Toll-like receptor 2 (TLR2) is a critical component of the innate immune system. TLR2, a membrane-bound receptor, is expressed on the surface of specific immune cells, where it is involved in recognizing microbial components and initiating signaling cascades that activate immune cells, thereby playing a pivotal role in early pathogen detection and activating immune responses. This study investigated the effect of Toll-like receptor 2 gene polymorphism on the expression of genes associated with innate immune cell activation. The PCR-RFLP method was used to study single nucleotide polymorphism in exon 2 of the TLR2 gene 385 T/G (NC_037344.1 rs55617172) in 103 Holstein cows. The frequency of the TLR2T allele was 0.74, and that of the TLR2TT genotype was 53.4%. The effect of the TLR2 gene polymorphism on the expression of the genes of proinflammatory cytokines tumor necrosis factor (TNF), interleukin 6 (IL6), and interleukin 12B (IL12B) in peripheral blood mononuclear cells (PBMCs) from 29 cows was shown when PBMCs were stimulated with the TLR2 agonist lipoteichoic acid (LTA) or the Toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS). PBMCs obtained from animals carrying the G allele of the TLR2 gene showed significantly increased expression of the proinflammatory cytokine genes TNF, IL6, and IL12B following stimulation with TLR2 or TLR4 agonists. The TLR2 genotype did not significantly affect the expression of genes encoding the Toll-like receptors TLR2 and TLR4 or the adaptor molecules MYD88 and TICAM2.

This study introduces and validates dynamic multiplex PCR, a newly developed multiplex PCR method designed to overcome the inherent limitations of conventional multiplexing, particularly the challenges associated with heterogeneous primer melting temperatures (Tm) among multiple primer pairs in a single reaction. While traditional protocols rely on a single, fixed annealing temperature (Ta) per cycle, the dynamic multiplex PCR approach applies four sequential annealing temperatures (52°C, 54°C, 57°C, and 60°C) within each individual PCR cycle. This novel intra-cycle multi-temperature strategy was evaluated using an 8-plex microsatellite panel in European sea bass (Dicentrarchus labrax) samples and compared against three established protocols: multiplex touchdown PCR and two conventional PCR setups at fixed temperatures of 54°C and 57°C. The performance of the methods was assessed through a dual-platform quantitative analysis. Amplification yields were first quantified via densitometric analysis of agarose gel electrophoresis, while capillary fragment analysis was utilized to evaluate peak height, total peak area, quality scores, and fragment sizing precision. Sizing accuracy was rigorously determined as the absolute deviation between observed fragment sizes and assigned allele values. Statistical evaluations were conducted using a General Linear Model (GLM) and ANOVA framework to examine the effects of the PCR method, operator, locus, and biological sample on amplification performance and genotyping reliability. The results demonstrate that dynamic multiplex PCR significantly outperforms both conventional and multiplex touchdown PCR protocols by providing more balanced and robust amplification across all eight loci. By accommodating diverse primer requirements within a single cycle, this method eliminates the need for extensive trial-and-error optimization, thereby substantially reducing the consumption of laboratory consumables and total processing time. These findings establish dynamic multiplex PCR as a highly reliable, cost-effective, and reproducible multiplex PCR approach, offering a significant methodological advancement for high-quality multi-locus genotyping and fragment analysis workflows.

Lake Sinai virus (LSV) is an RNA virus that affects honey bee health and is considered a significant contributor to colony losses. In this study, the prevalence of LSV was investigated in adult bees and capped broods collected from 32 apiaries in İzmir and Muğla, Türkiye. Using RT-PCR, LSV was detected in 68.7% of adult bees and 46.8% of capped broods, indicating a widespread presence in the studied regions. Phylogenetic analysis was conducted to determine the genetic diversity of LSV strains. In the variant typing study, LSV-2 was the most commonly detected variant, while LSV-4 was identified at a lower frequency. However, LSV-1, LSV-3, and LSV-5 were not detected in the analyzed samples. According to the phylogenetic analysis, high genetic heterogeneity was observed among LSV strains, with no distinct clustering by geographical distribution. These findings suggest that LSV is prevalent in Türkiye and exhibits genetic diversity independent of its location. This study provides RdRp-based phylogenetic data on honey bee LSV from Türkiye, contributing valuable molecular evidence to ongoing efforts to understand LSV distribution and evolution. Further research is necessary to assess the pathogenicity of different LSV variants and their potential impact on colony health.

Pseudomonas aeruginosa (P. aeruginosa) is an opportunistic and versatile pathogen that causes infections in both humans and animals. It is one of the ESKAPE pathogens, characterized by the ability to rapidly acquire resistance to antimicrobials. In recent years, multidrug-resistant (MDR), extensively drug-resistant (XDR), and pandrug-resistant (PDR) P. aeruginosa have been reported worldwide in humans and chickens. In poultry, P. aeruginosa is frequently isolated and spreads easily due to its multiple transmission routes and its ability to infect birds of all ages. As a zoonotic pathogen, it poses a risk to humans via the consumption of contaminated poultry products. This study aimed to isolate P. aeruginosa from a layer chicken flock in Türkiye and investigate its antimicrobial resistance (AMR) and biofilm formation ability. Isolation and identification were performed by conventional bacteriological analysis and sequence analysis of the 16S rRNA gene. Following phenotypic AMR testing, biofilm formation was quantitatively analyzed, and eleven AMR genes were screened by PCR. The recovered P. aeruginosa isolate exhibited resistance to aztreonam, colistin, and meropenem, thereby meeting the criteria for MDR. It also tested positive for blaOXA and mcr-2 resistance genes and was defined as a weak biofilm producer. To the best of our knowledge, this is the first report of blaOXA and mcr-2 genes co-existing in an MDR P. aeruginosa isolate recovered from a layer chicken flock in Türkiye. These findings highlight the potential risk posed by MDR and biofilm forming P. aeruginosa in poultry environments to both human and animal health.

Valproic acid (2-propyl valeric acid, VPA) is the drug of choice for the treatment of migraine, bipolar disorder, and epileptic disorders in both children and adults. Although VPA has beneficial effects, long-term administration is reported to damage many tissues and organs. VPA administration increases free-radical production. Edaravone (3-methyl-1-phenyl-2-pyrazolin-5-one, EDA) is a potent antioxidant that protects against oxidative stress. The aim of our study was to investigate the protective effects of EDA against brain damage in VPA-treated rats. Male Sprague Dawley rats were used in our study. The rats were randomly divided into four groups: control, EDA, VPA, and VPA+EDA. EDA and VPA were administered intraperitoneally at doses of 30 mg/kg and 0.5 g/kg, respectively daily for 7 days. On day 8, all animals were sacrificed under anesthesia, and brain tissues were removed. VPA caused decreases in the levels of reduced glutathione (GSH, P

Valproic acid (VPA) is a short-chain fatty acid used to treat disorders of the brain. When VPA is used for long term, it causes damage, particularly through the formation of free radicals. The lungs are among the organs most affected by long-term VPA treatment. Some antioxidant substances are used to prevent damage caused by free radicals. One of these substances is alpha-lipoic acid (ALA), which occurs naturally. In our study, the protective effect of ALA against VPA-induced lung injury was investigated. In this study, female Sprague-Dawley rats were divided into four groups. The 1st group served as the control (1 mL olive oil); the 2nd group received ALA (50 mg/kg/day) for 15 days; the 3rd group received VPA (0.5 g/kg/day) for 15 days; and the 4th group received both VPA and ALA at the same doses and for the same duration. ALA and VPA were dissolved in olive oil. On day 16, all groups were euthanized under anesthesia. The lung tissues were collected and homogenized. In the prepared supernatants, the levels of reduced glutathione and total antioxidant status, and the activities of glutathione reductase, glutathione peroxidase, catalase, paraoxonase, carbonic anhydrase, aryl esterase, and superoxide dismutase were decreased in the VPA group, while sialic acid, lipid peroxidation, reactive oxygen species, total oxidant, oxidative stress index, hydroxyproline, glycoprotein levels, and xanthine oxidase activities were increased in the VPA group. These values were reversed in ALA when VPA was administered. As a result, it may be concluded that ALA exerts protective effects against VPA-induced lung damage.

The welfare of working dogs during training is crucial for their health, learning capacity, and long-term performance. This study investigated the efficacy of the Dog Appeasing Pheromone (DAP) in mitigating acute stress responses in Belgian Malinois during a structured 7-day basic obedience training protocol. Thirty-three dogs were randomly allocated to three groups: Group 1 (control, no training or DAP), Group 2 (training without DAP), and Group 3 (training with DAP). Physiological indicators (heart rate, body temperature, salivary cortisol) and behavioral stress markers (ethogram-coded postures) were recorded daily. Dogs in Group 2 exhibited significantly elevated stress responses across all parameters compared to Groups 1 and 3 (P < 0.001). Group 3, treated with DAP, showed physiological and behavioral measures not statistically different from the control group, indicating effective stress mitigation. Specific stress behaviors—such as ear retraction, lowered head, and semi-low posture—were markedly reduced in DAP-treated dogs. Strong positive correlations were observed between physiological and behavioral indicators (e.g., cortisol and body posture: r=0.503; cortisol and tail position: r=0.744), supporting the reliability of behavioral observations in stress assessment. This is the first study to integrate behavioral and physiological metrics to demonstrate the stress-buffering effect of DAP during active training. Given its non-invasive and non-sedative nature, DAP represents a promising tool for improving welfare in working dogs without compromising performance. Incorporating DAP into training protocols may support ethical handling practices and enhance the well-being and functional success of dogs in demanding service roles.

This case presents a 2-year-old spayed female cat that developed a complete esophageal stricture following 14 days of oral dry doxycycline therapy. Following treatment, the patient developed symptoms of regurgitation, loss of appetite, and vomiting. Direct radiography revealed marked aerophagia in the esophagus, while contrast radiography showed esophageal dilatation at the level of the first thoracic vertebra (T1) and complete stricture preventing the contrast medium from passing into the stomach. Endoscopic examination verified a complete stricture in the middle esophagus, and computed tomography (CT) revealed that the stricture was localized at T1; no extraluminal mass or vascular ring anomaly was identified. The absence of a history of anesthesia, foreign body ingestion, or trauma in the medical history confirms that the stricture developed due to chemical damage to the esophageal mucosa caused by doxycycline in capsule form. Reports in the literature indicate that administering doxycycline capsules to cats without adequate fluids increases the risk of esophageal ulceration and scarring. Unfortunately, because of the poor prognosis of the complete stricture in the patient, the patient's poor general condition, and the owner's reluctance to pursue treatment, the decision was made to euthanize the patient. In conclusion, the presented case highlights the diagnostic value of combined radiography, contrast esophagography, endoscopy, and computed tomography for diagnosing esophageal strictures that develop after oral administration of doxycycline. Furthermore, it is essential to consider the pharmaceutical formulation and administration technique for oral drug delivery to cats to prevent potential complications.

This study evaluated the effects of dietary sanguinarine supplementation on performance, serum biochemical parameters, antioxidant status, immune responses, meat quality, and intestinal morphology in broilers. A total of 84 one-day-old male Ross 308 broilers were randomly assigned to four dietary treatments containing sanguinarine at 0, 0.5, 1.0, or 1.5 g/kg for a 35-day experimental period. On day 28, an immune challenge was applied by intraperitoneal injection of 0.1 mL lipopolysaccharide (LPS; Escherichia coli serotype O127:B8) to three birds per group, while the remaining birds received sterile saline. Broilers fed sanguinarine-supplemented diets showed significantly higher final body weight and body weight gain, as well as improved feed conversion ratio. Sanguinarine supplementation significantly increased serum glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), and total superoxide dismutase (T-SOD) activities, while reducing malondialdehyde (MDA) concentrations. Moreover, sanguinarine effectively attenuated LPS-induced oxidative stress by preventing reductions in antioxidant enzyme activities and excessive MDA production. Higher serum concentrations of IgA, IgM, and IgG were observed in broilers receiving sanguinarine supplementation. In addition, dietary sanguinarine reduced serum levels of interleukin (IL)-1β, IL-6, IL-4, IL-10, and tumor necrosis factor-alpha (TNF-α). In LPS-challenged birds, IL-1β, IL-6, and TNF-α concentrations were markedly decreased by sanguinarine supplementation. Furthermore, lower serum levels of creatinine, urea, total cholesterol, high-density lipoprotein, and low-density lipoprotein were detected. Broilers fed sanguinarine-supplemented diets exhibited increased villus height and reduced crypt depth in the small intestine. Results suggest that dietary sanguinarine is an effective feed additive for improving growth performance, antioxidant capacity, immune response, and intestinal health in broilers.

The aim of the present study was to investigate the in vitro effects of two lysozyme doses (10 and 50 FIP U/mL) on rumen fermentation and microbial populations using the artificial rumen system Rusitec with an 80:20 concentrate-to-forage diet. The ruminal pH, the total number of protozoa, total and individual volatile fatty acid (VFA) production, and methane (CH4) release were not significantly affected by the tested quantities of lysozyme. However, the ammonia nitrogen (NH3-N) concentration was significantly increased by supplementation with Lysozyme-50 (P < 0.05). The dry matter digestibility (DMD) was significantly higher in Lysozyme-50 than in Lysozyme-10 (P < 0.05). The total bacterial count decreased linearly in response to the addition of Lysozyme-50 (P < 0.05). Furthermore, in the presence of Lysozyme-50, the abundance of Butyrivibrio fibrisolvens decreased in both linear and quadratic manners (P < 0.05), and the abundance of Fibrobacter succinogenes decreased in a linear manner (P < 0.05). A linear decline tendency in the cell numbers of Ruminococcus albus (P = 0.092) and Megasphaera elsdenii (P = 0.085) was observed with Lysozyme-50 treatment. These findings suggest that while core fermentation parameters such as VFA production and digestibility remained stable, the elevated ammonia nitrogen concentration may reflect reduced nitrogen utilization efficiency, particularly at high doses. Additionally, since lysozyme has limited selectivity for certain rumen bacteria within the Rusitec system, lysozyme may influence rumen microbes differently from conventional antibiotics, warranting further in vitro and in vivo studies across diverse diets and dose ranges.