Potent drug delivery enhancement of betulinic acid and NVX-207 into equine skin in vitro - a comparison between a novel oxygen flow-assisted transdermal application device and microemulsion gels.

BACKGROUND: Gray horses are predisposed to equine malignant melanoma (EMM) with advancing age. Depending on the tumor's location and size, they can cause severe problems (e.g., defaecation, urination, feeding). A feasible therapy for EMM has not yet been established and surgical excision can be difficult depending on the location of the melanoma. Thus, an effective and safe therapy is needed. Naturally occurring betulinic acid (BA), a pentacyclic triterpene and its synthetic derivate, NVX-207 (3-acetyl-betulinic acid-2-amino-3-hydroxy-2-hydroxymethyl-propanoate) are known for their cytotoxic properties against melanomas and other tumors and have already shown good safety and tolerability in vivo. In this study, BA and NVX-207 were tested for their permeation potential into equine skin in vitro in Franz-type diffusion cell (FDC) experiments after incubation of 5 min, 30 min and 24 h, aiming to use these formulations for prospective in vivo studies as a treatment for early melanoma stages. Potent permeation was defined as reaching or exceeding the half maximal inhibitory concentrations (IC50) of BA or NVX-207 for equine melanoma cells in equine skin samples. The active ingredients were either dissolved in a microemulsion (ME) or in a microemulsion gel (MEG). All of the formulations were transdermally applied but the oil-in-water microemulsion was administered with a novel oxygen flow-assisted (OFA) applicator (DERMADROP TDA). RESULTS: All tested formulations exceeded the IC50 values for equine melanoma cells for BA and NVX-207 in equine skin samples, independently of the incubation time NVX-207 applied with the OFA applicator showed a significant time-dependent accumulation and depot-effect in the skin after 30 min and 24 h (P < 0.05). CONCLUSIONS: All tested substances showed promising results. Additionally, OFA administration showed a significant accumulation of NVX-207 after 30 min and 24 h of incubation. Further in vivo trials with OFA application are recommended.

Preparation and PET/CT imaging of implant directed 68Ga-labeled magnetic nanoporous silica nanoparticles.

BACKGROUND: Implant infections caused by biofilm forming bacteria are a major threat in orthopedic surgery. Delivering antibiotics directly to an implant affected by a bacterial biofilm via superparamagnetic nanoporous silica nanoparticles could present a promising approach. Nevertheless, short blood circulation half-life because of rapid interactions of nanoparticles with the host's immune system hinder them from being clinically used. The aim of this study was to determine the temporal in vivo resolution of magnetic nanoporous silica nanoparticle (MNPSNP) distribution and the effect of PEGylation and clodronate application using PET/CT imaging and gamma counting in an implant mouse model. METHODS: PEGylated and non-PEGylated MNPSNPs were radiolabeled with gallium-68 (68Ga), implementing the chelator tris(hydroxypyridinone). 36 mice were included in the study, 24 mice received a magnetic implant subcutaneously on the left and a titanium implant on the right hind leg. MNPSNP pharmacokinetics and implant accumulation was analyzed in dependence on PEGylation and additional clodronate application. Subsequently gamma counting was performed for further final analysis. RESULTS: The pharmacokinetics and biodistribution of all radiolabeled nanoparticles could clearly be visualized and followed by dynamic PET/CT imaging. Both variants of 68Ga-labeled MNPSNP accumulated mainly in liver and spleen. PEGylation of the nanoparticles already resulted in lower liver uptakes. Combination with macrophage depletion led to a highly significant effect whereas macrophage depletion alone could not reveal significant differences. Although MNPSNP accumulation around implants was low in comparison to the inner organs in PET/CT imaging, gamma counting displayed a significantly higher %I.D./g for the tissue surrounding the magnetic implants compared to the titanium control. Additional PEGylation and/or macrophage depletion revealed no significant differences regarding nanoparticle accumulation at the implantation site. CONCLUSION: Tracking of 68Ga-labeled nanoparticles in a mouse model in the first critical hours post-injection by PET/CT imaging provided a better understanding of MNPSNP distribution, elimination and accumulation. Although PEGylation increases circulation time, nanoparticle accumulation at the implantation site was still insufficient for infection treatment and additional efforts are needed to increase local accumulation.

Nurse Scheduling with State-of-the Art Open-Source Tools.

Nurse scheduling is still an unsolved issue, as it is NP-hard and highly context-dependent. Despite this fact, the practice needs guidance on how to tackle this problem without using costly commercial tools. Concretely, we have the following use case: a Swiss hospital is planning a new station designed for nurse training. The capacity planning is finished, and the hospital wants to assess whether shift planning with known constraints leads to valid solutions. Here, a mathematical model is combined with a genetic algorithm. We trust the solution of the mathematical model more, but if it does not provide a valid solution, we try out an alternative. Our solutions indicate that actual capacity planning together with the hard constraints cannot lead to valid staff schedules. The central conclusion is that more degrees of freedom are necessary and that open-source tools OMPR and DEAP are valuable alternatives to commercial products such as Wrike or Shiftboard, in which the degree of freedom of customization is reduced in favor of easiness of use.

In Vitro Antimicrobial Activity of N-Acetylcysteine against Pathogens Most Commonly Associated with Infectious Keratitis in Dogs and Cats.

To determine the in vitro antimicrobial activity of N-acetylcysteine (NAC) against common pathogens associated with infectious keratitis in dogs and cats, clinical isolates of Staphylococcus (S.) pseudintermedius (n = 20), Streptococcus (St.) canis (n = 10) and Pseudomonas (P.) aeruginosa (n = 7) of canine and feline infectious ulcerative keratitis and a quality control strain (P. aeruginosa DSM 19880) were tested. The minimal inhibitory concentration (MIC) of NAC concentrations was determined using microdilution methodology. For S. pseudintermedius and P. aeruginosa, NAC concentrations in the range of 1.56 mg/mL (0.156%) to 100 mg/mL (10%), and for St. canis, concentrations ranging from 0.195 mg/mL (0.0195%) to 6.25 mg/mL (0.625%) were tested. For S. pseudintermedius, the MIC was 3.12 mg/mL (0.312%) for all tested isolates. For P. aeruginosa isolates and the quality control strain, the MIC ranged from 3.12 mg/mL (0.312%) to 6.25 mg/mL (0.625%). For St. canis, the MIC ranged from 1.56 mg/mL (0.156%) to 3.12 mg/mL (0.312%). NAC has an in vitro antimicrobial activity against three bacterial species commonly found in infectious keratitis in dogs and cats and therefore may be a promising alternative or adjuvant to topical antibiotics. The results warrant a clinical pilot study to assess the potential of NAC to reduce or replace the use of topical antibiotics in line with the One Health approach.

Influence of PEG Chain Length of Functionalized Magnetic Nanoparticles on the Cytocompatibility and Immune Competence of Primary Murine Macrophages and Dendritic Cells.

A major drawback of nanoparticles (NPs) for biomedical applications is their preferential phagocytosis in immune cells, which can be avoided by surface modifications like PEGylation. Nevertheless, examinations of different polyethylene glycol (PEG) chain lengths on the competence of immune cells as well as possible immunotoxic effects are still sparse. Therefore, primary murine macrophages and dendritic cells were generated and incubated with magnetic nanoporous silica nanoparticles (MNPSNPs) modified with different mPEG chains (2 kDa, 5 kDa, and 10 kDa). Cytotoxicity, cytokine release, and the formation of reactive oxygen species (ROS) were determined. Immune competence of both cell types was examined and uptake of MNPSNPs into macrophages was visualized. Concentrations up to 150 µg/mL MNPSNPs showed no effects on the metabolic activity or immune competence of both cell types. However, ROS significantly increased in macrophages incubated with larger PEG chains, while the concentration of cytokines (TNF-α and IL-6) did not indicate a proinflammatory process. Investigations on the uptake of MNPSNPs revealed no differences in the onset of internalization and the intensity of intracellular fluorescence. The study gives no indication for an immunotoxic effect of PEGylated MNPSNPs. Nevertheless, there is still a need for optimization regarding their internalization to ensure an efficient drug delivery.

Stability of Important Veterinary Antibiotics Amoxicillin, Sulfadiazine, and Trimethoprim in Practice-Relevant Model Solutions.

Due to the frequent use of veterinary drugs in animal husbandry, it is important to know their environmental behavior. In this context, little attention has been paid to the stability of the active ingredients in solutions prepared for administration. This is particularly problematic for antibiotics that trigger resistance when administered subtherapeutically. In order to investigate a possible influence of the preparation and storage of veterinary drugs on compound stability, three widely used antibiotics (amoxicillin, sulfadiazine, trimethoprim) were prepared in different model solutions. Depending on their individual stabilities, the incubation period lasted up to 70 days. Samples were analyzed at regular intervals by high-performance liquid chromatography-diode array detection and ultraviolet spectrophotometry. Following official recommendations, the investigations covered various parameters, e.g., pH, buffer substances, influence of light, and temperature. Sulfadiazine was incubated together with trimethoprim at concentrations of 120 mg L-1 and 80 mg L-1 for 70 days. Both compounds proved to be very stable under all experimental conditions and between 92 and 100% of the active ingredients remained. In 0.1% formic acid, a transformation product was found with less than 5% of the parent substance. In contrast, amoxicillin (500 mg L-1) was instable in almost all solutions under investigation. Within 17 days, the concentration of AMO decreased to 72% in ultrapure water. With the exception of a physiological saline solution, the amount of amoxicillin dropped below 10% or even below the detection limit. Thus, a physiological saline solution is best suited for the storage of dissolved amoxicillin for later administration.

Argon cold atmospheric plasma eradicates pathogens in vitro that are commonly associated with canine bacterial keratitis.

Purpose: To investigate the antimicrobial effect of cold atmospheric plasma (CAP) on pathogens associated with canine bacterial keratitis. Materials and methods: Pseudomonas aeruginosa, Staphylococcus pseudintermedius, and Streptococcus canis strains, which were obtained from dogs with infectious keratitis, were subjected to testing. For each species, four isolates and a reference strain were cultivated on Columbia sheep blood agar and treated with the kiNPen Vet® plasma pen from Neoplas GmbH, Greifswald, Germany. Various continuous treatment durations (0.5, 2, and 5 min) were applied, along with a 0.5-min treatment repeated four times at short intervals. These treatments were conducted at distances of 3 and 18 mm between the agar surface and the pen. Results: CAP treatment reduced bacterial growth in all three species. The most effective treatment duration was 5 min at 3 mm distance, resulting in inhibition zones ranging from 19 to 22 mm for P. aeruginosa, 26-45 mm for S. pseudintermedius and an overall reduction of bacterial growth for Str. canis. Inhibition zones were smaller with decreasing treatment duration and larger distance. Treatment times of 30 s repeated four times and 2 min showed comparable results. Treatment with argon alone did not lead to visible reduction of bacterial growth. Conclusion: Argon cold atmospheric plasma demonstrated a potent in vitro antimicrobial effect on P. aeruginosa, S. pseudintermedius and Str. canis strains with the latter showing the highest sensitivity.

Effect of an Oxygen-Based Mechanical Drug Delivery System on Percutaneous Permeation of Various Substances In Vitro.

Transdermal drug administration is an elegant method to overcome various side effects of oral or parenteral drug administration. Nevertheless, due to an effective skin barrier, which is provided by the stratum corneum, transdermal drug delivery is sometimes very slow and ineffective. Thus, the effect of a medical device (DERMADROP TDA) for transdermal penetration of drugs in conjunction with a special vehicle emulsion on percutaneous permeation of several substances (with different physicochemical properties) was investigated in Franz-type diffusion cells with porcine skin over 28 h. This medical device disperses pharmaceutical agents via oxygen flow through an application system, which is used in conjunction with specially developed vehicle substances. Substance permeation of various substances with different physicochemical properties (diclofenac, enrofloxacin, flufenamic acid, indomethacin, and salicylic acid) was examined after application with a pipette and with the medical device. Therefore, acceptor media samples were collected up to 28 h after drug administration. Drug concentration in the acceptor medium was determined via high-performance liquid chromatography. Enhanced permeation was observed for diclofenac, enrofloxacin, flufenamic acid, indomethacin, and salicylic acid after oxygen-based administration. This correlates negatively with the molecular weight. Thus, drug administration can effectively be enhanced by a medical device using oxygen.

FeMn with Phases of a Degradable Ag Alloy for Residue-Free and Adapted Bioresorbability.

The development of bioresorbable materials for temporary implantation enables progress in medical technology. Iron (Fe)-based degradable materials are biocompatible and exhibit good mechanical properties, but their degradation rate is low. Aside from alloying with Manganese (Mn), the creation of phases with high electrochemical potential such as silver (Ag) phases to cause the anodic dissolution of FeMn is promising. However, to enable residue-free dissolution, the Ag needs to be modified. This concern is addressed, as FeMn modified with a degradable Ag-Calcium-Lanthanum (AgCaLa) alloy is investigated. The electrochemical properties and the degradation behavior are determined via a static immersion test. The local differences in electrochemical potential increase the degradation rate (low pH values), and the formation of gaps around the Ag phases (neutral pH values) demonstrates the benefit of the strategy. Nevertheless, the formation of corrosion-inhibiting layers avoids an increased degradation rate under a neutral pH value. The complete bioresorption of the material is possible since the phases of the degradable AgCaLa alloy dissolve after the FeMn matrix. Cell viability tests reveal biocompatibility, and the antibacterial activity of the degradation supernatant is observed. Thus, FeMn modified with degradable AgCaLa phases is promising as a bioresorbable material if corrosion-inhibiting layers can be diminished.

Treatment of pigs with enrofloxacin via different oral dosage forms - environmental contaminations and resistance development of Escherichia coli.

BACKGROUND: Antibacterial agents play important roles in the treatment of bacterial infections. However, the development of antimicrobial resistance (AMR) and carry-over of substances into the environment are several problems arising during oral treatment of bacterial infections. We assessed AMR development in commensal Escherichia coli (E. coli) in enrofloxacin treated and untreated animals. In addition, we examined fluoroquinolone in the plasma and urine of treated and untreated animals, and in sedimentation dust and aerosol. METHODS: In each trial, six pigs were treated with enrofloxacin via powder, granulate or pellet forms in two time periods (days 1-5 and 22-26). Four pigs served as untreated controls. The minimum inhibitory concentration (MIC) was determined to evaluate AMR development. Analysis of enro- and ciprofloxacin was performed with high performance liquid chromatography. RESULTS: Non-wildtype E. coli (MIC > 0.125 µg/mL) was detected in the pellet treated group after the first treatment period, whereas in the other groups, non-wildtype isolates were found after the second treatment period. E. coli with MIC > 4 µg/mL was found in only the pellet trial. Untreated animals showed similar susceptibility shifts several days later. Bioavailability differed among the treatment forms (granulate > pellet > powder). Enro- and ciprofloxacin were detected in aerosols and sedimentation dust (granulate, powder > pellet). CONCLUSIONS: This study indicates that the kind of the oral dosage form of antibiotics affects environmental contamination and AMR development in commensal E. coli in treated and untreated pigs.

Innovative Perspectives on Biofilm Interactions in Poultry Drinking Water Systems and Veterinary Antibiotics Used Worldwide.

Prudent use of antibiotics in livestock is widely considered to be important to prevent antibiotic resistance. This study aimed to evaluate the interactions between biofilms and veterinary antibiotics in therapeutic concentrations administrated via drinking water through a standardized experimental setup. In this context, two biofilms formed by pseudomonads (Pseudomonas (P.) aeruginosa or P. fluorescens) and a susceptible Escherichia (E.) coli strain were developed in a nutrient-poor medium on the inner surface of polyvinyl chloride pipe pieces. Subsequently, developing biofilms were exposed to sulfadiazine/trimethoprim (SDZ/TMP) or tylosin A (TYL A) in dosages recommended for application in drinking water for 5 or 7 days, respectively. Various interactions were detected between biofilms and antibiotics. Microbiological examinations revealed that only TYL A reduced the number of bacteria on the surface of the pipes. Additionally, susceptible E. coli survived both antibiotic treatments without observable changes in the minimum inhibitory concentration to 13 relevant antibiotics. Furthermore, as demonstrated by HPLC-UV, the dynamics of SDZ/TMP and TYL A in liquid media differed between the biofilms of both pseudomonads over the exposure period. We conclude that this approach represents an innovative step toward the effective evaluation of safe veterinary antibiotic use.

Effects of Topically Applied Betulinic Acid and NVX-207 on Melanocytic Tumors in 18 Horses.

The naturally occurring betulinic acid (BA) and its derivative NVX-207 induce apoptosis in equine melanoma cells in vitro. After topical application, high concentrations of the substances can be reached in healthy equine skin. With the aim to investigate the effect and safety of topically applied BA and NVX-207 in horses with melanocytic tumors, the longitudinal, prospective, randomized, double-blind, placebo-controlled study protocol included eighteen Lipizzaner mares with early-stage cutaneous melanoma assigned to three groups. Melanocytic lesions were topically treated either with a placebo, 1% BA or 1% NVX-207 twice a day for 91 days. Caliper measurements, clinical examinations and blood tests were performed to assess the effects and safety of the medication. The topical treatment was convenient and safe. The volumes of tumors treated with BA were significantly reduced over time as compared to tumors treated with the placebo from day 80 of the study. Although treatment with NVX-207 seemed to decrease tumor volume, these results did not reach statistical significance. The findings must be regarded as preliminary due to the limited group size and need to be replicated in a larger cohort with modified pharmaceutical test formulations. Accordingly, the treatment protocol cannot yet be recommended in its current form.

Influence of the H1 Antihistamine Mepyramine on the Antibacterial Effect of Florfenicol in Pigs.

The effect of florfenicol against Escherichia coli (E. coli) was investigated in vivo to confirm results of an in vitro study of Bruer et al. (2019), which has shown positive effects of various antibacterial agents in combination with the antihistamine mepyramine (MEP). Therefore, pigs were treated in three different settings: An untreated control group, 10 mg/kg florfenicol (FFC) and 10 mg/kg FFC in combination with 20 mg/kg MEP. E. coli were isolated from faecal samples and analyzed in growth quantity and resistance to FFC. The FFC medication induced an increased number of resistant E. coli strains isolated from faecal samples. The number of colonies detected after cultivation of animal samples treated with 10 mg/kg FFC was higher than the number of colonies after treatment with 10 mg/kg FFC in combination with of FFC and MEP. Furthermore, the effect of both compounds was examined on bacterial susceptibility of Pasteurella multocida in vitro, where the combination of FFC with MEP resulted in a diminished minimum inhibitory concentration. We confirmed the development of bacterial resistance in the intestine as non-target tissue caused by the use of the antibacterial agent florfenicol. Moreover, the combination of FFC with an antihistamine like MEP offers a possibility to enhance the efficacy of an antibacterial treatment and modifies the effect on gut microbiota.

Precision-cut bovine udder slices (PCBUS) as an in-vitro-model of an early phase of infection of bovine mastitis.

BACKGROUND: The aim of this study was to establish precision-cut bovine udder slices (PCBUS) as an in-vitro-model to investigate pathophysiological processes in the early phase of mastitis in order to have the possibility to investigate new therapeutic approaches for the treatment of such udder inflammation in later studies. Furthermore, this model should contribute to substitute in-vivo-experiments. Bovine mastitis is one of the most common and costly infectious diseases in the dairy industry, which is largely associated with the use of antimicrobial agents. Given this problem of antimicrobial resistance, it is essential to step up research into bacterial infectious diseases. Thus, the transfer of the in-vitro-model of precision-cut tissue slices to the bovine udder enables broad research into new therapeutic approaches in this area and can also be used to address issues in basic research or the characterisation of complex pathophysiological processes. RESULTS: A stimulation with LPS, PGN or the combination of both substances (LPS:PGN) demonstrates the ability of the PCBUS to react with a significant secretion of IL-1ß, TNF-α and PGE2. CONCLUSION: The slices represent an instrument for investigating pharmacological interactions with udder tissue, which can be useful for studies on pharmacological questions and the understanding of complex pathophysiological processes of infection and inflammation.

In vitro assessment of triterpenoids NVX-207 and betulinyl-bis-sulfamate as a topical treatment for equine skin cancer.

Equine sarcoid (ES) is the most prevalent skin tumor in equids worldwide. Additionally, aging grey horses frequently suffer from equine malignant melanoma (EMM). Current local therapies targeting these skin tumors remain challenging. Therefore, more feasible topical treatment options should be considered. In order to develop a topical therapy against ES and EMM, betulinyl-bis-sulfamate and NVX-207, derivatives of the naturally occurring betulin and betulinic acid, respectively, were evaluated for their antiproliferative (crystal violet staining assay), cytotoxic (MTS assay) and apoptotic (AnnexinV staining, cell cycle investigations) effects on primary ES cells, EMM cells and equine dermal fibroblasts in vitro. The more potent derivative was assessed for its in vitro penetration and permeation on isolated equine skin within 30 min and 24 h using Franz-type diffusion cells and HPLC analysis. Betulinyl-bis-sulfamate and NVX-207 inhibited the proliferation and metabolism in ES cells, EMM cells and fibroblasts significantly (p < 0.001) in a time- and dose-dependent manner. NVX-207 had superior anticancer effects compared to betulinyl-bis-sulfamate. Both compounds led to the externalization of phosphatidylserines on the cell membrane and DNA fragmentation, demonstrating that the effective mode of action was apoptosis. After 48 h of treatment with NVX-207, the number of necrotic cells was less than 2% in all cell types. Detected amounts of NVX-207 in the different skin layers exceeded the half-maximal inhibitory concentrations calculated by far. Even though data obtained in vitro are auspicious, the results are not unconditionally applicable to the clinical situation. Consequently, in vivo studies are required to address the antitumoral effects of topically applied NVX-207 in ES and EMM patients.

Betulinic acid shows anticancer activity against equine melanoma cells and permeates isolated equine skin in vitro.

BACKGROUND: Equine malignant melanoma (EMM) is a frequently occurring dermoepidermal tumor in grey horses. Currently available therapies are either challenging or inefficient. Betulinic acid (BA), a naturally occurring triterpenoid, is a promising compound for cancer treatment. To evaluate the potential of BA as a topical therapy for EMM, its anticancer effects on primary equine melanoma cells and dermal fibroblasts and its percutaneous permeation through isolated equine skin were assessed in vitro. RESULTS: BA showed antiproliferative and cytotoxic effects on both primary equine melanoma cells and fibroblasts in a time- and dose-dependent manner. The lowest half-maximal inhibitory concentrations were obtained 96 h after the beginning of drug exposure (12.7 µmol/L and 23.6 µmol/L for melanoma cells eRGO1 and MelDuWi, respectively, in cytotoxicity assay). High concentrations of the compound were reached in the required skin layers in vitro. CONCLUSION: BA is a promising substance for topical EMM treatment. Further clinical studies in horses are necessary to assess safety and antitumoral effects in vivo.

Low-frequency electromagnetic fields as an alternative to sanitize water of drinking systems in poultry production?

Low-frequency electromagnetic fields (LF-EMF) may present an alternative to conventional sanitation methods of water supply lines in animal production. The objective of this study was to evaluate the effect of the application of LF-EMF on bacterial concentrations and biofilms at scale-models of different drinking systems (circulating and non-circulating) conventionally used in poultry holdings. Treated systems were equipped with commercial devices producing pulsed electromagnetic signals of low frequency up to 10,000 Hz; max. 21 mT. Exposure of water to LF-EMF resulted in changes of the culturable bacterial counts, although with high standard deviations. Differing between systems types, LF-EMF treatment seemed to be responsible either for a limitation or for an increase of colony forming unit counts, with partly statistically significant differences, especially in early stages of treatment. In contrast, neither biofilm formation nor counts of cells suspended in water differed between treated and control lines over 28 days of experiment, as determined by fluorescence microscopy. Although this study indicates that LF-EMF may influence culturability of water microorganisms, no clear inhibitory effects on bacterial biofilm formation or on planktonic microbes by LF-EMF treatment were confirmed in the experiments.

In vitro and in vivo accumulation of magnetic nanoporous silica nanoparticles on implant materials with different magnetic properties.

BACKGROUND: In orthopedic surgery, implant-associated infections are still a major problem. For the improvement of the selective therapy in the infection area, magnetic nanoparticles as drug carriers are promising when used in combination with magnetizable implants and an externally applied magnetic field. These implants principally increase the strength of the magnetic field resulting in an enhanced accumulation of the drug loaded particles in the target area and therewith a reduction of the needed amount and the risk of undesirable side effects. In the present study magnetic nanoporous silica core-shell nanoparticles, modified with fluorophores (fluorescein isothiocyanate/FITC or rhodamine B isothiocyanate/RITC) and poly(ethylene glycol) (PEG), were used in combination with metallic plates of different magnetic properties and with a magnetic field. In vitro and in vivo experiments were performed to investigate particle accumulation and retention and their biocompatibility. RESULTS: Spherical magnetic silica core-shell nanoparticles with reproducible superparamagnetic behavior and high porosity were synthesized. Based on in vitro proliferation and viability tests the modification with organic fluorophores and PEG led to highly biocompatible fluorescent particles, and good dispersibility. In a circular tube system martensitic steel 1.4112 showed superior accumulation and retention of the magnetic particles in comparison to ferritic steel 1.4521 and a Ti90Al6V4 control. In vivo tests in a mouse model where the nanoparticles were injected subcutaneously showed the good biocompatibility of the magnetic silica nanoparticles and their accumulation on the surface of a metallic plate, which had been implanted before, and in the surrounding tissue. CONCLUSION: With their superparamagnetic properties and their high porosity, multifunctional magnetic nanoporous silica nanoparticles are ideal candidates as drug carriers. In combination with their good biocompatibility in vitro, they have ideal properties for an implant directed magnetic drug targeting. Missing adverse clinical and histological effects proved the good biocompatibility in vivo. Accumulation and retention of the nanoparticles could be influenced by the magnetic properties of the implanted plates; a remanent martensitic steel plate significantly improved both values in vitro. Therefore, the use of magnetizable implant materials in combination with the magnetic nanoparticles has promising potential for the selective treatment of implant-associated infections.

Influence of dietary zinc on the claw and interdigital skin of sheep.

Claw diseases like interdigital dermatitis and footrot threaten sheep health and are major welfare issues. Several studies mainly done in cattle suggested that zinc (Zn) supplementation may improve claw integrity. However, Zn supplements may differ markedly regarding Zn bioavailability. Zn bound to single amino acids has been shown to be more bioavailable than inorganic Zn sources. The aim of this study was to determine the effect of different Zn supplements on the integrity of the claw and interdigital skin of healthy sheep. At weaning 30 Merino lambs were randomly allocated to three different dietary treatments which were provided through the pelleted concentrates as follows: 1) no supplemental Zn (Zn0); 2) addition of 40 mg/kg Zn as Zn sulphate (ZnS); 3) addition of 40 mg/kg organic Zn as Zn amino acid complex (CZn). Barley straw and pelleted concentrates were given ad-libitum. The calculated Zn concentration of the total diet (roughage and concentrate) without supplemental Zn (Zn0) was 38 mg Zn/kg DM. The concentrates were formulated to meet the nutritional requirements for growing lambs and contained 207 g/kg DM crude protein and 12.4 MJ/kg DM metabolizable energy. After 8 weeks the lambs were slaughtered and the following specimens were collected: blood serum, liver, sole and coronary band of the claw, and interdigital skin. Serum and tissue Zn and copper (Cu) concentrations and claw hardness were determined. Routine pathohistology and electron microscopy were conducted. Franz diffusion cell system and Ki-67 immunostaining were used to determine the permeability of the interdigital skin and the keratinocyte proliferation in the basal layer of sole horn, coronary band and interdigital skin, respectively. The concentrations of Zn and Cu in serum and liver tissue as well as the Zn concentration in claw horn were not affected by dietary treatment. Zn0 lambs showed higher (p < 0.05) Cu concentrations in claw horn compared to both Zn supplemented groups. Routine pathohistology as well as electron microscopy did not show significant morphological differences between the three groups. Franz diffusion cell system proved to be a suitable method examining the interdigital skin permeability, but the group differences in this study were not significant. Dietary treatment did not affect keratinocyte proliferation in the coronary band. In the sole keratinocyte proliferation was significantly higher (p < 0.05) in the Zn0 group compared to CZn with ZnS being intermediate. Keratinocyte proliferation in the interdigital skin was significantly higher (p < 0.05) in the CZn group compared to the Zn0 with ZnS being intermediate. The results of the current experiment indicate that serum and tissue Zn concentrations and horn hardness are not affected by adding a moderate amount of Zn sulphate or Zn amino acid complex to a basal diet. However, supplemental Zn amino acid complex seems to affect keratinocyte proliferation of interdigital skin and sole horn of lambs. Effects on skin permeability should be retested using a higher number of animals prospectively.

Resistance of Escherichia coli in Turkeys after Therapeutic or Environmental Exposition with Enrofloxacin Depending on Flooring.

Gaining knowledge about the spread of resistance against antibacterial agents is a primary challenge in livestock farming. The purpose of this study was to test the effect of double antibiotic treatment (at days 10⁻14 and days 26⁻30) with enrofloxacin or solely environmental exposition (identical times, directly into the litter) on resistance against antibacterial agents in commensal Escherichia coli in comparison with the control (without treatment), depending on different flooring. A total of 720 Big 6 turkeys participated in three trials. Four different flooring designs were examined: An entire floor pen covered with litter, a floor pen with heating, a partially slatted flooring including 50% littered area, and a fully slatted flooring with a sand bath. A total of 864 Escherichia coli isolates were obtained from cloacal swabs and poultry manure samples at days 2, 9, 15, 21, and 35. The broth microdilution method (MIC) was used to determine the resistance of isolates to enrofloxacin and ampicillin. A double antibiotic treatment with enrofloxacin reduced the proportion of susceptible Escherichia coli isolates significantly in all flooring designs. Simulation of water losses had no significant effect, nor did the flooring design. Ampicillin-resistant isolates were observed, despite not using ampicillin.