The activity of monocyte-derived macrophages after stimulation with platelet-rich and platelet-poor concentrates. Study on an ovine model of insertion of a tibial implant coated with silicon-doped diamond-like carbon.

Introduction: Macrophages are crucial immune cells that play a role in tissue repair and can exhibit pro- or anti-inflammatory behaviour based on environmental stimulation. Their functional phenotype can be affected by platelet-derived products as determined by those products' composition. When the inflammatory response caused by implantation is excessive, it can lead to rejection of the implant. Therefore, a thorough evaluation of implant haemocompatibility is necessary to minimise undesirable consequences. Material and Methods: In an in vitro study, monocyte-derived macrophages (MDMs) were obtained from the whole blood of sheep after a silicon-doped diamond-like carbon-coated implant insertion. These MDMs were then exposed to autologous platelet-derived products for functional marker analysis. Results: Platelet-poor plasma (PPP) and pure platelet-rich plasma (P-PRP) stimulation increased arginase-1 activity, while leukocyte-rich PRP stimulation produced a mixed response involving higher O2- (6.49 ± 2.43 nM vs non-stimulated 3.51 ± 1.23 nM, P-value < 0.05) and NO (3.28 ± 1.38 µM vs non-stimulated 2.55 ± 0.32µM, P-value < 0.05) generation. Conclusion: Using PPP and P-PRP stimulation in post-implantation procedures may contribute to the polarisation of macrophages towards the M2-like pro-resolving phenotype, thereby accelerating wound healing. This would also prevent implant degradation due to an excessive inflammatory process.

Selected bacteria in sheep stool depending on breed and physiology state.

One of the important factors influencing the microbial community of ruminants, besides environment or diet, are breed and physiology. Therefore, the purpose of this study was to assess these changes in the levels of basic microbial phyla and families. For this study, qPCR analysis was performed to determine the level of bacteria (Firmicutes, Bacteroidetes, Actinobacteria, Proteobacteria clusters and Clostridiaceae, Lactobacillaceae families) in the feces of ewes of three native Polish sheep breeds (Polish Lowland Sheep (PON), Swiniarka Sheep (SW), and synthetic line BCP) at different physiological periods (conception, early pregnancy, lambing, end of lactation). The animals were kept in the same environment and were at the same age (2-years). The results showed a significant effect of both breed (p = 0.038) and physiological period (p < 0.05, p < 0.01) on the levels of bacteria analyzed. The breed showed differences across physiological periods. The influence of the race factor was noted primarily between the BCP synthetic line and the other two breeds (differences in terms of all analyzed clusters and families except Actinobacteria phyla). In the case of SW and PON, however, the observed differences were only at the level of Proteobacteria cluster and Clostridiaceae family. On the other hand, the early pregnant and lambing periods were the most microbiologically diverse in terms of the analyzed clusters and families of bacteria.

Combined Effect of Acid Whey Addition and Ultrasonic Treatment on the Chemical and Microbiological Stability of Lamb Stuffing.

The microbiological and chemical stability of stuffing is crucial in meat processing. Small ruminant (lamb) meat has many nutritional advantages (fatty acid composition and ratio, high biological value of protein, source of zinc, and selenium) but is poorly utilized in processing. In this experiment, we determined the effects of physical (sonication), chemical (salt, curing salt, and air access), and microbiological (acid whey) factors on the microbiological and chemical stability of lamb meat stuffing. Proximate composition and fatty acids profile, pH, water activity, lipid oxidation, color parameters, and microbiology examinations were performed in fresh meat stuffing and on the fifth day of refrigerated storage. Ultrasound treatment of the meat with whey reduced its acidity and increased the oxidative stability of fats but did not modify the water activity and color of the stuffing. Stuffing sonication did not affect the growth of LAB but reduced the number of Entereobacteriace, especially in the presence of whey and salt. The treatment of lamb meat stuffing with low-frequency and medium-intensity ultrasound assisted by the addition of acid whey and salt is a technique conducive to reducing the use of nitrates in meat technology and their consumption by consumers.

Phylogenetic Analysis of Small Ruminant Lentiviruses Originating from Naturally Infected Sheep and Goats from Poland Based on the Long Terminal Repeat Sequences.

Introduction: Previous gag and env sequence studies placed Polish small ruminant lentiviruses (SRLVs) isolated from sheep and goats in subtypes B1, B2, A1, A5, A12, A13, A16-A18, A23, A24 and A27. This study extended the genetic/phylogenetic analysis of previously identified Polish SRLV strains by contributing long terminal repeat (LTR) sequences. Material and Methods: A total of 112 samples were analysed. Phylogenetic analyses were carried out on the LTR fragment using the neighbour-joining, maximum likelihood, and unweighted pair group method with arithmetic mean methods. Results: Polish caprine and ovine LTR sequences clustered within group A and grouped in at least 10 clusters (subtypes A1, A5, A12, A13, A16-A18, A23, A24 and A27). Most of the Polish strains (78%) belonged to the same subtype by the indication of the gag, env and LTR genomic regions. Discrepancies in affiliation depending on the particular sequence were observed in 24 (21%) strains, most of which came from mixed-species flocks where more than one SRLV genotype circulated. Sequences of the LTR reflected subtype-specific patterns. Several subtype-specific markers were identified, e.g. a unique substitution of T to A in the fifth position of the TATA box in A17, A27, A20 and B3. Conclusion: This study provides valuable insights into the genetic diversity of SRLV field strains in Poland, their phylogenetic relationships and their position in the recently established SRLV classification. Our results confirmed the existence of the ten subtypes listed and the readier emergence of new SRLV variants in mixed-species flocks.

Effects of Topical Treatment of Foot Rot in Sheep Using Ozonated Olive Ointment.

INTRODUCTION: Foot rot in small ruminants is highly contagious, causes severe lameness, and impairs fertility and wool and meat production. It is usually treated with parenteral antibiotics, with attendant antibiotic resistance risk, and with bactericidal footbaths, potentially harmful to humans and the environment. An alternative treatment in sheep is proposed based on repeated topical ozonated ointment application. Its effectiveness and safety were evaluated by estimation of acute-phase response, biochemical indicators of organic damage, and antioxidant/oxidant balance (AOB). MATERIAL AND METHODS: The study was conducted on ten sheep with Egerton scale 2-3 lesions. Ozone application was repeated every day for seven days. Blood was drawn first (T0) after foot cleaning and before ozonation, then (T1) seven days after the first ozone application, and finally (T2) four days after the last application. RESULTS: High clinical effectiveness was observed, with total recovery by 28 days from the start of treatment. A significant increase in antiradical activity was noted on the basis of a 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) assay from 1.16 ± 0.04 µmolTe/mL at T0 to 1.23 ± 0.03 µmolTe/mL at T1, with a slight decrease in oxidative stress. Calculated on the basis of antiradical capacity, AOB was higher at T1 (130 ± 19%) and decreased to 110 ± 16% at T2. Calculated on the basis of reducing power, it was 169 ± 22% at T1 and 131 ± 17% at T2. CONCLUSION: These results indicated that the AOB is efficient enough to prevent oxidative organ injury and the applied doses of ozone are safe for animals.

Age-Related Changes in Acute Phase Reaction, Cortisol, and Haematological Parameters in Ewes in the Periparturient Period.

A well-functioning immune system is the basis for protection against infectious and metabolic diseases, and a smooth return to homeostasis. The periparturient period is considered critical because major changes in the endocrine, behavioural, digestive, and immune systems dysregulate immune function, leading to immunosuppression. With age, the immune system could become dysregulated. The purpose of the present investigation was to compare changes in plasma concentrations of acute phase proteins, cortisol, and haematological parameters in the peripheral blood of two age-related groups of healthy ewes to get a better understanding of changes around lambing. Two groups of ewes were enrolled in the study: 3-year-old (young; n = 9) and 7-year-old ewes (old; n = 9). All females were synchronised and inseminated. In blood plasma, serum amyloid A (SAA) and cortisol concentrations were measured using ELISA tests, a spectrophotometric method to determine haptoglobin (Hp), and a thrombin clottable estimation to determine the fibrinogen (Fb) concentration. The blood parameters were examined using an automated haematological analyser. In clinically healthy ewes, no significant effect of age was observed in SAA, Hp, Fb and cortisol concentration in most of analysed terms. SAA, Hp, Fb, and cortisol fluctuations typical for the periparturient period were observed. There were no age-associated differences in red or white blood cell parameters.

The Effect of Neutrophil-Derived Products on the Function of Leukocytes Obtained after Titanium Implantation in the Ovine Model.

Titanium (Ti) is currently the most common biomaterial used for orthopedic implants; however, these implants may cause deleterious immune response. To investigate the possible mechanisms involved in excessive inflammation, we assessed the activity of neutrophils and monocyte-derived macrophages (MDMs) during the insertion of the Ti implant in a sheep model. The study was conducted on 12 sheep, 4 of which were control animals and 8 were in the experimental group with inserted Ti implant. Neutrophil secretory response was estimated at two time points T0 before surgery and T1 1 h after implantation and was based on the release of enzymes from neutrophil granules and reactive oxygen and nitrogen species (RONS) generation. MDM function was evaluated 5 months after implantation, on the basis of RONS generation arginase activity and morphological changes. Moreover, the influence of some autologous neutrophil derived products, namely, antimicrobial neutrophil extract (ANE) and neutrophil degranulation products (DGP) on leukocytes was estimated. Our study revealed that Ti implant insertion did not cause any adverse effects up to 5 months after surgical procedure. Stimulation of neutrophil cultures with ANE decreased the enzyme release as well as superoxide generation. Treatment of MDM with ANE diminished superoxide and NO generation and increased arginase activity. On the other hand, MDM stimulated with DGP showed elevated superoxide and NO generation as well as decreased arginase activity. To summarize, ANE exerted an anti-inflammatory and pro-resolving effect on studied leukocytes, whereas DGP acted as pro-inflammatory.

The Level of Selected Bacterial Phyla on the Skin Surface of Small Ruminants According to the Breed and Species.

For decades, skin has been assigned the main role of an insulator of the inside of the body from the external environment, but it also plays a role in maintaining homeostasis. In this study, the level of selected bacterial phyla (Firmicutes, Bacteroidetes, Proteobacteria and Actinobacteria) was assessed in three sheep breeds (Swiniarka sheep, Uhruska sheep and BCP line (synthetic sheep breed; n = 6) and in two breeds of goats (Boer, Saenian; n = 6) living in the same environment and fed on the same feed, where the aim was to identify differences in terms of race, species and individual differences. Significant differences were found in Firmicute, Actinobacteria and Proteobacteria phyla (p ≤ 0.05). Statistically significant and positive correlations were demonstrated between Actinobacteria and Proteobacteria, Proteobacteria and Bacteroidetes or Firmicutes and Bacteroidetes. The obtained results suggest that the species and racial differences in the level of the studied bacterial phyla may also result from the physicochemical differences of the skin surface, as they could exacerbate the variations in humidity, temperature, composition of antimicrobial peptides (AMP) and lipid content. In addition, individual differences were observed, which indicate a similar effect of an individual on the microbiological composition of its organism.

Long-term Interactions of Circulating Neutrophils with Titanium Implants, the Role of Platelets in Regulation of Leukocyte Function.

Despite the fact that different biomaterials are widely used in many biomedical applications, they can still cause side effects. Therefore, our aim was to assess neutrophil activity during the inflammatory phase of the repair process and long-term interactions between circulating neutrophils and Titanium (Ti) implants. Additionally, neutrophil in vitro response after stimulation by the extract of antimicrobial peptides (AMP extract), pentoxifylline (PTX) and some platelet-rich (L-PRP and PURE PRP) and platelet-poor (PPP) concentrates were tested. The study was conducted on eight sheep after Ti implant insertion into the tibia and revealed that the Ti implant did not cause any side effects during the course of experiment. After addition of L-PRP into neutrophils, culture activity of these cells significantly increased (p < 0.01), whereas treatment with AMP extract, PURE PRP, PPP or PTX caused decrease in neutrophil enzymatic response (on the basis of elastase, myeloperoxidase and alkaline phosphatase release) and free radical generation. These effects were observed in neutrophils isolated during the inflammatory phase as well as 4 and 10 months after implantation. Obtained results will be useful in regulation of inflammatory response during implantation of biomaterial and create possibility to modulate the cells response towards pro- or anti-inflammatory to reduce host tissue damage.

Prospects and Applications of Natural Blood-Derived Products in Regenerative Medicine.

Currently, there are a number of therapeutic schemes used for the treatment of various types of musculoskeletal disorders. However, despite the use of new treatment options, therapeutic failure remains common due to impaired and delayed healing, or implant rejection. Faced with this challenge, in recent years regenerative medicine started looking for alternative solutions that could additionally support tissue regeneration. This review aims to outline the functions and possible clinical applications of, and future hopes associated with, using autologous or heterologous products such as antimicrobial peptides (AMPs), microvesicles (MVs), and neutrophil degranulation products (DGP) obtained from circulating neutrophils. Moreover, different interactions between neutrophils and platelets are described. Certain products released from neutrophils are critical for interactions between different immune cells to ensure adequate tissue repair. By acting directly and indirectly on host cells, these neutrophil-derived products can modulate the body's inflammatory responses in various ways. The development of new formulations based on these products and their clinically proven success would give hope for significant progress in regenerative therapy in human and veterinary medicine.

Chemical composition and fatty acid content in lamb and adult sheep meat.

The aim of the present study was to compare the chemical composition and fatty acid (FA) content of the muscle tissues of lambs and adult sheep bred for meat production. Throughout the research period, the animals were managed in a confinement indoor system under uniform environmental conditions. After slaughter, meat samples collected from the musculus biceps femoris were used to determine the basic chemical composition and fatty acid content in the intramuscular fat. The analyses showed that, compared to lambs, meat from adult animals exhibited a more beneficial ratio of n-6 to n-3 FAs, as well as a favorable percentage of OFAs (dietary fatty acids having an undesirable hypercholesterolemic effect on humans). The results of this study support the inclusion of dietitian-recommended mutton into the human diet and the promotion of the production of meat from adult animals as a high-value product. The sheep industry could benefit economically, particularly in countries where this meat type has not enjoyed a high standing in consumer preferences.

Selected leukocyte subpopulations in peripheral blood and uterine washings in cows before and after intrauterine administration of cefapirin and methisoprinol.

The aim of the study was to evaluate the selected lymphocyte subpopulations TCD4, TCD8, BCD21, BCD25, CD18, CD11b, and MHC II in blood and uterine flush of cows with endometritis, before and after intrauterine (i.u.) administration of cefapirin and methisoprinol. The research was carried out on 28 cows with clinical endometritis. Animals were divided into four groups, each composed of seven cows, depending on the i.u. preparation used: Group A, cefapirin; Group B, methisoprinol; Group C, cefapirin and methisoprinol simultaneously; and a control group-without medication. The study was performed using flow cytometry method. Summarizing the results of the research, i.u. infusion of cefapirin caused a weakening of the effector phase of the local uterine immune response; however, it enhanced leukocyte chemotaxis and antigen presentation. After i.u. administration of methisoprinol, the stimulation of specific uterine immunity mechanisms was mainly observed. The use of both mentioned preparations showed the strengthening of specific uterine immunological mechanisms presumably caused by methisoprinol, despite the inhibitory effect of the antibiotic. Intrauterine use of immunostimulatory substances can improve the effectiveness of the endometritis treatment in cows by improving specific local mechanisms of uterine immunity. As a consequence, it may enhance the effector function of immune competent cells and finally eliminate inflammation.

Risk factors associated with small-ruminant lentiviruses in sheepfold buildings.

INTRODUCTION: Small-ruminant lentivirus (SRLV) infection is widespread across Europe. It causes substantial economic losses in sheep breeding. The main route of SRLV infection is through the mother's milk, especially colostrum However, infection can also occur via contact between infected and healthy animals. It should be noted that the mechanisms of contact infection are still relatively poorly understood. The virus can also spread through a flock via an aerogenic mechanism. OBJECTIVE: Due to the increased risk of SRLV infection in sheep bred in an alcove system, this study sought to define the effect of various selected factors associated with alcove breeding on the frequency of SRLV infection in sheep. MATERIAL AND METHODS: Risk factors associated with small-ruminant lentivirus (SRLV) infection were analyzed among flocks of sheep in central-eastern Poland. Ninety-eight sheep flocks were selected for detailed investigation and included 6,470 ewes and 15 breeds and lines. Serologic testing of blood samples was used to identify infected animals and evaluate the epidemiologic status of particular flocks. Specific antibodies for Maedi Visna Virus (MVV) were detected via ELISA. Questionnaires were used to gather information concerning risk factors. RESULTS: The study's results indicate that factors associated with environmental conditions under which sheep are kept play a significant role in determining the risk of SRLV infection. CONCLUSIONS: Special attention should be focused on airborne contamination associated with the technologies used in sheep breeding. Breeding technologies that limit airborne contamination in farm buildings should be employed. In developing programmes to eliminate SLRV in sheep flocks, improvement of zoohygenic conditions should also be considered.

Factors affecting prevalence and abundance of A.perfoliata infections in horses from south-eastern Poland.

Equine Anoplocephalosis constitute a significant problem in horses worldwide. The aim of this study was to analyse intrinsic (host age and sex) and extrinsic (management type, pasture type and moisture) factors that influence the prevalence and FEC of A. perfoliata infections. Faecal samples were collected from 994 horses managed in studs or individually between 2012 and 2014. The Sedimentation-flotation method was applied for coproscopic analysis, and faecal egg counts were calculated. The overall prevalence was 25.1% (21.4-29.0) with the highest prevalence (36.1% [28.1-44.8]) found in horses 10-20 years old. The individuals kept in studs showed three times higher A. perfoliata prevalence compared to the ones managed individually. The prevalence significantly differed between pasture types, with individuals kept in studs (37.6% [34.3-40.9]) showing four times higher prevalence than horses kept individually (9.2% [4.8-16.5]). More horses kept on watery (42.0% [36.6-47.6]) and semi-watery (35.9% [31.3-40.7]) pastures were infected than those on dry (6.6% [4.6-9.2]) pastures. The overall A. perfoliata FEC in all examined individual was 2.67 and differed within sex, with mares showing 4.3 - times higher FEC of infection than stallions. Horses bred in studs (3.65±0.289) showed higher FEC than these bred individually (1.28±0.198). There was the effect of pasture type on A. perfoliata FEC, with horses kept on joint pastures (4.06±0.29) showing higher FEC than individuals kept individually (0.88±0.23). Pasture moisture significantly affected A. perfoliata FEC with the highest FECs in horses from watery pastures. Horses bred on dry pastures showed 16 times lower FEC than horses bred on watery pastures. Host age also significantly affected A. perfoliata FEC, with the oldest individuals showing the highest mean FEC. The presented analysis of intrinsic and extrinsic factors may help to overcome A. perfoliata infections in horses in different breeding systems. Understanding the role of management and pasture type risk factors that influence this parasitosis may benefit both breeders and veterinary surgeons.

Risk factors associated with small ruminant lentivirus infection in eastern Poland sheep flocks.

An analysis of the risk factors for ovine lentivirus infection was performed in sheep flocks located throughout the central-eastern region of Poland. Here, we report the infection details for 98 flocks with a total of 6470 ewes, 15 sheep breeds. The identification of infected animals and a review of the epidemiological status of each flock were based on an evaluation of serological tests performed on collected blood serum samples. Blood for examination was obtained from 2925 ewes of the 98 flocks under observation. Specific antibodies for Maedi Visna Virus (MVV) were detected via ELISA. Data illustrating the conditions at each sheep farm were obtained through questionnaires completed by farmers, as well as observations, measurements, and breeding records that were available. These observations were used to assess risk factors contributing to small ruminant lentivirus (SRLV) infection in sheep flocks. It was found that both sheep flock size and the type of management system had a significant effect on the increased risk of lentiviral infection. In addition, we demonstrate that there is a significant (p<0.0001) relationship between the occurrence of mastitis (OR 2.01, CI: 1.55-2.61) and diarrhea (OR 4.22, CI: 3.30-5.39) with SRLV infection in the observed sheep. Additionally, the infection rate of the animals translated directly to an impaired physical condition. Notably, the risk of infection could potentially be reduced if sheep producers are further acquainted with SRLV detection and invoke a control program based on diagnostic tests. Moreover, marketing approval should be granted for solely SRLV-seronegative animals.