mice slightly outweighed the lower inputs in ML animals resulting from decreased maintenance energy requirements. Therefore, selection to reduce maintenance energy requirements may be more useful in terminal crosses or in a selection index to reduce possible negative effects on output, especially reproductive performance.

Due to widespread development of anthelmintic resistance in equine parasites, recommendations for their control are currently undergoing marked changes with a shift of emphasis toward more coprological surveillance and reduced treatment intensity. Denmark was the first nation to introduce prescription-only restrictions of anthelmintic drugs in 1999, but other European countries have implemented similar legislations over recent years. A questionnaire survey was performed in 2008 among Danish horse owners to provide a current status of practices and perceptions with relation to parasite control. Questions aimed at describing the current use of coprological surveillance and resulting anthelmintic treatment intensities, evaluating knowledge and perceptions about the importance of various attributes of parasite control, and assessing respondents' willingness to pay for advice and parasite surveillance services from their veterinarians. A total of 1060 respondents completed the questionnaire. A large majority of respondents (71.9%) were familiar with the concept of selective therapy. Results illustrated that the respondents' self-evaluation of their knowledge about parasites and their control associated significantly with their level of interest in the topic and their type of education (P<0.0001). The large majority of respondents either dewormed their horses twice a year and/or performed two fecal egg counts per horse per year. This approach was almost equally pronounced in foals, horses aged 1-3 years old, and adult horses. The respondents rated prevention of parasitic disease and prevention of drug resistance as the most important attributes, while cost and frequent fecal testing were rated least important. Respondents' actual spending on parasite control per horse in the previous year correlated significantly with the amount they declared themselves willing to spend (P<0.0001). However, 44.4% declared themselves willing to pay more than what they were spending. Altogether, results indicate that respondents were generally familiar with equine parasites and the concept of selective therapy, although there was some confusion over the terms small and large strongyles. They used a large degree of fecal surveillance in all age groups, with a majority of respondents sampling and/or treating around twice a year. Finally, respondents appeared willing to spend money on parasite control for their horses. It is of concern that the survey suggested that foals and young horses are treated in a manner very similar to adult horses, which is against current recommendations. Thus, the survey illustrates the importance of clear communication of guidelines for equine parasite control., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Anthelmintic resistance is becoming increasingly prevalent among equine nematode parasites. The first reports documenting resistance were published in the 1960s, just a short time after introduction of the first modern anthelmintics phenothiazine and thiabendazole. Several factors are known to influence development of resistance, but evidence specific to equine parasites is limited. Most current knowledge and applications have been extrapolated from research with trichostrongylid parasites of sheep. The number of cyathostomin species co-infecting horses adds to the complexity of investigating drug resistance but, given their apparent limited biological diversity, viewing these in a unispecific context remains a pragmatic approach. Factors affecting resistance development in cyathostomins include parasite seasonality, life span and fecundity, host immunity, and the existence of encysted stages. Further, parasite refugia have been shown to play a vital role in resistance development in other parasites, and likely is also important in equine parasites. Specific genetic factors for drug resistance and possible modes of inheritance have been identified for trichostrongylid nematodes, but it is widely accepted that several more remain undiscovered. Current evidence with equine and ruminant parasites suggests that fitness is not significantly compromised in drug resistant strains. Attempts to develop in vitro and molecular assays for diagnosing anthelmintic resistance in equine nematodes have had only limited success, standardized guidelines are sorely needed for performing the fecal egg count reduction test in horse populations. Taken together, this review illustrates the complexity of understanding anthelmintic resistance in equine nematodes, and emphasizes the need for further research., (Copyright © 2013 Elsevier B.V. All rights reserved.)

In this study, we investigated the time course of attentional bias for threat-related (angry) facial expressions under conditions of high versus low cognitive (working memory) load. Event-related potential (ERP) and reaction time (RT) data were recorded while participants viewed pairs of faces (angry paired with neutral face) displayed for 500 ms and followed by a probe. Participants were required to respond to the probe while performing a concurrent task of holding in working memory a sequence of digits that were either in the same order (low memory load) or in a random mixed order (high memory load). The ERP results revealed that higher working memory load resulted in enhanced lateralized neural responses to threatening relative to neutral faces, consistent with greater initial orienting of attention to threatening faces (early N2pc: 180-252 ms) and enhanced maintenance of processing representations of threat (late N2pc, 252-320 ms; SPCN, 320-500 ms). The ERP indices showed significant positive relationships with each other, and also with the behavioral index of attentional bias to threat (reflected by faster RTs to probes replacing angry than neutral faces at 500 ms), although the latter index was not significantly influenced by memory load. Overall, the findings indicate that depletion of cognitive control resources, using a working memory manipulation, increases the capacity of task-irrelevant threat cues to capture and hold attention.

Since the 1960s equine parasite control has relied heavily on frequent anthelmintic treatments often applied with frequent intervals year-round. However, increasing levels of anthelmintic resistance in cyathostomins and Parascaris equorum are now forcing the equine industry to change to a more surveillance-based treatment approach to facilitate a reduction in treatment intensity. The principle of selective therapy has been implemented with success in small ruminant parasite control, and has also found use in horse populations. Typically, egg counts are performed from all individuals in the population, and those exceeding a predetermined cutoff threshold are treated. Several studies document the applicability of this method in populations of adult horses, where the overall cyathostomin egg shedding can be controlled by only treating about half the horses. However, selective therapy has not been evaluated in foals and young horses, and it remains unknown whether the principle is adequate to also provide control over other important parasites such as tapeworms, ascarids, and large strongyles. One recent study associated selective therapy with increased occurrence of Strongylus vulgaris. Studies are needed to evaluate potential health risks associated with selective therapy, and to assess to which extent development of anthelmintic resistance can be delayed with this approach. The choice of strongyle egg count cutoff value for anthelmintic treatment is currently based more on tradition than science, and a recent publication illustrated that apparently healthy horses with egg counts below 100 eggs per gram (EPG) can harbor cyathostomin burdens in the range of 100,000 luminal worms. It remains unknown whether leaving such horses untreated constitutes a potential threat to equine health. The concept of selective therapy has merit for equine strongyle control, but several questions remain as it has not been fully scientifically evaluated. There is a great need for new and improved methods for diagnosis and surveillance to supplement or replace the fecal egg counts, and equine health parameters need to be included in studies evaluating any parasite control program., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Obesity in human populations, currently a serious health concern, is considered to be the consequence of an energy imbalance in which more energy in calories is consumed than is expended. We used interval mapping techniques to investigate the genetic basis of a number of energy balance traits in an F11 advanced intercross population of mice created from an original intercross of lines selected for increased and decreased heat loss. We uncovered a total of 137 quantitative trait loci (QTLs) for these traits at 41 unique sites on 18 of the 20 chromosomes in the mouse genome, with X-linked QTLs being most prevalent. Two QTLs were found for the selection target of heat loss, one on distal chromosome 1 and another on proximal chromosome 2. The number of QTLs affecting the various traits generally was consistent with previous estimates of heritabilities in the same population, with the most found for two bone mineral traits and the least for feed intake and several body composition traits. QTLs were generally additive in their effects, and some, especially those affecting the body weight traits, were sex-specific. Pleiotropy was extensive within trait groups (body weights, adiposity and organ weight traits, bone traits) and especially between body composition traits adjusted and not adjusted for body weight at sacrifice. Nine QTLs were found for one or more of the adiposity traits, five of which appeared to be unique. The confidence intervals among all QTLs averaged 13.3 Mb, much smaller than usually observed in an F2 cross, and in some cases this allowed us to make reasonable inferences about candidate genes underlying these QTLs. This study combined QTL mapping with genetic parameter analysis in a large segregating population, and has advanced our understanding of the genetic architecture of complex traits related to obesity.

Optic disc oedema describes the nonspecific, localized swelling of the optic nerve head regardless of aetiology. Therefore, differentiating among the various aetiologies depends on a thorough history and knowledge of the clinical characteristics of the underlying conditions. Papilloedema strictly refers to optic disc oedema as a consequence of elevated intracranial pressure. It is usually a bilateral condition and visual function is preserved until late. Optic disc oedema caused by an anterior optic neuropathy is usually unilateral and accompanied by the loss of visual function.

Changes in maintenance energy requirements and in feed efficiency have been achieved by divergent selection for heat loss in mice in 3 replicates, creating high heat loss, high maintenance (MH) and low heat loss, low maintenance (ML) lines and an unselected control (MC). However, feed intake has mainly been measured in mature animals and not during growth or reproduction. Additionally, there is evidence that reducing maintenance energy will increase fat content, an undesirable result. To evaluate if selection has altered body composition and lifecycle feed intake, mating pairs were continuously mated and maintained for up to 1 yr unless culled. Offspring pairs were sampled from each line at each parity and maintained from 21 to 49 d of age. Feed intake was recorded for mating pairs throughout the year and on offspring pairs. Body weight was recorded on all animals at culling as well as percent fat, total fat, and total lean, measured by dual X-ray densitometry. Average daily gain was also recorded for offspring. Energy partitioning was achieved using 2 approaches: Approach I regressed energy intake of the pair on sum of daily metabolic weight and total gain to obtain maintenance (bm) and growth (bg) coefficients for each line, replicate, feeding period, and sex (offspring pairs only); Approach II calculated bm for each pair assuming constant energy values for lean and fat gain. Energy coefficients and body composition traits were evaluated for effect of selection (MH vs. ML) and asymmetry of selection ([MH + ML]/2 vs. MC). Both MC mating and offspring pairs tended to have greater BW than the average of the selection lines (P < 0.08). Males of offspring pairs weighed more than females (P < 0.01), while females of mating pairs weighed more than males (P < 0.01). Line was insignificant (P > 0.15) for body composition traits. Using Approach I, MH mice had a greater bm than ML mice for mating pairs (P = 0.03) but not offspring pairs (P = 0.50). For Approach II, MH had a greater bm than ML mice for both mating (P = 0.01) and offspring pairs (P = 0.01). The effect of selection for heat loss on body composition was smaller than previously reported and unlikely to outweigh the benefit of reduced feed intake, which was shown to be maintained throughout an entire lifecycle that included reproducing animals. Additionally, the reduction in energy intake seems primarily due to reduced maintenance energy costs, validating the success of the selection procedure.

Parascaris equorum and cyathostomins are currently considered the most important parasites of horses and have traditionally been controlled with anthelmintics belonging to three drug classes: benzimidazoles, the tetrahydropyrimidine pyrantel, and macrocyclic lactones. Unfortunately, resistance to benzimidazoles, and to a lesser extent pyrantel, is widespread in cyathostomins around the world. Furthermore, resistance to macrocyclic lactones appears to be in the early stages of development in cyathostomins in multiple locations. In contrast, P. equorum populations have remained susceptible to the three anthelmintic drug classes for a considerably longer period of time. However, over the last 10 years, resistance to macrocyclic lactones has been described in multiple countries. In contrast, resistance to pyrantel has only been described in the USA; resistance to benzimidazoles has yet to be reported. Despite the large number of reports of anthelmintic resistance in both cyathostomins and P. equorum, there are presently no reports that definitively link anthelmintic resistance with clinical problems in horses. However, that generally appears to be due to a publication bias toward well managed horse farms and the lack of appropriate diagnostic methods for rapidly quantifying anthelmintic resistance in these parasites. Management-based, and other, reasons likely responsible for this apparent anomaly are also discussed. Finally, future research priorities in this area, identified from a discussion at the 9th International Conference on Equine Infectious Diseases, are summarized., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Migrating Strongylus vulgaris and encysted cyathostomin larvae cause a localized inflammatory response in horses. It is unknown whether these larvae elicit a systemic acute phase response (APR), evidenced by changes in serum amyloid A (SAA), haptoglobin (Hp), iron (Fe), albumin, or albumin/globulin (A/G) ratio. In this study, 28 horses were randomly allocated to receive either pyrantel tartrate or a pelleted placebo formulation in their daily feed. Concurrent with treatment, all the horses were administered 5000 pyrantel-susceptible cyathostomin infective larvae once daily, 5 days a week, for 24 weeks. Beginning in the fifth week, the horses also received 25 S. vulgaris larvae once weekly for the remainder of the study. At regular biweekly intervals, fecal samples were collected for quantitative egg counts, and whole blood and serum samples were collected for measurement of packed cell volume, total protein, albumin, globulin, A/G ratio, SAA, Hp, and Fe. On days 161-164, all the horses were euthanatized and necropsied. Samples were collected for enumeration of total luminal worm burdens, encysted cyathostomin larval populations, and migrating S. vulgaris larvae. Concentrations of Hp, Fe, and A/G ratio were associated significantly with strongyle burdens. Only treated male horses had significant increases in serum albumin. Larval S. vulgaris did not associate with Fe, whereas Fe was associated negatively with both total cyathostomin burdens and encysted L4s. The A/G ratios differed significantly between the two treatment groups. Significant differences between groups and individual time points were also observed for Hp and Fe, whereas SAA concentrations remained low throughout the study. In general, this study illustrated that experimental inoculations with S. vulgaris and cyathostomins may be associated with changes in Hp, Fe, and serum proteins, but not with SAA. Overall, these changes suggest that mixed strongyle infections elicit a mild acute phase reaction., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Strongylus vulgaris is regarded as the most pathogenic helminth parasite infecting horses. Migrating larvae cause pronounced endarteritis and thrombosis in the cranial mesenteric artery and adjacent branches, and thromboembolism can lead to ischemia and infarction of large intestinal segments. A recently developed serum ELISA allows detection of S. vulgaris-specific antibodies during the six-month-long prepatent period. A population of horses has been maintained at the University of Kentucky without anthelmintic intervention since 1979, and S. vulgaris has been documented to be highly prevalent. In 2012, 12 foals were born in this population, and were studied during a 12-month period (March-March). Weekly serum samples were collected to monitor S. vulgaris specific antibodies with the ELISA. Nine colts underwent necropsy at different time points between 90 and 300 days of age. At necropsy, Strongylus spp. and Parascaris equorum were identified to species and stage and enumerated. Initial statistical findings indicate a significant interaction between foal age and ELISA results (p<0.042). All foals had initial evidence of S. vulgaris-directed maternal antibodies transferred in the colostrum, but then remained ELISA negative during their first three months of life. Foals born in February and March became ELISA positive at about 12 weeks of age, while those born in April and May went positive at about 15 and 21 weeks, respectively. Foal date of birth was significantly associated with ELISA results (p<0.0001). This could be explained by birth date-dependent differences in parasite exposure. One foal remained ELISA-negative throughout the course of 30 weeks during the study. A significant association was found between ELISA values and larval S. vulgaris burdens (p<0.0001) as well as a three-way interaction between S. vulgaris, S. edentatus, and P. equorum burdens (p<0.001). A plateau with a subsequent decline in ELISA values corresponded with S. vulgaris larvae leaving the bloodstream and migrating back to the intestine., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Divergent selection for heat loss was implemented in mice creating maintenance high (MH) and low maintenance (ML) lines and an unselected control (MC) in 3 independent replicates. Mice from the ML line have improved feed efficiency, due to decreased maintenance energy requirement, but there is potential for a correlated decline in reproductive performance and survivability. Number fully formed (NFF), number born alive (NBA), number weaned (NW), litter weaning weight (LWW), pup weaning weight (PWW), fraction alive at birth (FAB), fraction alive at weaning, and birth interval were recorded at every parity on 21 mating pairs from each line × replicate combination cohabitated at 7 wk of age and maintained for up to 1 yr. Traits were summed over parities to evaluate lifetime production. Pairs were culled due to death or illness, no first parity by 42 d cohabitation, 2 consecutive litters with none born alive, 3 consecutive litters with none weaned, 42 d between parities, or average size of most recent 2 litters less than half the average of first 3 litters. Survival probabilities were produced and evaluated for each line and used to calculate mean number of parities using a Markov-chain algorithm assuming a maximum of 4, 6, 8, 10, or 12 parities or 1 yr. Line was insignificant for all litter traits while NFF, NW, and FAB decreased with parity (P < 0.05) and PWW tended to increase (P < 0.07). The MC mice had higher lifetime NW, LWW, and PWW (P < 0.04). Birth interval showed that MH mice had increasingly larger intervals while remaining the same in ML mice (P < 0.01). In the survival analysis, MC mice had the greatest survival rates overall, but ML mice had the greatest rates in the period up to 5 parities while MH mice had the greatest rates in later parities. This resulted in greater mean number of parities for ML mice up to maximum of 8 parities and higher means for MH mice when the maximum number of allowed parities was 10 or higher. Reproductive performance was not substantially affected by changing maintenance energy requirements. The ML animals appear to survive well in early parities and produce more parities when a low number of maximum parities is enforced, but this benefit declines in later parities and MH animals survive better and increase mean number of parities when turnover rates are low. Therefore, selection for low maintenance animals may be beneficial for systems desiring a short generation interval but less so for systems desiring longevity.

The method presented in this study allows the screening and quantification of 96 drugs, from different groups: opiates, amphetamines, hallucinogens, benzodiazepines, antihistamines, antidepressants, antipsychotics, barbiturates and other sedatives, muscle relaxants, etc. in hair. Drugs are extracted from 10mg of washed hair in 18h by a mixture of methanol:acetonitrile:ammonium formate (pH 5.3). Absolute recovery ranged from 70% to 106% for 75% of the analytes. The limits of detection in the low pg/mg range, may allow the detection of single dose drug exposure, with possible application in drug facilitated assaults (DFA); however, chronic use (compliance) can also be examined. The method has been fully validated for the drugs included in the study. The accuracy of the method was demonstrated by the analysis of certified authentic hair samples containing common drugs of abuse. The hair-method has broad potential as the measuring range is wide for the target analytes and new drugs can easily be added to the method due to the versatility of the extraction procedure and chromatographic system., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Assessment of total uncertainty of analytical methods for the measurements of drugs in human hair has mainly been derived from the analytical variation. However, in hair analysis several other sources of uncertainty will contribute to the total uncertainty. Particularly, in segmental hair analysis pre-analytical variations associated with the sampling and segmentation may be significant factors in the assessment of the total uncertainty budget. The aim of this study was to develop and validate a method for the analysis of 31 common drugs in hair using ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) with focus on the assessment of both the analytical and pre-analytical sampling variations. The validated method was specific, accurate (80-120%), and precise (CV≤20%) across a wide linear concentration range from 0.025-25 ng/mg for most compounds. The analytical variation was estimated to be less than 15% for almost all compounds. The method was successfully applied to 25 segmented hair specimens from deceased drug addicts showing a broad pattern of poly-drug use. The pre-analytical sampling variation was estimated from the genuine duplicate measurements of two bundles of hair collected from each subject after subtraction of the analytical component. For the most frequently detected analytes, the pre-analytical variation was estimated to be 26-69%. Thus, the pre-analytical variation was 3-7 folds larger than the analytical variation (7-13%) and hence the dominant component in the total variation (29-70%). The present study demonstrated the importance of including the pre-analytical variation in the assessment of the total uncertainty budget and in the setting of the 95%-uncertainty interval (±2CVT). Excluding the pre-analytical sampling variation could significantly affect the interpretation of results from segmental hair analysis., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Background: Methadone blood concentrations in fatal cases are highly variable and there is an appreciable overlap between therapeutic methadone concentrations and the concentrations detected in fatalities. As with other opioids, the background of these methadone-related deaths is unclear. The aim of this study was to investigate if short-time abstinence was contributing to the cause of death in methadone-related deaths by evaluation of the EDDP/methadone ratio in blood and urine., Methods: Samples of blood and urine were collected from 103 autopsy cases and analysed for the concentrations of methadone and its main metabolite EDDP. The cases were divided into three groups according to the cause of death: cases where methadone was the cause of death (N=67), cases where poly-drug poisoning including methadone was the cause of death (N=24) and cases where death were caused by other factors (N=12). Urine samples from 11 living persons receiving methadone were also included., Results: In general, a substantial overlap of the methadone concentrations in blood and urine was seen between the groups. There was a tendency of lower median EDDP/methadone urinary ratios in the methadone poisoning group (median: 1.0), poly-drug poisoning group (median: 0.94) and in the fatalities not related to methadone (median: 1.1) compared to the living subjects in methadone treatment (median: 1.6), although the differences were not significant., Conclusion: It was not possible to reveal a possible abstinence period prior to death by using the EDDP/methadone ratio in blood and urine in methadone-related deaths., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Anthelmintic resistance is an increasing challenge for the control of equine parasites. The fecal egg count reduction test (FECRT) is the practical gold standard method for evaluating reduction in anthelmintic efficacy, but the interpretation is complicated due to high levels of variability. A hierarchical statistical model was described for analysis of FECRT data from multiple farms to evaluate the role of biological factors in determining the strongyle efficacy of pyrantel pamoate in a study performed in Denmark. The model was then used to describe two notions of farm efficacy, namely conditional and marginal efficacy. The median of the lower prediction limits was used to describe a robust classification rule. The performance of the methodology was evaluated using Monte Carlo simulations. The field study was performed on 64 Danish horse farms of different breeds. Of 1644 horses, 614 had egg counts ≥ 200 eggs per gram (EPG) and were treated. Individual coprocultures were performed for identification of Strongylus vulgaris from all horses pre-treatment. Thirty-one farms (48.4%) were positive for S. vulgaris, but pyrantel efficacy was unaffected by the presence of this parasite in the statistical model. Further, there were no significant effects of age, gender, or interactions between these, while the pre-treatment egg count was negatively associated with the egg count reduction. The statistical model classified 81.3%, 10.9%, and 7.8% of farms as no signs of resistance (NR), suspect resistance (SR), and resistance (RE), respectively. In comparison, arithmetic calculations classified 68.8%, 17.2%, and 14.1% in the same categories. Using 10,000 simulated data sets, the methodology provided a classification of farms into different efficacy categories with a false discovery of reduced farm efficacy rate equaling 8.74%. In addition, model-classification was unaffected by presence of single outlier horses in a separate simulation study., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Cyathostomins can cause a severe inflammation of equine large intestine characterized by substantial ventral edema and pronounced protein loss. Anthelmintic treatment of horses can result in a localized inflammatory response in the colonic mucosa of clinically normal horses. The aim of this study was to evaluate the systemic inflammatory response of ponies naturally infected with cyathostomins to single dose representatives of three anthelmintic drug classes, namely, oxibendazole, pyrantel pamoate, and moxidectin. Thirty ponies aged between 1 and 18 years of age were allocated to one of three anthelmintic treatments groups. Anthelmintic efficacy was evaluated using the fecal egg count reduction test performed weekly between 2 and 8 weeks post-treatment. Inflammatory responses were evaluated on days 0, 1, 3, 5, and 14 after treatment using hematology, measurement of the acute phase inflammatory markers serum amyloid A, fibrinogen, haptoglobin, and iron, and real-time PCR measurement of expression of the genes for interleukins 1-β and -10, tumor necrosis factor-α, and interferon-γ. There were subtle inflammatory responses to treatment, but cytokine expression was significantly associated with the interaction term between treatment group and anthelmintic efficacy (P<0.05). Of the acute phase markers, only fibrinogen associated with treatment group. The findings suggest that systemic inflammatory responses subsequent to anthelmintic treatment of cyathostomin infection are minimal. It is possible that this response is 'buffered' by anti-inflammatory products of the parasites and/or the anti-inflammatory effects of the macrocyclic lactones., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Decades of intensive anthelmintic treatment has caused equine large strongyles to become quite rare, while the cyathostomins have developed resistance to several drug classes. The larval culture has been associated with low to moderate negative predictive values for detecting Strongylus vulgaris infection. It is unknown whether detection of other large strongyle species can be statistically associated with presence of S. vulgaris. This remains a statistical challenge because of the rare occurrence of large strongyle species. This study used a modified Poisson regression to analyse a dataset for associations between S. vulgaris infection and simultaneous occurrence of Strongylus edentatus and Triodontophorus spp. In 663 horses on 42 Danish farms, the individual prevalences of S. vulgaris, S. edentatus and Triodontophorus spp. were 12%, 3% and 12%, respectively. Both S. edentatus and Triodontophorus spp. were significantly associated with S. vulgaris infection with relative risks above 1. Further, S. edentatus was associated with use of selective therapy on the farms, as well as negatively associated with anthelmintic treatment carried out within 6 months prior to the study. The findings illustrate that occurrence of S. vulgaris in larval cultures can be interpreted as indicative of other large strongyles being likely to be present.

Background: The purpose of the present study was to quantify renal cell injury after ischemia and reperfusion in a pig model using (99m)Tc-lactadherin as a marker of apoptosis and (99m)Tc-sestamibi as a marker of mitochondrial dysfunction., Methods: Thirty-four pigs were randomized into unilateral renal warm ischemia of 120 (WI120) or 240 min (WI240). The glomerular filtration rate (GFR) was calculated by renal clearance of (51)Cr-ethylenediaminetetraacetic acid, and apoptosis was quantified by immunohistochemical detection of caspase-3. After 240 min of reperfusion, intravenous (99m)Tc-lactadherin or (99m)Tc-sestamibi was injected simultaneously with (153)Gd microspheres into the aorta. Ex-vivo static planar images of the kidneys were acquired for determination of the differential renal function of tracer distribution using a gamma camera., Results: In WI120, there was no significant difference in the uptake of microspheres in the ischemic and contralateral normal kidney indicating adequate perfusion (uptake in ischemic kidney relative to the sum of uptake in both kidneys; 46% ± 12% and 51% ± 5%). In WI240, the uptake of microspheres was severely reduced in both groups (17% ± 11% and 27% ± 17%). GFR was severely reduced in the post ischemic kidney in both groups. In both groups, the uptake of lactadherin was reduced (41% ± 8%, 17% ± 13%) but not different from the uptake of (153)Gd microspheres. Caspase-3-positive cell profiles were increased in the post-ischemic kidneys (p < 0.001) and increased as the length of ischemia increased (p = 0.003). In both WI120 and WI240, the amount of (99m)Tc-sestamibi in the ischemic kidney was significantly lower than the amount of (153)Gd microspheres (40 ± 5 versus 51 ± 5 and 20 ± 11 versus 27 ± 17; p < 0.05)., Conclusions: In an established pig model with unilateral renal warm ischemia, we found significantly reduced (99m)Tc-sestamibi uptake relative to perfusion in the kidneys exposed to ischemia indicating a potential ability to detect renal ischemic and reperfusion injuries. However, apoptosis was not detected using (99m)Tc-lactadherin in the post-ischemic kidneys despite increased number of caspase-3-positive cell profiles., Trial Registration: This study is approved by the Danish Inspectorate of Animal Experiments (2010/561-1837).

Locomotor activity, body temperature, feed intake, and BW were measured on 382 mature male mice sampled from lines previously selected (25 generations) for either high (MH) or low (ML) heat loss and an unselected control (MC). Animals were from all 3 independent replicates of the 3 lines and across 4 generations (68 through 71). Locomotor activity and body temperatures were obtained using implanted transmitters with data collection over 4 d following a 3-d postsurgery recovery period. Data were collected every minute and then averaged into 30-min periods, thus providing 192 data points for each mouse. Least-squares means for feed intake adjusted for BW (Feed/BW, feed·BW(-1)·d(-1), g/g) were 0.1586, 0.1234, and 0.1125 (±0.0022) for MH, MC, and ML, respectively, with line being a highly significant source of variation (P < 0.0003). Line effects for locomotor activity counts, transformed to the 0.25 power for analysis, were significantly different, with MH mice being 2.1 times more active than ML mice (P < 0.003); MC mice were intermediate. Differences in body temperature were significant for both line (P < 0.03) and day effects (P < 0.001), with a 0.32°C difference between the MH and ML lines. Fourier series analysis used the combined significant periodicities of 24, 18, 12, 9, 6, and 3 h to describe circadian cycles for activity and body temperature. All 3 lines expressed daily peaks in body temperature and locomotor activity ∼3 h into darkness and ∼2 h after lights were turned on. There was a stronger relationship between locomotor activity and Feed/BW (P < 0.0001) than between body temperature and Feed/BW (P < 0.01); differences between lines in locomotor activity and body temperature explained 17% and 3%, respectively, of differences between lines in Feed/BW. Thus, line differences in locomotor activity contribute to line differences in maintenance, but approximately 80% of the differences between the MH and ML selection lines in Feed/BW remains independent of differences in locomotor activity.

Extreme heat and cold events can create deleterious physiological changes in cattle as they attempt to cope. The genetic background of animals can influence their response to these events. The objective of the current study was to determine the impact of myostatin genotype (MG) on body temperature during periods of heat and cold stress. Two groups of crossbred steers and heifers of unknown pedigree and breed fraction with varying percentages of Angus, Simmental, and Piedmontese were placed in a feedlot over 2 summers and 2 winters. Before arrival, animals were genotyped for the Piedmontese-derived myostatin mutation (C313Y) to determine their MG as either homozygous normal (0 copy; n = 84), heterozygous (1 copy; n = 96), or homozygous for inactive myostatin (2 copy; n = 59). Hourly tympanic and vaginal temperature measurements were collected for steers and heifers, respectively, for 5 d during times of anticipated heat and cold stress. Mean (±SD) ambient temperature for summer and winter stress events were 24.4 (±4.64) and -1.80 (±11.71), respectively. A trigonometric function (sine + cosine) with periods of 12 and 24 h was used to describe the diurnal cyclical pattern. Hourly body temperature was analyzed within a season, and fixed effects included MG, group, trigonometric functions nested within group, and interaction of MG with trigonometric functions nested within group; random effects were animal and residual (Model [I]). A combined analysis of season and group was also investigated with the inclusion of season as a main effect and the nesting of effects within both group and season (Model [C]). In both models, the residual was fitted using an autoregressive covariance structure. A 3-way interaction of MG, season, and trigonometric function periodicities of 24 h (P < 0.001) and 12 h (P < 0.02) for Model [C] indicate that a genotype × environment interaction exists for MG. For MG during summer stress events the additive estimate was 0.10°C (P < 0.01) and dominance estimate was -0.12°C (P < 0.001). During winter stress events the additive estimate was 0.10°C (P < 0.001) and dominance estimate was 0.054°C (P > 0.05). The current study illustrated that a genotype × environment interaction exists for MG and 1-copy animals were more robust to environmental extremes in comparison with 0- or 2-copy animals.

Selection was practiced for high (MH) and low (ML) heat loss using direct calorimetry to create lines of mice differing in feed intake per unit average BW (FI/BW). A total of 25 generations of selection was practiced during maintenance of an unselected control (MC). As a percentage of MC, mice of MH and ML lines differed in heat loss and FI/BW by 56 and 34%, respectively, when selection ceased. The purpose of this study was to determine hepatic mitochondrial efficiency in mice at maintenance across the genetic lines. After measurement of feed intakeand BW over a 2-wk period, mice were euthanized, and their livers were extracted. Livers were homogenized and mitochondria were isolated in buffer for measurement of oxygen consumption and, hence, mitochondrial activity using a Clark-type oxygen electrode. Mitochondria of ML mice expressed greater (P < 0.03) respiratory control ratio by 22% than that for MH mice, revealing greater degree of uncoupling during electron transport in MH mice. Difference in FI/BW (P < 0.001) was large with MH mice consuming 34% more than ML mice. The respiratory control ratio differed between mice of MH and ML lines as expected; however, regression analysis revealed that it explained essentially no variation in feed intake per BW.

Background: Strongyle parasites are ubiquitous in grazing horses. Strongylus vulgaris, the most pathogenic of the large strongyles, is known for its extensive migration in the mesenteric arterial system. The lifecycle of S. vulgaris is characterised by a long prepatent period where the migrating larvae are virtually undetectable as there currently is no test available for diagnosing prepatent S. vulgaris infection. Presence of S. vulgaris larvae in the arterial system causes endarteritis and thrombosis with a risk of non-strangulating intestinal infarctions. Emergence of anthelmintic resistance among cyathostomins has led to recommendations of reduced treatment intensity by targeting horses that exceed a predetermined strongyle faecal egg count threshold. One study suggests an apparent increase in prevalence of S. vulgaris on farms where reduced anthelmintic treatment intensity has been implemented. These issues highlight the need for an accurate and reliable assay for diagnosing prepatent S. vulgaris infection., Methods: Immunoscreening of a larval S. vulgaris cDNA library using hyperimmune serum raised against S. vulgaris excretory/secretory antigens was performed to identify potential diagnostic antigens. Immunoreactive clones were sequenced, one potential antigen was characterised, expressed as a recombinant protein, initially evaluated by western blot (WB) analysis, the diagnostic potential of the IgG subclasses was evaluated by ELISA, and the diagnostic accuracy evaluated using serum from 102 horses with known S. vulgaris infection status., Results: The clone expressing the potential antigen encoded a S. vulgaris SXP/RAL2 homologue. The recombinant protein, rSvSXP, was shown to be a potential diagnostic antigen by WB analysis, and a target of serum IgGa, IgG(T) and total IgG in naturally infected horses, with IgG(T) antibodies being the most reliable indicator of S. vulgaris infection in horses. Evaluation of diagnostic accuracy of the ELISA resulted in a sensitivity of 73.3%, a specificity of 81.0%, a diagnostic odds ratio of 11.69; a positive likelihood ratio (LR) of 3.85 and a negative LR was 0.33. The area under the ROC curve was 0.820., Conclusion: IgG(T) antibodies to recombinant SvSXP show potential for use as an antigen for prepatent diagnosis of migrating stages of S. vulgaris with moderate to good diagnostic accuracy.

Previous research has demonstrated that emotional material is more likely to be remembered than neutral material (Hamann, 2001). The present study employed the item-method of directed forgetting in order to examine whether emotionally negative words are not only easier to remember, but also harder to forget. Event-related potentials (ERPs) were additionally measured in order to investigate the processes of selective rehearsal and active inhibition in directed forgetting. The results demonstrated directed forgetting effects for both neutral and negative words, with a stronger effect for negative items. Late positive potentials (LPPs) for 'to-be-remembered' (TBR) relative to 'to-be-forgotten' (TBF) cues were enhanced when the cues followed negative in comparison to neutral words, indicating the greater selective rehearsal of TBR negative items. Frontal positivities to TBF relative to TBR cues were not modulated by word valence, indicating that inhibitory processes were unaffected by emotion. Taken together, the present research demonstrates for the first time that, not only are emotionally negative words prone to the same directed forgetting effects as neutral words, but that these effects are in fact enhanced for negative words and due to increased selective rehearsal of TBR negative items. The discrepancies between the present findings and those of previous studies are discussed., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Introduction: The objective of this study was to evaluate the Danish Brain Cancer Program by examining the criteria for admission to the program and the results of magnetic resonance imaging (MRI) of the brain in 359 patients referred to the program at the Odense University Hospital during one year. The admission criteria given by the Danish Health and Medicines Authority are as follows: 1. Prior computed tomography or MRI indicating tumour. 2. Progressive focal neurological deficits. 3. Epileptic seizure in adults. 4. Change in behaviour or cognition showing progression. 5. Headache with progression over 3-4 weeks., Material and Methods: This was a retrospective analysis of the cerebral MRI of 359 patients. The patients were categorized by admission criteria and MRI outcome. The findings were grouped into four main outcomes: 1. Primary malignant intracerebral tumour. 2. Intracranial tumour -(including meningeoma and metastasis). 3. Acute pathology in total (including tumours and other acute pathologies). 4. Findings of no consequence., Results: We found 46 acute/subacute pathologies including 21 tumours of which eight were primary intracerebral malignant tumours and 313 scans did not have findings of any consequence. In the group with monosymptomatic headache, we found significantly fewer tumours (p = 0.0066, Fisher's exact test) and acute pathologies (p = 0.0008) than in the remaining groups. In the group with change in behaviour or cognition, we found significantly more primary intracerebral malignant tumours (p = 0.0002), tumours in all (p = 0.0001) and acute pathologies (p = 0.0002) than in the other groups., Conclusion: Fewer tumours than expected were found. Significantly fewer pathologies were found in the group with monosymptomatic headache than in the remaining groups., Funding: not relevant., Trial Registration: not relevant.

Parasites infecting horses are ubiquitous and clinically important across the world. The major parasitic threats to equine health are cyathostomins, Parascaris equorum, Anoplocephala perfoliata, and Strongylus vulgaris. Increasing levels of anthelmintic resistance reported world wide in equine parasites have led to recommendations of constructing sustainable parasite control programmes based on systematic surveillance of parasite levels. Regulations at the European Union level now make anthelmintics available on prescription-only basis and disallow prophylactic treatment. This emphasizes the needs for reliable and practical diagnostic tools for detection of major parasites infecting equines. The current, widely used coprological techniques are important and useful, but they do have considerable limitations as they are incapable of diagnosing the pathogenic migrating stages. Species-specific molecular assays have been developed for diagnosing patent infections with 21 cyathostomin species, A. perfoliata, and S. vulgaris, but none of these have found use in practice. An antibody-directed enzyme-linked immunosorbent assay (ELISA) has been developed, validated and made commercially available for diagnosing A. perfoliata infection, but interpretation is complicated by the fact that horses not harbouring tapeworms can maintain elevated antibody titres. Recent work with a coproantigen ELISA has shown promise for reliable detection of current A. perfoliata infection. Perhaps most remarkable is the fact that the pathogenic larval stages of cyathostomins and large strongyles cannot be detected by any of the available diagnostics. With the lengthy prepatency periods characterizing these parasites, there is a huge need for developing such assays. The recent identification of a possible diagnostic marker for encysted cyathostomins holds great promise, and could become very useful in clinical practice. Several attempts have been made to construct assays for diagnosing the highly pathogenic migrating larvae of S. vulgaris, but none of these have performed sufficiently to make a useful test. The present review illustrates that classical coprological techniques remain the cornerstone of equine parasitology diagnosis and surveillance, and will remain so in a foreseeable future. However, promising progress has been made for developing assays capable of diagnosing prepatent stages of strongyle infection, and there is reason to hope for validated and useful assays in the relative near future., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Strongyle and trichostrongyle parasites are ubiquitous nematodes of grazing livestock. Several molecular diagnostic tests are based upon measuring and quantifying DNA obtained from parasite eggs. It is well known that such eggs undergo development during storage, but it remains unknown to which extent developmental stages can affect the variation of diagnostic test results. This study investigated the influence of developmental stages of strongyle eggs on the variation real-time polymerase chain reaction (PCR) results. Mixed species strongyle eggs were obtained from the faeces of a naturally infected horse. Eggs were isolated and placed in microtiter plates with demineralized water. A total of 25 wells containing 100 eggs each were set up and kept refrigerated for up to five days. Once daily, five wells were examined on an inverted microscope at 100× magnification, where the developmental stages of the eggs were noted, and then eggs harvested for DNA extraction. The protocol was repeated three times. Genomic DNA was extracted using a commercial kit previously validated for strongyle type eggs. PCR reactions were performed with a primer set specific for the ribosomal DNA region for all strongyle type parasites (NC1, NC2). SYBR Green Real-Time PCRs were performed in triplicates. Results revealed a statistically significant increase in PCR yield after three days, which was statistically associated with beginning embryonation of the eggs. In conclusion, storage time and developmental stage of strongyle eggs are significant sources of error in studies based on quantitative real-time PCR analysis. This study suggests that for refrigerated storage of more than three days, eggs should be inactivated and preserved for further analysis., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The large roundworm of horses, Parascaris equorum is considered ubiquitous in breeding operations, and is regarded as a most important helminth pathogen of foals. Over the past decade, this parasite has been reported increasingly resistant to anthelmintic drugs worldwide. This paper reports analysis of the population genetic structure of P. equorum. Adult parasites (n=194) collected from Sweden, Norway, Iceland, Germany, Brazil and the USA were investigated by amplified restriction fragment length polymorphism (AFLP) analysis. The genetic variation was low (Hj=0.12-0.4), for the global population of worms. This was accompanied by a weak degree of population structure (Fst=0.2), low gene flow (Nm=1.0) and low mutation rate (4 Nμ=0.07). Thus, the low genetic diversity is probably a result of a low mutation rate in DNA, although the gene flow (due to global movement of horses) is large enough to allow the spread of novel mutations. Surprisingly, isolates from Icelandic horses were not found to be different from other isolates, in spite of the fact that these have been isolated for thousands of years. The study indicates that the global P. equorum population is essentially homogenous, and continents do not appear to be strong barriers for the population structure of this species. Consequently, the potential spread of rare anthelmintic resistance genes may be rapid in a homogenous population., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Strongyle parasites are ubiquitous in grazing horses, and the large strongyle Strongylus vulgaris is considered the most pathogenic helminth parasite of horses. Recent investigations have suggested an association between occurrence of this parasite and usage of selective therapy based on regular fecal egg counts. The established diagnostic method for S. vulgaris involves larval culture and subsequent morphological identification of third stage larvae under the microscope. Recently, a real-time PCR assay was developed and validated for the detection and semi-quantification of S. vulgaris eggs in equine fecal samples. The purposes of the present study were (a) to determine the presence of S. vulgaris by real-time PCR in Danish and American horses on farms using vastly different anthelmintic treatment regimens and (b) to evaluate the association between larval culture results and the PCR. A total of 991 horses representing 53 different horse farms in Denmark and Central Kentucky were studied. Fresh fecal samples were collected from all horses, and strongyle eggs retrieved for DNA extraction and subsequent real-time PCR analysis. Individual larval cultures were performed on the Danish part of the data set (663 horses on 42 farms). On the Danish farms, the S. vulgaris PCR prevalence was found to be 9.2% on farms not basing parasite control on fecal egg counts, and 14.1% on farms using selective therapy. No horses were PCR positive in the American part of the study (328 horses on 11 farms). Kappa-values indicated a moderate agreement between PCR and larval culture results, while McNemar tests revealed no statistical difference between the paired proportions. Significant associations were found between PCR cycle of threshold (Ct) value groups and larval culture counts. Results indicate that both diagnostic methods can be useful for determining the occurrence of S. vulgaris on horse farms, but that they both are affected by potential sources of error. The PCR results confirmed previous findings suggesting that S. vulgaris can reemerge under selective therapy regimens., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Nematodes belonging to the order Strongylida are ubiquitous in grazing horses, and the large strongyle Strongylus vulgaris is considered the most pathogenic. This parasite was originally described widely prevalent in equine establishments, but decades of frequent anthelmintic treatment appears to have reduced the prevalence dramatically. Increasing levels of anthelmintic resistance in cyathostomin parasites have led to implementation of selective therapy to reduce further development of resistance. It has been hypothesized that S. vulgaris could reoccur under these less intensive treatment circumstances. The aim with the present study was to evaluate the occurrence of S. vulgaris and the possible association with usage of selective therapy. A total of 42 horse farms in Denmark were evaluated for the presence of S. vulgaris using individual larval cultures. Farms were either using a selective therapy principle based on regular fecal egg counts from all horses, or they treated strategically without using fecal egg counts. A total of 662 horses were included in the study. Covariate information at the farm and horse level was collected using a questionnaire. The overall prevalence of S. vulgaris was 12.2% at the individual level and 64.3% at the farm level. Farms using selective therapy had horse and farm prevalences of 15.4% and 83.3%, respectively, while the corresponding results for farms not using selective therapy were 7.7% and 38.9%. These findings were found statistically significant at both the horse and the farm level. Stud farms using selective therapy were especially at risk, and occurrence of S. vulgaris was significantly associated with the most recent deworming occurring more than six months prior. The results suggest that a strict interpretation of the selective therapy regimen can be associated with an increased prevalence of S. vulgaris. This suggests that modifications of the parasite control programs could be considered on the studied farms, but it remains unknown to which extent this can be associated with increased health risks for infected horses., (Copyright © 2012 Elsevier B.V. All rights reserved.)

An ultra-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of 25 common pharmaceuticals in whole blood. The selected pharmaceuticals represent the most frequently detected drugs in our forensic laboratory with basic properties such as analgesics, antidepressants, antihistamines, antihypertensives, antipsychotics and β-blockers. Whole blood samples were extracted with butyl acetate after adjusting pH with 2M NaOH. The target analytes were separated on a 100 × 2.1 mm ACQUITY BEH 1.7 µm C18 column by a formic acid/acetonitrile gradient elution using a Waters ACQUITY Ultra-Performance Liquid Chromatography system. Quantification was performed on a Waters tandem quadrupole ACQUITY TQD using multiple reaction monitoring in positive mode. The analytes were eluted within 11 min. The limit of quantification (LOQ) ranged from 0.002 to 0.01 mg/kg depending on the analyte. A good linear behavior was achieved for all analytes in the range from LOQ to 1.0 or 2.0 mg/kg blood. The absolute recoveries were between 55-87% for all compounds except norfluoxetine (44%). The method showed acceptable precision and accuracy for almost all analytes. Only unstable compounds like levomepromazine, methylphenidate, mirtazapine, norfluoxetine and zuclopenthixol deviated more. The method was successfully applied to more than 200 authentic blood samples within a year from forensic investigations.

Avermectins and milbemycins (AM) are potent compounds against all major nematode parasites, but their continuous usage has led to the development of widespread resistance in many of the important species of ruminant and equine parasites. The exception to this has been the cyathostomins, where AM resistance was recently first reported only after decades of drug exposure. Data from a Brazilian study suggests that AM resistance has developed in cyathostomins and reports of shortened egg reappearance periods after ivermectin treatment have been published recently from USA and Germany. Thus, AM resistance in cyathostomins is an emerging worldwide concern, but there is only limited amount data on the extent of this problem. To limit the development and spread of AM-resistant cyathostomins the equine industry must implement new strategies for worm control, and the veterinary parasitology community must develop and validate improved protocols for detecting anthelmintic resistance in the field., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Clinically important equine parasites are ubiquitous in managed horse populations. The traditional approach to parasite control is frequent administration of anthelmintics to all horses on a farm. However, increasing levels of anthelmintic resistance is forcing horse owners and veterinarians to shift this control paradigm. Treatment regimens involving routine deworming of all horses throughout the year are now being replaced by more sustainable approaches, which take in to account the importance of maintaining adequate parasite refugia. The selective therapy principle has been recommended for more than 15 years, but there is limited experience with this approach. The relative magnitude of the faecal egg count for an individual horse is a consistent trait, and this provides a reliable basis for selective therapy. But no studies have evaluated the consequences of selective therapy in the long-term, and such studies are strongly needed to validate this approach. Importantly, it remains unclear how selective therapy may affect the prevalence and intensity of other parasites of significant pathogenic potential (e.g. Strongylus vulgaris), which have become uncommon due to years of intensive chemotherapy. Consequently, a selective approach requires vigilant surveillance of the parasite fauna and intensity. This places a demands for reliable diagnostic tools. Also noteworthy is the fact that the majority of equine nematode parasites are more pathogenic during their larval stages, when they cannot be detected by traditional egg counting techniques. Consequently, parasite-specific diagnostic tools capable of assessing prepatent parasite burdens, and able to differentiate between strongyle species of different pathogenic potentials, would be of great value to the equine clinician. Tools for detecting infections with the tapeworm Anoplocephala perfoliata are laborious, difficult to interpret, and at present there is no established method to evaluate treatment efficacy. Thus, better diagnostic tools are needed for tapeworms as well. Biological control, especially the predacious fungi have demonstrated good potential as an adjunct for strongyle control and such a product could easily have a market in equine establishments. In summary, there is general agreement that the traditional treat-all at frequent interval approach should be abandoned, and that optimal parasite control can be maintained with far fewer anthelmintic treatments. But better diagnostic techniques and more evidence documenting the long-term consequences of selective therapy programs are needed to develop and validate systems for sustainable equine parasite control., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Strongyle parasites are ubiquitous in grazing horses. Of these, the bloodworm Strongylus vulgaris is regarded as most pathogenic. Increasing levels of anthelmintic resistance in strongyle parasites has led to recommendations of decreased treatment intensities, and there is now a pronounced need for reliable tools for detection of parasite burdens in general and S. vulgaris in particular. The only method currently available for diagnosing S. vulgaris in practice is the larval culture, which is laborious and time-consuming, so veterinary practitioners most often pool samples from several horses together in one culture to save time. Recently, molecular tools have been developed to detect S. vulgaris in faecal samples. The aim of this study was to compare the performance of a conventional polymerase chain reaction (PCR) assay with the traditional larval culture and furthermore test the performance of pooled versus individual PCR for farm screening purposes. Faecal samples were obtained from 331 horses on 18 different farms. Farm size ranged from 6 to 56 horses, and horses aged between 2 months and 31 years. Larval cultures and PCR were performed individually on all horses. In addition, PCR was performed on 66 faecal pools consisting of 3-5 horses each. Species-specific PCR primers previously developed were used for the PCR. PCR and larval culture detected S. vulgaris in 12.1 and 4.5% of individual horses, respectively. On the farm level, eight farms tested positive with the larval culture, while 13 and 11 farms were positive with the individual and pooled PCRs, respectively. The individual PCR method was statistically superior to the larval culture, while no statistical difference could be detected between pooled and individual PCR for farm screening. In conclusion, pooled PCR appears to be a useful tool for farm screening for S. vulgaris., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Growth, feed intake, and temperament indicator data, collected over 5 yr on a total of 1,141 to 1,183 mixed-breed steers, were used to estimate genetic and phenotypic parameters. All steers had a portion of Hereford, Angus, or both as well as varying percentages of Simmental, Charolais, Limousin, Gelbvieh, Red Angus, and MARC III composite. Because the steers were slaughtered on various dates each year and the animals thus varied in days on feed, BW and feed data were adjusted to a 140-d feeding period basis. Adjustment of measures of feed efficiency [G:F or residual feed intake (RFI), intake adjusted for metabolic body size, and BW gain] for body fatness recorded at slaughter had little effect on the results of analyses. Average daily gain was less heritable (0.26) than was midtest BW (MBW; 0.35). Measures of feed intake had greater estimates of heritability, with 140-d DMI at 0.40 and RFI at 0.52; the heritability estimate for G:F was 0.27. Flight speed (FS), as an indicator of temperament, had an estimated heritability of 0.34 and a repeatability of 0.63. As expected, a strong genetic (0.86) correlation was estimated between ADG and MBW; genetic correlations were less strong between DMI and ADG or MBW (0.56 and 0.71). Residual feed intake and DMI had a genetic correlation of 0.66. Indexes for phenotypic RFI and genotypically restricted RFI (no correlation with BW gain) were compared with simple economic indexes incorporating feed intake and growth to elucidate expected selection responses under different criteria. In general, few breed differences were detected across the various measurements. Heterosis contributed to greater DMI, RFI, and MBW, but it did not significantly affect ADG, G:F, or FS. Balancing output (growth) with input costs (feed) is needed in practicing selection, and FS would not be recommended as an indicator trait for selection to change feed efficiency. An index including BW gain and RFI produced the best economic outcome.

Horses, mules and donkeys are indispensable farming and working animals in many developing countries, and their health status is important to the farmers. Strongyle parasites are ubiquitous in grazing horses world-wide and are known to constitute a threat to equine health. This study determined the prevalence of strongyle infection, the efficacy of ivermectin and fenbendazole treatment, and strongyle re-infection rates of working horses during the dry months in Nicaragua. One hundred and five horses used by farmers for transport of people and goods were randomly allocated into three treatment groups, i.e., the IVM group treated with ivermectin, the FBZ group treated with fenbendazole and the control group treated with placebo. Determined by pre-treatment faecal egg counts (FECs), horses showed a high prevalence (94%) of strongyle parasites with high intensities of infection (mean FEC of 1117 eggs per gram (EPG) with an SD of 860 EPG, n=102). Body condition scores of all horses ranged from 1.5 to 3.5 with a mean of 2.4 (scales 1-5). Fourteen days after treatment faecal egg count reductions (FECRs) were 100% and 94% in the IVM and the FBZ groups, respectively. The egg reappearance period (ERP) defined as the time until the mean FEC reached 20% of the pre-treatment level, was estimated as 42 days for the FBZ group and 60 days for the IVM group. Individual faecal cultures were set up and the larval differentiation revealed a 36% prevalence of Strongylus vulgaris before treatment (n=45). In the FBZ group, 25% of the horses were S. vulgaris-positive 70 days post treatment compared to 11% in the IVM group. Our results indicate that strongyle infection intensities in Nicaragua are high and that S. vulgaris is endemic in the area. Furthermore, efficacies and ERPs of IVM and FBZ were within the expected range with no signs of anthelmintic resistance., (Copyright © 2011 Elsevier B.V. All rights reserved.)

In Denmark the prevalence of both asthma and caesarean section has increased during the last 20 years. This review investigates the coherence between caesarean section and the development of asthma. Twelve epidemiological articles about children over three years were included. Eight of 12 studies discovered no association between caesarean section and asthma. There is no difference in the quality of the studies. We can conclude that it is uncertain if caesarean section is a risk factor to the development of asthma, but it cannot be eliminated that asthma is one of many consequences of caesarean section.

Reasons for Performing Study: Strongyle worm burdens are assumed to subclinically affect equine performance. This assumption appears to be particularly pronounced in the equine racing industry., Hypothesis: Race results of Standardbred trotters are negatively affected by high strongyle faecal egg count levels., Methods: Faecal samples were obtained from 213 racing Standardbred trotters, aged ≥ 2 years, and stabled at training facilities of 21 professional trainers with license at racecourses in Denmark. Strongyle egg counts were generated using a McMaster technique. Race results were recorded as the finishing position of the horse (position 1-3 vs. finishing lower) and winning purse. The effect of strongyle egg counts on performance was assessed using regression analyses., Results: Strongyle egg counts ranged from 0-3500 with a mean of 319 and a median of 150 eggs/g. Finishing in positions 1-3 was significantly associated with higher egg counts., Conclusions: Race performance of the population of professionally trained Danish Standardbred trotters was not negatively affected by higher strongyle faecal egg count levels., Potential Relevance: The traditional frequent anthelmintic treatments of racehorses may be inordinate., (© 2011 EVJ Ltd.)

The effects of individual SNP and the variation explained by sets of SNP associated with DMI, metabolic midtest BW, BW gain, and feed efficiency, expressed as phenotypic and genetic residual feed intake, were estimated from BW and the individual feed intake of 1,159 steers on dry lot offered a 3.0 Mcal/kg ration for at least 119 d before slaughter. Parents of these F(1) × F(1) (F(1)(2)) steers were AI-sired F(1) progeny of Angus, Charolais, Gelbvieh, Hereford, Limousin, Red Angus, and Simmental bulls mated to US Meat Animal Research Center Angus, Hereford, and MARC III composite females. Steers were genotyped with the BovineSNP50 BeadChip assay (Illumina Inc., San Diego, CA). Effects of 44,163 SNP having minor allele frequencies >0.05 in the F(1)(2) generation were estimated with a mixed model that included genotype, breed composition, heterosis, age of dam, and slaughter date contemporary groups as fixed effects, and a random additive genetic effect with recorded pedigree relationships among animals. Variance in this population attributable to sets of SNP was estimated with models that partitioned the additive genetic effect into a polygenic component attributable to pedigree relationships and a genotypic component attributable to genotypic relationships. The sets of SNP evaluated were the full set of 44,163 SNP and subsets containing 6 to 40,000 SNP selected according to association with phenotype. Ninety SNP were strongly associated (P < 0.0001) with at least 1 efficiency or component trait; these 90 accounted for 28 to 46% of the total additive genetic variance of each trait. Trait-specific sets containing 96 SNP having the strongest associations with each trait explained 50 to 87% of additive variance for that trait. Expected accuracy of steer breeding values predicted with pedigree and genotypic relationships exceeded the accuracy of their sires predicted without genotypic information, although gains in accuracy were not sufficient to encourage that performance testing be replaced by genotyping and genomic evaluations.

Ivermectin resistance has recently been described in Parascaris equorum and there have been reports from several countries of a shortened egg reappearance period (ERP) following ivermectin treatment for cyathostomins. This study was aimed at determining the efficacy of ivermectin in treating cyathostomins and P. equorum in Danish horses. A total of 196 animals were selected from 52 farms, all of which were using a selective anthelmintic treatment strategy. ERP was investigated with weekly samples from 96 horses from nine farms. Horses were treated with ivermectin oral paste by their owners at an estimated dose rate of 0.2mg/kg. Overall, faecal egg counts were reduced 10-14 days after treatment by 96.9% and 100% for P. equorum and cyathostomins, respectively. Mean faecal egg count reductions at 4 and 6 weeks post treatment were 99.5% and 96.9%, respectively. No signs of developing ivermectin resistance were found in either cyathostomins or P. equorum in the studied horses., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Background: Disulfiram is used to a great extent in Denmark to treat alcoholism but the evidence is limited., Aim: To study the effect of supervised disulfiram treatment in alcohol dependence. Subjects were recruited from a psychiatric emergency ward following alcohol withdrawal treatment., Methods: A total of n=39 patients were openly randomized to either disulfiram 800 mg twice a week for 26 weeks (n=19) or no disulfiram (n=20). All patients were also treated with cognitive behavioural therapy (CBT) in groups., Results: The rate of abstinence was 20% and 26% in the control and disulfiram group, respectively. This difference was not statistically significant (NS). A trend towards increased mean time to first drink was found in the disulfiram group (96 vs. 76 days in the control group, NS), while fewer patients in this group completed CBT group therapy (41% vs. 67% in the control group, NS). Alcohol-free days were 100 days in both groups (NS)., Conclusion: Supervised disulfiram administration did not have any major impact on the treatment outcome.

Increasing levels of anthelmintic resistance in equine nematodes have led to recommendations of more sustainable anthelmintic treatment protocols with emphasis on parasite surveillance and diagnosis, rather than prophylactic calendar-based treatments. This requires knowledge of the diagnostic test performance of techniques for counts of eggs per gram of feces (EPG) as well as methods for culturing, counting and identifying third stage (L(3)) strongyle larvae per gram of feces (LPG). For horses, such information does not exist in the published literature. The aim of this study was to examine the relationship between worm count and fecal egg count (FEC) data for strongyle and Parascaris equorum infections as well as larval culture counts for diagnosing Strongylus spp. infections. Necropsy data from 693 horses used for critical or controlled tests, including information on total worm counts, fecal egg counts (FEC) and larval culture results collected at the University of Kentucky over a period of 50 years were analyzed. Sensitivity, specificity, positive and negative predictive values, and receiver operator characteristic (ROC) curves were generated for the larval cultures and ascarid egg counts. For the strongyle egg counts, potential FEC cutoff values for treatment were evaluated statistically by comparing the total strongyle worm counts below and above chosen cutoff values. All tests had high positive predictive values (>0.95), but moderate negative predictive values (<0.70). The negative predictive values of the larval counts were negatively affected by increasing egg count levels. Strongyle FEC cutoff values up to the level of 500 EPG yielded significantly higher strongyle worm counts in the treatment group, whereas no differences were found at higher cutoffs. This supports usage of cutoffs for treatment in the 0-500 EPG range. Altogether, the present study yields unique and useful information of widely used methods for parasite surveillance and diagnosis in equine establishments., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Introduction: Barbiturates are potent drugs for treatment of alcohol withdrawal symptoms, but they entail a risk of over-dosage and respiratory depression. The purpose of the present study was to investigate the correlation between phenobarbital dose and phenobarbital blood concentration in patients withdrawing from long-term alcohol intoxication., Material and Methods: A total of 497 patients who were hospitalized for treatment of alcohol withdrawal symptoms during an 18-month period were enrolled in the study. Phenobarbital 200 mg was administered orally every 30 or 60 minutes in response to the observed symptoms. Within the first 24 hours after admission, i.e. at 8 AM, blood was collected for determination of phenobarbital concentration, and the cumulated dose of phenobarbital at the time of the blood sampling was registered., Results: The mean cumulated phenobarbital dose at the time of the blood sampling was 877 mg +/- 557 mg, while the mean plasma phenobarbital concentration was 104 micromol/l +/- 62 micromol/l. A statistically significant linear correlation between phenobarbital dose and concentration was found for both males and females as 83% and 84% of the variation in drug concentration, respectively, could be explained by the phenobarbital dose. We observed no serious complications of the phenobarbital treatment--including respiratory problems or severe sedation., Discussion: The strong linear correlation between phenobarbital dose and concentration suggests that absorption of plasma phenobarbital from the gastrointestinal system is highly predictable.

For 9 consecutive months (March-November 2008), faecal samples were collected monthly from 129 horses residing within 40 km of Salzburg, Austria. Samples were analysed quantitatively using a modified McMaster egg counting technique. Whenever a faecal egg count (FEC) result exceeded 250 eggs per gram (EPG), the horse was treated with pyrantel, ivermectin or moxidectin. In 52 of 129 horses (40.3%), no strongyle eggs were ever detected over the course of 9 months. In 39 horses (30.2%), strongyle eggs were detected in at least 1 sample, but the egg count never exceeded 250 EPG. The remaining 38 (29.5%) horses were treated at least once in response to a FEC that exceeded 250 EPG. As a result of this selective anthelmintic scheme, the total number of anthelmintic treatments was reduced to 54% of the number of treatments administered to the same horses in the previous year. Both the maximum and mean FEC dropped significantly after initiation of the study. A statistically significant, negative correlation was demonstrated between the maximum and mean FEC of a horse and its age. Pasture hygiene appeared to reduce FECs, but the effect was not statistically significant. The magnitude of the initial FEC was significantly correlated with the maximum FECs in the subsequent 8 months (p<0.01). The same relationship was observed for the maximum FEC of the first 2 samples. Furthermore, horses which required several anthelmintic treatments had a higher initial FEC and a greater maximum FEC in the first 2 samples than horses which received only one or no treatment. These results suggest that selective anthelmintic treatment accomplished a reduced pasture contamination with strongyle eggs, while simultaneously decreasing the number of anthelmintic treatments. Sustained implementation of a selective treatment strategy has the potential to reduce selection pressure for anthelmintic resistance. These results reported herein will assist equine practitioners in designing and monitoring sustainable anthelmintic treatment programs., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

Introduction: Wernicke's encephalopathy (WE) is caused by lack of thiamin. In Denmark, WE most often occurs in alcoholics. In the present study, we wanted to investigate the frequency of WE in patients hospitalized for treatment of alcohol withdrawal symptoms in a psychiatric emergency ward., Material and Methods: Prospective registration of symptoms and treatment regimen in all patients admitted for alcohol withdrawal treatment in the period 22.02.2007-31.08.2008., Results: Symptoms of WE occurred in 52 of a total of 497 patients (11%). Ataxia of gait was the predominant symptom, succeeded by cognitive impairment, whereas ocular palsy was unusual. Patients with WE were on average administered approximately 50% more phenobarbital than patients without symptoms of WE (p < 0.01), probably reflecting a more severe withdrawal reaction. In 21 of the 52 patients with WE (41%) the preceding period of alcohol intake was shorter than 14 days., Discussion: WE is a frequently occurring condition among patients treated for alcohol withdrawal symptoms in psychiatric wards. A possible pathogenetic factor is the alcohol withdrawal reaction, as cerebral hyperactivity may lead to increased thiamin consumption and thus depletion of the depots. To avoid insufficient treatment, routine administration of large doses of intravenous thiamine to all patients admitted with alcohol withdrawal symptoms should be considered.