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Precise recognition of individual ovine specimens plays a pivotal role in implementing smart agricultural platforms and optimizing herd management systems. With the development of deep learning technology, sheep face recognition provides an efficient and contactless solution for individual sheep identification. However, with the growth of sheep, their facial features keep changing, which poses challenges for existing sheep face recognition models to maintain accuracy across the dynamic changes in facial features over time, making it difficult to meet practical needs. To address this limitation, we propose the lifelong biometric learning of the sheep face network (LBL-SheepNet), a feature decoupling network designed for continuous adaptation to ovine facial changes, and constructed a dataset of 31,200 images from 55 sheep tracked monthly from 1 to 12 months of age. The LBL-SheepNet model addresses dynamic variations in facial features during sheep growth through a multi-module architectural framework. Firstly, a Squeeze-and-Excitation (SE) module enhances discriminative feature representation through adaptive channel-wise recalibration. Then, a nonlinear feature decoupling module employs a hybrid channel-batch attention mechanism to separate age-related features from identity-specific characteristics. Finally, a correlation analysis module utilizes adversarial learning to suppress age-biased feature interference, ensuring focus on age-invariant identifiers. Experimental results demonstrate that LBL-SheepNet achieves 95.5% identification accuracy and 95.3% average precision on the sheep face dataset. This study introduces a lifelong biometric learning (LBL) mechanism to mitigate recognition accuracy degradation caused by dynamic facial feature variations in growing sheep. By designing a feature decoupling network integrated with adversarial age-invariant learning, the proposed method addresses the performance limitations of existing models in long-term individual identification. Full article

Wool has distinctive biological, physical, and chemical properties that contribute to its value both for the sheep and in global fibre and textile markets. Its fibres are primarily composed of proteins, principally keratin and keratin-associated proteins (KAPs). To better comprehend the genes that underpin key wool traits, this study examined the keratin-associated protein 36-1 gene (KRTAP36-1) in Chinese Tan lambs. We identified three previously reported alleles of the gene (named A, B and C) that were present in the lambs studied, with genotype frequencies as follows: 2.0% (n = 5; AA), 6.9% (n = 17; AB), 13.8% (n = 34; AC), 8.9% (n = 22; BB), 33.4% (n = 82; BC) and 35.0% (n = 86; CC). The frequencies of the individual alleles in the Chinese Tan lambs were 12.4%, 29.1% and 58.5% for alleles A, B and C, respectively. The three alleles were in Hardy–Weinberg Equilibrium. In an association analysis, it was revealed that allele C was associated with variation in the mean fibre curvature of the fine wool of the Chinese Tan lambs, but this association was not observed in their heterotypic hair fibres. This finding suggests that KRTAP36-1 might be differentially expressed in the wool follicles that produce the two fibre types, and that along with other KRTAP genes, it may be involved in determining fibre curvature and the distinctive curly coat of the lambs. Full article

To achieve the goals of productivity and sustainability across diverse livestock systems, reproductive factors play a pivotal role. Historically, reproductive research has primarily focused on females, as they are responsible for maintaining pregnancy and delivering offspring following oocyte fertilization. However, since the early 2000s, the biological significance of sperm RNAs has been increasingly recognized in various livestock species. These RNAs contribute both genetically and epigenetically at the time of fertilization and during early embryonic development. Multiple types of sperm RNA have been identified in bovine, porcine, ovine, buffalo, and caprine spermatozoa. Notably, transcriptomic profiling has shown potential to differentiate between high- and low-fertility males, even when conventional semen quality values appear normal in both groups. This opens the possibility for more accurate identification of highly fertile sires. Nevertheless, a definitive marker or set of markers has yet to be established, likely due to the transcriptome’s sensitivity to environmental conditions and to the variability in evaluation methodologies. Therefore, global scientific efforts should aim to establish standardized, robust protocols, as sperm RNA represents a promising avenue for enhancing the sustainability of animal production systems. Full article

Seasonal reproduction in sheep reduces reproductive efficiency. Melatonin (MT) plays a crucial role in reproductive processes. The purpose of this study was to assess the effects of a 5-day MT implant pretreatment on estrus synchronization and reproductive performance in sheep during seasonal anestrus. A total of 40 multiparous Mongolian sheep were selected and randomly divided into two groups. In the MT group (n = 20), the ewes received an MT implant for 5 days, and then, they were given a progesterone (P4)-containing vaginal sponge for 14 days with equine chorionic gonadotropin (eCG) administered (330 I.U. per ewe; I.M.) at sponge removal. Control (CON) ewes (n = 20) were similarly treated but did not receive MT implants. The results demonstrated that MT implantation significantly improved serum levels of total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), and glutathione peroxidase (GSH-Px), increased post-ovulatory luteal diameter and serum P4 levels, and reduced ovarian apoptosis. Compared with the CON group, the MT group showed significantly higher pregnancy (68.23% vs. 50.59%) and lambing rates (63.53% vs. 47.06%; number of lambed ewes/number of total ewes) following cervical-timed artificial insemination. Ovarian transcriptome analysis revealed 522 differentially expressed genes (DEGs) in the MT group compared with the CON group, including 355 upregulated and 167 downregulated DEGs. In addition, MT significantly enhanced proliferation and inhibited apoptosis in cultured granulosa cells (GCs) and luteal cells (LCs) in vitro. Moreover, it enhanced the antioxidant capacity of GCs and LCs probably by activating the NRF2 signaling pathway as well as stimulating steroid hormone synthesis. In conclusion, MT implantation 5 days before applying the conventional P4-eCG protocol enhances ovine reproductive outcomes during seasonal anestrus. MT implantation has a beneficial role on the growth and function of ovarian cells. These findings offer novel evidence supporting the functional role of MT in mammalian reproduction, and would be informative for optimizing estrus synchronization in sheep. Full article

A novel composite patch osteosynthesis technique (CPT) has demonstrated promising ex vivo biomechanical performance in small tubular bones. To bridge the gap toward clinical evaluations, this study compared the stability of the CPT to a stainless-steel locking plate (LP) in an experimental in vivo ovine bilateral proximal phalanx fracture model. Eight sheep underwent a midline osteotomy with a 4.5 mm circular unicortical defect in the lateral proximal phalanx of both front limbs, treated with the CPT (n = 8) or the LP (n = 8). A half-limb walking cast, or a custom off-loading hoof shoe, was used for postoperative protection. Implant stability was assessed by post-surgery X-ray evaluations and post-euthanasia (16 weeks) dual-energy X-ray absorptiometry (DXA). At week one, all CPT implants demonstrated mechanical failure, while all LPs remained overall intact. Mean BMD was 0.45 g/cm² for CPT and 0.60 g/cm² for LP in the fracture area (p = 0.078), and 0.37 g/cm² vs. 0.41 g/cm² in the distal epiphysis (p = 0.016), respectively. In conclusion, the CPT demonstrated indications of inferior stability compared to the LP in this fracture model, which may limit its clinical applicability in weight-bearing or high-load scenarios and in non-compliant patients. Full article

This study identified four new keratin-associated protein genes (KRTAP19-n) in sheep: sKRTAP19-1, sKRTAP19-2, sKRTAP19-4, and sKRTAP19-6. These genes are closely related to the previously identified sheep genes KRTAP19-3 and KRTAP19-5, as well as to human KRTAP19-n genes. However, no clear orthologous relationships were found, suggesting complex evolutionary dynamics for this gene family. Extensive nucleotide sequence variation was observed across the four genes. sKRTAP19-1 had four variants, defined by four synonymous single-nucleotide polymorphisms (SNPs) and a variable number of “GGCTAC” hexanucleotide repeats. sKRTAP19-2 had five variants involving seven SNPs, three of which were non-synonymous. sKRTAP19-4 had five variants with nine SNPs (three being non-synonymous) and a three-nucleotide deletion. sKRTAP19-6 had eight variants, defined by 13 SNPs and a two-nucleotide consecutive substitution, with four of the SNPs being non-synonymous. One distinct variant each of sKRTAP19-4 and sKRTAP19-6 was found exclusively in Yanchi Tan sheep, with seven unique nucleotide differences compared to other variants. These unique variants were identical to the Romanov sheep genome in the region amplified (excluding the primer binding regions), suggesting a shared ancestral origin. The findings highlight considerable genetic diversity in ovine KRTAP19-n and lay a foundation for future research into their role in regulating wool fibre characteristics. Full article

Ovine herpesvirus 2 (OvHV-2) causes the fatal veterinary disease malignant catarrhal fever (MCF). Fusion is an essential step in the host cell entry of enveloped viruses and is an important target for vaccine development. OvHV-2 cannot be propagated in vitro, so a robust virus-free cell–cell membrane fusion assay is necessary to elucidate its entry mechanism. OvHV-2 cell–cell fusion requires three conserved herpesviral envelope glycoproteins: gB, gH, and gL. OvHV-2 fusion activity is detectable but low. We hypothesize that enhancing the cell surface expression of gB, which is the core herpesviral fusogen, will increase cell–cell fusion. We generated C-terminal truncation mutants of gB and determined their cell surface expression, subcellular distribution, and fusion activity. Two mutants, including one that lacked the entire cytoplasmic tail domain, failed to function in the cell–cell fusion assay, despite wild-type levels of surface expression. This suggests that the OvHV-2 gB cytoplasmic tail is critical for fusion. A gB mutant truncated at amino acid 847 showed increased surface expression and fusion relative to the wild type. This suggests that the robust fusion activity of gB847 is the result of increased surface expression. gB847 may be used in place of wild-type gB in an improved, more robust OvHV-2 fusion assay. Full article

Ιn this study, we evaluated the genetic resources of nine Greek sheep breeds. The genotyping data of 292 animals were acquired from Illumina’s OvineSNP50 Genotyping BeadChip. The genetic diversity and inbreeding levels were evaluated using the observed and expected heterozygosity indices, the F_IS inbreeding coefficient, and runs of homozygosity (ROH). The genetic differentiation of breeds was assessed using the F_ST index, whereas their population structure was analyzed using admixture and principal components analysis (PCA). Historical recombination patterns and genetic drift were evaluated based on linkage disequilibrium, effective population sizes, and gene flow analysis to reveal migration patterns. PCA revealed distinct clusters mostly separating mountainous, insular, and lowland breeds. The F_ST value was the lowest between Serres and Karagouniko breeds (0.050). Admixture analysis revealed a genetic substructure for Serres and Kalarritiko breeds, while Chios, followed by Katsika, demonstrated the highest within-breed genetic uniformity. ROH analysis revealed low levels of inbreeding for all breeds. Genetic introgression from both Anatolia and Eastern Europe has been evidenced for Greek sheep breeds. The results also revealed that Greek sheep breeds maintain adequate levels of genetic diversity, without signs of excessive inbreeding, and can serve as valuable resources for the conservation of local biodiversity. Full article

The assessment of milk coagulation properties (MCPs) is crucial for enhancing goat cheese production and quality. In this study, 501 bulk goat milk samples were collected from various farms to evaluate the MCPs. Traditionally, cheesemaking aptitude is evaluated using lactodynamographic analysis, a reliable but time-consuming laboratory method. Mid-infrared spectroscopy (MIRS) offers a promising alternative for the large-scale prediction of goat milk’s technological traits. Reference MCP measurements were paired with mid-infrared spectra, and prediction models were developed using partial least squares regression, with accuracy evaluated through cross- and external validation. The ability of MIRS to classify milk samples by coagulation aptitude was evaluated using partial least squares discriminant analysis. Only the model for rennet coagulation time obtained sufficient accuracy to be applied for screening (R²_CrV = 0.68; R²_Ext = 0.66; RPD = 2.05). Lower performance was observed for curd-firming time (R²_CrV = 0.33; R²_Ext = 0.27; RPD = 1.42) and curd firmness (R²_CrV = 0.55; R²_Ext = 0.43; RPD = 1.35). Classification of high coagulation aptitude achieved balanced accuracy values of 0.81 (calibration) and 0.74 (validation). With further model refinement and larger calibration datasets, MIRS may become a resource for the dairy-goat sector to monitor and improve milk suitability for cheesemaking. Full article

The increasing number of acetabular revision total hip arthroplasties requires the evaluation of alternative materials in addition to established standards using a defined animal experimental defect that replicates the human acetabular revision situation as closely as possible. Defined bone defects in the load-bearing area of the acetabulum were augmented with various materials in an ovine periacetabular defect model (Group 1: NanoBone^® (artificial hydroxyapatite-silicate composite; Artoss GmbH, Germany); Group 2: autologous sheep cancellous bone; Group 3: Tutoplast^® (processed allogeneic sheep cancellous bone; Tutogen Medical GmbH, Germany)) and bridged with an acetabular reinforcement ring of the Ganz type. Eight months after implantation, a μ-CT examination (n = 8 animals per group) was performed. A μ-CT analysis of the contralateral acetabula (n = 8, randomly selected from all three groups) served as the control group. In a defined volume of interest (VOI), bone volume (BV), mineral volume (MV), and bone substitute volume (BSV), as well as the bone surface (BS) relative to the total volume (TV) and the surface-to-volume ratio (BS/BV), were determined. To assess the bony microarchitecture, trabecular thickness (Tb.Th), trabecular separation (Tb.Sp), and trabecular number (Tb.N), as well as connectivity density (Conn.D), the degree of anisotropy (DA), and the structure model index (SMI), were evaluated. The highest BV was observed for NanoBone^® (Group 1), which also showed the highest proportion of residual bone substitute material in the defect. This resulted in a significant increase in BV/TV with a significant decrease in BS/BV. The assessment of the microstructure for Groups 2 and 3 compared to Group 1 showed a clear approximation of Tb.Th, Tb.Sp, Tb.N, and Conn.D to the microstructure of the control group. The SMI showed a significant decrease in Group 1. All materials demonstrated their suitability by supporting biological defect reconstruction. NanoBone^® showed the highest rate of new bone formation; however, the microarchitecture indicated more advanced bone remodeling and an approximate restoration of the trabecular structure for both autologous and allogeneic Tutoplast^® cancellous bone when using the impaction bone grafting technique. Full article

Mesenchymal stem cells (MSCs) represent a promising cell source for cartilage tissue engineering due to their chondrogenic potential. However, current differentiation protocols result in limited efficiency. This study assessed the combined effects of transforming growth factor-beta 3 (TGF-β3) and fibroblast growth factor-2 (FGF-2) on the morphology, proliferation, chondrogenic differentiation, chondrogenic gene expression, and cytokine profile of ovine bone marrow-derived MSCs (BM-MSCs). BM-MSCs were cultured under four conditions: control (αMEM) or αMEM supplemented with FGF-2, TGF-β3, or TGF-β3 + FGF-2. Morphological and proliferation analyses, Alcian blue staining in 2D and 3D, and real-time PCR for early (Chad, Comp, and Sox 5) and late (Agg, Col IX, Sox 9, and Fmod) chondrogenic markers were performed. Cytokine secretion profiles were analyzed using multiplex assay. TGF-β3 induced morphological changes indicative of early chondrogenesis, while FGF-2 enhanced proliferation. The combination of both cytokines led to a synergistic increase in cell proliferation, early and late chondrogenic gene expression, and glycosaminoglycans (GAG) deposition. Cytokine analysis revealed that TGF-β3 enhanced the immunomodulatory and angiogenic profile of BM-MSCs, whereas co-treatment with FGF-2 yielded a balanced and potentially regenerative secretome. Dual stimulation with TGF-β3 and FGF-2 significantly improves the chondrogenic differentiation of ovine BM-MSCs by enhancing both molecular and functional markers of cartilage formation. Full article

Tusavirus 1 of species Protoparvovirus incertum 1 (family Parvoviridae) was first identified in humans and later in small ruminants (caprine and ovine). This study reports the full-length coding sequences (~4400–4600 nt) of three novel tusavirus-related protoparvoviruses from ovine (“misavirus”, PV540792), for the first time bovine (“sisavirus”, PV540793) and subsequently from caprine (“gisavirus” PV540850/51) fecal samples, using next-generation sequencing (NGS) and PCR techniques. Their NS1, VP1 and VP2 proteins shared 61–63% amino acid identities with each other and with tusaviruses, suggesting these three viruses belong to three novel species in the genus Protoparvovirus. Phylogenetic analyses placed them with tusaviruses on a separate main branch, implying a shared origin among these most likely ruminant protoparvoviruses. A small-scale epidemiological investigation on 318 ruminant enteric samples using novel generic NS1 primers found misavirus in 14/51 (27.5%) ovine and sisavirus in 19/203 (9.4%) bovine samples from multiple Hungarian farms. Tusavirus was present in 5/51 (9.8%) ovine and 15/62 (24.2%) caprine samples, all from one farm. The highest prevalences for all three viruses were found in animals aged 2–12 months, though sporadic cases were also found in other age groups. Partial NS and VP sequence-based phylogenetic trees showed virus-specific lineages for misa-, sisa-, gisa- and tusaviruses, with various strains forming sub-lineages. These findings suggest the presence of multiple genotypes and/or members of additional species, which was supported by a VP sequence-based hierarchical cluster analysis. The study’s viruses were mostly phylogenetically separated by host; however, two bovine sisavirus strains with diverse phylogenetic localizations in the NS (belonging to bovine sisaviruses) and VP1 trees (distantly related to ovine misaviruses) could indicate previous (interspecies?) recombination events. Full article

Objectives: The study investigates runs of homozygosity (ROH) and heterozygosity (ROHet), and their patterns in nine sheep breeds (772 animals in total) maintained in Poland (native and conserved), corresponding to their genetic diversity, inbreeding levels, and selection signatures. Methods: Genotypes were obtained using the Illumina OvineSNP50 BeadChip and quality-filtered SNPs were used to detect ROH and ROHet segments with the detectRUNS R package, following stringent parameters for segment length, SNP density, and genotype quality. Results: Significant variation in ROH characteristics was observed across breeds. Short ROH segments were predominant in all breeds, indicating historical inbreeding events. In contrast, longer ROH segments signified recent inbreeding, particularly in Swiniarka (SW) and Polish Merino of Colored Variety (MPC). The ROH-based genomic inbreeding coefficient (F_ROH) varied across breeds, with SW exhibiting the highest levels, suggesting reduced genetic diversity. ROHet analysis revealed that Uhruska (UHR) had the highest heterozygous segments span, while Black-headed (BH) sheep exhibited the lowest ROHet extent. ROH islands identified across breeds revealed regions under selection, associated with traits such as reproductive performance, wool quality, and body condition. Genes located within these islands (e.g., U6, SPP1, ABCG2) were linked to economically significant traits including milk production, growth, and carcass quality. Conclusions: The presented results highlight the genetic adaptations shaped by selection pressures, while also providing insights into the genetic architecture of sheep breeds maintained in Poland. Full article

The increasing interest in fermented foods stems from their health benefits, mediated by foodborne microorganisms. This study aimed to characterize the fermentative microbiota of Pecorino di Picinisco, a traditional Italian cheese made from ovine raw milk, and to evaluate the probiotic and technological potential of selected lactic acid bacteria strains. Three strains representative of the different species found (Lactococcus lactis, Lactiplantibacillus plantarum and Latilactobacillus curvatus) were chosen and analyzed. All three strains were able to adhere to human intestinal Caco-2 cells, were resistant to simulated in vitro digestion and significantly prolonged the lifespan of Caenorhabditis elegans, used as a simplified in vivo model, with respect to the commercial probiotic strain Lacticaseibacillus rhamnosus GG. The L. plantarum Pic37.4 strain was particularly promising; therefore, its cell-free supernatant was employed to evaluate the antimicrobial activity against indicator strains of foodborne and intestinal pathogens or spoilage bacteria. The results demonstrated the effectiveness of the supernatant against all strains tested, with the strongest effect on the intestinal pathogen enterotoxigenic Escherichia coli K88. In addition, the inhibitory effect on pathogen adhesion to intestinal mucosa was investigated on Caco-2 cells, resulting in a significant reduction in adhesion mediated by the L. plantarum Pic37.4 supernatant. The antimicrobial properties of the L. plantarum strain were confirmed in vivo in C. elegans. These promising results lay the ground for further investigations aimed at substantiating the probiotic and technological potential of the L. plantarum Pic37.4 investigated in this work. Full article

Background/Objective: Virus-based vaccine vectors have been widely utilised in commercial vaccines, predominantly for virus infections. They also offer promise for bacterial diseases, for which many vaccines are sub-optimal or ineffective. It is well-established for chlamydial infections, including ovine enzootic abortion, that the major outer membrane protein (MOMP) antigen is protective. Immune responses strongly associated with controlling Chlamydiae include cellular interferon-gamma (IFN-γ) production. Methods: A study was conducted to compare the ability of a modified Orf virus vector directly with a modified sheep maedi visna virus vector to deliver the C. abortus antigen ompA and stimulate vaccine-induced responses in sheep. The Orf virus-based vaccine (mORFV-ompA) was found to be more effective in stimulating MOMP-specific antibodies and cellular antigen-driven IFN-γ in immunised sheep. This mORFV-ompA vaccine was assessed in a follow-up immunogenicity investigation in sheep, where the cellular and humoral immune responses elicited following immunisation with the live or inactivated vaccine were determined. Sheep were immunised intramuscularly with a live mORFV-ompA (n = 10) or an inactivated mORFV-ompA (n = 10). An additional group of 10 sheep served as unvaccinated controls. Results: Serological anti-MOMP antibodies and cellular recall responses of peripheral blood mononuclear cells to the native C. abortus antigen were assessed. Immunisation with either the live or inactivated mORFV-ompA-induced anti-MOMP immunoglobulin-G. Antigen-specific cellular responses, characterised by the secretion of IFN-γ and interleukin (IL)-17A, with negligible IL-10 and no IL-4, were detected in lymphocyte stimulation assays from both mORFV groups. No antibody responses to the mORFV platform were detected following immunisations. Conclusions: Both live and inactivated vaccines have the potential to be a platform technology for deployment in sheep. This addresses a notable gap in veterinary vaccine development where the induction of both humoral responses and cellular responses is required without using an adjuvant. The successful use of the MOMP candidate antigen suggests potential utility for bacterial disease deployment. Full article