Volume-Controlled 19 F MR Ventilation Imaging of Fluorinated Gas.

BACKGROUND: 19 F MRI of inhaled gas tracers has developed into a promising tool for pulmonary diagnostics. Prior to clinical use, the intersession repeatability of acquired ventilation parameters must be quantified and maximized. PURPOSE: To evaluate repeatability of static and dynamic 19 F ventilation parameters and correlation with predicted forced expiratory volume in 1 second (FEV1 %pred) with and without inspiratory volume control. STUDY TYPE: Prospective. POPULATION: A total of 30 healthy subjects and 26 patients with chronic obstructive pulmonary disease (COPD). FIELD STRENGTH/SEQUENCE: Three-dimensional (3D) gradient echo pulse sequence with golden-angle stack-of-stars k-space encoding at 1.5 T. ASSESSMENT: All study participants underwent 19 F ventilation MRI over eight breaths with inspiratory volume control (w VC) and without inspiratory volume control (w/o VC), which was repeated within 1 week. Ventilated volume percentage (VVP), fractional ventilation (FV), and wash-in time (WI) were computed. Lung function testing was conducted on the first visit. STATISTICAL TESTS: Correlation between imaging and FEV1 %pred was measured using Pearson correlation coefficient (r). Differences in imaging parameters between first and second visit were analyzed using paired t-test. Repeatability was quantified using intraclass correlation coefficient (ICC) and coefficient of variation (CoV). Minimum detectable effect size (MDES) was calculated with a power analysis for study size n = 30 and a power of 0.8. All hypotheses were tested with a significance level of 5% two sided. RESULTS: Strong and moderate linear correlations with FEV1 %pred for COPD patients were found in almost all imaging parameters. The ICC w VC exceeds the ICC w/o VC for all imaging parameters. CoV was significantly lower w VC for initial VVP in COPD patients, FV, CoV FV, WI and standard deviation (SD) of WI. MDES of all imaging parameters were smaller w VC. DATA CONCLUSION: 19 F gas wash-in MRI with inspiratory volume control increases the correlation and repeatability of imaging parameters with lung function testing. EVIDENCE LEVEL: 2 TECHNICAL EFFICACY: Stage 2.

Differentiation of movement behaviour in an adaptively diverging salamander population.

Dispersal is considered to be a species-specific trait, but intraspecific variation can be high. However, when and how this complex trait starts to differentiate during the divergence of species/lineages is unknown. Here, we studied the differentiation of movement behaviour in a large salamander population (Salamandra salamandra), in which individual adaptations to different habitat conditions drive the genetic divergence of this population into two subpopulations. In this system, salamanders have adapted to the deposition and development of their larvae in ephemeral ponds vs. small first-order streams. In general, the pond habitat is characterized as a spatially and temporally highly unpredictable habitat, while streams provide more stable and predictable conditions for the development of larvae. We analysed the fine-scale genetic distribution of larvae, and explored whether the adaptation to different larval habitat conditions has in turn also affected dispersal strategies and home range size of adult salamanders. Based on the genetic assignment of adult individuals to their respective larval habitat type, we show that pond-adapted salamanders occupied larger home ranges, displayed long-distance dispersal and had a higher variability of movement types than the stream-adapted individuals. We argue that the differentiation of phenotypically plastic traits such as dispersal and movement characteristics can be a crucial component in the course of adaptation to new habitat conditions, thereby promoting the genetic divergence of populations.

Zebra finch males compensate in plumage ornaments at sexual maturation for a bad start in life.

BACKGROUND: An individual's fitness in part depends on the characteristics of the mate so that sexually attractive ornaments, as signals of quality, are used in mate choice. Often such ornaments develop already early in life and thus are affected by nutritional conditions experienced then. Individuals thus should benefit by compensating as soon as possible for poor initial development of ornaments, to be attractive already at sexual maturity. Here, we tested whether early nutritional stress affects the cheek patch size of male Zebra finches (Taeniopygia guttata), which are important in mate choice, and whether a small cheek patch size early on is compensated at sexual maturation. Furthermore we tested whether exploration behaviour is affected by such a compensation, as shown for other compensatory growth trajectories. RESULTS: Zebra finch males which were raised under poorer nutritional conditions initially expressed smaller cheek patches at day 50 post-hatching but then compensated in cheek patch size already at 65 days, i.e. when becoming sexually mature. Furthermore, compensatory growth in cheek patch during adolescence was negatively correlated with activity and exploration behaviour, measured in a novel environment. CONCLUSION: This compensation in cheek patch size benefits male attractiveness but also was related to less exploration behaviour, an established proxy for avian personality traits. We discuss the possibility that compensatory priorities exist so that not all deficits from a bad start are caught-up at the same time. Resource allocation to compensate for poorly expressed traits is likely to have evolved to optimise traits by the time they are most beneficial.

The more the better - polyandry and genetic similarity are positively linked to reproductive success in a natural population of terrestrial salamanders (Salamandra salamandra).

Although classically thought to be rare, female polyandry is widespread and may entail significant fitness benefits. If females store sperm over extended periods of time, the consequences of polyandry will depend on the pattern of sperm storage, and some of the potential benefits of polyandry can only be realized if sperm from different males is mixed. Our study aimed to determine patterns and consequences of polyandry in an amphibian species, the fire salamander, under fully natural conditions. Fire salamanders are ideal study objects, because mating, fertilization and larval deposition are temporally decoupled, females store sperm for several months, and larvae are deposited in the order of fertilization. Based on 18 microsatellite loci, we conducted paternity analysis of 24 female-offspring arrays with, in total, over 600 larvae fertilized under complete natural conditions. More than one-third of females were polyandrous and up to four males were found as sires. Our data clearly show that sperm from multiple males is mixed in the female's spermatheca. Nevertheless, paternity is biased, and the most successful male sires on average 70% of the larvae, suggesting a 'topping off' mechanism with first-male precedence. Female reproductive success increased with the number of sires, most probably because multiple mating ensured high fertilization success. In contrast, offspring number was unaffected by female condition and genetic characteristics, but surprisingly, it increased with the degree of genetic relatedness between females and their sires. Sires of polyandrous females tended to be genetically similar to each other, indicating a role for active female choice.

Exploratory Study on Individual Locomotor Activity in Local Dual-Purpose and Commercial Breeder Pullets.

Improving animal welfare is a prerequisite for the societal acceptance of poultry production. Support for improvements requires practical tools to quantify animal welfare and identify predispositions at the individual level, where possible. In this study, the activities and behavior of dual-purpose chickens (N = 245) and commercial breeders (N = 224) were analyzed. The general locomotor activity (GLA) data were collected using an RFID system over five days with 9-to-14-week-old animals. The results show that the animals of comparable age and stocking density differed from each other in their activity (p ≤ 0.001) according to breed, but no sex differences were observed (p = 0.159). No correlations were found between GLA and plumage condition (p > 0.05). The individual variations within the breeds are presented and discussed on an animal-by-animal level, providing new insights into the individual behavioral variability of chickens. The RFID systems can reliably generate GLA data that help to understand the potential interplay between behavior and animal welfare. The technology is also suitable for creating individual (personality) profiles that can be used for breeding. With a better understanding of the role of activity, husbandry and management practices can be adapted to improve animal welfare.

Olfactory kin recognition in a songbird.

The ability to recognize close relatives in order to cooperate or to avoid inbreeding is widespread across all taxa. One accepted mechanism for kin recognition in birds is associative learning of visual or acoustic cues. However, how could individuals ever learn to recognize unfamiliar kin? Here, we provide the first evidence for a novel mechanism of kin recognition in birds. Zebra finch (Taeniopygia guttata) fledglings are able to distinguish between kin and non-kin based on olfactory cues alone. Since olfactory cues are likely to be genetically based, this finding establishes a neglected mechanism of kin recognition in birds, particularly in songbirds, with potentially far-reaching consequences for both kin selection and inbreeding avoidance.

The Academic, Societal and Animal Welfare Benefits of Open Science for Animal Science.

Animal science researchers have the obligation to reduce, refine, and replace the usage of animals in research (3R principles). Adherence to these principles can be improved by transparently publishing research findings, data and protocols. Open Science (OS) can help to increase the transparency of many parts of the research process, and its implementation should thus be considered by animal science researchers as a valuable opportunity that can contribute to the adherence to these 3R-principles. With this article, we want to encourage animal science researchers to implement a diverse set of OS practices, such as Open Access publishing, preprinting, and the pre-registration of test protocols, in their workflows.

Plumage and Fat Condition Scores as Well-Being Assessment Indicators in a Small Passerine Bird, the Zebra Finch (Taeniopygia guttata).

The well-being of animals kept by humans in laboratories, farms, or as pets should always be of the highest importance, and a prerequisite for this is adequate housing. To assess the potential indicators for animal well-being, ideally simple, non-invasive, and reliable methods are necessary. Here, we propose a novel plumage scoring system for small songbirds, using the example of the Zebra Finch, and examine its reliability in comparison with a well-known body condition index, that is, the fat score. We used up to five different observers of different experience levels to assess inter- and intra-observer reliability of the proposed plumage score and also the fat score. We found substantial inter-observer reliability for the proposed novel plumage score, and lower inter-observer reliability for the fat score, which seems to require more training of observers. The intra-observer reliability of the experienced observer who trained the others also showed a very strong reliability for the plumage score and for the fat score. Thus, we conclude that our proposed novel plumage score is a simple, reliable, and non-invasive way to estimate an important indicator of captive Zebra Finches' well-being. Furthermore, the plumage score can be reliably taught to other observers. The plumage score, maybe in combination with the fat score, may be an important tool to reliably assess well-being on a regular basis in captive populations in zoos, laboratories, or pet stocks.

Productive performance, perching behavior, keel bone and other health aspects in dual-purpose compared to conventional laying hens.

Several alternatives to avoid killing male day-old chicks are available. One of these alternatives is to keep dual-purpose chicken strains. The aim of this study was to compare dual-purpose hens (Lohmann Dual, LD) with conventional laying hens (Lohmann Tradition, LT) in terms of performance, animal welfare parameters such as keel bone state and foot pad dermatitis, and perching behavior. We expected a generally equal or even better performance of the dual-purpose hens except for laying performance. Four hundred female day-old chicks were housed in 6 pens (3 pens per strain) and reared until 54 wk of age. Each pen offered a littered area, elevated slatted manure pit, elevated wooden frame with perches or grids and nest boxes on the manure pit. The wooden frame was alternately equipped with perches or grids. The elevated manure pit as well as the elevated structure was accessible via ramp. Productive performance parameters like mortality, total number of eggs and body weight were assessed periodically. In week 49, 132 hens (66 hens per strain) were randomly selected for radiography of the keel bone and assessment of plumage and foot pad state. Perching behavior was analyzed via scan sampling during rearing and laying period, respectively. Statistical analyzes were done with Linear Mixed Effect Model and General Linear Mixed Model. LD had a higher radiographic density than LT hens (P = 0.0016), other keel bone parameters (fracture score, P = 0.36; deformation, P = 0.83) showed no differences. The vast majority of fractures occurring in both strains were located in the caudal part of the keel bone. During the laying period, usage of elevated structures was higher with grids compared to perches (P < 0.001) and in LD compared to LT (P = 0.01). Some animal welfare problems were less frequent in LD compared to LT hens while other problems did not differ between the 2 strains or were even more frequent in LD hens. Grids may be more suitable as resting area than perches and may possibly help to decrease the prevalence of keel bone damage.

Brain gain-Is the cognitive performance of domestic hens affected by a functional polymorphism in the serotonin transporter gene?

The serotonin transporter (5-HTT) plays an important role in regulating serotonergic transmission via removal of serotonin (5-HT) from synaptic clefts. Alterations in 5-HTT expression and subsequent 5-HT transmission have been found to be associated with changes in behaviour, such as fearfulness or activity, in humans and other vertebrates. In humans, alterations in 5-HTT expression have been suggested to be able to lead to better learning performance, with more fearful persons being better at learning. Similar effects of the variation in the 5-HTT on fearfulness have been found in chickens, and in this study, we investigated effects on learning. Therefore, we tested 52 adult laying hens, differing in their functional 5-HTT genotype (W/W, W/D and D/D) in an operant learning paradigm in three different phases (initial learning, reversal learning and extinction) and in a tonic immobility test for fearfulness. We found that the 5-HTT polymorphism affects the initial learning performance of laying hens, with homogeneous wild-type (W/W) hens being the slowest learners, and the most fearful birds. W/W hens, showed significantly more choices to solve the initial learning task (LME, p = 0.031) and had the highest latencies in a tonic immobility test (p = 0.039), indicating the highest fearfulness. Our results provide interesting first insights into the role of 5-HTT in chickens and its sensitive interaction with the environment. We further suggest that the 5-HTT gene can be an interesting target gene for future breeding strategies as well as for further experimental studies.

Computer Vision for Detection of Body Posture and Behavior of Red Foxes.

The behavior of animals is related to their health and welfare status. The latter plays a particular role in animal experiments, where continuous monitoring is essential for animal welfare. In this study, we focus on red foxes in an experimental setting and study their behavior. Although animal behavior is a complex concept, it can be described as a combination of body posture and activity. To measure body posture and activity, video monitoring can be used as a non-invasive and cost-efficient tool. While it is possible to analyze the video data resulting from the experiment manually, this method is time consuming and costly. We therefore use computer vision to detect and track the animals over several days. The detector is based on a neural network architecture. It is trained to detect red foxes and their body postures, i.e., 'lying', 'sitting', and 'standing'. The trained algorithm has a mean average precision of 99.91%. The combination of activity and posture results in nearly continuous monitoring of animal behavior. Furthermore, the detector is suitable for real-time evaluation. In conclusion, evaluating the behavior of foxes in an experimental setting using computer vision is a powerful tool for cost-efficient real-time monitoring.

Seven steps to enhance Open Science practices in animal science.

The Open Science movement aims at ensuring accessibility, reproducibility, and transparency of research. The adoption of Open Science practices in animal science, however, is still at an early stage. To move ahead as a field, we here provide seven practical steps to embrace Open Science in animal science. We hope that this paper contributes to the shift in research practices of animal scientists towards open, reproducible, and transparent science, enabling the field to gain additional public trust and deal with future challenges to guarantee reliable research. Although the paper targets primarily animal science researchers, the steps discussed here are also applicable to other research domains.

Odour-based natal nest recognition in the zebra finch (Taeniopygia guttata), a colony-breeding songbird.

Passerine birds have an extensive repertoire of olfactory receptor genes. However, the circumstances in which passerine birds use olfactory signals are poorly understood. The aim of this study is to investigate whether olfactory cues play a role in natal nest recognition in fledged juvenile passerines. The natal nest provides fledglings with a safe place for sleeping and parental food provisioning. There is a particular demand in colony-breeding birds for fledglings to be able to identify their nests because many pairs breed close to each other. Olfactory orientation might thus be of special importance for the fledglings, because they do not have a visual representation of the nest site and its position in the colony when leaving the nest for the first time. We investigated the role of olfaction in nest recognition in zebra finches, which breed in dense colonies of up to 50 pairs. We performed odour preference tests, in which we offered zebra finch fledglings their own natal nest odour versus foreign nest odour. Zebra finch fledglings significantly preferred their own natal nest odour, indicating that fledglings of a colony breeding songbird may use olfactory cues for nest recognition.

Tasty straw pellets - Exploration of flavoured rooting material by pigs.

Tail biting is a common problem in pigs kept in conventional fully slatted pens. Suitable enrichment materials can help to prevent the occurrence of this behavioural disorder by encouraging pigs to increase exploration behaviour. We investigated whether additional flavours can increase exploration behaviour in undocked pigs. Therefore, we offered straw pellets flavoured with either fried onion (FO), strawberry (SB), ginger, almond (AL), vanilla or without flavour (control) during rearing (eight groups in total) and fattening (16 groups in total). Flavoured pellets were offered in an altering order during intervals of 1 week in material dispensers. Exploration duration at the material dispensers was continuously recorded via an ultra-high-frequency radio-frequency identification system. Pigs were weighed weekly and their tail lengths and tail injuries were scored in four categories. For analysis, changes in tail length scores compared to the previous week were calculated as Δ-tail length. The different flavours affected pigs' exploration durations in both rearing (factor flavour, P < 0.0001) and fattening (factor flavour, P < 0.0001). Highest exploration durations during rearing were recorded when straw pellets were flavoured with FO and AL compared to all others. During fattening, exploration duration was highest towards controls without significant difference to SB. Exploration durations additionally were affected by temporal effects, i.e. week and day during rearing (both factors: P < 0.0001) and fattening (both factors: P < 0.0001). During rearing, highest exploration durations were recorded in the first week and on the first day within week after changing the flavour of the straw pellets. During fattening, exploration durations varied between weeks and within weeks. The highest durations were recorded at the end of weeks, i.e. on the fifth and seventh days after material change. During rearing, tail injuries were affected by week (P < 0.0001). From the fourth week of rearing, the prevalence of tail injuries significantly increased. During the fattening period, tail injuries were affected by flavour (P < 0.05). Fattening pigs had fewest tail injuries after straw pellets flavoured with AL were offered. Altogether, based on the exploration durations, rearing pigs showed different preferences for the flavoured straw pellets but highly explored flavours differed between rearing and fattening pigs. Despite a weekly change of the flavour of straw pellets, exploration durations decreased during rearing but increased again in the transition between rearing and fattening. Thus, straw pellets with alternating flavours are a suitable possibility to provide environmental enrichment to pigs but will not prevent tail biting.

Application of YOLOv4 for Detection and Motion Monitoring of Red Foxes.

Animal activity is an indicator for its welfare and manual observation is time and cost intensive. To this end, automatic detection and monitoring of live captive animals is of major importance for assessing animal activity, and, thereby, allowing for early recognition of changes indicative for diseases and animal welfare issues. We demonstrate that machine learning methods can provide a gap-less monitoring of red foxes in an experimental lab-setting, including a classification into activity patterns. Therefore, bounding boxes are used to measure fox movements, and, thus, the activity level of the animals. We use computer vision, being a non-invasive method for the automatic monitoring of foxes. More specifically, we train the existing algorithm 'you only look once' version 4 (YOLOv4) to detect foxes, and the trained classifier is applied to video data of an experiment involving foxes. As we show, computer evaluation outperforms other evaluation methods. Application of automatic detection of foxes can be used for detecting different movement patterns. These, in turn, can be used for animal behavioral analysis and, thus, animal welfare monitoring. Once established for a specific animal species, such systems could be used for animal monitoring in real-time under experimental conditions, or other areas of animal husbandry.

Suggestions to Derive Maximum Stocking Densities for Layer Pullets.

Stocking densities for domestic chickens (Gallus gallus domesticus) are regulated by the Council Directives of the European Union for both laying hens and broiler chickens. For layer pullets no regulation of stocking density has been established yet. Based on the existing Council Directives for laying hens (1999/74/EC), broiler chickens (2007/43/EC) and calculations of the floor space that is required for the respective chicken's body, we exemplarily calculated maximum stocking densities for layer pullets. Based on the calculations we obtained absolute additional spaces for birds of different live body mass classes, i.e., useable floor space that the birds have additionally available to the space covered by their body. This allowed us to calculate the relative additional space per individual. We suggest the relative additional space to be a key parameter to derive requirements for a maximum stocking density in layer pullets. We analysed several scenarios for pullets under consideration of the Council Directives for laying hens and for broiler chickens, coming to the conclusion that layer pullets at the end of their rearing period should be provided ideally with a relative additional space of about 40-60%.

Behavioural consequences of divergent selection on general locomotor activity in chickens.

General locomotor activity is a highly variable phenotypic trait of animals. In domestic chickens it is different within and between breeds. The general locomotor activity is a substantially heritable trait and has been shown to be correlated with several other behavioural traits, such as for example feather pecking and anxiety in chickens. However, whether there is a relation between different levels of general locomotor activity and behavioural changes remained unclear. Therefore, a selection line model system has been established, where hens from the same founder population were selected over eight generations for either high or low general locomotor activity. The selection led to significant increases, respectively decreases in general locomotor activity and differences in growth. We here tested 128 hens of the 8th generation in three behavioural tests. We assumed fearfulness to be affected from selection on general locomotor activity, which we tested in a tonic immobility test. Socio-positive and socio-negative behaviours were tested in respective test paradigms. Fearfulness was higher in hens selected for high general locomotor activity. Social behavioural traits and feather pecking were not affected by selection for general locomotor activity. Evolutionary mechanisms that link fear and general locomotor activity are discussed and also why social behaviours and feather pecking seems not to be affected from selection on general locomotor activity. Our results provide interesting new insights on how selection on one trait, general locomotor activity, affects the behavioural phenotype in other dimensions too.

Fear but not social behaviour is affected by a polymorphism in the 5'-flanking region of the serotonin transporter (5-HTT) gene in adult hens.

The serotonin transporter gene (5-HTT) is involved in the regulation of the neural serotonin. Polymorphisms in the 5-HTT gene have been described in many species to be involved in physiological processes and emotions. A functional polymorphism in the 5´-flanking region of the 5-HTT gene is known from chickens, with a deletion-allele (D), which is associated with an increased 5-HTT expression, in comparison to the wild-type-allele (W). In domestic populations, the majority of hens carry the W-allele. The regulatory changes of the 5-HTT are accompanied in chickens, as in humans, by modulations of fear. Beside these effects on fear, the understanding of potential functional consequences on the social behaviour in the gregarious chicken is lacking. Thus, we here investigated whether the 5-HTT polymorphism with three genotypes (WW, WD, DD), is not only linked to fear-related behaviour, but affects also socio-positive and -negative behaviours of adult hens. Our data confirmed the effects on fear-related behaviour. WW hens showed highest levels of fear. Interestingly, no differences in the social behaviours were present between the hens of the different 5-HTT genotypes. We further discuss implications for potential evolutionary pathways via natural selection and / or artificial selection through domestication of the 5-HTT polymorphism, which might have enabled a stable social lifestyle in the wild ancestors of modern chickens.