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Linkage disequilibrium (LD) maps are fundamental tools for exploring the genetic basis of traits of interest in any species. Quantifying LD patterns in cattle breeds has been made possible due to the availability of huge quantities of SNPs through modern sequencing technology. The present research aims to determine and compare linkage disequilibrium levels at different distances on the genome of Sistani domestic cattle and subspecies of Bos taurus and Bos indicus based on genome SNP data. A total of 60 Bos indicus Sahiwal (dairy) and Nellore (beef), Bos taurus Hereford (beef) and Holstein (dairy), and Sistani cattle were sampled and genotyped using Illumina Bovine HD 770 k chip. To ensure the caliber of the sequencing, 10 samples (genetically sequenced cattle) were randomly chosen among all breeds represented. LD was evaluated at distances of 1–50 Kb, 50–100 Kb, 100–500 Kb, and 0.5–1 Mb, and average r² values for all autosomes were calculated within distance classes. For all breeds, the average r² was over 0.2 at distances less than 100 Kb, while for Sistani, Nellore, and Sahiwal, the average r² was above 0.2 between 100 and 500 Kb. Furthermore, for all breeds, the average r² exceeding 0.3 was noted at distances smaller than 50 Kb, while this amount for Holstein and Hereford was observed at distances between 50 and 100 Kb. In various breeds, greater changes in LD levels were observed (at <10 Kb distance). In this study, the Sistani breed showed LD decay patterns similar to indicine cattle (Nellore and Sahiwal), which may be due to the geographic proximity of the Sistan and Baluchestan province to Pakistan, the origin of indicine breeds (they may have had genetic or kinship relationships over a long historical period), or due to ascertainment bias in the SNP chips used. Full article

The digestion and absorption properties of 1,3-dipalmitoyl-2-oleoyl glycerol (POP)-rich lipids was evaluated using in vitro gastrointestinal digestion and a Caco-2 cell-mediated coupled model. Caco-2 cell viability and monolayer integrity were assessed by an MTT assay and transepithelial electrical resistance. The IC50 for bile salts, pancreatin, and free fatty acid (FFA) were 0.22 mM, 0.22 mg/mL, and 1.47 mM, respectively, and no cytotoxicity was observed for bovine serum albumin (0.01–0.20 mM) or triacylglycerol (1.00–10.00 mM). The in vitro-digested POP-rich lipid containing FFA > 2.95 mM caused the disruption of monolayer tight junctions in Caco-2 cells. The major triacylglycerols (TAG) of POP-rich lipids were POP (50.8%), POO (17.8%), POL/OPL/PLO (7.6%), PPO (7.1%), and PLP (6.8%). Following digestion and uptake into Caco-2 cells, the resynthesized TAGs included PPO (20.6%), PPP (15.9%), POO (14.0%), POL/OPL/PLO (12.2%), POP (10.9%), OOO (7.5%), OPO (7.0%), OOL/OLO (6.7%), PLP (3.1%), and PPL (2.2%). The secreted major TAGs were POL/OPL/PLO (50.8%), PPP (11.1%), and OOL/OLO (8.4%), indicating a diverse TAG profile in newly synthesized lipids. This study provides a coupled model for lowering cytotoxicity and maintaining the monolayer in Caco-2 cells, and for evaluating the digestion and absorption properties of functional lipids containing specific fatty acids incorporated into TAG. Full article

Kobuviruses (KoVs) are a group of small, non-enveloped RNA viruses classified in the genus Kobuvirus within the Picornaviridae family, comprising Aichivirus species A to F. KoVs have been identified in humans and several mammals, including domestic ungulates. This study investigated the presence of KoVs in a collection of bovine stool samples (n = 38) obtained from animals with enteritis or without clinical signs. By RT-PCR screening, KoV RNA was detected in 10/ 38 animals (26.3%). Six of the ten positive animals had enteric signs. On sequence analysis of the amplicons, eight strains were related to species Aichivirus B, commonly identified in cattle. In contrast, two strains (ITA/2019/572-1 and ITA/2020/bovine/30-2), displayed the highest nt identity (up to 97.1%) to cattle, yak, and goat Aichivirus D strains. On whole genome analysis, strains ITA/2019/572-1 and ITA/2020/30-2 showed 88.9% nt identity to each other and 87.8–90.3% nt to the bovine kobuvirus strain CHN/2021/ON730709 identified in China. Interestingly these three Aichivirus D strains showed a recombinant makeup, clustering with D1 genotype in the capsid region and with D2 genotype in the non-structural genes. These findings suggest that Aichivirus D KoVs are common components of livestock virome. Understanding the genetic diversity of KoVs in animals will be useful to improve the diagnostics and gather epidemiological data. Full article

Non-bovine dairy animals, commonly referred to as non-traditional dairy species, include goats, sheep, yaks, buffalo, donkeys, alpacas, llamas, and other less commonly farmed species. These animals have been integral to livestock systems since ancient times, providing milk and other essential products. Despite their historical significance, dairy production from many of these species remains predominantly confined to rural areas in developing countries, where scientific advancements and technical improvements are often limited. As a consequence of this, the scientific literature and technological developments in the processing and characterization of dairy products from these species have lagged behind those for cow’s milk. This review aims to compile and analyze existing research on dairy products derived from non-traditional animals, focusing on their molecular characteristics, including proteins (alpha, beta, kappa, and total casein), fats (cholesterol and total fat), lactose, albumin, ash, total solids, and somatic cell count, among others, for each of these species. Additionally, we discuss emerging technologies employed in their processing, encompassing both non-thermal methods (such as high-pressure processing, pulsed electric fields, ultrasound processing, UV-C irradiation, gamma radiation, microfiltration, and cold plasma processing) and thermal methods (such as ohmic heating). This review also explores the specific potential applications and challenges of implementing these technologies. By synthesizing recent findings, we aim to stimulate further research into innovative technologies and strategies that can enhance the quality and yield of non-bovine dairy products. Understanding the unique properties of milk from these species may lead to new opportunities for product development, improved processing methods, and increased commercialization in both developing and developed markets. Full article

In the context of revitalizing the use of traditional plant species as remarkable sources of bioactive compounds, the determination of their biological effects is of utmost importance. Among Lamiaceae species, Teucrium montanum (Mountain Germander) represents understudied Mediterranean plant species; it is rich in polyphenols, which are well-studied biologically active compounds for human disease prevention and the reduction of oxidative stress, i.e., phenolic acids, phenylethanoid glycosides, and flavonoids. For that purpose, the aim of this study was to investigate the antioxidant, cytotoxic, and genotoxic effects of Mountain Germander (MG) polyphenolic extract (0.025, 0.050, 0.150, and 0.500 mg extract mL⁻¹) on the hepatocellular (HepG2), tongue (CAL 27), gastric (AGS), and colorectal (Caco-2) continuous human cancer cell lines, as well as its bacteriostatic potential on representative members of human microbiota. In addition, the antioxidant potential of the MG polyphenolic extract was determined using bovine serum album and DNA plasmid as cellular model macromolecules. In vitro analysis revealed a significant cytotoxic effect of all MG extract concentrations on AGS and Caco-2 cell lines after prolonged treatment (24 h). In addition, treatment with 0.500 mg extract mL⁻¹ showed the most pronounced antioxidant effect under prolonged treatment (24 h) on CAL 27 and HepG2 cell lines. All of the applied MG extract concentrations seem to have a genoprotective effect on DNA plasmid. Furthermore, a significant inhibitory effect on E. coli was detected upon the treatment with 0.150 mg extract mL⁻¹, reducing the cell viability by 56%. Full article

The invasive blue crab is challenging the Mediterranean basin, progressively declining local populations. This reflects a lower prey availability and suitability of dietary nutrients (mainly n-3 polyunsaturated fatty acids, PUFA). The present study aimed to challenge blue crab males and females with a feed source low in n-3 PUFA with respect to one showing a proper fatty acid profile and to investigate the responses in terms of growth, welfare, lipid characterization of target tissues, and reproductive status. Blue crabs were divided into three groups as follows: (i) Marine: crabs fed sardinella (Sardinella aurita) fillet for 60 days; (ii) Mix: crabs fed bovine heart for the first 40 days and sardinella fillet for the following 20 days; and (iii) Terrestrial: crabs fed bovine heart for 60 days. The diet did not alter the health status but reflected the fatty acid profile of muscle and ovary of the blue crabs. In each group, males and females showed a proper hepatopancreas structure, with comparable levels of lipid reserves. This properly supported gonad maturation in both sexes. However, males and females from the group fed the terrestrial diet were characterized by reduced body weight, revealing that blue crabs prioritize reproductive investment rather than growth by directing crucial nutrients to reproductive organs when a suboptimal diet is available. Full article

The present work is devoted to the synthesis and analysis of the physicochemical properties of new functionalized asymmetric Gemini surfactants. Herein, alkyl- and azide-substituted surfactants with symmetric and asymmetric substituents were synthesized by using the click-reaction method. The critical aggregation concentration values of Gemini surfactants were determined. The binding processes of functionalized Gemini surfactants with bovine serum albumin were evaluated by fluorescence spectroscopy. Also, using the temperature dependences of the binding constants, the mechanism of Gemini surfactants binding with bovine serum albumin was studied. The hydrodynamic diameters of the formed bovine serum albumin/surfactant aggregates were analyzed. Based on electrophoretic light scattering, the ability of the synthesized Gemini surfactants to form associates was analyzed. The possibility of changing the mechanism of interaction in the 15c/bovine serum albumin system was shown. Based on the results obtained using different light scattering techniques and fluorescence spectroscopy, the mechanisms of interaction between bovine serum albumin and surfactants were determined. Full article

Bovine respiratory syncytial virus (BRSV) is an enveloped RNA virus that utilizes clathrin-mediated endocytosis for cell entry and is a significant pathogen in bovine respiratory disease (BRD). Heat shock protein family A member 4 (HSPA4), a member of the HSP70 family, is known to be involved in the progression of various cancers. However, its role in virus entry has not been previously explored. Through experiments involving Western blot analysis, virus titer, and virus copies analysis, we demonstrated that HSPA4 can regulate BRSV entry and replication. The specific regulation mode is to enhance BRSV entry by promoting clathrin-mediated endocytosis. We used Western blot, virus titer, virus copies analysis, and IFA to demonstrate that HSPA4 can promote clathrin heavy chain protein (CHC) expression and further promote BRSV entry by activating the PI3K–Akt signaling pathway. Furthermore, we observed that HSPA4 boosts the efficiency of clathrin-mediated endocytosis by increasing the ATPase activity of heat shock cognate protein 70 (HSC70), thereby facilitating BRSV entry. Additionally, our investigation into the impact of HSPA4 on the entry of other viruses revealed that HSPA4 can facilitate the entry of a variety of viruses into host cells. Full article

Approximately 30 distinct Mycoplasma species have been isolated from cattle, but only a few are pathogenic and can cause serious respiratory diseases. Consequently, this study aimed to identify Mycoplasma spp. infections in cattle with bovine respiratory disease (BRD), considering factors such as animal demographics, concurrent infections with other pathogens, post-mortem clinical findings and histological examinations, and seasonality. A total of 326 samples were collected from 322 cattle that had died from BRD in Northwestern Italy. A total of 54 animals (16.8%) tested positive for Mycoplasma spp., and Mycoplasma bovis (n = 22, 40.7%) and Mycoplasma dispar (n = 13, 24.1%) were the most frequently detected species among the examined cattle. Among positive cattle, those aged five months or younger were approximately five times more likely to be infected by Mycoplasma dispar than by Mycoplasma bovis compared to those older than five months (proportional incidence ratio: 5.1, 95% CI 1.2–21.2). The main bacterial pathogens identified in cattle exhibiting co-infection was Pasteurella multocida, whereas the main viral pathogens were BRSV and BoHV-1. Histopathological investigations predominantly revealed catarrhal bronchopneumonia or purulent catarrhal bronchopneumonia among the examined cattle. Finally, Mycoplasma hyopharyngis, a species isolated from the pharyngeal and nasal cavities of pigs so far, was detected for the first time in the pneumonic lung of a bovine infected with BRD. Further investigations are necessary to thoroughly characterize its host range and pathogenic potential. Full article

At present, provisional restorative materials are widely used for both short- and long-term treatment purposes. In esthetic dentistry, for the simulation of natural teeth, fluorescence plays a crucial role in enhancing the elegant, vibrant, and life-like appearance of restorative teeth, making them more closely resemble natural teeth. To achieve ideal esthetic outcomes, restorative materials must replicate the characteristics of natural teeth to provide the most realistic appearance possible. Thus, this study aims to compare the color difference in fluorescence (∆E₀₀) between a normal bovine tooth and eight provisional restorative materials under ultraviolet radiation. Eight provisional restorative materials were fabricated, for a total of 80 specimens (size 13.0 mm × 8.0 mm × 1.0 mm). A sound bovine tooth incisor was collected and used as a control for both enamel and dentine conditions. The color difference in fluorescence (∆E₀₀) was assessed using a spectrophotometer. A one-way ANOVA and Tukey’s test (p < 0.05) were used to analyze the resulting data. The results revealed that the color difference in fluorescence (∆E₀₀) between the normal bovine tooth and eight provisional restorative materials used in this study was higher than the 50% acceptability threshold, indicating a clear mismatch between the fluorescence of the materials and that of the normal bovine tooth. Unifast Trad had the lowest ∆E₀₀ among the provisional restorative materials, while Luxatemp Fluorescence had the highest color difference in fluorescence (∆E₀₀). The color difference in fluorescence (∆E₀₀) between Unifast Trad and Luxatemp Fluorescence is statistically significant. However, no significant difference was observed between Temporary CB, Vipi Block Trilux, and Protemp 4, or between Luxatemp Star and Luxatemp Fluorescence. It can thus be concluded that the ∆E₀₀ mostly differs among the different materials. Luxatemp Star and Luxatemp Fluorescence exhibited the highest ∆E₀₀ by a significant margin compared to the other groups. Full article

Subacute rumen acidosis (SARA) is a significant concern in dairy cattle fed grain-rich diets. To elucidate the underlying pathophysiological mechanisms, ruminal papilla biopsies are often used. This study aimed to assess how the sampling site along the ruminal papilla influences gene expression profiles in rumen epithelium during SARA. Rumen biopsies from five ruminal-cannulated non-lactating Holstein cows were collected during feeding of a forage diet (FD) and seven (wk1) and 21 days (wk3) after transition to high-grain (HG) feeding. Gene expression in apical (AP), basal (BP), and total length (TP) papillae were compared using RT-qPCR. Significant diet-induced effects were observed in AP for DSG1 (wk3, p = 0.0317), ZO1 (wk1 and wk3, p = 0.0159), GLUT3 (wk3, p = 0.0159), TLR4 (wk3, p = 0.0635), and NFKB (wk1, p = 0.0159), but hardly in BP or TP. Within wk1, TP showed higher transcript levels of ZO1 and TLR4 (p = 0.0079) and SGLT1 (p = 0.0317) compared to AP and BP independently from diet effects. These findings suggest that the apical parts of rumen papillae biopsies are most suitable for gene expression analyses to investigate diet-induced effects on rumen physiology and underscore the importance of considering the sampling site for accurate gene expression studies in rumen epithelium during SARA, providing valuable insights for future research and diagnostic approaches in managing rumen health in dairy cattle. Full article

Lameness, also attributed to musculoskeletal disorders of the back, is a major issue in dairy cow farming, affecting both animal welfare and economics. Chiropractic techniques are well studied in humans and horses for the treatment of musculoskeletal disorders. The anatomy of the bovine spine differs significantly from that of horses, but the knowledge of the animal’s specific anatomy is crucial for chiropractic treatment. This report explores how chiropractic methods could be adapted to cattle, considering their unique anatomy, and potentially integrated into standard veterinary care. Five case reports and an example of measuring the alignment of the facet joints in the spine are presented and discussed. Chiropractic treatments showed success in alleviating lameness and spinal issues in dairy cows, particularly when combined with other interventions. However, the role of chiropractic treatment in cattle as a complementary therapy needs further research. Full article

There is an increasing understanding that some mitral valve pathologies have developmental origins. The time course of valvulogenesis varies by animal model; in cattle, the branched chordae tendineae architecture becomes fully developed at full term. The mechanism by which chordae tendineae bifurcate during fetal development remains unknown. The current study presents a detailed description of bovine chordae tendineae formation and bifurcation during fetal development. Analysis of Movat Pentachrome-stained histological sections of the developing mitral valve apparatus was accompanied by micro-CT imaging. TEM imaging of chordae branches and common trunks allowed the measurement of collagen fibril diameter distributions. We observed a proteoglycan-rich “transition zone” at the junction between the fetal mitral valve anterior leaflet and chordae tendineae with “perforations” lined by MMP1/2 and Ki-67 expressing endothelial cells. This region also contained clusters of proliferating endothelial cells within the bulk of the tissue. We hypothesize this zone marks a region where chordae tendineae bifurcate during fetal development. In particular, perforations created by localized MMP activity serve as a site for the initiation of a “split” of a single chordae attachment into two. This is supported by TEM results that suggest a similar population of collagen fibrils runs from the branches into a common trunk. A clear understanding of normal mitral valvulogenesis and its signaling mechanisms will be crucial in developing therapeutics and/or tissue-engineered valve replacements. Full article

Mycoplasma bovis (M. bovis) is capable of causing pneumonia, arthritis, mastitis, and various other ailments in cattle of all age groups, posing a significant threat to the healthy progression of the worldwide cattle industry. The invasion of non-phagocytic host cells serves as a pivotal mechanism enabling M. bovis to evade the immune system and penetrate mucosal barriers, thereby promoting its spread. To investigate the differences in M. bovis invasion into four types of non-phagocytic cells (Madin–Darby bovine kidney (MDBK) cells, embryonic bovine lung (EBL) cells, bovine embryo tracheal (EBTr) cells and bovine turbinate (BT) cells) and further elucidate its invasion mechanism, this study first optimized the experimental methods for M. bovis invasion into cells. Utilizing laser scanning confocal microscopy, transmission electron microscopy, and high-content live-cell imaging systems, the invasion process of M. bovis into four types of non-phagocytic cells was observed. The invasion rates of three different strains of M. bovis (PG45, 07801, 08M) were quantified through the plate counting method. In order to clarify the specific pathway of M. bovis invasion into cells, chlorpromazine (CPZ), amiloride (AMI), and methyl-β-cyclodextrin (M-β-CD) were used to inhibit CLR-mediated clathrin-dependent endocytosis (CDE) pathway, macropinocytosis, and lipid raft pathway, respectively. Subsequently, the invasion rates of PG45 into these four types of cells were measured. Using siRNA technology, the expression of clathrin (CLR) in EBL cells was knocked down to further verify the role of CLR in the invasion process of M. bovis. The results showed that the optimal conditions for M. bovis to invade non-phagocytic cells were a multiplicity of infection (MOI) of 1000 and an optimal invasion time of 4 h. All three strains of M. bovis have the ability to invade the four types of non-phagocytic cells, yet their invasion abilities vary significantly. Observations from transmission electron microscopy further confirmed that at 120 min post-infection, PG45 had successfully invaded EBL cells and was present within endocytic vesicles. It is noteworthy that almost all PG45 successfully escaped from the endocytic vesicles after 240 min of infection had passed. Through chemical inhibition experiments and CLR protein knockdown experiments, it was found that when the CDE and lipid raft pathways were blocked or CLR protein expression was reduced, the invasion rates of PG45, 07801, and 08M in MDBK, EBL, EBTr, and BT cells were significantly decreased (p < 0.05). The above results indicate that M. bovis can invade all types of non-phagocytic cells through endocytic pathways involving CDE (clathrin-dependent endocytosis) or lipid raft-mediated endocytosis, and possesses the ability to escape from phagosomes. Full article

Milk-derived extracellular vesicles (mEVs) are emerging as promising therapeutic candidates due to their unique properties and versatile functions. These vesicles play a crucial role in immunomodulation by influencing macrophage differentiation and cytokine production, potentially aiding in the treatment of conditions such as bone loss, fibrosis, and cancer. mEVs also have the capacity to modulate gut microbiota composition, which may alleviate the symptoms of inflammatory bowel diseases and promote intestinal barrier integrity. Their potential as drug delivery vehicles is significant, enhancing the stability, solubility, and bioavailability of anticancer agents while supporting wound healing and reducing inflammation. Additionally, bovine mEVs exhibit anti-aging properties and protect skin cells from UV damage. As vaccine platforms, mEVs offer advantages including biocompatibility, antigen protection, and the ability to elicit robust immune responses through targeted delivery to specific immune cells. Despite these promising applications, challenges persist, including their complex roles in cancer, effective antigen loading, regulatory hurdles, and the need for standardized production methods. Achieving high targeting specificity and understanding the long-term effects of mEV-based therapies are essential for clinical translation. Ongoing research aims to optimize mEV production methods, enhance targeting capabilities, and conduct rigorous preclinical and clinical studies. By addressing these challenges, mEVs hold the potential to revolutionize vaccine development and targeted drug delivery, ultimately improving therapeutic outcomes across various medical fields. Full article