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MDPI AG

Preprints posted in the last 90 days, ranked by how well they match Animals's content profile, based on 20 papers previously published here. The average preprint has a 0.04% match score for this journal, so anything above that is already an above-average fit.

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Effects of periodic group housing opportunities on reproductive performance and welfare in sows

Shimasaki, T.; Yoyou, K.; Kojima, T.; Huang, C.-Y.; Kato, H.; Ishida, M.; Takeda, K.

2026-05-21 animal behavior and cognition 10.64898/2026.05.19.726187 medRxiv
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ObjectiveStall housing of pregnant sows raises welfare concerns, whereas conventional group housing systems often reduce space efficiency. This study evaluated the effects of periodic group housing (PG) on reproductive performance and welfare compared with continuous stall housing (CS). MethodsSows in the CS group (n = 15) were continuously housed in stalls. In the PG group (n = 15), sows were housed in groups of three and allocated 1 day of group housing and 6 days of stall housing per week over 10 weeks. During group housing sessions, the sows had access to a group housing area containing sawdust. Behavioral observations and salivary cortisol measurements were conducted on the first day of the stall housing session in weeks 1, 6, and 10. Behavioral indices were expressed as proportions based on 90 sampling points recorded at 1-min intervals. ResultsThe number of stillbirths was significantly lower in the PG group than in the CS group (0.63 vs. 1.49 piglets per litter). whereas other reproductive outcomes, including total litter size and average birth weight, did not differ. In older parity sows, the PG treatment markedly increased the proportion of time spent lying, suggesting reduced discomfort associated with restricted movement. Furthermore, the proportion of exploratory behavior decreased markedly, and drinking behavior showed a decreasing trend across parity levels in the PG group, suggesting partial satisfaction of motivations for environmental exploration and oral manipulation. The proportion of oral abnormal behavior showed a pronounced interaction between housing treatment and experimental week, increasing from week 1 to week 6 in the PG group. Salivary cortisol concentrations did not differ between the groups. ConclusionPG may improve reproductive performance and partially satisfy the behavioral motivations restricted under continuous stall housing. This system may represent a practical alternative for improving animal welfare while minimizing economic losses.

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Morphometric Characterization and Variation of Somali Dromedary Camels (Camelus dromedarius) in a selected district of Benadir Region

Barre, A.

2026-04-15 animal behavior and cognition 10.64898/2026.04.12.718019 medRxiv
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Camels are one of the hardy animal species known for their high adaptability to extreme climatic conditions. However, scientific studies on this important animal species are limited. This study aims to investigate the statistical relationships between some body measurements of dromedary camels with different characteristics raised in the Mogadishu, Somalia. The research was carried out on a total of 248 camels (102 males, 146 females) of different age groups, belonging to the SiifDacar, Hoor, and Eyddimo types, raised in camel farms in Dharkenley, Dayniile, and Hodan districts of Mogadishu. Body measurements were taken manually using a standard measuring tape, and the collected data were subjected to statistical analysis. As a result of the analyses, significant differences were found between the camels named with the local language as mentioned above in terms of withers height, rump height, body length, and shoulder width in female camels. In male camels, it was determined that there was a statistically significant difference among the camels named in terms of rump width. When evaluated specifically for female camels, it was found that the camels of the SiifDacar had higher withers and rump height compared to the camels of the Hoor, and were also longer than the other two different camels. Although no notable difference in shoulder width was found between the Hoor and Eyddimo camels, Siif Dacar females exhibited broader shoulders than both. In male camels, Siif Dacar had a significantly wider rump compared to those of the Hoor camels. Furthermore, the study identified varying degrees of statistically significant correlations among different body measurements in the camels. Overall, it was observed that female camels of the Siif Dacar were larger in size compared to those of the Hoor and Eyddimo camels, and that male camels generally exhibited greater body dimensions than females. These findings contribute to the registration of the different-named camel types in Somalia as separate breeds and to a better understanding of the morphological characteristics of camels.

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Transvaginal ovum retrieval in scimitar horned oryx (Oryx dammah) and roan antelope (Hippotragus equinus)

Pennington, P. M.; Gillis, J. D.; Tourzani, D. A.; Lambert, C. J.; Nguyen, T. Q.; Metzler, S.; Citino, S. B.; James, M.; Penfold, L. M.; Herrick, J. R.

2026-05-05 physiology 10.64898/2026.04.30.721932 medRxiv
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Development and use of assisted reproductive technologies (ARTs) in non-domestic species provides novel tools for species conservation. As a first step towards in vitro embryo production, we developed an OPU technique for two antelope species, scimitar horned oryx (Oryx dammah) and roan antelope (Hippotragus equinus) utilizing a custom-made needle guide and existing OPU equipment utilized by livestock and human practitioners. Females were anesthetized and placed in sternal recumbency for transvaginal OPUs. Prior to OPUs (36 - 45 hours), SHO and roan were either hormonally stimulated with follicle stimulating hormone (FSH, 140 or 250IU) as a single injection or not. A total of 32 and 26 OPUs were completed in SHO (n=10) and roan (n=7), respectively, representing one to four OPUs per animal at monthly intervals. A total of 141 oocytes were recovered from 215 follicles in SHO and 31 oocytes from 58 follicles in roan. FSH dose (250IU) increased (P<0.05) the number of follicles aspirated and the number of oocytes recovered in SHO. No effects of FSH were observed in roan (P>0.05). Good quality oocytes were recovered from all females and procedures were conducted in four consecutive months with no evidence of scar tissue buildup or reduced capacity to recover quality oocytes. These ARTs can be used to develop in vitro embryo production tools for population management and the preservation of female genetics; bolstering genetic diversity and guarding against extinction.

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Pathologies and causes of death in stranded cetaceans in the Canary Islands (2013-2018)

Diaz Santana, P. J.; Arbelo, M.; Diaz-Delgado, J.; Groch, K.; Suarez-Santana, C.; Consoli, F.; Bernaldo de Quiros, Y.; Quesada-Canales, O.; Sierra, E.; Fernandez, A.

2026-04-05 pathology 10.64898/2026.04.01.715953 medRxiv
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Cetacean pathology is a cornerstone for population and marine ecosystem health monitoring, allowing clear differentiation among natural and anthropogenic threats. Previous studies in the Canary Islands reported natural causes of death in 59.4% (1999-2005) and 81% (2006-2012) of stranded cetaceans, versus anthropogenic causes in 33.3% and 19%, respectively. This study aimed to determine the causes of death (CD), pathologic findings, and epidemiological patterns of 316 cetaceans stranded in the Canary Islands between 2013 and 2018. The CDs were classified in pathologic entities (PEs) emphasizing natural versus anthropic origins. Of 316 animals, 224 (70.9%) from 18 species were suitable for pathological investigations. Among natural PEE, natural pathology associated with good nutritional status (NP-GNS) and natural pathology associated with significant loss of nutritional status (NP-LNS) represented 43/224 (19.2%) and 36/224 (16%) cases, respectively. Natural pathology with undetermined nutritional status (NP-UNS) occurred in 19/224 (8.5%) animals. Intra- and interspecific traumatic interactions (ITI) represented 30/224 (13.4%) cases, followed by neonatal/perinatal pathology (NPN) 19/224 (8.5%) and live-stranding stress and/or capture myopathy (LS-CM) 18/224 (8%). Infectious and parasitic diseases predominated in natural PEs. Anthropogenic PEs included interaction with fishing activities (IFA) in 17/224 (7.6%) cases, vessel collisions (VC) in 9/22 (4%) cases, and foreign body-associated pathology (FBAP) in 3/224 (1.3%) animals. Overall, anthropogenic causes accounted for 12.9% of deaths, natural causes for 73.6%, and the CD could not be established in 30/194 (13.4%) cases. This study reaffirms the trends concerning recognized PEs (NP-GNS, NP-LNS, NP-UNS, ITI, NPN, LS-CM, IFA, VC, and FBAP), expands the body of knowledge on cetacean pathology in the Canary Islands, and reports novel findings including mixed infections, clostridiosis in uncommon species, uremic syndrome secondary to urethral nematodiasis, gas embolism in unusual species, epibiont stomatitis, congenital musculo-skeletal malformations, or neoplastic processes. These findings advance understanding of cetacean mortality patterns and support conservation and management strategies.

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Statistical invisibility of working equids in Mexico: Dissecting the gap between global diagnostics and official data (1970-2022).

Garcia-Seco, E.; Diaz, M. A.; Tadich Gallo, T.; Toribio, R. E.; Galindo Maldonado, F.; Hernandez-Gil, M.

2026-04-17 scientific communication and education 10.64898/2026.04.15.718791 medRxiv
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BackgroundWorking equids are fundamental to the socioeconomic structure of Mexicos small-scale agricultural sector, which accounts for 71.2% of the countrys active Agricultural Production Units (APUs). Despite their critical role in human rural livelihoods, food security, and sustainable development, these animals face systemic "statistical invisibility" within national and international productive frameworks. This study evaluates the long-term population dynamics and geographical distribution of working equids to analyze their current status amidst agricultural modernization. MethodsA retrospective analysis was conducted using national census data from 1970 to 2022 provided by the National Institute of Statistics, Geography, and Informatics (INEGI). Population trends for horses, donkeys, and mules were calculated using the Average Annual Variation Rate (AAVR). The severity of population declines was classified according to an adaptation of the International Union for Conservation of Nature (IUCN) criteria. Finally, national census records from INEGI, Agri-food and Fisheries Information Service (SIAP) and The Ministry of Agriculture and Rural Development (SADER) were contrasted with FAOSTAT database estimates to identify reporting discrepancies. ResultsBetween 1970 and 2022, the total equine population in Mexico decreased by 76.5%, falling from 6.8 to 1.6 million. However, a "paradox of modernization" was identified: while total numbers plummeted, the proportion of equids used specifically for work reached a historical peak of 81% in 2022, effectively having doubled from the 44% recorded in 2007. While donkeys and mules have suffered drastic total reductions (87% and 88%, respectively), working horses experienced a resilient 37% recovery between 2007 and 2022 (+3.71% AAVR). Furthermore, a staggering 710.8% discrepancy was found between national census data and FAOSTAT estimates, representing an overestimation of 11.3 million animals in international records. ConclusionsThe persistence and recent recovery of working equids reflect a "resilience of necessity" for approximately 500,000 APUs that depend exclusively on animal traction and packing due to economic constraints and complex topography. These findings challenge the narrative of total agricultural mechanization and highlight an urgent need for evidence-based public policies that address the statistical invisibility of working equids as indispensable drivers of rural sustainability and food security.

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Investigating cognitive enrichment for dairy calves through behavioral measures of participation and engagement: a pilot study

Amarioarei, G.; Cellier, M.; Aigueperse, N.; Wolfe, T.; Shepley, E.; Diallo, A. B.; Vasseur, E.

2026-04-04 zoology 10.64898/2026.04.01.715895 medRxiv
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Introducing cognitive enrichment from an early age has the potential to enhance an animals capacity to learn both simple and complex tasks, promote neural plasticity, and support cognitive development. This is applicable for young cattle who are at a critical stage in their development and could benefit from the influence cognitive enrichment has on their behavioral expression. This study aims to explore the effects cognitive enrichment has on weaned dairy calves through analyzing behavioral measures of voluntary participation and short-term behavioral reactions to enrichment exposure. Our study involved a total of five pairs of weaned calves (n=8 treatment; n=2 control). The treatment groups were presented with three variations of a puzzle box, each equipped with unique challenges that offer different solutions (push, slide, pull). These boxes were provided to the calves twice daily over the span of nine days in an isolated corridor located behind their pen. We hypothesized that motivated calves would consistently engage with cognitive enrichment voluntarily over time and express directed natural behaviors, reflecting sustained participation across repeated trials. Results demonstrated that calves consistently visited the cognitive enrichment area across trials, with an average latency of 75.7 {+/-} 47.0s from the pen to the enrichment. Secondly, the calves spent a significant proportion of trial time within the enrichment area at 65% (870.1 {+/-} 21s). Lastly, all calves expressed a broad range of behaviors in line with their natural exploration within the enrichment area, while the puzzle box treatment groups expressed higher durations of behavioral expressions when compared to the control (F=11.7, p<0.0001). Combined, these results indicate the calves motivations to voluntarily participate in a cognitive challenge. While these are promising findings for cognitive enrichment and its applicability to dairy calves, further work is needed to understand broader parameters. Specifically, how can social dynamics influence enrichment interaction in groups, how can this type of enrichment be implemented on farms, and what are the long-term effects to providing cognitive enrichment in the early stages of development.

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Ruffled minds? First insights into restlessness as a potential novel indicator of impaired welfare in bulls fattened for meat production

Hintze, S.; Wildemann, T.; Krottenthaler, F.; Winckler, C.

2026-03-31 animal behavior and cognition 10.64898/2026.03.29.715061 medRxiv
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Restlessness is a symptom of chronic boredom in humans and a behavioural phenomenon anecdotally described as a concern in bulls raised for fattening purposes, but it has so far not been addressed in research. The two studies presented in this paper aimed to gain first insights into restlessness in bulls. We operationally defined restlessness by the number of transitions between behaviours in a given time period, and quantified restlessness in bulls of different weight classes (300, 400, 500 kg) on farms keeping bulls on fully-slatted floors (n=8, Study 1) as well as across three different husbandry systems (fully-slatted floor (FS, n=4), straw-based (SB, n=4) and organic pasture (OP, n=3), Study 2). All farms were visited twice, and the behaviour of different individuals was continuously recorded for 15 minutes each between 9 a.m. and 5 p.m. (Study 1) and for 8 minutes each between 6 a.m. and 10 p.m. (Study 2). The effects of weight class and husbandry system were analysed using generalised linear mixed-effects models, and we ran a sequence analysis to cluster observations by the sequence, frequency, and duration of bulls behaviours in Study 1. Bulls kept in fully-slatted floor systems in Study 1 changed their behaviour on average 48.3 times per 10 minutes, with high variability both within and across farms. Weight class did not have a statistically supported effect on the number of transitions, and the sequence analysis revealed four clusters that differed in sequence and in the number of transitions. In Study 2, OP bulls showed fewer transitions than SB and FS bulls (X22 = 23.6, p < 0.001), while SB and FS bulls did not differ. While SB pens were more structured and offered more space per animal, both SB and FS systems can be characterised by monotony, which may explain the similar level of restlessness in both systems. Alternatively, or in addition, the high feeding intensity in SB and FS systems may have caused the higher number of transitions compared to the OP system, potentially elicited by subacute ruminal acidosis and/or laminitis and the resulting pain. However, these explanations are speculative and require systematic disentanglement in future studies. This study provides initial insights into restlessness in bulls and lays the groundwork for future research to identify the causes underlying restlessness and investigate its association with bull welfare.

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Different Paradigms from Computer Vision Align with Human Assessment of the Mouse Grimace Scale

Reimann, M.; Aloui, J.; Obländer, N.; Andresen, N.; Hohlbaum, K.; Hellwich, O.; Reiske, P.

2026-03-06 animal behavior and cognition 10.64898/2026.03.04.709492 medRxiv
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Animal welfare is a central aspect in animal-based research where mice are most commonly used. Their facial expression can be analyzed to assess their well-being status using the Mouse Grimace Scale. However, its manual application becomes increasingly impractical when used on a large number of animals. This lead to the ongoing integration of computer vision methods to automate the analysis. While such methods have proven effective qualitatively, a systematic assessment to verify their reliability largely remains an open research gap. In this work, we attempted to close this gap as we evaluated three dominant paradigms (i.e., classification from supervised learning features, self-supervised learning features, or landmark locations) for the binary (i.e., well-being un-/impaired) classification of facial mouse images. Our quantitative results showed that such methods can be employed successfully with as low as 16% type II error rates. For qualitative assessment, we visualized the decision-making process and demonstrated that mainly pixels associated with the mouse rather than its environment are used. We further discovered that visual characteristics of the mice beyond those described by the Mouse Grimace Scale contributed to the classification. Our work showed that the automated well-being status assessment in mice is trustworthy and urges towards widespread adoption.

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Semen collection, short term storage, and cryopreservation in the Texas horned lizard (Phrynosoma cornutum)

Julien, A. R.; Griffioen, J. A.; Perry, S. M.; Doege, R.; Burger, I. J.; Barber, D. R.

2026-04-06 zoology 10.64898/2026.04.03.716302 medRxiv
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As global reptile populations continue to decline, improving reproductive success in managed populations of listed species, such as Phrynosoma cornutum (the Texas horned lizard) has become increasingly critical for species survival. One understudied area of reproductive research in reptile species is gamete collection and storage, a crucial component for maintaining genetic diversity. In Texas, semen was collected from wild P. cornutum (n = 20) in June 2025. Semen collection was performed via electroejaculation (EEJ) under alfaxalone anesthesia. Prior to semen collection, snout-vent-lengths (SVL) and weights were recorded and testes measurements were taken using a portable ultrasound. Average sperm motility and concentration across all lizards was 83.7% and 85.7 x 106 sperm/mL, respectively. While lizards with longer SVLs had higher sperm motility, weight and testis size did not affect sperm parameters. Samples were extended in INRA96 and divided for use in cold-storage longevity or cryopreservation trials. Samples under cold-storage conditions were assessed for motility daily for 10 days. Motility was not significantly reduced until 48 hours post-collection and maintained 19% motility at day 10. For cryopreservation, samples were diluted 1:1 in INRAFreeze cryopreservation media and frozen in liquid nitrogen, then immediately thawed. Average post-thaw sperm motility was 13.9%, with the highest post-thaw motility recorded at 38.2%. This is the first report of semen storage and cryopreservation in Phrynosoma and provides valuable insight into semen storage potential in reptile species.

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Large-scale automated detection of gray whales off California in panchromatic and multispectral satellite imagery.

HOUEGNIGAN, L.; Cuesta Lazaro, E.

2026-04-19 bioinformatics 10.64898/2026.04.15.718679 medRxiv
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Increasing human activities along the US west coast are of concern for populations of cetaceans and particularly for a number of large whale species that are recovering from overexploitation during the era of commercial whaling. New rapid monitoring tools, such as satellite imagery analysis powered by recent advances in artificial intelligence, have potential to provide additional broad-scale and near real-time capacities for survey and monitoring. This paper investigates and demonstrates the feasibility of automatic detection of gray whales in sub-meter satellite imagery off the coast of California, USA. Observations and statistical analysis of regional imagery allowed not only an assessment of their detectability but also the development of robust signal processing and machine learning-based solutions for automated detection. To that end, a regional dataset of 221 gray whales was created using signal processing to inform a deep-learning-based detection framework, and 20 different large neural network architectures for feature extraction followed by a support vector machine algorithm for classification were evaluated for their detection performance. Neural network backbones included 19 convolutional neural networks and 1 transformer network. The best architecture generally achieved satisfying performance with an average balanced accuracy reaching up to 99.90%. It is also demonstrated that panchromatic imagery, in spite of the lesser amount of information provided, can be used to perform detection with a relatively high accuracy of 87.05%, allowing wider spatial and temporal coverage. Large-scale deployment of the best performing models over a broad range of regional satellite imagery resulted in the detection of 3353 gray whales, as well as opportunistic detections of humpback, blue and fin whales, in and going from December 28th 2009 to March 26th 2023. It also provided meaningful data points concerning the migration routes of gray whales within the Channel Islands and Southern California Bight. The large number of high-confidence detections indicates the capacity for a large-scale monitoring approach to support state and federal conservation policies such as gear mitigation, vessel speed reduction programs, or shipping lane redefinition that could also be expanded to other areas and for other species.

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Accurate estimation of canine inbreeding using ultra low-coverage whole genomesequencing

Pellegrini, M.; Kim, R.; Rubbi, L.; Kislik, G.; Smith, D.

2026-04-07 bioinformatics 10.64898/2026.04.04.716453 medRxiv
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The measurement of inbreeding has gained significance across diverse fields, including population and conservation genetics, agricultural genetics, breeding programs for animals and plants, and wildlife management. This is due to the fact that inbreeding leads to increased homozygosity and results in lower genetic diversity, rendering populations more vulnerable to environmental changes, diseases, and other stressors. High or mid-coverage whole genome sequencing (WGS) has been widely used for inbreeding estimation, but it is resource-intensive. We aimed to investigate the use of ultra low-coverage whole genome sequencing (ulcWGS) as a cost-effective alternative for inbreeding analysis. Domestic dogs were used for our study as their extensive breeding histories lead to populations with a wide range of inbreeding levels. We constructed a multi-breed reference panel from high-coverage WGS samples. Inbreeding in independent ulcWGS samples was then estimated using runs of homozygosity (RoH) and inbreeding coefficients (F). We modeled the relationship between these measures and sequencing depth using nonlinear regression, to generate inbreeding estimates relative to sequencing depth. Resulting relative RoH and F measurements were significantly correlated, with purebred dogs exhibiting more runs of homozygosity and higher inbreeding coefficients compared to mixed-breed dogs. Our findings demonstrate that ulcWGS can provide reliable and economical estimations of inbreeding, expanding accessibility to genetic monitoring.

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On the intra-laboratory replicability of results in animal cognition research, obtained in animals that were not exposed to, or unaffected by, experimental manipulations - Exemplified by spatial learning in pigs (Sus Scrofa Domesticus) across 12 holeboard studies

van der Staay, F. J.; Antonides, A.; Dwulit, A. K.; Fijn, L.; Gieling, E. T.; Grimberg-Henrici, C. G. E.; Meijer, E.; Roelofs, S.; Vernooij, J. C. M.; Witjes, V. L.; Arndt, S. S.

2026-04-22 animal behavior and cognition 10.64898/2026.04.19.718108 medRxiv
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The replicability of experimental results is considered a cornerstone of empirical research. However, the reproducibility of results from groups that have not undergone experimental manipulation -- the standard against which comparisons in an experiment are made -- has been almost entirely neglected in animal research. Our aim is to address this gap by exemplarily determining within-laboratory replicability using research in pigs, an increasingly popular large animal model species. Drawing on data from twelve independent porcine holeboard studies conducted in our laboratory, we examine the replicability of groups that were not subjected to experimental manipulation (typically the control group), eventually including groups on which the experimental treatments had no effect. These analyses were also performed on all eight studies involving the Terra x Finnish Landrace x Duroc pig breed, with all other breeds excluded to increase genetic uniformity. The holeboard is a complex spatial discrimination task in which an animal must learn to find food at four of sixteen possible locations (holes) arranged in a 4 x 4 matrix. The main variables measured are spatial working memory (WM), reference memory (RM) and the inter-visit interval (IVI), which serves as an index of motivation. All studies showed predominantly linear improvements in WM and learning rates across successive trial blocks. IVI showed greater variation across trialblocks, but this did not affect WM and RM learning, which are robust and replicable indices of spatial learning in pigs. Assessing replicability provides relevant information, such as whether behavioural and physiological traits in a model species are stably expressed and robust across studies. Including replicability research and publishing its results can stimulate the development and use of more replicable methods and workflows, thereby increasing scientific rigour. Provided the data are available and accessible, the next step should be to expand replicability studies to include those conducted in different laboratories.

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Reverse gingival venipuncture: a refined technique for serial blood collection in small rodents

Luapan, J.; Johnson, H.; Seiberlich, M.; Brayton, C. A.; Jimenez, I. A.

2026-04-28 zoology 10.64898/2026.04.24.720739 medRxiv
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ObjectiveTo assess the safety, efficacy, and repeatability of a novel blood collection technique, percutaneous reverse gingival venipuncture (RGV), across multiple rodent species, and to characterize the associated anatomy through dissection and histopathology. MethodsSuccess rate and complications of RGV were evaluated at a private practice between December 2024 and September 2025 in client-owned chinchillas (Chinchilla lanigera) (n=102), guinea pigs (Cavia porcellus) (n=78), Syrian hamsters (Mesocricetus auratus) (n=32), dwarf hamsters (Phodopus campbelli and P. sungorus) (n=4), squirrels (Callosciurus erythraeus, C. finlaysonii) (n=7), prairie dogs (Cynomys ludovicianus) (n=2), a capybara (Hydrochoerus hydrochaeris) (n=1) and a Patagonian mara (Dolichotis patagonum) (n=1). An experimental study was conducted in laboratory rats (Rattus norvegicus) (n=5), Chinese hamsters (Cricetulus griseus) (n=12), and chinchillas (n=11) from February 2026 to March 2026 to evaluate RGV success rate, serial hematology, and histopathology. ResultsThe success rate of RGV was 100% in most rodent species, but was lower in guinea pigs (44.87%, n=78) and chinchillas (64.60%, n=113). No animals experienced clinical complications. No significant changes in serial hematology were appreciated in Chinese hamsters (n=6) on days 0, 7, and 14. Histopathology did not reveal any complications. ConclusionsRGV in anesthetized rodents is safe, effective, minimally invasive, and repeatable, yielding clinically relevant blood volumes with precise control and minimal risks. Clinical RelevanceRGV may facilitate more routine hematology and chemistry analysis in rodents by veterinary practitioners, with few risks and complications. In research settings, RGV may improve animal welfare and contribute to refinement and reduction.

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The lost vultures of Romania: reconstructing two centuries of decline from historical records (Gyps fulvus, Aegypius monachus, Neophron percnopterus, Gypaetus barbatus)

Osvath, G.; Denes, A. L.; Kovacs, Z.; Birau, A. C.; Papp, E.; Jako, G. V.; Zeitz, R.

2026-05-18 zoology 10.64898/2026.05.13.723308 medRxiv
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Romania represents one of the few European Union member states in which all four Old World vulture species historically maintained breeding populations: the Griffon Vulture (Gyps fulvus), Cinereous Vulture (Aegypius monachus), Egyptian Vulture (Neophron percnopterus) and Bearded Vulture (Gypaetus barbatus). Until the 2026 reintroduction efforts initiated by Foundation Conservation Carpathia and Rewilding Romania, Romania remained the last EU country whose former vulture guild had not been targeted for active recovery. Despite this exceptional significance in a European conservation context, no comprehensive synthesis of the historical and contemporary distribution of these species in Romania has been undertaken. We conducted a comprehensive review to gather all available vulture occurrence data and present a fully georeferenced database of 1,170 occurrence records spanning 1818-2025. We systematically searched museum collections, historical ornithological literature, modern field surveys and citizen-science platforms. The database documents substantial breeding populations across the Carpathian arc and Dobrogea until the early twentieth century, followed by near-total breeding collapse between the 1920s and 1960s driven by persecution, secondary poisoning and agrarian transformation. In total, 149 confirmed or probable breeding records have been documented for the four species combined, with the most recent confirmed breeding records dating to 1929 (Gyps fulvus), 1929 (Gypaetus barbatus), 1942 (Aegypius monachus) and 1966 (Neophron percnopterus). Non-breeding occurrences increase markedly after 2010, consistent with dispersal from expanding Balkan source populations. The F[a]g[a]ra {square} and Retezat Mountains emerge as the historically most important breeding strongholds for all four species. Our dataset constitutes the most detailed historical baseline currently available for vulture conservation in Romania and is intended to identify key historical sites with high potential for future reintroduction and recovery. Our results show that Romania historically supported the full guild of European obligate scavengers, and that its collapse occurred within barely four decades (1920s-1960s). The dataset highlights the value of reconstructing historical baselines in regions where functional extinction preceded the onset of modern monitoring, and provides an empirical foundation for reassembling a keystone scavenger guild at a continental scale.

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Forensic Identification of Confiscated Helmeted Hornbill (Rhinoplax vigil) Casques and Implications for Individual Quantification in Wildlife Crimes

Shen, Y.; He, K.; Wang, W.; Huang, L.; Chen, J.

2026-04-06 zoology 10.64898/2026.04.02.715475 medRxiv
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In wildlife forensic practice, species identification and estimation of the Minimum Number of Individuals (MNI) for highly processed specimens have long relied on weight-based conversion methods, which may result in underestimation of the number of individuals involved in a case. Focusing on confiscated casque products of the helmeted hornbill (Rhinoplax vigil), this study combines macroscopic morphological examination with mitochondrial DNA barcoding (16S rRNA, COI, and Cytb) to explore a more robust approach for individual quantification. The results demonstrate that the conventional "weight-based" approach overlooks critical biological information contained in anatomical structures and cannot accurately reflect the actual number of individuals involved. Based on this, we propose an anatomy-based criterion centered on the principle of structural uniqueness: specimens retaining biologically unique beak or casque structures should be directly assigned to a single individual, whereas weight-based estimation should only be applied when original anatomical features are entirely absent. In addition, considering material loss during processing, we propose approximately 85 g as a reference threshold for estimating the number of individuals in heavily processed solid casque products. This approach improves the scientific rigor and accuracy of forensic identification and provides reliable technical support for the conviction, sentencing, and law enforcement of wildlife trafficking cases involving helmeted hornbill and other endangered species.

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Effect of dietary Chitosan supplementation on Intestinal Barrier Function and Growth Performance in weaned piglets challenged by Enterohemorrhagic haemolytic Escherichia coli

Liu, J.; De Blassio, S.; Xie, K.; Li, X.; Luo, Y.; Zheng, P.; Mao, X.; Yan, H.; Wang, Q.; Good, L.; Pelligand, L.; He, J.

2026-03-25 microbiology 10.64898/2026.03.23.713631 medRxiv
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Enterohemorrhagic Escherichia coli O157:H7 (EHEC) is an important zoonotic pathogen that disrupts intestinal epithelial barrier integrity and induces excessive inflammatory responses, thereby leading to impaired growth performance and intestinal injury. EHEC is also an important cause Hemolytic Uremic Syndrome (HUS) in children and older adults. In pig production, chitosan is considered a promising alternative to antibiotics due to its bioadhesive and antimicrobial properties, but the effects and underlying mechanisms of chitosan (COS) under pathogenic challenge remain to be elucidated. One hundred and eight pigs were randomly divided into three treatments: an unchallenged control group (CON), an EHEC-challenged control group (ECON), and an EHEC-challenged group supplemented with 100 mg/kg COS (ECOS). Results show that EHEC challenge increased the feed conversion ratio (FCR), increased inflammatory cytokine levels, disrupted intestinal morphology, and downregulated tight junction and nutrient transporter gene expression (P<0.05). Dietary COS supplementation significantly improved average daily gain (ADG) and FCR during day 6-14 (P<0.05). Moreover, COS reduced fecal shedding of total E. coli (P = 0.085) and EHEC, attenuated systemic inflammation by decreasing serum TNF- and IL-6 levels, and enhanced humoral immunity as indicated by increased IgA and IgM concentrations (P<0.05). Importantly, COS alleviated EHEC-induced intestinal injury by restoring villus height and villus-to-crypt ratio, with enhanced mucosal digestive enzyme activities, and upregulated expression of tight junction proteins (ZO-1 and occludin) and nutrient transporters (SGLT-1 and PEPT1) (P<0.05). In conclusion, these findings indicate that dietary COS improves growth performance in EHEC-challenged weaned pigs, with enhanced intestinal barrier integrity and nutrient transport capacity.

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Application of modern mathematical methods for species discrimination in the water fleas (Cladocera: Branchiopoda) that appear similar to the human eye: case of Bosmina (Bosmina) longirostris (O.F. Muller, 1776) from European Eurasia and Sakhalin Island

Garibian, P.; Rubleva, V.; Burlakov, A.; Valeyev, V.; Kasatkina, A.; Kirova, V.

2026-05-22 zoology 10.64898/2026.05.20.726562 medRxiv
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4.1%
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Intraspecific morphological variability presents a complex challenge for biological systematics and biomonitoring, particularly for organisms with high phenotypic plasticity, such as zooplankton. Morphological differences between individuals of the water flea species Bosmina longirostris (Crustacea: Cladocera) are difficult to distinguish visually, parthenogenetic females look morphologically uniform within the species; nevertheless, they demonstrate differences attributable to their geographic origin and developmental stage. A reference dataset of microscopic images was created for the study, including populations from two geographically separated regions (seven ones from European Russia and seven ones from Sakhalin Island in the Pacific Ocean (Far East of Russia) and two age groups, demonstrating the ability of a neural network classify to successfully the intraspecific morphological variation. This study demonstrates that deep learning methods are prospective for the detection and understanding of fine morphological intraspecific differences in the cladocerans.

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First hematological and biochemical data in a rehabilitated giant pangolin (Smutsia gigantea) from southern Cameroon

Montblanc, M.; Harvey-Carroll, J.; Vanassche, J.; Donaldson, M.; Connelly, E.; Hywood, L.

2026-05-03 zoology 10.64898/2026.04.29.721582 medRxiv
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Giant pangolin (Smutsia gigantea) is one of the least studied pangolin species worldwide, with no published hematological and biochemical data available. We report the first blood parameters from a rehabilitated adult male from Campo Maan National Park (southern Cameroon). Hematological and biochemical findings are described and discussed in relation to available data from other pangolin species. These preliminary results provide the first reference framework for this species and highlight their relevance for clinical assessment, health monitoring, and conservation management.

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Automated morphometry and weight prediction of juvenile Chinook Salmon leveraging open-source deep learning models

Knight, B.; Jeffres, C.

2026-03-12 ecology 10.64898/2026.03.10.710725 medRxiv
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Minimizing handling of threatened and endangered fish has become increasingly important as populations have dwindled. To minimize handling in morphometric measurements, the HandsFreeFishing program has been developed for juvenile Chinook Salmon (Oncorhynchus tshawytscha). By segmenting a 2D image many morphometric measurements are able to be estimated; from these measurements a weight prediction model is built based on fish whose ground truthed weights were measured using a digital scale. While many segmentation methods may be used, here Metas Segment Anything model (SAM) is employed to produce segmentation masks of raw images. This model is open-source and easily used on any image (of any size) with good performance. In the proposed framework, the user supplies a bounding box around a target fish along with minimal orientation data (left or right facing, upside down or right-side up); the rest of the segmentation, feature extraction, and final weight prediction is completely automated. A main goal of the segmentation is to estimate the surface area of the side profile of the fish. Then, assuming an ellipsoidal shape, this surface area can be related to the volume of the fish, which is directly proportional to the weight. Even on a relatively small dataset of 149 images (fork length 27-90mm) our results confirm the predictive qualities of the morphometric features measured. The model achieved weight prediction with a mean absolute error of 0.16 g with a mean absolute percentage error of 12%, and an r-squared value of 0.99, on fish ranging from 0.31g - 7.74g. The raw images come from a variety of fish viewers, the design of which is relatively inexpensive and reproducible, and, in conjunction with the HandsFreeFishing program, allows for minimal handling compared to traditional length and weight measurement methods.

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Daily feeding rhythms may play a role in the genetic variability of feed efficiency in growing pigs

Gilbert, H.; Foury, A.; Agboola, L.; Devailly, G.; Gondret, F.; Moisan, M.-P.

2026-04-21 zoology 10.64898/2026.04.17.719142 medRxiv
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AO_SCPLOWBSTRACTC_SCPLOWImproving feed efficiency in pigs is essential for reducing production costs and environmental impacts. This study examines the influence of circadian feeding rhythms and genetic polymorphisms on feed efficiency variability using two pig lines divergently selected for Residual Feed Intake (RFI) over ten generations. Feeding behavior was monitored using automatic concentrate dispensers, recording 6,494,097 visits from 3,824 pigs to analyze meal frequency, duration, and diurnal patterns. LRFI pigs ate less frequently, with larger meals and longer durations, they exhibited two distinct feeding peaks: one around 8:00 AM and a higher one at 5:00 PM and they consumed more feed during the diurnal period and less at night. HRFI pigs showed a smoother, less rhythmic feeding behavior with increased nocturnal intake. The differences between the two RFI lines became more pronounced as the number of generations of selection increased, suggesting a genetic basis. Feeding behaviors, including intake during the two main diurnal peaks, were found to be heritable (heritability estimates: 0.30-0.40) and genetic correlations were observed between feed intake and RFI, especially for intake between the two peaks. Then, we investigated the evolution of allele frequencies of single nucleotide polymorphisms (SNPs) in DNA sequences surrounding 10 core clock genes (ARNTL, CLOCK, CRY1, CRY2, NPAS2, NR1D1, PER1, PER2, PER3, RORA) along generations of selection. SNPs with significant frequency changes were mapped to regulatory regions and transposable elements, especially in HRFI line, suggesting potential functional impacts on circadian regulation. These results underscore the role of feeding behavior and genetic variation in feed efficiency, offering insights for breeding programs aimed at improving metabolic efficiency and sustainability in pig production.