Genotoxic Effect of Atrazine, Arsenic, Cadmium and Nitrate, Individually and in Mixtures at Maximum Contaminant Levels on mammalian Breast Cell Lines

There is strong evidence that hormonally active agents (HAAs) such as Atrazine (ATZ), Cadmium (Cd), Arsenic (As) and Nitrate (NO3) have both estrogenic activity and carcinogenic potential. Atrazine has clastogenic effects and may also act as tumor promoter as it induces the aromatase enzyme. Arsenic and Cadmium have been implicated in the etiology of skin, lung, prostate and liver cancers. Nitrate in drinking water has been found to increase the risk of bladder cancer.This study examined the genotoxicity of the aforementioned HAAs alone and in mixtures using mammalian breast cell lines, MCF-7 and MCF-10A, which are estrogen receptorpositive (ER+) and estrogen receptor-negative (ER-), respectively. To study the clastogenic potential by whole cell and flow karyotype damage, cells were exposed to environmentally relevant concentrations of ATZ, Cd, As and NO3 for 4 and 7 days.Results indicated that all treatments induced whole cell clastogenicity in MCF-7 cells; except Cd and NO3 after 4 and 7 days as well as the 10% quaternary As mixture after 1 week. In MCF-10A cells, all treatments except the 10% mixture induced whole cell clastogenicity after 4 days, where flow karyotype damage was detected in all treatments except for the 10% mixture after 1 week. Estrogen caused whole cell damage but not flow karyotype damage in MCF-7. On the other hand, estrogen caused flow karyotype damage and not whole cell damage in MCF-10A cells, suggesting that estrogen receptor modulated the genotoxicity of estrogen. Cd caused flow karyotype damage but not whole cell damage in MCF-7 indicating that Cd’s gentoxicity is not related to its estrogenic activity.Keywords: HAAs, clastogenicity, flow-karyotype, genotoxicity, MCF-7, MCF-10

Stranger danger? An investigation into the influence of human-horse bond on stress and behaviour

Human-animal bond is receiving increasing attention and is thought to confer benefits on well-being and performance in working animals. One important benefit of bonding is the "safe base" an attachment figure provides, which manifests in better coping and increased exploration during potential threat. However, there is limited research exploring the existence or benefits of human-horse bonds, though bonding is sought after by both pleasure and elite riders. The purpose of the current study was to determine whether the presence of horses' owners confers a safe-base, therefore improving horse behaviour and physiological stress responses during novel handling tests. Horses completed two different handling tests, one with their owner and the other with an unfamiliar experimental handler (n = 46). Test and handler order was randomised and handlers were double blind to the performance of the horse with the alternate handler. Time taken to complete the tests and proactive behaviour were measured as indicators of performance and compliance. Core temperature, discrepancy in eye temperature, heart rate and heart rate variability were recorded to assess stress responses. If horses experience a "safe base" effect in the vicinity of their owner, they would be expected to show lower stress responses and greater behavioural compliance, compared to being handled by a stranger. There was no difference in behaviour or any physiological stress response between the handlers. This indicates that a calm, competent, but unknown handler may be equally effective to an owner during stressful procedures as neither equine performance nor affective state supported a safe-base effect. This supports previous research suggesting that the level of bond between human and horse may not be the most salient factor in coping or compliance during training and handling. These findings have implications for veterinary and clinical behaviour counselling, where novel human handlers must modify behaviour under potentially stressful circumstances

Poker face: discrepancies in behaviour and affective states in horses during stressful handling procedures

Correct assessment of stress in horses is important for both horse welfare and handler safety during necessary aversive procedures. Handlers depend on behaviour when judging how well an individual is tolerating stressful procedures such as loading or veterinary intervention. However, evidence suggests that behaviour may not accurately reflect affective states in horses. This may be explained by individual differences in coping styles, which have tentatively been identified in horses. The current study assessed whether behaviour during two novel handling procedures was associated with physiological indicators of stress. Core temperature, discrepancy in eye temperature and heart rate variability (HRV) were compared with compliance and proactivity shown by horses during two novel handling tests (n = 46). Test A required subjects to cross a large blue tarpaulin on the ground. Test B required subjects to walk through plastic streamers suspended overhead. Physiological indicators of stress did not correlate with time taken to complete the handling tests. This indicates some subjects crossed an object they found aversive. Crossing time may be influenced more by stimulus-control than the level of aversion experienced. The level of proactivity shown was not associated with HRV, HR, core temperature or the discrepancy in temperature between eyes. This suggests that proactive horses, which appear more stressed, show similar stress responses to more reactive individuals. These findings support previous research indicating that behaviour commonly used within the equestrian industry may not provide reliable indicators of a horse’s ability to tolerate a stressful procedure. The influence of training and the extent to which a horse is under stimulus-control may over-shadow inherent emotional responses, with implications for handler safety and horse welfare

Dually noted: the effects of a pressure headcollar on compliance, discomfort and stress in horses during handling

Horse handlers often encounter problem behaviour resulting from a lack of stimulus control. Handlers are often only 15% of the weight of horses, which evolved strong flight responses. Therefore, many riders and handlers resort to the use of "aids" to maintain control of their animals. However, there are increasing concerns about the efficacy and welfare implication of such devices, particularly when applied to sensitive facial structures. One such device is a Dually® headcollar which aims to increase compliance. Despite its popularity, little is known about the effects of this aid on behaviour or stress. The aim of the current study was to determine whether the use of a Dually headcollar improves compliance during handling and, if so, whether this might be achieved with concomitant increases in stress or discomfort. Subjects completed two novel handling tests, one wearing a Dually with a line attached to the pressure mechanism and one attached to the standard ring as a Control. Crossing time and proactive behaviour were recorded as indicators of compliance. Core temperature and the discrepancy between eye temperatures were measured using IRT before and after testing as an indicator of stress. The Horse Grimace Scale (HGS) was used to measure discomfort caused by each configuration of the device. The Dually did not result in more compliant behaviour, compared to the Control (p=0.935; p=0.538). However, the Dually configuration did result in a significantly higher HGS scores (p=0.034). This may indicate that there is an impact on animal welfare by using this device that is not justified by improved behaviour. However, IRT readings of core temperature (p=0.186) and discrepancy between the eyes (p=0.972) did not indicate the Dually increased stress in subjects. Taken together, this suggests the Dually is ineffective in naïve horses but causes increased discomfort

Zylkéne to load? The effects of alpha-casozepine on compliance and coping in horses during loading

Horses are routinely transported for access to safe off-road riding, veterinary care, breeding, sale or moving to a new home environment. However, transport is a known stressor in horses. For this reason, problem behavior when loading is a commonly reported issue which presents risks to handlers and horse welfare. Existing literature and manufacturers recommendations suggests that alpha-casozepine may be effective in improving the behavior and welfare of horses during loading onto a vehicle for transport. The current paper aims to assess the behavioral and physiological effects of a commercially available alpha-casozepine feed supplement (Zylkéne Equine) in horses during loading and confinement on a transport lorry. Subjects (n = 10) were loaded once with the supplement and once without, in a balanced random order with each subject acted as their own control. The handler was blind to treatment. Time to load onto the lorry, and movement of feet, licking and chewing, and vocalising within the lorry, were recorded as behavioral indicators of compliance and coping. Heart rate, heart rate variability, salivary cortisol, and infrared thermography of both core temperature and the discrepancy between eyes, were measured as indicators of arousal. There were no significant differences in physiology between Treatment and Control (P > 0.05). Treatment resulted in a significantly shorter Loading Time than control (P = 0.04), however, the actual difference in median time was only 0.45 seconds. No other behavioral indicator differed between Treatment and Control (P > 0.05). Power analysis revealed the sample was sufficient to detect a significant effect. Where modest effects were observed for a small number of variables, Treatment effect contradicted predictions. Taken together, this indicates that alpha-casozepine does not affect a horse’s ability to cope with loading and confinement in a horse lorry. Further work is required to ascertain whether the maximum dosage – twice that used here – might affect coping and behavior in horses