Animals

23 pages, 2383 KB

Open AccessArticle

A New Family-Based Approach for Detecting Allele-Specific Expression and for Mapping Possible eQTLs

by Maher Alnajjar, Zsófia Fekete, Tibor Nagy, Zoltán Német, Agshin Sakif, Nóra Ninausz, Péter Fehér, Viktor Stéger and Endre Barta

Animals 2025, 15(18), 2766; https://doi.org/10.3390/ani15182766 - 22 Sep 2025

Viewed by 504

Abstract

Allele-specific expression (ASE) reflects the unequal expression of the parental alleles and can imply functional variants in cis-regulatory elements. The conventional ASE detection methods often depend on the presence of heterozygous variants in transcripts or sequencing a large number of individuals, both of [...] Read more.

Allele-specific expression (ASE) reflects the unequal expression of the parental alleles and can imply functional variants in cis-regulatory elements. The conventional ASE detection methods often depend on the presence of heterozygous variants in transcripts or sequencing a large number of individuals, both of which are often limited. In this study, we present a family-based strategy for detecting ASE and potential cis-regulatory elements utilizing both RNA-seq and whole-genome sequencing (WGS) from a pedigree. Using a rabbit family consisting of two divergent parents and their eight offspring, we identified 913 ASE genes by analyzing inheritance patterns of gene expression levels. Expression was classified into three levels—high, medium, and low—and used to define seven distinct expression groups across the family (e.g., H_L: high in the mother, low in the father, and intermediate in the offspring). Many ASE genes lacked heterozygous exonic variants, and inference was achieved via RNA read count patterns. We also pinpointed conserved transcription factor binding sites (TFBS) with sequence variants showing similar inherited genotypic patterns (e.g., AAxBB), suggesting their regulatory roles as eQTLs. Differential gene expression (DEG) analysis between the parents highlighted some candidate genes related to meat production and quality traits. Our findings show that the family-based method using RNA-seq and WGS data is promising for exploring ASE and mapping possible eQTLs. Full article

(This article belongs to the Special Issue Livestock Genetic Evaluation and Selection)

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15 pages, 4822 KB

Open AccessArticle

Transcriptomic Analysis Reveals Developmental Toxicity Induced by Environmentally Relevant Concentrations of Fenvalerate and Sulfamethoxazole in Embryo and Juvenile Marine Medaka (Oryzias melastigma, McClelland, 1839)

by Shuyuan Chen, Meina Duan, Qian Liu, Yuna Huang and Dong Sun

Animals 2025, 15(18), 2765; https://doi.org/10.3390/ani15182765 - 22 Sep 2025

Viewed by 340

Abstract

The abuse of pesticides and antibiotics has increased the potential risks for coastal marine organisms. Thus, the evaluation of the toxic effects on and mechanisms of both marine organisms is very important. This study investigated the developmental toxicity and mechanisms of FEN and [...] Read more.

The abuse of pesticides and antibiotics has increased the potential risks for coastal marine organisms. Thus, the evaluation of the toxic effects on and mechanisms of both marine organisms is very important. This study investigated the developmental toxicity and mechanisms of FEN and SMX in embryo and juvenile marine medaka. The results showed that when embryos were exposed to FEN (0.3, 30 μg·L⁻¹) or SMX (1, 10 μg·L⁻¹) for 12 days, there were no significant effects on hatching, malformation, or mortality rates. However, the combined exposure to F-S (F-0.3 + S-10) significantly reduced the hatching rate and increased the malformation rate and mortality rate. Histological analysis revealed that there were no pathological changes in the tissues of juvenile marine medaka, indicating no sensitivity at low doses. A transcriptome analysis of tissues exposed to F-S identified 1135 differentially expressed genes (DEGs), which was significantly more than that for single exposure groups (FEN: 499 DEGs; SMX: 138 DEGs). KEGG enrichment analysis highlighted that growth-related pathways were disturbed, such as steroid biosynthesis, the cell cycle, fatty acid metabolism, etc. Our findings suggest that the combined exposure to FEN and SMX induces developmental toxicity by regulating the dysregulation of metabolic and proliferative pathways, highlighting the ecological risks posed to marine organisms in polluted estuaries. Full article

(This article belongs to the Special Issue Ecotoxicology in Aquatic Animals: 2nd Edition)

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21 pages, 1523 KB

Open AccessSystematic Review

Effects of Palm Kernel Cake on Nutrient Utilization and Performance in Confined Cattle, Sheep and Goats: A Comparative Meta-Analytical Approach

by Julián Andrés Castillo Vargas and Anaiane Pereira Souza

Animals 2025, 15(18), 2764; https://doi.org/10.3390/ani15182764 - 22 Sep 2025

Viewed by 537

Abstract

This meta-analysis explored the relationship of palm kernel cake inclusion level (PKC_Inclusion) with nutrient utilization and performance in cattle, goats and sheep under confinement. For this purpose, a dataset with 51 studies was constructed by using the PRISMA (Preferred Reporting Items [...] Read more.

This meta-analysis explored the relationship of palm kernel cake inclusion level (PKC_Inclusion) with nutrient utilization and performance in cattle, goats and sheep under confinement. For this purpose, a dataset with 51 studies was constructed by using the PRISMA (Preferred Reporting Items for Systematic Review and Meta-Analysis) Protocol. Relationships of PKC_Inclusion with dry matter (DM), crude protein (CP), EE (ether extract), NDF (neutral detergent fiber) and TDN (total digestible nutrients) intake and digestibility, as well as with ADG (average daily gain) and FE (feed efficiency), were explored under a mixed model approach, considering the species and study as fixed and random effects, respectively. Data revealed independent (p < 0.075) relationships of EE and TDN intake and digestibility with PKC_Inclusion for cattle, goats and sheep. However, the relationship of CP intake and digestibility with PKC_Inclusion did not differ (p > 0.114) between ruminant species. Goats and sheep demonstrated similar quantitative patterns for DM and NDF intake but different quantitative patterns (p < 0.037) from those observed for cattle with the increase in PKC_Inclusion in the diet. Regarding performance, FE responses were similar between cattle and sheep but differed (p < 0.001) from those observed for goats; however, ADG demonstrated similar (p = 0.243) decreasing rates among ruminant species in function of dietary PKC_Inclusion. In conclusion, PKC_Inclusion has differential effects on the intake and digestibility of DM and most of the nutritional components in confined cattle, goats and sheep, except for CP. The data reported herein could be used in future nutritional models to allow for the better use of alternative feedstuffs, such as PKC in productive ruminants under confinement. Full article

(This article belongs to the Collection Use of Agricultural By-Products in Animal Feeding)

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10 pages, 2208 KB

Open AccessArticle

Rapid and Visually Specific Detection of Sarcocystis miescheriana and Sarcocystis suihominis Infections in Domestic Pigs (Sus scrofa domesticus) Using Loop-Mediated Isothermal Amplification

by Zhun Hu, Tao Qin, Luyao Qian, Lu Xu, Liwu Zhang, Shuangsheng Deng, Jianping Tao and Junjie Hu

Animals 2025, 15(18), 2763; https://doi.org/10.3390/ani15182763 - 22 Sep 2025

Viewed by 326

Abstract

The domestic pig (Sus scrofa domesticus) serves as an intermediate host for two Sarcocystis species: the non-zoonotic S. miescheriana and the zoonotic S. suihominis, both of which threaten animal and human health and contribute to economic losses in swine production. [...] Read more.

The domestic pig (Sus scrofa domesticus) serves as an intermediate host for two Sarcocystis species: the non-zoonotic S. miescheriana and the zoonotic S. suihominis, both of which threaten animal and human health and contribute to economic losses in swine production. Existing diagnostic methods, such as microscopy and PCR, suffer from limitations regarding their sensitivity, cost, and field applicability, especially in resource-constrained settings. To address these challenges, we developed a highly specific and ultrasensitive loop-mediated isothermal amplification (LAMP) assay targeting the mitochondrial cytochrome c oxidase subunit I (cox1) gene for rapid detection. The optimized protocol (62 °C for 40 min) showed absolute specificity, with no cross-reactivity to related coccidia. The assay exhibited remarkable sensitivity, detecting as little as 6.7 × 10⁻⁶ ng/μL (S. miescheriana) and 5.4 × 10⁻⁷ ng/μL (S. suihominis), representing 10-fold and 10,000-fold improvements over conventional PCR, respectively. With a simple visual readout, the proposed LAMP assay eliminates the need for sophisticated equipment, making it an ideal field-deployable diagnostic tool for basic laboratories and under-resourced regions. Full article

(This article belongs to the Section Pigs)

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20 pages, 313 KB

Open AccessArticle

Egg Production and Biochemical Evaluation of Laying Quails Fed Diets Containing Phytase Overdosage Under Different Thermal Conditions

by Amana Fernandes Maia, Apolônio Gomes Ribeiro, Raiane dos Santos Silva, Edijanio Galdino da Silva, Luiz Arthur dos Anjos Lima, Edilson Paes Saraiva, Felisbina Luisa Pereira Guedes Queiroga, Ana Cristina Silvestre Ferreira, Xavière Rousseau, Fernando Guilherme Perazzo Costa and Ricardo Romão Guerra

Animals 2025, 15(18), 2762; https://doi.org/10.3390/ani15182762 - 22 Sep 2025

Viewed by 415

Abstract

This study evaluated the effects of increasing phytase levels on serum biochemical parameters and renal and hepatic changes in Japanese quails (Coturnix japonica) under different temperature conditions. A total of 720 quails were distributed in a 5 × 3 factorial design [...] Read more.

This study evaluated the effects of increasing phytase levels on serum biochemical parameters and renal and hepatic changes in Japanese quails (Coturnix japonica) under different temperature conditions. A total of 720 quails were distributed in a 5 × 3 factorial design with five phytase levels (0, 500, 1000, 1500, and 3000 FTU/kg) and three temperature ranges (24, 30, and 36 °C), totaling 15 treatments with six replicates of eight birds each. Data were collected in cycles 2 and 4. The parameters measured included eggshell thickness (ST), total egg production (TEP), liver weight (LW), gamma-glutamyl transferase (GGT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (FA), phosphorus (P), calcium (Ca), urea (URE), creatine kinase (CK), and uric acid (UA). PRO was higher at 30 °C compared to 24 °C and 36 °C. EC showed a significant interaction between phytase levels and temperature, with phytase improving EC, especially at 1500 FTU and 36 °C. PF was not affected in the second cycle but had a quadratic effect in the fourth cycle. Phytase supplementation improved shell thickness and regulated biochemical parameters, especially at high temperatures. Based on these results, a phytase level of 1500 FTU/kg is recommended to optimize performance and mitigate the negative effects of high temperatures on quail health. Full article

(This article belongs to the Section Poultry)

11 pages, 221 KB

Open AccessArticle

An Increase in Dietary Net Energy Concentration Affects Nutrient Digestibility and Noxious Gas Emissions and Reveals a Better Growth Rate in Growing–Finishing Pigs

by Usman Kayode Kolawole and In Ho Kim

Animals 2025, 15(18), 2761; https://doi.org/10.3390/ani15182761 - 22 Sep 2025

Viewed by 511

Abstract

This experiment was performed to evaluate the optimum level of net energy (NE) to improve growth performance and nutrient digestibility, and reduce noxious gas emissions in growing–finishing pigs. A total of 150 pigs ([Yorkshire × Landrace] × Duroc) with an initial average body [...] Read more.

This experiment was performed to evaluate the optimum level of net energy (NE) to improve growth performance and nutrient digestibility, and reduce noxious gas emissions in growing–finishing pigs. A total of 150 pigs ([Yorkshire × Landrace] × Duroc) with an initial average body weight (BW) of 32.64 ± 1.49 kg were randomly assigned to one of five treatments for 112 days (16 weeks: growing stage, initial–week 6; finishing stage, 6-finish/week 16). There were five treatment groups with six replicates and five pigs (three males and two females) per pen. The dietary treatment includes CON, a basal diet (NE 2475 kcal/kg), TRT1, basal diet −5.0% NE (2353 kcal/kg), TRT2, basal diet −2.5% NE (2414 kcal/kg), TRT3, basal diet +2.5% NE (2537 kcal/kg), and TRT4, basal diet +5.0% NE (2599 kcal/kg). Through the experiment, increasing NE by +5.0% increased average daily gain and bodyweight, having the highest value (p < 0.05) and a decreased feed conversion ratio (p < 0.05), whereas decreasing NE by −5% decreased average daily gain (p < 0.05) and increased feed conversion ratio (p < 0.05) at week 6 with no effects on nutrient digestibility and noxious gas emission. Hence, integrating the increasing level of net energy (NE) with +5.0% into the pig diet during the growing–finishing phase can be considered the appropriate approach for enhancing both average daily gain and feed efficiency in pigs. A +5% (2599 kcal/kg) increase in NE has the potential to reduce costs by lowering the amount of feed needed while either sustaining or improving growth, as the use of energy-dense ingredients becomes more cost effective. Full article

(This article belongs to the Section Animal Nutrition)

12 pages, 242 KB

Open AccessArticle

Serum Creatinine, Cystatin C and Symmetric Dimethylarginine Concentrations and Relationship Between Them in Healthy Small and Miniature Dogs: A Preliminary Study

by Julia Rafalska, Agnieszka Rusiecka and Jolanta Bujok

Animals 2025, 15(18), 2760; https://doi.org/10.3390/ani15182760 - 22 Sep 2025

Viewed by 3981

Abstract

Currently, there are common reference intervals for creatinine levels in dogs, despite different weight categories. This can result in missing the diagnosis of kidney disease at an early stage in small dogs. The aim of this study was to determine and compare available [...] Read more.

Currently, there are common reference intervals for creatinine levels in dogs, despite different weight categories. This can result in missing the diagnosis of kidney disease at an early stage in small dogs. The aim of this study was to determine and compare available markers of the GFR (serum cystatin C, creatinine and SDMA) in healthy small and miniature dogs. Each dog underwent the same protocol, including physical examination, blood pressure measurement and ultrasound of the urinary system. The serum filtration markers determined in the study were creatinine, cystatin C and SDMA. Urinalysis was performed, including urine specific gravity (USG) and the urine protein/creatinine ratio (UPC). Forty samples were included. The median values with the IQR (interquartile range) for creatinine, SDMA and cystatin C levels in the examined dogs were 63.8 µmol/L (16.7), 10.63 µg/dL (4.64) and 0.5 mg/L (0.2), respectively. Compared with Yorkshire Terriers and Chihuahuas, Maltese dogs had significantly higher median serum creatinine concentration (p < 0.001). Combining results of GFR markers, USG, UPC, patient’s clinical condition as well as the breed is mandatory when assessing renal function in small dogs. Maltese dogs may have physiologically higher serum creatinine levels. Full article

(This article belongs to the Section Veterinary Clinical Studies)

15 pages, 1109 KB

Open AccessArticle

Genetic Diversity and Core Germplasm Identification in Penaeus japonicus Using Whole-Genome Resequencing

by Dingyuan Zhang, Jikang Shentu, Weijian Liu, Yanxia Wang, Minjun Zhu, Zhiming Yang and Liegang Si

Animals 2025, 15(18), 2759; https://doi.org/10.3390/ani15182759 - 22 Sep 2025

Viewed by 372

Abstract

The kuruma shrimp (Penaeus japonicus), a globally high-value aquaculture species, faces critical challenges in sustainable development due to genetic diversity degradation and declining disease resistance. This study employed whole-genome resequencing (WGRS) to systematically assess genetic diversity, population structure, and core germplasm [...] Read more.

The kuruma shrimp (Penaeus japonicus), a globally high-value aquaculture species, faces critical challenges in sustainable development due to genetic diversity degradation and declining disease resistance. This study employed whole-genome resequencing (WGRS) to systematically assess genetic diversity, population structure, and core germplasm characteristics across 20 geographically distinct populations from Zhejiang, Fujian (China), and introduced Japanese stocks. Using 343.40 Gb of high-quality sequencing data (average depth: 12.44×), we identified 9,146,248 single nucleotide polymorphisms (SNPs), with 6.32% located in exon regions, while intergenic (56.75%) and intronic regions (30.99%) showed the highest polymorphism density. Principal component analysis (PCA) and phylogenetic tree construction revealed two major clades: Fujian (FJ) and Japan-introduced (RB) populations clustered closely due to shared artificial breeding backgrounds, whereas Zhejiang (XS) and Fujian (LS) populations displayed genetic heterogeneity driven by adaptive divergence. Core germplasm screening via the CoreHunter algorithm selected four representative individuals (FJ4-M, LS1-M, XS1-M, XS6-M), with the modified Rogers’ distance (0.34) and allele coverage (0.93) confirming effective preservation of original genetic diversity. This study provides genomic insights and technical frameworks for germplasm conservation, precision breeding, and genetic improvement in P. japonicus. Full article

(This article belongs to the Special Issue Genetics, Breeding, and Farming of Aquatic Animals)

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20 pages, 4700 KB

Open AccessArticle

Computer-Aided Diagnosis of Equine Pharyngeal Lymphoid Hyperplasia Using the Object Detection-Based Processing Technique of Digital Endoscopic Images

by Natalia Kozłowska, Marta Borowska, Tomasz Jasiński, Małgorzata Wierzbicka and Małgorzata Domino

Animals 2025, 15(18), 2758; https://doi.org/10.3390/ani15182758 - 22 Sep 2025

Viewed by 403

Abstract

In human medicine, computer-aided diagnosis (CAD) is increasingly employed for screening, identifying, and monitoring early endoscopic signs of various diseases. However, its potential—despite proven benefits in human healthcare—remains largely underexplored in equine veterinary medicine. This study aimed to quantify endoscopic signs of pharyngeal [...] Read more.

In human medicine, computer-aided diagnosis (CAD) is increasingly employed for screening, identifying, and monitoring early endoscopic signs of various diseases. However, its potential—despite proven benefits in human healthcare—remains largely underexplored in equine veterinary medicine. This study aimed to quantify endoscopic signs of pharyngeal lymphoid hyperplasia (PLH) as digital data and to assess their effectiveness in CAD of PLH in comparison and in combination with clinical data reflecting respiratory tract disease. Endoscopic images of the pharynx were collected from 70 horses clinically assessed as either healthy or affected by PLH. Digital data were extracted using an object detection-based processing technique and first-order statistics (FOS). The data were transformed using linear discriminant analysis (LDA) and classified with the random forest (RF) algorithm. Classification metrics were then calculated. When considering digital and clinical data, high classification performance was achieved (0.76 accuracy, 0.83 precision, 0.78 recall, and 0.76 F1 score), with the highest importance assigned to selected FOS features: Number of Objects and Neighbors, and Tracheal Auscultation. The proposed protocol of digitizing standard respiratory tract diagnostic methods provides effective discrimination of PLH grades, supporting the clinical value of CAD in veterinary medicine and paving the way for further research in digital medical diagnostics. Full article

(This article belongs to the Special Issue Animal–Computer Interaction: New Horizons in Animal Welfare)

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14 pages, 1642 KB

Open AccessFeature PaperArticle

Evaluation of the Effects of Enlarged Housing on Social Play and Reward Seeking in Rats

by E. J. Marijke Achterberg, Anne-Marie J. M. Baars, Daphne A. van Hal, Heidi M. B. Lesscher and Pascalle L. P. Van Loo

Animals 2025, 15(18), 2757; https://doi.org/10.3390/ani15182757 - 22 Sep 2025

Viewed by 484

Abstract

The legal minimum dimensions for laboratory rat housing prevent rats from fully performing their behavioural repertoire. In the regular cages, rats cannot run, climb, or even stand upright. The need to house rats in larger cages has been recognized internationally from a welfare [...] Read more.

The legal minimum dimensions for laboratory rat housing prevent rats from fully performing their behavioural repertoire. In the regular cages, rats cannot run, climb, or even stand upright. The need to house rats in larger cages has been recognized internationally from a welfare perspective, which has led to the development of several commercial cage systems. It is, however, important to evaluate the effect of changes in housing conditions on behaviour in existing animal models. The aim of the study was to determine whether housing animals in larger cages with mandatory enrichment or with additional enrichment would affect play behaviour or sucrose reward sensitivity. This paper describes the effect of housing rats in enlarged cages, the EC4Rats, on social play and for reward seeking. A total of 48 male Lister Hooded rats, PND21-23 at the start of the experiment, were assigned to one of three experimental groups, i.e., (1) standard housing with standard enrichment, (2) EC4Rats housing with standard enrichment and (3) EC4Rats housing with EC4Rats enrichment. The results of the study show that despite subtle differences in social play behaviour, housing rats in these larger cages did not affect reward sensitivity nor the motivation for sucrose self-administration in rats. Taken together, these findings suggest that housing systems that provide rats with more opportunities to engage in natural behaviours do not affect social play or reward-related behaviour. Full article

(This article belongs to the Section Human-Animal Interactions, Animal Behaviour and Emotion)

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16 pages, 1531 KB

Open AccessReview

Antimicrobial Resistance and Residues from Biofilms in Poultry, Swine, and Cattle Farms: A Scoping Review

by Zehra Irshad, Andrea Laconi, Ronald Vougat Ngom, Roberta Tolosi and Alessandra Piccirillo

Animals 2025, 15(18), 2756; https://doi.org/10.3390/ani15182756 - 22 Sep 2025

Viewed by 451

Abstract

Background: The use of antibiotics in livestock has contributed to the spread of antimicrobial resistance (AMR) and biofilms can play a role in its emergence and dissemination. This review aimed to map the literature on AMR, antimicrobial resistance genes (ARGs), and antibiotic residues [...] Read more.

Background: The use of antibiotics in livestock has contributed to the spread of antimicrobial resistance (AMR) and biofilms can play a role in its emergence and dissemination. This review aimed to map the literature on AMR, antimicrobial resistance genes (ARGs), and antibiotic residues (ARs) in biofilms from drinking water distribution systems in poultry, swine, and cattle farms. Methods: The review was conducted according to the PRISMA-ScR extension. Four databases (PubMed, Scopus, Agricola, and Web of Science) were searched. Studies were screened in Rayyan. Results: The search yielded 1242 studies. After screening 732 studies, only 4 met the inclusion criteria. These studies focused on poultry (n = 3) and dairy cattle (n = 1), not on swine. Isolation relied on plating methods. Two studies complemented culturing with 16S rRNA sequencing. No studies applied culture-independent techniques. The number of biofilm-derived isolates across studies ranges from 6 to 102. Three studies performed antimicrobial susceptibility testing, using a wide range of antibiotics (16 to 31). One study analyzed ARGs; none quantified ARs. Conclusions: The limited number of studies and lack of standardized methods hinder the generalizability of the findings, underscoring the need for research to clarify biofilms’ role in AMR dissemination in livestock farms. Full article

(This article belongs to the Section Animal System and Management)

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20 pages, 7319 KB

Open AccessArticle

Modification of Brassica rapa L. Polysaccharide by Selenylation and Its Immune-Enhancing Activity When Combined with a Live-Attenuated Newcastle Disease Vaccine in Poultry

by Sijia Wang, Jungang Wang and Hong Shen

Animals 2025, 15(18), 2755; https://doi.org/10.3390/ani15182755 - 21 Sep 2025

Viewed by 417

Abstract

Background: The quest for effective immunoenhancers is central to improving vaccine efficacy, especially against avian viruses such as Newcastle disease (ND) virus. Selenized polysaccharides integrate bioactive polysaccharides with selenium’s immunoenhancing properties while reducing selenium toxicity, making them promising candidates for the development [...] Read more.

Background: The quest for effective immunoenhancers is central to improving vaccine efficacy, especially against avian viruses such as Newcastle disease (ND) virus. Selenized polysaccharides integrate bioactive polysaccharides with selenium’s immunoenhancing properties while reducing selenium toxicity, making them promising candidates for the development of a novel vaccine immunoenhancer. Aim: This study aimed to develop an efficient selenized Brassica rapa L. polysaccharide (sBRP) and evaluate its potential to enhance the immunogenicity of a live-attenuated ND vaccine in poultry. Methods: Selenization was achieved via nitrite-assisted selenization of Brassica rapa L. polysaccharide (BRP). In vivo, 180 yellow-feathered broilers were divided into six groups: control (Con), vaccine-only (Vac), BRP (20 mg/kg), and low/medium/high-dose sBRP (sBRP-L/M/H: 5/10/20 mg/kg). On days 14 and 28, all groups except Con were vaccinated against ND via drinking water. Concurrently, the BRP and sBRP-L/M/H groups received their respective polysaccharides via oral gavage. Parameters assessed included immune organ indices, lymphocyte proliferation, serum antibody titers (HI), cytokine levels (IL-2/IL-6/IFN-γ), and densities of intestinal intraepithelial lymphocytes (IELs) and goblet cells (GCs). Results: sBRP exhibited a selenium content of 30.6 mg/g, with Se-O-C covalent modification confirmed. The sBRP-H group significantly enhanced immune organ indices, lymphocyte proliferation, Newcastle disease virus HI antibody titers, and serum IL-2/IL-6/IFN-γ levels. The sBRP-M group increased IEL and GC densities in the intestine. Conclusions: sBRP acts synergistically with the vaccine to enhance vaccine-induced cellular, humoral, and mucosal immunity, demonstrating promise as a novel oral vaccine immunoenhancer. Full article

(This article belongs to the Section Animal Nutrition)

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14 pages, 1841 KB

Open AccessFeature PaperArticle

Do Hatchery-Reared Southern Pygmy Perch (Nannoperca australis) Develop Effective Survival Behaviour in a Soft-Release Site?

by James King, Peter Rose, Amina Price and Rafael Freire

Animals 2025, 15(18), 2754; https://doi.org/10.3390/ani15182754 - 21 Sep 2025

Viewed by 335

Abstract

The captive breeding and release of threatened small-bodied freshwater fish is a common conservation method, yet many of these fish lack the behavioural profile to survive in the wild. Soft-release sites that provide wild-like experiences with minimal threat to survival can improve post-release [...] Read more.

The captive breeding and release of threatened small-bodied freshwater fish is a common conservation method, yet many of these fish lack the behavioural profile to survive in the wild. Soft-release sites that provide wild-like experiences with minimal threat to survival can improve post-release outcomes. Here, we investigated whether captive-bred first generation Southern pygmy perch (Nannoperca australis) exposed to six months of soft-release experience develop natural behaviour. In laboratory tests, we compared the behaviour of fish from a hatchery, a soft-release site, or the wild in emergence, exploration, habitat choice, predator response, and novel food tests. As predicted, we found that fish from the soft-release site showed similar behavioural responses to wild-caught fish. However, soft-release fish were significantly larger (14.6 mm, 1.6 g advantage) and made greater use of refuge structures (basket ledges). Also, while trends suggested altered anti-predator responses, statistical support was limited, warranting further investigation. We conclude that soft-release experience enhances growth and shelter-seeking behaviour in this species, though further tests in a more natural environment should be undertaken to confirm ecologically important experience-dependent changes in behaviour. We recommend prioritising soft-release programmes that maximise natural foraging opportunities, reduce competition, and provide complex structures to support shelter use. Finally, the overall similarity in behaviour of fish from different environments suggests that, in this species, behaviour appears mostly inherited. Full article

(This article belongs to the Collection Behavioral Ecology of Aquatic Animals)

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17 pages, 994 KB

Open AccessReview

Genetic Strategies for Improving Pig Robustness: Reducing Antibiotic Use Through Enhanced Resilience and Disease Resistance

by László Gombos, László Búza, Ferenc Szabó and László Varga

Animals 2025, 15(18), 2753; https://doi.org/10.3390/ani15182753 - 20 Sep 2025

Viewed by 495

Abstract

This review investigates genetic strategies aimed at improving robustness in pigs to enhance disease resistance and reduce reliance on antibiotics. Robustness refers to a pig’s ability to stay healthy and productive under stressful or challenging conditions. The review outlines current breeding practices focused [...] Read more.

This review investigates genetic strategies aimed at improving robustness in pigs to enhance disease resistance and reduce reliance on antibiotics. Robustness refers to a pig’s ability to stay healthy and productive under stressful or challenging conditions. The review outlines current breeding practices focused on key traits such as maternal ability, growth, immune function, and survival, and highlights that these robustness-related traits show measurable heritability, making them suitable for genetic improvement. Special attention is given to resistance against porcine reproductive and respiratory syndrome (PRRS), a major disease in swine. We also evaluate breed-specific differences, environmental influences, and immune response profiles, emphasizing their impact on breeding outcomes. The development of robust pig lines emerges as a sustainable strategy to reduce antibiotic dependence and enhance herd health. A distinctive contribution of this work is the integration of genetic robustness and resilience strategies with antibiotic stewardship objectives. We link genomic selection, advanced phenotyping, and targeted management interventions within a One Health framework to outline actionable, system-level pathways for reducing antimicrobial inputs. To our knowledge, this combined genetic and public health perspective has not been comprehensively addressed previously. Full article

(This article belongs to the Topic Sustainability and Innovation in Agriculture for the Food Production of Tomorrow)

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12 pages, 238 KB

Open AccessArticle

The Effect of Lysolecithin Supplementation on Growth Performance, Nutrient Digestibility, Intestinal Morphology, and Lipid Metabolism in Yellow-Feathered Broilers Fed Diets with Different Fat Levels

by Ying Zhang, Leilei Zhu, Zheng Luo and Jie Feng

Animals 2025, 15(18), 2752; https://doi.org/10.3390/ani15182752 - 20 Sep 2025

Viewed by 367

Abstract

This study was conducted to investigate the interaction of dietary fat level and lysolecithin (LL) supplementation on the growth performance, nutrient digestibility, intestinal morphology, intestinal lipase activity, and lipid metabolism in yellow-feathered broilers (YFBs) during day 1–50. A total of 384 one-day-old male [...] Read more.

This study was conducted to investigate the interaction of dietary fat level and lysolecithin (LL) supplementation on the growth performance, nutrient digestibility, intestinal morphology, intestinal lipase activity, and lipid metabolism in yellow-feathered broilers (YFBs) during day 1–50. A total of 384 one-day-old male broilers were assigned to a 2 × 2 factorial design, including dietary fat level (Normal-fat, NF: 3% lard, day 1–21; 4.5% lard, day 22–50/Low-fat, LF: 2.4% lard, day 1–21; 3.6% lard, day 22–50) and LL supplementation (0% or 0.05%). Each treatment had six replicates with 16 birds each. The trial lasted 50 days from day 1 to day 50. During the starter phase (day 1–21), birds fed the NF diet exhibited significantly greater average daily gain (ADG) and a lower feed conversion ratio (FCR) compared to those fed the LF diet (p < 0.05). Supplementation with LL significantly improved ADG and reduced FCR throughout all phases (day 0–21, day 22–50, and day 0–50) (p < 0.05). At day 21, LL also increased the apparent metabolizable energy (AME) and the digestibility of crude protein and ether extract (p < 0.05). LL supplementation increased the villus height-to-crypt depth (VH:CD) ratio in the small intestine and elevated intestinal lipase activity during the experimental period (p < 0.05). Regarding lipid metabolism, LL supplementation decreased serum total cholesterol (TC) and the LDL:HDL ratio, while increasing liver total lipase (TL) activity at both day 21 and day 50 (p < 0.05). These findings suggest that dietary supplementation with 0.05% LL improves growth performance, enhances nutrient digestibility and intestinal absorptive capacity, and positively modulates lipid metabolism in YFBs. Full article

(This article belongs to the Section Poultry)

13 pages, 1412 KB

Open AccessArticle

Molecular and Serological Detection of Leishmania spp. in Mediterranean Wild Carnivores and Feral Cats: Implications for Wildlife Health and One Health Surveillance

by Francesca Suita, Víctor Lizana, Jordi Aguiló-Gisbert, Jordi López-Ramon, João Torres Da Silva, Eduardo A. Díaz and Jesús Cardells

Animals 2025, 15(18), 2751; https://doi.org/10.3390/ani15182751 - 20 Sep 2025

Viewed by 538

Abstract

Leishmaniasis is a zoonotic disease caused by protozoan parasites of the genus Leishmania, transmitted by phlebotomine sandflies. While domestic dogs are the main hosts in the Mediterranean basin, wild carnivores have also been proposed as potential reservoirs. This study assessed the presence [...] Read more.

Leishmaniasis is a zoonotic disease caused by protozoan parasites of the genus Leishmania, transmitted by phlebotomine sandflies. While domestic dogs are the main hosts in the Mediterranean basin, wild carnivores have also been proposed as potential reservoirs. This study assessed the presence of Leishmania spp. in 250 animals from the Valencian Community, eastern Spain—an endemic region—using TaqMan qPCR on spleen samples from 216 wild carnivores and sera from 34 feral cats, and ELISA serology on 174 wild carnivores. DNA of Leishmania spp. was detected in 14 out of 250 individuals (5.6%), with red foxes representing most positive cases (10/102; 9.8%). Seropositivity was observed only in red foxes, with 5 out of 174 individuals testing positive (2.9%). Most qPCR-positive animals had high Ct values, consistent with low parasite loads. One fox, positive by both methods, showed advanced skin lesions and was later diagnosed with sarcoptic mange, suggesting possible interaction with Leishmania infection. The overall low prevalence and parasite burden suggest limited circulation in the surveyed wildlife. These findings contribute to understanding the epidemiological role of wild mesocarnivores and highlight the relevance of wildlife monitoring within a One Health approach. Full article

(This article belongs to the Special Issue Pathogeny, Diagnosis, and Management Strategies in Diseases Threatening Terrestrial Wildlife)

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13 pages, 410 KB

Open AccessArticle

Mapping PRNP Polymorphisms in Portuguese Serra da Estrela Ovine Populations: Insights into Scrapie Susceptibility and Farm Animal Improvement

by Soraia Rodrigues, Guilherme Moreira, Sérgio Santos-Silva, Sara Gomes-Gonçalves, Maria Aires Pereira, Alexandra Baptista, Rita Cruz, Fernando Esteves and João R. Mesquita

Animals 2025, 15(18), 2750; https://doi.org/10.3390/ani15182750 - 20 Sep 2025

Viewed by 408

Abstract

Scrapie (classical and atypical) susceptibility in sheep is strongly influenced by PRNP gene polymorphisms. In Portugal, limited data exist for native breeds such as Serra da Estrela, despite their relevance to animal conservation and food production. The full coding region of PRNP gene [...] Read more.

Scrapie (classical and atypical) susceptibility in sheep is strongly influenced by PRNP gene polymorphisms. In Portugal, limited data exist for native breeds such as Serra da Estrela, despite their relevance to animal conservation and food production. The full coding region of PRNP gene of 92 Serra da Estrela sheep was sequenced and SNP frequencies were analysed. The predicted functional impact of nonsynonymous SNPs was assessed using PolyPhen-2 and AMYCO. A total of 27 SNPs were identified, including 20 nonsynonymous variants. Thirteen major haplotypes were observed. The ARR allele, which provides resistance to classical scrapie, was present in 58.7% of the population, with 18.5% of animals being homozygous. Several previously unreported SNPs were identified, and their impact on prion protein aggregation propensity and structure was explored. The high frequency of the ARR allele without full ARR fixation suggests that no selective breeding for scrapie resistance has been applied. These results support the adoption of gradual selection strategies that preserve genetic variability and promote farmer compliance, while increasing classical and atypical scrapie resistance. Full article

(This article belongs to the Special Issue Technological and Statistical Advances in Breeding and Genetics of Sheep and Goat)

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16 pages, 1882 KB

Open AccessArticle

Effects of Chronic Thermal Stress on the Physiology, Metabolism, Histology, and Gut Microbiota of Juvenile Schizothorax grahami

by Shuangqian Bai, Tingyin Li, Lingfu Kong, Baoliang Bi and Qing Hu

Animals 2025, 15(18), 2749; https://doi.org/10.3390/ani15182749 - 20 Sep 2025

Viewed by 432

Abstract

Temperature is a critical factor influencing fish health and aquaculture success. This study investigates the physiological, histological, and microbiota responses of juvenile Schizothorax grahami to chronic thermal stress across six temperature treatments. We found that exposure to temperatures exceeding 27 °C resulted in [...] Read more.

Temperature is a critical factor influencing fish health and aquaculture success. This study investigates the physiological, histological, and microbiota responses of juvenile Schizothorax grahami to chronic thermal stress across six temperature treatments. We found that exposure to temperatures exceeding 27 °C resulted in complete mortality in juvenile S. grahami, with the upper thermal tolerance range between 24 °C and 27 °C. Chronic thermal stress caused a significant decline in serum glucose (Glu), triglycerides (TG), and total cholesterol (TC) at 24 °C, indicating rapid energy depletion, while reduced serum enzyme activity of catalase (CAT) and total superoxide dismutase (T-SOD) at 24 °C group further reflected a decrease in antioxidant capacity. Histological examination revealed adaptive intestinal villus hypertrophy with increased length and muscularis thickness at temperature under 24 °C. Furthermore, the relative abundance of Cetobacterium and Fusobacteriota suggested either adaptive responses or stress-related dysbiosis that may contribute to weakened host immune function. Overall, our findings highlight the vulnerability of juvenile S. grahami to rising temperatures, with moderate thermal stress inducing adaptive responses, while higher temperatures impair metabolism, gut integrity, microbial health, and may even lead to lethality. These results underscore the need for effective conservation strategies to protect this critically endangered species in the face of climate change. Full article

(This article belongs to the Section Aquatic Animals)

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19 pages, 3616 KB

Open AccessFeature PaperArticle

Effects of Partial Replacement of Alfalfa Hay with Alfalfa Silage in Dairy Cows: Impacts on Production Performance and Rumen Microbiota

by Tian Xia, Zixin Liu, Ziyan Yang, Aoyu Jiang, Chuanshe Zhou and Zhiliang Tan

Animals 2025, 15(18), 2748; https://doi.org/10.3390/ani15182748 - 19 Sep 2025

Cited by 1 | Viewed by 431

Abstract

As an important feed source for ruminants, alfalfa’s rational and efficient utilization is of great significance for the production and economic benefits of pastures. This study focuses on Sanhe dairy cows and includes a control group (CON group, alfalfa in the diet is [...] Read more.

As an important feed source for ruminants, alfalfa’s rational and efficient utilization is of great significance for the production and economic benefits of pastures. This study focuses on Sanhe dairy cows and includes a control group (CON group, alfalfa in the diet is hay) and an experimental group (AS group, alfalfa silage partially replaces alfalfa hay of equal dry weight). The feeding experiment lasted for 60 days. The results revealed that, compared with the CON group, the AS group exhibited increased milk yield, milk protein, and milk fat. There were no significant differences in apparent digestibility, serum biochemical indicators, and volatile fatty acid (VFA) levels between the two groups. However, the microbial composition of the rumen differed significantly between the two groups of cows based on β-diversity. On the genus level, compared with the CON group, the relative abundance of Erysipelatoclostridium, Pseudoflavonifractor, and Candidatus Saccharimonas in the AS group was significantly reduced. In summary, partially replacing alfalfa hay with alfalfa silage feed is beneficial for improving the production performance of cows and changing rumen microbial diversity. These findings provide a basis for the effective utilization of alfalfa. Full article

(This article belongs to the Special Issue Maximizing Fiber Utilization for Sustainable, Efficient Ruminant Production)

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19 pages, 5232 KB

Open AccessArticle

Whole Genome Resequencing Reveals the Genetic Basis of Desert Arid Climate Adaptation in Lop Sheep

by Chenchen Yang, Changhai Gong, Abliz Khamili, Xiaopeng Li, Qifeng Gao, Hong Chen, Xin Xiang, Jieru Wang, Chunmei Han and Qinghua Gao

Animals 2025, 15(18), 2747; https://doi.org/10.3390/ani15182747 - 19 Sep 2025

Viewed by 385

Abstract

The Lop sheep (LOP), a unique local breed from Xinjiang, exhibits remarkable resilience to the harsh conditions of a desert arid climate and frequent sandstorms, alongside notable fecundity characteristics. This study aims to investigate the adaptability of LOP within this challenging environment by [...] Read more.

The Lop sheep (LOP), a unique local breed from Xinjiang, exhibits remarkable resilience to the harsh conditions of a desert arid climate and frequent sandstorms, alongside notable fecundity characteristics. This study aims to investigate the adaptability of LOP within this challenging environment by collecting whole blood samples from 110 LOP individuals in the Lop Nur region of Xinjiang for genome resequencing. The resulting data will be compared with whole genome resequencing information from 22 local sheep breeds worldwide to analyze the origin and evolution of LOP. Additionally, comparisons will be made with HUS sheep from warm and humid regions to identify genomic differences through selection signal analysis, thereby assessing the impact of a desert arid climate on the extreme living conditions of LOP. Finally, qPCR was used to preliminarily analyse the impact of the desert arid climate on the genome of the Bactrian sheep. Genetic diversity results indicate that LOP exhibits a relatively stable genetic structure alongside high genetic diversity. The results of population structure analysis and gene flow indicate that we can tentatively posit that LOP is a breed that originated from the Middle East, subsequently mixing with MGS upon its arrival in Xinjiang. Chinese local sheep breeds trace their origins to AMS, with the gene flow evolving from west to east, progressing through mountainous hills (BSBS), basins (LOP, HTS, CLHS, DLS), plains (MGS, TANS), and coastal areas (HUS). LOP is associated with ALTS, BSBS, HTS, CLHS, and DLS, as well as with MGS, HUS, TANS, WDS, and SSSP, in a context of gene exchange, with the degree of exchange diminishing in that order. Selection signal analysis revealed that the candidate genes identified are closely related to adaptation to desert arid climates and disease resistance (PDGFD, NDUFS3, ATP1B2, ITGB8, and CD79A), using HUS as the reference group. qPCR results demonstrated that LOP was significantly upregulated in cardiac, splenic, and lung tissues compared to HUS, suggesting that LOP plays a crucial role in cardiac function, immune response, and respiratory capacity. Full article

(This article belongs to the Section Animal Genetics and Genomics)

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15 pages, 1527 KB

Open AccessArticle

Effects of Fungal Probiotics on Rumen Fermentation and Microbiota in Angus Cattle

by Lijun Wang, Maolong Li, Chaoqi Liu, Xinxin Li, Ping Wang, Juan Chang, Sanjun Jin, Qingqiang Yin, Qun Zhu, Xiaowei Dang and Fushan Lu

Animals 2025, 15(18), 2746; https://doi.org/10.3390/ani15182746 - 19 Sep 2025

Viewed by 411

Abstract

The potential of fungal probiotics as ruminant feed additives has not been extensively studied. This study aimed to evaluate the effect of A. oryzae and T. longibrachiatum supplementation on Angus cattle during the early stages of fattening. In this study, 80 Angus cattle [...] Read more.

The potential of fungal probiotics as ruminant feed additives has not been extensively studied. This study aimed to evaluate the effect of A. oryzae and T. longibrachiatum supplementation on Angus cattle during the early stages of fattening. In this study, 80 Angus cattle aged approximately 9~10 months (40 males and 40 females), with an average initial body weight (BW) of 276.46 ± 27.92 kg, were randomly assigned to four groups. Each group included 4 replicates (2 replicates of males and 2 replicates of females). Each replicate contained 5 male or 5 female Angus cattle. Cattle in the control group received a total mixed ration (TMR) without additives, while those in test groups 1, 2, and 3 received a TMR supplemented with complex probiotics (CPs) at 1.0, 2.0, and 3.0 g·kg⁻¹ of feed dry matter (DM), respectively. The adaptation and experimental periods were 7 and 60 days, respectively. Compared with those in the control group, the apparent digestibility of ether extract, calcium, and acid detergent fiber (ADF) was higher in test group 3. Test group 2 exhibited increased apparent digestibility of hemicellulose. Meanwhile, test group 3 exhibited increased levels of acetate, propionate, butyrate, and total volatile fatty acids and decreased pH. CPs increased the abundance of the SR1 phylum, Solibacillus, Lysinibacillus, and Planococcaceae_Bacillus and decreased the proportions of Lactococcus, Ruminococcus, and Ophryoscolex. Solibacillus was associated with the apparent digestibility of CP, Ca, and ADF. Planococcaceae_Bacillus was associated with increased apparent digestibility of CP, ADF, and hemicellulose. This suggests that CPs improve crude protein and cellulose digestion by increasing the proportions of Solibacillus and Planococcaceae_Bacillus. Therefore, the optimal CP dietary supplementation dose for Angus cattle was 3 g·kg⁻¹ of DM. Full article

(This article belongs to the Section Cattle)

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20 pages, 7870 KB

Open AccessArticle

A New Species of Boulenophrys (Megophridae) from Mt. Hengshan, Hunan Province, China, with Re-Description on B. hengshanensis

by Dai-Yong Kuang, Yi-Fu Wei, Yi-Sha Luo, Kang-Wen Pei, Ying-Yue Cao, Meng-Fei Zhang, Tai-Fu Huang, Ling Pu and Sheng-Chao Shi

Animals 2025, 15(18), 2745; https://doi.org/10.3390/ani15182745 - 19 Sep 2025

Cited by 1 | Viewed by 1155

Abstract

Boulenophrys gutu sp. nov. was described from Mt. Hengshan, and B. hengshanensis was re-described based on the holotype and newly collected specimens using phylogenetic and morphological evidence. The new species forms an independent clade, and it is diagnosed by a combination of [...] Read more.

Boulenophrys gutu sp. nov. was described from Mt. Hengshan, and B. hengshanensis was re-described based on the holotype and newly collected specimens using phylogenetic and morphological evidence. The new species forms an independent clade, and it is diagnosed by a combination of following characters: (1) male SVL 34.4–44.7 mm (n = 7), female SVL 36.2–52.8 mm (n = 8); (2) dorsal surface of head, body, and limbs relatively smooth; (3) vomerine ridge weak, vomerine teeth absent; (4) narrow lateral fringes on toes; (5) heels moderate long, meeting when thighs are positioned at right angles to body; (6) supratympanic fold behind tympanum thick, distinctly enlarged with thickness near diameter of tympanum; (7) inner metatarsal tubercle small (IMT/SVL 4.4–5.2%); (8) several large dark brown patches along both ventrolateral sides of abdomen; (9) coloration of inner and outer metacarpal tubercle, inner metatarsal tubercle, and tip of digits ivory. Phylogenetic analyses based on 16S rRNA and COI genes revealed that B. hengshanensis is sister to B. wugongensis. Morphological comparisons showed that B. hengshanensis is diagnosed by a combination of following characters: (1) moderate body size, male SVL 34.4–38.0 mm (n = 9), female SVL 48.4 mm (n = 1); (2) weak vomerine ridge, absence of vomerine teeth; (3) tongue not notched behind; (4) a small horn-like tubercle on upper eyelid; (5) rudimentary webbing between toes; (6) narrow lateral fringes on toes; (7) heels relatively short, not meeting when thighs are positioned at right angles to body; (8) supratympanic fold behind tympanum narrow, not distinctly enlarged; (9) a pair of dark brown broad stripes along ventrolateral sides of abdomen; (10) dense creamy white dots present on lower abdomen, merge with deep brown patches without clear boundary. Full article

(This article belongs to the Special Issue Evolution, Diversity, and Conservation of Herpetofauna: Second Edition)

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12 pages, 437 KB

Open AccessFeature PaperReview

Speculative Review on the Feasibility of Porcine Circovirus 2 Elimination

by Joaquim Segalés and Marina Sibila

Animals 2025, 15(18), 2744; https://doi.org/10.3390/ani15182744 - 19 Sep 2025

Viewed by 542

Abstract

Porcine circovirus 2 (PCV2) is still infecting pigs after almost 20 years of massive vaccination all over the world. Vaccines are highly effective at counteracting the clinical signs of systemic disease caused by PCV2 and can significantly reduce the number of subclinically infected [...] Read more.

Porcine circovirus 2 (PCV2) is still infecting pigs after almost 20 years of massive vaccination all over the world. Vaccines are highly effective at counteracting the clinical signs of systemic disease caused by PCV2 and can significantly reduce the number of subclinically infected pigs. However, current vaccination programs based on one single dose in piglets are insufficient to prevent infection in a proportion of animals. Moreover, systematic vaccination of the herd changes viral epidemiology and, consequently, can cause modifications in infection timing. Such a scenario may prompt intrauterine and piglet early infections, thus facilitating viral circulation even before vaccination takes place. Considering the demonstrated high vaccine efficacy, it would be legitimate to explore the possibility of eliminating PCV2 from swine herds, but only one attempt to eliminate the virus from a herd has been published so far. The present speculative review evaluates the existing scientific literature regarding the feasibility of getting rid of this virus under commercial farm conditions. The use of PCV2 vaccination in all swine populations within a herd and the implementation of regional or national control programs are foreseen as compulsory for the eventual successful elimination of this endemic viral infection. Full article

(This article belongs to the Special Issue Pathogen Elimination in Animal Populations)

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31 pages, 1942 KB

Open AccessArticle

IECA-YOLOv7: A Lightweight Model with Enhanced Attention and Loss for Aerial Wildlife Detection

by Wenyue Ke, Tengfei Liu and Xiaohui Cui

Animals 2025, 15(18), 2743; https://doi.org/10.3390/ani15182743 - 19 Sep 2025

Viewed by 362

Abstract

Grassland ecosystems are vital for global biodiversity, yet traditional wildlife monitoring methods are often labor-intensive and costly. Although drone-based aerial surveys provide a scalable alternative, they face significant challenges such as detecting extremely small targets, handling complex backgrounds, and operating under strict computational [...] Read more.

Grassland ecosystems are vital for global biodiversity, yet traditional wildlife monitoring methods are often labor-intensive and costly. Although drone-based aerial surveys provide a scalable alternative, they face significant challenges such as detecting extremely small targets, handling complex backgrounds, and operating under strict computational constraints. To address these issues, this study proposes IECA-YOLOv7, a lightweight detection model that incorporates three key innovations: an Improved Efficient Channel Attention (IECA) module for enhanced feature representation, a content-aware CARAFE upsampling operator for improved detail recovery, and a Normalized Wasserstein Distance (NWD) loss function for robust small-target localization. Evaluated on a dedicated grassland wildlife dataset (GWAID), the model achieves a mAP@0.5 of 86.6% and a mAP@0.5:0.95 of 47.2%, outperforming the baseline YOLOv7-tiny by 2.9% in Precision and 1.8% in Recall. Furthermore, it surpasses non-YOLO architectures such as RetinaNet, EfficientDet-D0, and DETR by significant margins, demonstrating superior performance in small-object detection under complex conditions. Cross-dataset validation on VisDrone, CARPK, and DOTA demonstrates a strong generalization capability. With a model size under 5 MB, IECA-YOLOv7 effectively balances accuracy and efficiency, offering a practical solution for real-time wildlife monitoring via drones under challenging environmental constraints such as variable lighting, occlusion, and limited computational resources, thereby supporting broader conservation efforts. Full article

(This article belongs to the Section Animal System and Management)

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16 pages, 8750 KB

Open AccessArticle

Tissue Cytometry Assay with Nuclear Segmentation for Quantifying NETotic Cells in Neutrophils Stimulated by Spermatozoa in Veterinary Species

by Rodrigo Rivera-Concha, Marion León, Nikol Ponce-Rojas, Aurora Prado-Sanhueza, Pamela Uribe, Anja Taubert, Carlos Hermosilla, Raúl Sánchez and Fabiola Zambrano

Animals 2025, 15(18), 2742; https://doi.org/10.3390/ani15182742 - 19 Sep 2025

Viewed by 339

Abstract

Upon activation, neutrophils perform three distinct functions: phagocytosis, degranulation of antimicrobial substances into the extracellular medium, and release of neutrophil extracellular traps. Determination of the nuclear area expansion of neutrophils activated to release neutrophil extracellular traps has become critical in demonstrating early neutrophil [...] Read more.

Upon activation, neutrophils perform three distinct functions: phagocytosis, degranulation of antimicrobial substances into the extracellular medium, and release of neutrophil extracellular traps. Determination of the nuclear area expansion of neutrophils activated to release neutrophil extracellular traps has become critical in demonstrating early neutrophil activation and has become standard. Here, we demonstrate an automated method for measuring nuclear area expansion in two different mammalian species: canine and bovine. For both species, neutrophils were isolated from peripheral blood and co-incubated with fresh spermatozoa for up to 120 min for canine neutrophil–spermatozoa and recently thawed cryopreserved spermatozoa up to 240 min for bovine neutrophil–spermatozoa. Fluorescence images were acquired using a TissueFAXS microscope and then analyzed using StrataQuest v.7.0 software. The images show the release of neutrophil extracellular traps upon activation with spermatozoa for both species, as evidenced by the co-localization of neutrophil elastase and DNA staining. Neutrophil nuclei were expanded as early as 15 min and were detected at up to 120 min in both species. Analysis by nuclei segmentation showed that the data sets generated for both species were reliable and consistent with previously published methods. The method was developed as an automated alternative for measuring the area expansion of neutrophil nuclei in different species. Full article

(This article belongs to the Special Issue Molecular Mechanisms, Immunological, and Physiological Aspects Impacting Animal Fertility)

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21 pages, 4076 KB

Open AccessArticle

Comparative Transcriptomics of Olfactory Rosettes Reveals Expression Divergence and Adaptive Evolution in Herbivorous and Carnivorous Xenocyprididae Fishes

by Hua Xue, Hailong Gu, Liu Yang, Jingchen Chen and Wenqiao Tang

Animals 2025, 15(18), 2741; https://doi.org/10.3390/ani15182741 - 19 Sep 2025

Viewed by 384

Abstract

Olfaction plays a crucial role in fish feeding behaviors and ecological adaptation. However, systematic studies on its transcriptional regulation and molecular evolutionary mechanisms in herbivorous and carnivorous fishes remain scarce. In this study, we analyzed four Xenocyprididae species: two herbivorous (Ctenopharyngodon idella [...] Read more.

Olfaction plays a crucial role in fish feeding behaviors and ecological adaptation. However, systematic studies on its transcriptional regulation and molecular evolutionary mechanisms in herbivorous and carnivorous fishes remain scarce. In this study, we analyzed four Xenocyprididae species: two herbivorous (Ctenopharyngodon idella and Megalobrama amblycephala) and two carnivorous (Elopichthys bambusa and Culter alburnus), using olfactory rosette transcriptome sequencing and cross-species comparisons. The number of unigenes per species ranged from 40,229 to 42,405, with BUSCO completeness exceeding 89.2%. Functional annotation was performed using six major databases. Olfactory-related candidate genes were identified based on Pfam domains (7tm_4) and KEGG pathways (ko04740), revealing 8–19 olfactory receptor genes per species. These candidate genes were predominantly enriched in the olfactory transduction and neuroactive ligand–receptor interaction pathways. A total of 3681 single-copy orthologous genes were identified, and their expression profiles exhibited clear interspecific divergence without forming strict clustering by dietary type. High-threshold differentially expressed trend genes (|log₂FC| ≥ 4) were enriched in pathways related to RNA processing, metabolite transport, and xenobiotic metabolism, suggesting that the olfactory system may participate in diverse adaptive responses. Ka/Ks analysis indicated that most homologous genes were under purifying selection, with only 0.87–2.07% showing positive selection. These positively selected genes were enriched in pathways related to immune response and neural regulation, implying potential roles in adaptive evolution associated with ecological behavior. Furthermore, the olfactory-related gene oard1 exhibited Ka/Ks > 1 in the E. bambusa vs. C. idella comparison. qRT-PCR validation confirmed the reliability of the RNA-Seq data. This work is the first to integrate two complementary indicators—expression trends and evolutionary rates—to systematically investigate the transcriptional regulation and molecular evolution of the olfactory system in Xenocyprididae species under the context of dietary differentiation, providing valuable reference data for understanding the perceptual basis of dietary adaptation in freshwater fish. Full article

(This article belongs to the Section Aquatic Animals)

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24 pages, 1536 KB

Open AccessArticle

Effects of a Multi-Strain Lactic and Propionic Acid Bacteria Inoculant on Silage Quality, Methane Emissions, Milk Composition, and Rumen Microbiome

by Segun Olorunlowu, Pola Sidoruk, Julia Sznajder, Jakub Szczesny, Dorota Lechniak, Piotr Pawlak, Marcin Ryczek, Haihao Huang, Lingyan Li, Agung Irawan, Jolanta Komisarek, Malgorzata Szumacher-Strabel and Adam Cieslak

Animals 2025, 15(18), 2740; https://doi.org/10.3390/ani15182740 - 19 Sep 2025

Viewed by 535

Abstract

Ensiling grass with microbial inoculants is a promising strategy to enhance forage quality, animal performance, and environmental sustainability. This study evaluated the effects of a multi-strain inoculant (Lactobacillus plantarum, L. buchneri, Propionibacterium acidipropionici, and P. thoeni) on silage [...] Read more.

Ensiling grass with microbial inoculants is a promising strategy to enhance forage quality, animal performance, and environmental sustainability. This study evaluated the effects of a multi-strain inoculant (Lactobacillus plantarum, L. buchneri, Propionibacterium acidipropionici, and P. thoeni) on silage fermentation, nutrient digestibility, milk production, methane emissions, and rumen microbiota in dairy cows. In a 2 × 2 crossover design, 24 lactating Polish Holstein–Friesians were fed total mixed rations differing only in grass silage treated with or without inoculant. Inoculated silage had lower pH (4.56 vs. 5.06; p = 0.02) and higher crude protein (129 vs. 111 g/kgDM; p < 0.05). Cows fed inoculated silage showed higher ruminal propionate (28.3 vs. 26.3 mM; p = 0.03), reduced ammonia (7.61 vs. 8.67 mM; p = 0.02), and fewer protozoa (1.21 vs. 1.66 × 10⁵/mL; p = 0.03). Nutrient digestibility improved (p < 0.05), while methane emissions declined both per cow (368 vs. 397 g/d; p = 0.01) and per kgDMI (15.1 vs. 16.5; p = 0.01). Milk yield increased (p = 0.04), and the fatty acid profile improved. Our study revealed that cows fed inoculated silage had higher nutrient digestibility, lower methane emissions, and microbial shifts in the rumen detected by 16S rRNA sequencing (p < 0.05). Full article

(This article belongs to the Special Issue Impacts of Silage-Based Forages on Ruminant Health and Welfare)

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60 pages, 1328 KB

Open AccessReview

A Comprehensive Review: Current Strategies for Detoxification of Deoxynivalenol in Feedstuffs for Pigs

by Changning Yu, Peter Plaizier, Joshua Gong, Chengbo Yang and Song Liu

Animals 2025, 15(18), 2739; https://doi.org/10.3390/ani15182739 - 19 Sep 2025

Viewed by 660

Abstract

Deoxynivalenol (DON) is a mycotoxin produced by Fusarium graminearum and Fusarium culmorum, primarily contaminating wheat, corn, and their derivatives. Although less toxic than some other mycotoxins, DON is significant due to its abundance, particularly in wheat, and its resistance to in vitro [...] Read more.

Deoxynivalenol (DON) is a mycotoxin produced by Fusarium graminearum and Fusarium culmorum, primarily contaminating wheat, corn, and their derivatives. Although less toxic than some other mycotoxins, DON is significant due to its abundance, particularly in wheat, and its resistance to in vitro detoxification. Compared to other livestock, pigs are the most susceptible animals to DON’s presence due to its interaction with wheat-heavy diets. The best way to attenuate the effects of DON is to prevent its presence on cultivated land; however, given that it is difficult to prevent the occurrence of DON in freshly harvested cereals completely, other strategies must be applied. For pigs, chronic consumption of feed with DON concentrations below even the regulated limits can still negatively impact growth. Thus, DON-detoxifying agents including antidotes, DON binding agents, and exogenous in vivo enzyme additives, have been added to feed in an attempt to detoxify DON. This comprehensive review highlights the most recently developed DON-detoxification techniques and assesses each treatment’s viability and efficacy, with an emphasis on chemical and biochemical techniques and the rapid development of artificial intelligence (AI) technologies. Herein we explore the unmet needs and future directions of current technologies while discussing promising strategies that can advance the DON-detoxification field. Full article

(This article belongs to the Special Issue Advances in Pig Microbiome: Gut Influences and Beyond)

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18 pages, 2691 KB

Open AccessArticle

YOLOv8-DMC: Enabling Non-Contact 3D Cattle Body Measurement via Enhanced Keypoint Detection

by Zhi Weng, Wenwen Hao, Caili Gong and Zhiqiang Zheng

Animals 2025, 15(18), 2738; https://doi.org/10.3390/ani15182738 - 19 Sep 2025

Viewed by 419

Abstract

Accurate and non-contact measurement of cattle body dimensions is essential for precision livestock management. This study presents YOLOv8-DMC, a lightweight deep learning model optimized for anatomical keypoint detection in side-view images of cattle. The model integrates three attention modules—DRAMiTransformer, MHSA-C2f, and CASimAM—to improve [...] Read more.

Accurate and non-contact measurement of cattle body dimensions is essential for precision livestock management. This study presents YOLOv8-DMC, a lightweight deep learning model optimized for anatomical keypoint detection in side-view images of cattle. The model integrates three attention modules—DRAMiTransformer, MHSA-C2f, and CASimAM—to improve robustness under occlusion and lighting variability. Following keypoint prediction, a 16-neighborhood depth completion and pass-through filtering process are applied to generate clean, colored point clouds. This enables precise 3D localization of keypoints by matching them to valid depth values. The model achieves

A P_{@ 0.5}

of 0.931 and AP@[0.50:0.95] of 0.868 on a dataset of over 7000 images, improving baseline accuracy by 2.14% and 3.09%, respectively, with only 0.35 M additional parameters and 0.9 GFLOPs in complexity. For real-world validation, strictly lateral-view RGB-D images from 137 cattle were collected, with ground-truth manual measurements. Compared with manual measurements, the average relative errors are 2.43% for body height, 2.26% for hip height, 3.65% for body length, and 4.48% for cannon circumference. The system supports deployment on edge devices, providing an efficient and accurate solution for 3D cattle measurement in real-world farming conditions. Full article

(This article belongs to the Section Cattle)

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18 pages, 2902 KB

Open AccessFeature PaperArticle

Comparative Assessment of Sperm Morphology in Liquid-Preserved Boar Semen Using Cytological Stains

by Annika Braune, Axel Wehrend, Johannes Kauffold and Abbas Farshad

Animals 2025, 15(18), 2737; https://doi.org/10.3390/ani15182737 - 19 Sep 2025

Viewed by 553

Abstract

Accurate assessment of sperm morphology is essential for artificial insemination using liquid-preserved boar semen. This study compared nine commonly used staining techniques, eosin, eosin–nigrosin, Diff-Quick^®, Hemacolor^®, Sangodiff-G^®, Spermac^®, Formol–Citrate–Rose Bengal stain, Testsimplets^®, and [...] Read more.

Accurate assessment of sperm morphology is essential for artificial insemination using liquid-preserved boar semen. This study compared nine commonly used staining techniques, eosin, eosin–nigrosin, Diff-Quick^®, Hemacolor^®, Sangodiff-G^®, Spermac^®, Formol–Citrate–Rose Bengal stain, Testsimplets^®, and Methyl Violet, based on morphological assessment, cost, time efficiency, and storage stability. Each staining method was applied to 36 slides, totaling 324 samples, and evaluated four times each (1296 evaluations). Slides were analyzed four times: immediately after staining and after 1 day, 1 week, and 3 months of storage. The results indicated that Eosin was the fastest and most cost-effective method, providing strong contrast, though it increased detection of structural alterations. Eosin–nigrosin offered detailed morphology but formed colored crystals over time. Diff-Quick^® and Hemacolor^® showed good initial performance, but Hemacolor^® lost pigment clarity after 3 months (p = 0.0273). Sangodiff-G^® had poor contrast and reduced detection of abnormalities (p = 0.00229). Spermac^® delivered high contrast but was time-consuming. Formol–Citrate–Rose Bengal stain required extensive preparation and showed significant post-storage changes (p < 0.0001). Testsimplets^®, despite their high cost, suffered from declining interpretability (p < 0.0001). Methyl Violet lacked sufficient resolution and was highly unstable over time (p < 0.0001). In conclusion, Eosin emerged as the most practical and economical staining method for routine morphological evaluation of liquid-preserved boar semen. While eosin–nigrosin was also effective, its storage instability limits broader application. Other methods showed specific weaknesses, emphasizing the need to tailor stain selection to laboratory goals and constraints. Full article

(This article belongs to the Section Pigs)

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Animals, Volume 15, Issue 18 (September-2 2025) – 130 articles

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