• Wasserstein GAN based Chest X-Ray Dataset Augmentation for Deep Learning Models: COVID-19 Detection Use-Case

      Hussain, B. Zahid; Andleeb, Ifrah; Ansari, Mohammad Samar; Joshi, Amit Mahesh; Kanwal, Nadia (IEEE, 2022-07-11)
    • The effects of allogrooming and social network position on behavioural indicators of stress in female lion-tailed macaques (Macaca silenus)

      Yates, Kerrie; Stanley, Christina R.; Bettridge, Caroline; Manchester Metropolitan University; University of Chester; Bangor University (Elsevier, 2022-08-28)
      Allogrooming serves an important social function in primates and confers short term benefits such as parasite removal and stress-relief. There is currently mixed evidence as to the immediate impact of allogrooming on an individual’s stress levels, which may be influenced by their role in the grooming dyad, position in their social network, or their relationship with their grooming partner. In this study of seven captive adult female lion-tailed macaques (Macaca silenus) in a mixed sex group at Chester Zoo, UK, we found evidence to support a tension-reduction function of allogrooming. Focal animal sampling showed that the duration of self-directed behaviour (SDB), which indicates moderate to high levels of stress, was significantly lower in the five-minute period following allogrooming than the five-minute minute period preceding it for both recipients and groomers. However, when compared to match-control periods, both SDB rates and durations were significantly lower across all individuals in the five-minute period both before and after allogrooming, indicating that although allogrooming reduces stress, it is also more likely to occur when individuals are already in a relatively relaxed state. The rate and duration of SDB post-grooming did not correlate with the strength of a dyad’s bond (based on proximity). This suggests that it is the act of allogrooming itself, rather than the identity of the partner, that reduces stress for both parties. Analysis of the proximity network highlighted a clear cost to social integration; node strength, a measure of the number and strength of an individual’s direct relationships, positively correlated with the duration of self-directed behaviour, suggesting that more gregarious individuals may experience higher levels of stress. These findings add to the growing body of literature that examines the effect of the individual social environment on primate stress levels, and also highlight the need to further investigate the link between social integration and the stress experienced by group-living animals.
    • From dyads to collectives: a review of honeybee signalling

      Hasenjager, Matthew; Franks, Victoria; Leadbeater, Ellouise; University of Tennessee; University of Chester; Royal Holloway University of London (Springer, 2022-08-22)
      The societies of honeybees (Apis spp) are microcosms of divided labour where the fitness interests of individuals are so closely aligned that, in some contexts, the colony behaves as an entity in itself. Self-organization at this extraordinary level requires sophisticated communication networks, so it is not surprising that the celebrated “waggle dance”, by which bees share information about locations outside the hive, evolved here. Yet bees within the colony respond to several other lesser-known signalling systems, including the tremble dance, the stop signal and the shaking signal, whose roles in coordinating worker behaviour are not yet fully understood. Here, we firstly bring together the large but disparate historical body of work that has investigated the “meaning” of such signals for individual bees, before going on to discuss how network-based approaches can show how such signals function as a complex system to control the collective foraging effort of these remarkable social insect societies.
    • Peptide mass fingerprinting of preserved collagen in archaeological fish bones for the identification of flatfish in European waters

      Dierickx, Katrien; Presslee, Samantha; Hagan, Richard; Oueslati, Tarek; Harland, Jennifer; Hendy, Jessica; Orton, David; Alexander, Michelle; Harvey, Virginia L.; University of York; University of Lille (The Royal Society, 2022-07-27)
      Bones of Pleuronectiformes (flatfish) are often not identified to species due to the lack of diagnostic features on bones that allow adequate distinction between taxa. This hinders in-depth understanding of archaeological fish assemblages and particularly flatfish fisheries throughout history. This is especially true for the North Sea region, where several commercially significant species have been exploited for centuries, yet their archaeological remains continue to be understudied. In this research, 8 peptide biomarkers for 18 different species of Pleuronectiformes from European waters are described using MALDI-TOF MS and LC-MS/MS data obtained from modern reference specimens. Bone samples (n=202) from three archaeological sites in the UK and France dating to the medieval period (c. 7th–16th century CE) were analysed using ZooMS. Of the 201 that produced good quality spectra, 196 were identified as flatfish species, revealing a switch in targeted species through time and indicating that ZooMS offers a more reliable and informative approach for species identification than osteological methods alone. We recommend this approach for future studies of archaeological flatfish remains as the precise species uncovered from a site can tell much about the origin of the fish, where people fished and whether they traded between regions.
    • Collagen fingerprinting of Caribbean archaeological fish bones: Methodological implications for historical fisheries baselines and anthropogenic change

      Harvey, Virginia L.; LeFebvre, Michelle J.; Sharpe, Ashley E.; Toftgaard, Casper; DeFrance, Susan D.; Giovas, Christina M.; Fitzpatrick, Scott M.; Buckley, Michael; University of Manchester; University of Florida; Smithsonian Tropical Research Institute; National Museum of Denmark; Simon Fraser University; University of Oregon. (Elsevier, 2022-08-22)
      The Caribbean Sea is the most species-rich sea in the Atlantic, largely due to its vast coral reef systems. However, its high biodiversity and endemism face unprecedented anthropogenic threats, including synergistic modern pressures from overfishing, climate change and bioinvasion. Archaeological data indicate initial human settle- ment of the Caribbean ~7000 years before present (yr BP), with regionally variable human impacts on fisheries through time based on standard morphological identification of fish bone. Such studies, however, are challenged by the low taxonomic resolution of archaeological fish bone identifications due to high species diversity and morphological similarity between members of different families or genera. Here, we present collagen finger- printing (Zooarchaeology by Mass Spectrometry; ZooMS) as a method to overcome this challenge, applying it to 1000 archaeological bone specimens identified morphologically as ray-finned fish (superclass Actinopterygii) from 13 circum-Caribbean sites spanning ca. 3150–300 yr BP (years before present). The method successfully identified collagen-containing samples (n = 720) to family (21%), genus (57%), and species (13%) level. Of the 209 samples that were morphologically identified below superclass, collagen fingerprinting verified the taxo- nomic identity of 94% of these, but also refined the identifications to a lower [more precise] taxon in 45% of cases. The remaining 6% of morphological identifications were found to be incorrectly assigned. This study represents the largest application of ZooMS to archaeological fish bones to date and advances future research through the identification of up to 20 collagen biomarkers for 45 taxa in 10 families and 2 orders. The results indicate that refinement of ZooMS archaeological fish identifications in this study is limited not by the quality of the preserved collagen but by the extent of the available modern collagen reference collection. Thus, efforts should be directed towards expanding collagen fingerprint databases in the first instance. Significantly, the high- resolution taxonomic identifications of archaeological bone that ZooMS can offer make ancient fisheries data highly relevant to modern sustainability and conservation efforts in the Caribbean. Additionally, more precise identifications will allow archaeologists to address a variety of questions related to cultural fishing practices and changes in fish stocks through time. This study supports the use of ZooMS as an effective biochemical tool available for mass-taxonomic identification of archaeological fish bone samples spanning century to millennial time scales in the circum-Caribbean.
    • From dyads to collectives: a review of honeybee signalling

      Hasenjager, Matthew J.; Franks, Victoria R.; Leadbeater, Ellouise; orcid: 0000-0002-4029-7254; email: elli.leadbeater@rhul.ac.uk (Springer Berlin Heidelberg, 2022-08-22)
      Abstract: The societies of honeybees (Apis spp.) are microcosms of divided labour where the fitness interests of individuals are so closely aligned that, in some contexts, the colony behaves as an entity in itself. Self-organization at this extraordinary level requires sophisticated communication networks, so it is not surprising that the celebrated waggle dance, by which bees share information about locations outside the hive, evolved here. Yet bees within the colony respond to several other lesser-known signalling systems, including the tremble dance, the stop signal and the shaking signal, whose roles in coordinating worker behaviour are not yet fully understood. Here, we firstly bring together the large but disparate historical body of work that has investigated the “meaning” of such signals for individual bees, before going on to discuss how network-based approaches can show how such signals function as a complex system to control the collective foraging effort of these remarkable social insect societies.
    • Baseline Behavioral Data and Behavioral Correlates of Disturbance for the Lake Oku Clawed Frog (Xenopus longipes)

      Dias, Jemma E.; Ellis, Charlotte; Smith, Tessa E; Hosie, Charlotte A; Tapley, Benjamin; Michaels, Christopher J.; University of Chester; Zoological Sciety of London Outer Circle
      Animal behavior and welfare science can form the basis of zoo animal management. However, even basic behavioral data are lacking for the majority of amphibian species, and species-specific research is required to inform management. Our goal was to develop the first ethogram for the critically endangered frog Xenopus longipes through observation of a captive population of 24 frogs. The ethogram was applied to produce a diurnal activity budget and to measure the behavioral impact of a routine health check where frogs were restrained. In the activity budget, frogs spent the vast majority of time swimming, resting in small amounts of time devoted to feeding, foraging, breathing, and (in males) amplexus. Using linear mixed models, we found no effect of time of day or sex on baseline behavior, other than for breathing, which had a greater duration in females. Linear mixed models indicated significant effects of the health check on duration of swimming, resting, foraging, feeding, and breathing behaviors for all frogs. This indicates a welfare trade-off associated with veterinary monitoring and highlights the importance of non-invasive monitoring where possible, as well as providing candidates for behavioral monitoring of acute stress. This investigation has provided the first behavioral data for this species which can be applied to future research regarding husbandry and management practices.
    • Long term analysis of social structure: evidence of age-based consistent associations in male Alpine ibex

      Brambilla, Alice; von Hardenberg, Achaz; Canedoli, Claudia; Brivio, Francesca; Sueur, Cédric; Stanley, Christina R.; University of Zurich; University of Chester; University of Milano Bicocca; University of Sassari; University de Strasbourg; Institut Universitaire de France (Wiley, 2022-06-28)
      Despite its recognized importance for understanding the evolution of animal sociality as well as for conservation, long term analysis of social networks of animal populations is still relatively uncommon. We investigated social network dynamics in males of a gregarious mountain ungulate (Alpine ibex, Capra ibex) over ten years focusing on groups, sub-groups and individuals, exploring the dynamics of sociality over different scales. Despite the social structure changing between seasons, the Alpine ibex population was highly cohesive: fission–fusion dynamics lead almost every male in the population to associate with each other male at least once. Nevertheless, we found that male Alpine ibex showed preferential associations that were maintained across seasons and years. Age seemed to be the most important factor driving preferential associations while other characteristics, such as social status, appeared less crucial. We also found that centrality measures were influenced by age and were also related to individual physical condition. The multi-scale and long-term frame of our study helped us show that ecological constrains, such as resource availability, may play a role in shaping associations in a gregarious species, but they cannot solely explain sociality and preferential association that are likely also to be driven by life-history linked physiological and social needs. Our results highlight the importance of long-term studies based on individually recognizable subjects to help us build on our understanding of the evolution of animal sociality.
    • Pre- and peri-operative clinical information, physiological observations and outcome measures following flexible ureterorenoscopy (FURS), for the treatment of kidney stones. A single-centre observational clinical pilot-study in 51 patients

      Hughes, Stephen Fôn; orcid: 0000-0001-6558-9037; email: Stephen.hughes6@wales.nhs.uk; Moyes, Alyson Jayne; Jones, Kevin; Bell, Christopher; Duckett, Abigail; Moussa, Ahmed; Shergill, Iqbal (BioMed Central, 2022-07-14)
      Abstract: Background: Kidney stone disease contributes to a significant proportion of routine urological practice and remains a common cause of worldwide morbidity. The main aim of this clinical-pilot study was to investigate the effect of flexible ureterorenoscopy (FURS) on pre- and peri-operative clinical information, physiological observations and outcome measures. Methods: Included were 51 patients (31 males, 20 females), who underwent elective FURS, for the treatment of kidney stones. Pre-operative and peri-operative clinical information, and post-operative physiological observations and outcome measures were collected using a standard case report form. Pre-operative clinical information included age, gender, BMI, previous history of stone formation and hypertension. Pre-operative stone information included the size (mm), Hounsfield units (HU), laterality and intra-renal anatomical location. Peri-operative surgical details included surgical time in minutes; Laser use; Duration and energy of laser; and post-operative stenting. The physiological outcomes measured included systolic and diastolic blood pressure (mmHg), Likert pain score, temperature, heart rate (bpm) and respiration rate (bpm). Following initial descriptive analysis, a series of Pearson’s correlation coefficient tests were performed to investigate the relationship between surgical factors other variable factors. Results: A series of significant, positive correlations were observed between; age and surgical time (p = 0.014, r = 0.373); stone size and Hounsfield unit (p = 0.029, r = 0.406); surgical time and duration of laser (p < 0.001, r = 0.702); surgical time and BMI (p = 0.035, r = 0.322); baseline heart rate and Hounsfield unit (p = 0.026, r = − 0.414); base line heart rate and BMI (p = 0.030, r = 0.307).; heart rate at 120-min post FURS and age (p = 0.038, r = − 0.308); baseline pain score and BMI (p = 0.010, r = 0.361); baseline respiration rate and BMI (p = 0.037, r = 0.296); respiration rate at 240-min post FURS and BMI (p = 0.038, r = 0.329); respiration rate at 120 min post FURS and age (p = 0.022, r = − 0.330). Four patients developed post-operative complications (3—UTIs with urinary retention, 1–urosepsis). Conclusions: We report that following FURS there is an association between various physiological, clinical and surgical parameters. Although these correlations are weak, they warrant further investigation as these may be linked with untoward complications, such as infection that can occur following FURS. This data, however, will need to be validated and reproduced in larger multi-centre studies.
    • Social roles influence cortisol levels in captive Livingstone's fruit bats (Pteropus livingstonii)

      Edwards, Morgan J.; Stanley, Christina R.; Hosie, Charlotte A.; Richdon, Sarah; Price, Eluned; Wormell, Dominic; Smith, Tessa E.; University of Chester; Bristol Zoological Society; Durrell Wildlife Conservation Trust (Elsevier, 2022-06-27)
      A critical component of conserving and housing species ex situ is an explicit scientific understanding of the physiological underpinnings of their welfare. Cortisol has been repeatedly linked to stress, and therefore used as an indicator of welfare for many species. In order to measure cortisol in the Livingstone's fruit bat (Pteropus livingstonii; a critically endangered keystone species) without disturbing the captive population, we have developed and validated a non-invasive, novel hormone extraction procedure and faecal glucocorticoid assay. A total of 92 faecal samples, 73 from the P. livingstonii breeding colony at Jersey Zoo, Channel Islands and 19 samples from P. livingstonii housed at Bristol Zoological Gardens, UK, have been collected and analyzed. Mixed-effect modelling of the influence of physiological state variables on cortisol concentration revealed that lactating females had higher cortisol levels than non-lactating females, indicating that our assay is measuring biologically relevant hormone concentrations. Males and older bats also had higher cortisol than non-lactating females and younger individuals. Further analysis applied social network methodology to compare the cortisol levels of bats with different social roles. We found that individuals that linked social groups possessed higher than average cortisol levels and conversely, individuals with high-quality, positive relationships had lower cortisol levels. These results demonstrate, for the first time in a bat species, social mediation of stress hormones. Lastly, the frequency of vocalisation was found to positively correlate with cortisol concentration in males, suggesting that this behaviour may be used by animal management as a visual indicator of a bat's hormonal status. Hence, this research has provided unique insights and empirical scientific knowledge regarding the relationship between the physiology and social behaviour of P. livingstonii, therefore allowing for recommendations to be made to optimise bat welfare at the individual level.
    • Phylogenetic placement and life history trait imputation for Grenada Dove Leptotila wellsi

      Peters, Catherine; Geary, Matthew; Nelson, Howard; Rusk, Bonnie; von Hardenberg, Achaz; Muir, Anna P.; University of Chester; University of Cambridge; Grenada Dove Conservation Programme (Cambridge University Press, 2022-08-05)
      Phylogenetic analyses can be used to resolve taxonomic uncertainties and reconstruct a species’ evolutionary history. This can be combined with ecological data to predict missing life history traits which are important for creation of conservation management strategies. We investigated the evolutionary and life history of the Critically Endangered Grenada Dove Leptotila wellsi by estimating its phylogenetic placement and using this new phylogeny to test the accuracy of phylogenetic comparative methods for estimating both documented and unknown life history traits. We extracted DNA from two Grenada Dove samples and obtained sequences from three mitochondrial markers: Cytochrome oxidase I (COI), NADH dehydrogenase 2 (ND2) and Cytochrome b (Cyt b); and one nuclear marker: β-Fibrinogen intron 7 (β-FIB). We present the first genetic data obtained for the Grenada Dove. Our data identifies the Grey-Chested Dove Leptotila cassinii as the species which shares both a most recent common ancestor, with an estimated divergence of approximately 2.53 million years ago, and the smallest genetic distance (p=0.0303) with the Grenada dove. Life history trait values for the Grenada Dove predicted from our analyses using phylogenetic imputation are: clutch size=2 (±0.09) eggs, clutches per year=1.4 (±0.81), incubation time=14.2 (±0.75) days, hatching weight=3.8 (±1.05) grams and single imputation: fledging age (genus median) =15.5 days, longevity (genus median) =8.6 years. This study contributes novel information regarding evolutionary history and life history characteristics to inform long-term conservation actions for a Critically Endangered species.
    • New haplotypes found in stranded long-finned pilot whales (Globicephala melas) in the eastern North Atlantic and adjacent waters

      Ball, Rachel J.; Kitchiner, Ashleigh; Davison, Nicholas J.; Berrow, Simon; McKeown, Niall J.; IJsseldijk, Lonneke L.; Geary, Matthew; McDowall, Ian; Muir, Anna P.; University of Chester; University of Glasgow; Scottish Marine Animal Stranding Scheme; Irish Whale and Dolphin Group; Galway-Mayo Institute of Technology; Aberystwyth University; Utrecht University (Wiley, 2021-12-17)
      Long-finned pilot whale (Globicephala melas) mitochondrial (mtDNA) genetic diversity is considered low, especially in the North Atlantic, where only six haplotypes have been recorded using a 345bp portion of the control region. Previous studies have been based on a small number of samples and have not included samples from Ireland or the Netherlands. We utilized mtDNA control region sequencing of individuals stranded around Ireland, Scotland, and the Netherlands between 1995-2019 (n=180) to assess regional diversity. Following alignment of 631bp, nine haplotypes were identified, of which five were newly described (haplotype diversity (h)=0.511). Pairwise tests revealed significant differentiation between the Irish and Scottish samples. Potential confounding factors are discussed but, given the dangers of failing to recognize population structure this needs to be investigated further using nuclear markers and individual-based approaches before population isolation is assumed. The analyzed samples included six mass stranding events. One of the mass strandings reported two haplotypes, confirming mixing of matrilines within a North Atlantic stranding for the first time. This study shows that stranding sample databases are a useful tool for assessment of genetic diversity and provides new insights into genetic diversity of long-finned pilot whale haplotypes in the eastern North Atlantic and adjacent waters.
    • Anthropogenic influences on habitat use by African Houbaras Chlamydotis undulata on Lanzarote, Canary Islands

      Geary, Matthew; Cooper, Joseph R.; Collar, Nigel J.; University of Chester; BirdLife International (Elsevier, 2022-06-27)
      African Houbara Chlamydotis undulata is threatened in North Africa by unsustainable hunting and massive overuse of captive-bred birds to replace wild losses. A small population on the Canary Islands is protected from these threats, but the archipelago is economically dependent on tourism which has led to extensive land-use change, particularly close to the coasts. We investigated the drivers of houbara distribution and abundance in and around the large semi-desert El Jable region of northern Lanzarote in order to identify potential measures to conserve this important population. All houbaras seen during point counts in the centre of 30 tetrads (2 km ×2 km) were recorded, along with their location. We used negative binomial regression to evaluate the effects of land use and human activity on the abundance of birds at tetrad scale. At finer scale we used logistic regression to assess the effect of land use on the distribution of displaying males. We recorded 196 houbara sightings on our surveys, although only 10 males were observed displaying. Houbara abundance had a quadratic relationship with the proportion of huerta (agricultural gardens) in a tetrad. The distribution of male displays was positively related to the proportion of long-abandoned farmland within a 100 m radius of their display site. African Houbaras favour the vicinity of small-scale agriculture and abandoned farmland, but avoid areas with higher levels of human land-use. Reduction of extensive land-use change and disturbance in El Jable are key conservation measures.
    • Targeting burrows improves detection in giant pangolin Smutsia gigantea camera trap surveys

      Matthews, Naomi; Nixon, Stuart; von Hardenberg, Achaz; Geary, Matthew; University of Chester; North of England Zoological Society; Chester Zoo
      The Endangered giant pangolin (Smutsia gigantea) is rare and elusive across its central African range. Due to its solitary and nocturnal nature, the species is difficult to study and subsequently its ecology is poorly known. Pangolins are considered the World’s most-trafficked mammals. Therefore, accurately confirming presence and monitoring trends in distribution and abundance is essential to inform and prioritise conservation efforts. Camera traps are a popular tool for surveying rare and cryptic species. However non-targeted camera trap surveys yield low camera trapping rates for pangolins. Here we use camera trap data from surveys conducted within three protected areas in Uganda to test whether targeted placement of cameras improves giant pangolin detection probability in occupancy models. The results indicate that giant pangolin detection probability is highest when camera traps are targeted on burrows. The median number of days from camera deployment to first giant pangolin event was 12, with 97.5% of events captured within 32 days from deployment. The median interval between giant pangolin events at a camera trap site was 33 days. We demonstrate that camera trap surveys can be designed to improve detection of giant pangolins and outline a set of recommendations to maximise the effectiveness of efforts to survey and monitor the species.
    • Zooarchaeology through the lens of collagen fingerprinting at Denisova Cave

      Brown, Samantha; Wang, Naihui; Oertle, Annette; Kozlikin, Maxim B.; Shunkov, Michael V.; Derevianko, Anatoly P.; Comeskey, Daniel; Jope-Street, Blair; Harvey, Virginia L.; Chowdhury, Manasij Pal; et al. (Nature Research, 2021-07-29)
      Denisova Cave, a Pleistocene site in the Altai Mountains of Russian Siberia, has yielded significant fossil and lithic evidence for the Pleistocene in Northern Asia. Abundant animal and human bones have been discovered at the site, however, these tend to be highly fragmented, necessitating new approaches to identifying important hominin and faunal fossils. Here we report the results for 8253 bone fragments using ZooMS. Through the integration of this new ZooMS-based data with the previously published macroscopically-identified fauna we aim to create a holistic picture of the zooarchaeological record of the site. We identify trends associated with climate variability throughout the Middle and Upper Pleistocene as well as patterns explaining the process of bone fragmentation. Where morphological analysis of bones from the site have identified a high proportion of carnivore bones (30.2%), we find that these account for only 7.6% of the ZooMS assemblage, with large mammals between 3 and 5 more abundant overall. Our analysis suggests a cyclical pattern in fragmentation of bones which sees initial fragmentation by hominins using percussive tools and secondary carnivore action, such as gnawing and digestion, likely furthering the initial human-induced fragmentation.
    • Medieval fish remains on the Newport ship identified by ZooMS collagen peptide mass fingerprinting

      Buckley, Michael; Harvey, Virginia L.; Pettifer, David; Russ, Hannah; Wouters, Wim; Van Neer, Wim; University of Manchester; University of Birmingham; University of Wales Trinity Saint David; Royal Belgian Institute of Natural Sciences (Springer, 2022-02-11)
      Fish represent a key economic, social and ecological group of species that humans have exploited for tens of thousands of years. However, as many fish stocks are going into decline and with little known about the anthropogenic impacts on the health of the marine ecosystem pre-Industrial Revolution, understanding historical and archaeological exploitation of fish species is key to accurately modelling these changes. Here, we explore the potential of collagen peptide mass fingerprinting (also known as Zooarchaeology by Mass Spectrometry, or ZooMS) for identifying fish remains from the Medieval (fifteenth century) Newport ship wreck (Wales, UK), and in doing so we establish a set of biomarkers we consider useful in discriminating between European fish taxa through the inclusion of over 50 reference taxa. The archaeological results identified nine distinct taxonomic groups, dominated by ling (> 40%), and a substantial amount of cod (> 20%) and hake (~ 20%). The vast majority of samples (> 70%) were identified to species level, and the inability to identify the remaining taxonomic groups with confidence using ZooMS was due to the fact that the reference collection, despite being relatively large in comparison to those presented in mammalian studies, reflects only a small proportion of fish biodiversity from this region. Although the results clearly demonstrate the potential for ZooMS as a means of fish bone identification, the sheer number of different fish species that potentially make up ichthyoarchaeological assemblages leads to obvious requirements for the analysis on much greater numbers of modern reference specimens, or the acquisition of collagen sequences.
    • Extrinsic and intrinsic factors affecting the activity budget of alpine marmots ( Marmota marmota )

      Ferrari, Caterina; Pasquaretta, Cristian; Caprio, Enrico; Ranghetti, Luigi; Ranghetti, Luigi; Rolando, Antonio; Bertolino, Sandro; Bassano, Bruno; von Hardenberg, Achaz (Springer Berlin Heidelberg, 2022-03-15)
      Abstract: Extrinsic and intrinsic factors may influence the activity budget of wild animals, resulting in a variation in the time spent in different activities among populations or individuals of the same species. In this study, we examined how extrinsic and intrinsic factors affect the behaviour of the alpine marmot (Marmota marmota), a hibernating social rodent inhabiting high-elevation prairies in the European Alps. We collected behavioural observations during scan sampling sessions on marked individuals at two study sites with different environmental characteristics. We used Bayesian hierarchical multinomial regression models to analyse the influence of both intrinsic (sex and age-dominance status) and extrinsic (environmental and climatic variables) factors on the above-ground activity budget. Marmots spent most of their time above ground foraging, and were more likely to forage when it was cloudy. Extrinsic factors such as the site, period of the season (June, July–August, and August–September), and time of the day were all related to the probability of engaging in vigilance behaviour, which reaches its peak in early morning and late afternoon and during July, the second period included in the study. Social behaviours, such as affiliative and agonistic behaviours, were associated mostly with sex and age-dominance status, and yearlings were the more affiliative individuals compared to other status. Overall, our results suggest that in alpine marmots, intrinsic factors mostly regulate agonistic and affiliative behaviours, while extrinsic factors, with the unexpected exception of temperature, affect the probabilities of engaging in all types of behavioural categories.
    • Phylogenetic analyses of ray-finned fishes (Actinopterygii) using collagen type I protein sequences

      Harvey, Virginia L.; Keating, Joseph N.; Buckley, Michael; University of Manchester; University of Bristol (The Royal Society, 2021-08-11)
      Ray-finned fishes (Actinopterygii) are the largest and most diverse group of vertebrates, comprising over half of all living vertebrate species. Phylogenetic relationships between ray-finned fishes have historically pivoted on the study of morphology, which has notoriously failed to resolve higher order relationships, such as within the percomorphs. More recently, comprehensive genomic analyses have provided further resolution of actinopterygian phylogeny, including higher order relationships. Such analyses are rightfully regarded as the ‘gold standard’ for phylogenetics. However, DNA retrieval requires modern or well-preserved tissue and is less likely to be preserved in archaeological or fossil specimens. By contrast, some proteins, such as collagen, are phylogenetically informative and can survive into deep time. Here, we test the utility of collagen type I amino acid sequences for phylogenetic estimation of ray-finned fishes. We estimate topology using Bayesian approaches and compare the congruence of our estimated trees with published genomic phylogenies. Furthermore, we apply a Bayesian molecular clock approach and compare estimated divergence dates with previously published genomic clock analyses. Our collagen-derived trees exhibit 77% of node positions as congruent with recent genomic-derived trees, with the majority of discrepancies occurring in higher order node positions, almost exclusively within the Percomorpha. Our molecular clock trees present divergence times that are fairly comparable with genomic-based phylogenetic analyses. We estimate the mean node age of Actinopteri at ∼293 million years (Ma), the base of Teleostei at ∼211 Ma and the radiation of percomorphs beginning at ∼141 Ma (∼350 Ma, ∼250–283 Ma and ∼120–133 Ma in genomic trees, respectively). Finally, we show that the average rate of collagen (I) sequence evolution is 0.9 amino acid substitutions for every million years of divergence, with the α3 (I) sequence evolving the fastest, followed by the α2 (I) chain. This is the quickest rate known for any vertebrate group. We demonstrate that phylogenetic analyses using collagen type I amino acid sequences generate tangible signals for actinopterygians that are highly congruent with recent genomic-level studies. However, there is limited congruence within percomorphs, perhaps due to clade-specific functional constraints acting upon collagen sequences. Our results provide important insights for future phylogenetic analyses incorporating extinct actinopterygian species via collagen (I) sequencing.
    • Proteome Variation with Collagen Yield in Ancient Bone

      Harvey, Virginia L.; Procopio, Noemi; Hopkins, Rachel J. A.; Buckley, Michael; Northumbria University; University of Manchester; University of New Mexico (American Chemical Society, 2021-02-02)
      Isotope analyses are some of the most common analytical methods applied to ancient bone, aiding the interpretation of past diets and chronology. For this, the evaluation of “collagen yield” (as defined in radiocarbon dating and stable isotope research) is a routine step that allows for the selection of specimens that are deemed adequate for subsequent analyses, with samples containing less than ∼1% “collagen yield” normally being used for isotopic analysis but discounted for radiocarbon dating. The aims of this study were to use proteomic methods of MALDI–TOF (matrix assisted laser desorption ionization time-of-flight mass spectrometry) and LC−ESI−MS/MS (liquid chromatography electrospray ionization tandem mass spectrometry) to investigate the endogeneity of the dominant proteinaceous biomolecules within samples that are typically considered to contain poorly preserved protein. Taking 29 archaeological samples, we evaluated the proteome variability between different acid-soluble fractions removed prior to protein gelatinization and considered waste as part of the radiocarbon dating process. We then correlated these proteomes against the commonly used “collagen yield” proxy for preservation. We found that these waste fractions contained a significant amount of both collagenous and noncollagenous proteins (NCPs) but that the abundance of these was not correlated with the acquired “collagen yield”. Rather than a depleted protein load as would be expected from a low “collagen yield”, the variety of the extracted NCPs was comparable with that commonly obtained from ancient samples and included informative proteins useful for species identification, phylogenetic studies, and potentially even for isotopic analyses, given further method developments. Additionally, we did not observe any correlation between “collagen yield” and peptide mass fingerprint success or between the different fractions taken from the same sample but at different radiocarbon pretreatment stages. Overall, these findings highlight the value in retaining and analyzing sample fractions that are otherwise discarded as waste during the radiocarbon dating process but more importantly, that low “collagen yield” specimens that are often misinterpreted by archaeologists as being devoid of protein can still yield useful molecular sequence-based information.
    • Experimental taphonomy of fish bone from warm and cold water species: Testing the effects of amino acid composition on collagen breakdown in modern fish bone using thermal maturation experiments

      Harvey, Virginia L.; Wogelius, Roy A.; Manning, Phillip L.; Buckley, Michael; University of Manchester (Elsevier, 2021-01-12)
      Decay experiments have the potential to provide useful analogues in the interpretation of archaeological remains. Previous studies have focused on how physical properties or processing methods can influence fish bone distributions within archaeological sites. However, the means by which intrinsic chemical properties of fish bone, such as baseline collagen type I [‘collagen (I)’] chemistry, may affect both biomolecule and whole bone degradation has not been the focus of any prior study. The variation of facies and resulting impact on taphonomy is not a new concept, but an understanding of the discrete relationship between temperature and the breakdown of collagen (I) in bone material has not been well explored. Here, modern fish bone powder is subjected to differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and water-immersed heat experiments. This is to test whether taxa with less thermally stable collagen (I) configurations, such as cold-water species with reduced proline and hydroxyproline concentrations (Pro+Hyp), will generate collagen breakdown products (TOC and TN) more rapidly than those with more thermally stable arrangements, such as warm-water fish with typically increased Pro+Hyp. Our results show that bone collagen (I) in the cold-water fish species in this study (cod and herring with lower Pro+Hyp concentrations) display significantly increased decomposition rates than collagen (I) from the warm-water fishes in this study (amberjack and tilapia with higher Pro+Hyp concentrations), given the same experimental conditions (heating in water at 75 °C for up to eight days). Initial reaction rate estimates, based on TOC and TN product concentrations, suggest that cod bone (15.6% Pro+Hyp) reacts ~9 times faster than tilapia bone (20.3% Pro+Hyp). We suggest that the primary influencer of collagen (I) stability in bone is the concentration of Pro+Hyp residues and not a function of physical bone structure. Our results suggest that a reduction in collagen (I) stability is likely to lead to a decrease in whole bone stability following deposition, due to the intimate association between organic and inorganic phases of bone. Therefore, species composition based upon bone remains may vary in archaeological and palaeontological sites, as a function of the thermal stability of collagen (I).