Staphylococcus aureus AbcA transporter enhances persister formation under ß-lactam exposure.

We evaluated the role of Staphylococcus aureus AbcA transporter in bacterial persistence and survival following exposure to the bactericidal agents nafcillin and oxacillin at both the population and single-cell levels. We show that AbcA overexpression resulted in resistance to nafcillin but not oxacillin. Using distinct fluorescent reporters of cell viability and AbcA expression, we found that over 6-14 hours of persistence formation, the proportion of AbcA reporter-expressing cells assessed by confocal microscopy increased sixfold as cell viability reporters decreased. Similarly, single-cell analysis in a high-throughput microfluidic system found a strong correspondence between antibiotic exposure and AbcA reporter expression. Persister cells grown in the absence of antibiotics showed neither an increase in nafcillin MIC nor in abcA transcript levels, indicating that survival was not associated with stable mutational resistance or abcA overexpression. Furthermore, persister cell levels on exposure to 1×MIC and 25×MIC of nafcillin decreased in an abcA knockout mutant. Survivors of nafcillin and oxacillin treatment overexpressed transporter AbcA, contributing to an enrichment of the number of persisters during treatment with pump-substrate nafcillin but not with pump-non-substrate oxacillin, indicating that efflux pump expression can contribute selectively to the survival of a persister population.

Complete genomic characterization of bovine papillomavirus type 1 and 2 strains infers ongoing cross-species transmission between cattle and horses.

Infection with bovine papillomavirus (BPV) types 1 and 2 results in the most common skin tumor of horses, termed equine sarcoid. The persistent and recurrent nature of this tumor stands in contrast to the regressive nature of BPV-1/- 2 induced cutaneous papillomas in cattle. The circulation of horse-specific BPV-1/- 2 variants within equine populations has been suggested as a possible explanation for the difference in clinical presentation of BPV-1/- 2 infection between horses and cattle. In order to investigate this hypothesis, we identified 98 complete BPV-1/- 2 genomes using a Nanopore sequencing approach. Separate BPV-1/- 2 alignments were used to infer Bayesian phylogenetic trees. Phylogeny-trait association concerning host species was investigated using Bayesian Tip-association Significance software (BaTS) Overall, 179 unique BPV-1 and 128 BPV-2 substitutions were found. The E2 coding region in the viral genome exhibited an exceptionally high rate of non-synonymous mutations (81 %, n = 13/16). Interestingly, extensive deletions in the L1/L2 region (up to 1.5 kb) were found exclusively in horse-derived samples. Nevertheless, the most frequently detected single nucleotide polymorphisms were shared between equine and bovine hosts, which is in agreement with BaTS results indicating no phylogeny-host correlation. We found indications that horse-specific mutations might exist in subpopulations of equine derived BPV-1/- 2, but these did not result in horse-adapted genetic variants. Based on these observations, cross-species transmission from cattle to horses seems to be an ongoing process, rather than an ancient occurrence that has been followed by the circulation of horse-adapted BPV variants in the horse population..

Ocular adverse events associated with immune checkpoint inhibitors, a scoping review.

INTRODUCTION: Immune checkpoint inhibitors (ICIs) have become an important part of the treatment of multiple cancers, especially for advanced melanoma and non-small cell lung cancer. Some tumors are capable of escaping immunosurveillance by stimulating checkpoints on T-cells. ICIs prevent activation of these checkpoints and thereby stimulate the immune system and indirectly the anti-tumor response. However, the use of ICIs is associated with various adverse events. Ocular side effects are rare but may have a major impact on the quality of life of the patient. METHODS: A comprehensive literature search of the medical databases Web of Science, Embase and PubMed was performed. Articles that provided a comprehensive description of a case report containing 1) cancer patient(s) treated with (a combination of) immune checkpoint inhibitors, and 2) assessed occurrence of ocular adverse events, were included. A total of 290 case reports were included. RESULTS: Melanoma (n = 179; 61.7%) and lung cancer (n = 56; 19.3%) were the most frequent reported malignancies. The primary used ICIs were nivolumab (n = 123; 42.5%) and ipilimumab (n = 116; 40.0%). Uveitis was most the common adverse event (n = 134; 46.2%) and mainly related to melanoma. Neuro-ophthalmic disorders, including myasthenia gravis and cranial nerve disorders, were the second most common adverse events (n = 71; 24.5%), mainly related to lung cancer. Adverse events affecting the orbit and the cornea were reported in 33 (11.4%) and 30 cases (10.3%) respectively. Adverse events concerning the retina were reported in 26 cases (9.0%). CONCLUSION: The aim of this paper is to provide an overview of all reported ocular adverse events related to the use of ICIs. The insights retrieved from this review might contribute to a better understanding of the underlying mechanisms of these ocular adverse events. Particularly, the difference between actual immune-related adverse events and paraneoplastic syndromes might be relevant. These findings might be of great value in establishing guidelines on how to manage ocular adverse events related to ICIs.

Diffraction of Gaussian and Laguerre-Gauss beams from a circular aperture using the moment expansion method.

A method based on the distribution theory is introduced to compute the Fresnel diffraction integral. It is applied to the diffraction of Gaussian and Laguerre-Gauss beams by a circular aperture. Expressions of the diffracting field are recast into a perturbation series describing the near- and far-field regions.

Evidence of Partial Seniority Conservation in the πg_{9/2} Shell for the N=50 Isotones.

The reduced transition probabilities for the 4_{1}^{+}→2_{1}^{+} and 2_{1}^{+}→0_{1}^{+} transitions in ^{92}Mo and ^{94}Ru and for the 4_{1}^{+}→2_{1}^{+} and 6_{1}^{+}→4_{1}^{+} transitions in ^{90}Zr have been determined in this experiment making use of a multinucleon transfer reaction. These results have been interpreted on the basis of realistic shell-model calculations in the f_{5/2}, p_{3/2}, p_{1/2}, and g_{9/2} proton valence space. Only the combination of extensive lifetime information and large scale shell-model calculations allowed the extent of the seniority conservation in the N=50 g_{9/2} orbital to be understood. The conclusion is that seniority is largely conserved in the first πg_{9/2} orbital.

Design of an electrospun tubular construct combining a mechanical and biological approach to improve tendon repair.

Hand tendon injuries represent a major clinical problem and might dramatically diminish a patient's life quality. In this study, a targeted solution for flexor tendon repair was developed by combining a mechanical and biological approach. To this end, a novel acrylate-endcapped urethane-based polymer (AUP) was synthesized and its physico-chemical properties were characterized. Next, tubular repair constructs were developed using electrospinning of the AUP material with incorporated naproxen and hyaluronic acid (i.e. anti-inflammatory and anti-adhesion compounds, respectively), and with a tubular braid as mechanical reinforcement. Tensile testing of the repair constructs using ex vivo sheep tendons showed that the developed repair constructs fulfilled the required mechanical properties for tendon repair (i.e. minimal ultimate stress of 4 MPa), with an ultimate stress of 6.4 ± 0.6 MPa. Moreover, in vitro biological assays showed that the developed repair tubes and the incorporated bioactive components were non-cytotoxic. In addition, when equine tenocytes and mesenchymal stem cells were co-cultured with the repair tubes, an increased production of collagen and non-collagenous proteins was observed. In conclusion, this novel construct in which a mechanical approach (fulfilling the required mechanical properties) was combined with a biological approach (incorporation of bioactive compounds), shows potential as flexor tendon repair application. Graphical abstract.

Objective frequency-specific hearing thresholds definition for medicolegal purposes in case of occupational NIHL: ASSR outperforms CERA.

Audiological use of the 40 Hz-ASSR (auditory steady state responses) could be valuable for objectively estimating the frequency-specific threshold in adults undergoing an expertise examination for medicolegal and/or compensation purposes. The present prospective study was set up to clarify the relationship between the thresholds obtained by cortical evoked response audiometry (CERA) and by 40 Hz-ASSR, in the same ears, within a large homogeneous sample of 164 subjects (328 ears) with NIHL and well documented exposure to noise. All these subjects claimed financial compensation for occupational NIHL, and there was a suspicion of exaggeration of the reported NIHLs. ASSR thresholds show a good correlation with the CERA thresholds. However, a systematic shift is noticed, ASSR thresholds being on average (1-2 - 3 kHz) 4.38 dB lower (i.e. showing less hearing loss) than CERA thresholds. Moreover, the binaural multiple ASSR technique allows a considerable time gain when compared to the CERA.

Angular momentum generation in nuclear fission.

When a heavy atomic nucleus splits (fission), the resulting fragments are observed to emerge spinning1; this phenomenon has been a mystery in nuclear physics for over 40 years2,3. The internal generation of typically six or seven units of angular momentum in each fragment is particularly puzzling for systems that start with zero, or almost zero, spin. There are currently no experimental observations that enable decisive discrimination between the many competing theories for the mechanism that generates the angular momentum4-12. Nevertheless, the consensus is that excitation of collective vibrational modes generates the intrinsic spin before the nucleus splits (pre-scission). Here we show that there is no significant correlation between the spins of the fragment partners, which leads us to conclude that angular momentum in fission is actually generated after the nucleus splits (post-scission). We present comprehensive data showing that the average spin is strongly mass-dependent, varying in saw-tooth distributions. We observe no notable dependence of fragment spin on the mass or charge of the partner nucleus, confirming the uncorrelated post-scission nature of the spin mechanism. To explain these observations, we propose that the collective motion of nucleons in the ruptured neck of the fissioning system generates two independent torques, analogous to the snapping of an elastic band. A parameterization based on occupation of angular momentum states according to statistical theory describes the full range of experimental data well. This insight into the role of spin in nuclear fission is not only important for the fundamental understanding and theoretical description of fission, but also has consequences for the γ-ray heating problem in nuclear reactors13,14, for the study of the structure of neutron-rich isotopes15,16, and for the synthesis and stability of super-heavy elements17,18.

The use of equine chondrogenic-induced mesenchymal stem cells as a treatment for osteoarthritis: A randomised, double-blinded, placebo-controlled proof-of-concept study.

BACKGROUND: There is a need to improve therapies for osteoarthritis in horses. OBJECTIVES: To assess the efficacy of equine allogeneic chondrogenic-induced mesenchymal stem cells combined with equine allogeneic plasma as a novel therapy for osteoarthritis in horses. STUDY DESIGN: Randomised, double-blinded, placebo-controlled experiment. METHODS: In 12 healthy horses, osteoarthritis was induced in the metacarpophalangeal joint using an osteochondral fragment-groove model. Five weeks after surgery, horses were randomly assigned to either an intra-articular injection with chondrogenic-induced mesenchymal stem cells + equine allogeneic plasma (= intervention) or with 0.9% saline solution (= control). From surgery until the study end, horses underwent a weekly joint and lameness assessment. Synovial fluid was collected for cytology and biomarker analysis before surgery and at Weeks 5, 5 + 1d, 7, 9 and 11. At Week 11, horses were subjected to euthanasia, and the metacarpophalangeal joints were evaluated macroscopically and histologically. RESULTS: No serious adverse events or suspected adverse drug reactions occurred during the study. A significant improvement in visual and objective lameness was seen with the intervention compared with the control. Synovial fluid displayed a significantly higher viscosity and a significantly lower glycosaminoglycan concentration in the intervention group. Other biomarkers or cytology parameters were not significantly different between the treatment groups. Significantly less wear lines and synovial hyperaemia were present in the intervention group. The amount of cartilage oligomeric matrix protein, collagen type II and glycosaminoglycans were significantly higher in the articular cartilage of the intervention group. MAIN LIMITATIONS: This study assessed the short-term effect of the intervention on a limited number of horses, using an osteoarthritis model. This study also included multiple statistical tests, increasing the risk of type 1 error. CONCLUSIONS: Equine allogeneic chondrogenic-induced mesenchymal stem cells combined with equine allogeneic plasma may be a promising treatment for osteoarthritis in horses. The Summary is available in Spanish - see Supporting Information.

Limited parasite acquisition by non-native Lepomis gibbosus (Actinopterygii: Centrarchidae) at two ponds in the Upper Rhine basin, Germany.

Metazoan parasite communities of Lepomis gibbosus (Centrarchidae), one of the most successfully introduced fish species in Europe, were studied at two isolated ponds (Knielingen, Tropfen) along the Upper Rhine in Germany. Nine parasite taxa were observed, including North American species co-introduced to Europe (ancyrocephalid monogeneans, diplostomid trematodes), circumpolar species infecting L. gibbosus in both their native and non-native ranges (bothriocephalid cestodes) and locally acquired parasitic nematodes. Both parasite communities consisted predominantly of North American species. Acquisition of local parasites was not observed at Tropfen, where the fish community comprised just two species, with L. gibbosus dominant. Low prevalence and abundance of acquired parasites was found at Knielingen, which supported a diverse fish community. At Tropfen, a high abundance of the North American parasite Posthodiplostomum centrarchi probably contributed to the lower condition index, hepatomegaly and splenomegaly observed. Due to low local parasite competency, L. gibbosus appears to have no significant impact on parasite dynamics in affected habitats.

The clinical diagnosis of equine sarcoids-Part 2: Assessment of case features typical of equine sarcoids and validation of a diagnostic protocol to guide equine clinicians in the diagnosis of equine sarcoids.

Research has shown that the accuracy of the clinical diagnosis of equine sarcoids (ES) can be improved. Particularly, less experienced veterinarians are often mistaken in their clinical judgement despite a high level of diagnostic confidence. The aim of this study was to develop and assess the performance of a diagnostic protocol (DP) to improve diagnostic accuracy and identify diagnostically challenging cases. The design of the DP was based on typical clinical features of ES and its algorithm was optimised through repeated tests on clinical cases prior to validating its performance in a representative online examination. A total of 22 equine practitioners and 31 veterinary students used the DP to diagnose 40 standardised ES and non-ES cases in an online examination. Scores of these 53 respondents were compared to scores of 128 respondents of comparable levels of expertise, and 14 experts, all assessing the same cases without using the DP. Overall, respondents using the DP were significantly more likely (odds ratio (OR) 1.25; 95% confidence interval (95% CI) 1.09-1.43) to diagnose a case correctly compared to respondents not using the DP and felt significantly more confident of their diagnosis (OR 1.53; 95% CI 1.39-1.67). Thus, the DP proved to be a reliable tool to increase clinical diagnostic accuracy and diagnostic confidence. The DP algorithms may be further improved with experiences gained from its application in equine practice and clinicians will be able to optimise their diagnostic accuracy and selection of lesions requiring a biopsy.

The clinical diagnosis of equine sarcoids - Part 1: Assessment of sensitivity and specificity using a multicentre case-based online examination.

Equine clinicians and researchers often make the diagnosis of equine sarcoids (ES) on clinical assessment alone, without histopathological confirmation. However, the accuracy of the clinical diagnosis of ES has not been critically assessed. To assess sensitivity, specificity, positive and negative predictive values of the clinical diagnosis of ES, 40 clinical cases with histologically confirmed equine skin lesions (26 ES and 14 non-ES) were compiled in a representative online examination. For each case and lesion, at least one photograph and all relevant information were presented in a standardised format. This included the horse's signalment, lesion localisation, lesion progression, presence of other skin lesions, earlier treatments and response to treatment. No information relevant for the assessment of the lesion was intentionally withheld. Fourteen ES experts, 39 board-certified equine specialists, 103 equine practitioners and 25 novices categorised the cases into ES or non-ES and graded their level of diagnostic confidence on a scale from 1 to 6 for each case. The overall success rate was 82.0% while sensitivity and specificity were 83.3% and 79.6%, respectively. The positive and negative predictive values were 88.4% and 72.0%, respectively, in the tested population with a 66% prevalence of ES. However, less experienced veterinarians were frequently wrong in their clinical judgement despite a high level of diagnostic confidence. Therefore, the authors propose to develop a diagnostic tool to help clinicians optimise their selection of lesions requiring a biopsy. Ultimately, this will help reduce costs and morbidity generated by unnecessary diagnostic and therapeutic efforts.

The possible role of Stomoxys calcitrans in equine sarcoid transmission.

The association between bovine papillomavirus (BPV) and equine sarcoids is well established, but it is unclear how the virus spreads. Although evidence in support of viral spread through direct animal contact exists, this does not explain sarcoid development in isolated equids. BPV DNA has been detected in flies, which could indicate that these insects serve as a vector. This study aimed to investigate whether BPV-negative stable flies (Stomoxys calcitrans) become positive for BPV DNA after exposure to equine sarcoid or bovine papilloma tissue under experimental conditions and, if so, for how long. A total of 420 stable flies were caught alive and exposed to BPV positive equine sarcoid or bovine papilloma tissue. During the following week, dead flies were collected daily and BPV loads were determined by quantitative PCR. There was a significant rise in BPV load after tissue exposure both in sarcoid and papilloma exposed flies, but the viral load was higher and remained high for a longer time after exposure to papilloma tissue compared to sarcoid tissue. Within days, viral loads decreased again and became indifferent from loads before exposure. The results of these experiments indicate that BPV transmission by S. calcitrans seems possible and is more likely to occur after contact with bovine papillomas than with equine sarcoids. Transmission seems only possible shortly after tissue exposure. Further research could include experimental induction of sarcoids with BPV positive stable flies, or a repeat of the experiment with micro-dissection prior to PCR.

Equine allogeneic chondrogenic induced mesenchymal stem cells: A GCP target animal safety and biodistribution study.

The safety of the intra-articular use of mesenchymal stem cells (MSCs) is scarcely reported. Therefore, the goal of this study was to investigate the safety of a single intra-articular injection with allogeneic chondrogenic induced MSCs combined with equine plasma (=the investigational product: IVP) compared to a saline (0.9% NaCl) placebo control (=control product: CP). Sixteen healthy experimental horses were randomly assigned to receive a single intra-articular injection with either the IVP (n=8) or the CP (n=8) in the left metacarpophalangeal joint. All horses underwent a daily clinical assessment throughout the entire study period of 42days to assess adverse events. Additionally, a local joint assessment and a lameness examination were performed daily during the first two weeks, and weekly the following 4weeks. Blood samples were taken weekly for hematological and biochemical analysis. At the end of the study period, horses of the IVP group were euthanized for a thorough necropsy and to check for biodistribution. Tissue samples of the injected joint were collected for histological examination. In both CP and IVP treated horses a mild transient subjective increase in periarticular temperature and lameness was noted after the intra-articular injection with no significant differences between the treatment groups. No distribution of the cells was found using immunohistochemistry and no ectopic tissue formation or signs of inflammation were found on histology. A single intra-articular injection of allogeneic chondrogenic induced MSCs combined with allogeneic plasma in horses had the same clinical side effects as an intra-articular injection with saline solution.

Foramen epiploicum mesh closure (FEMC) through a ventral midline laparotomy.

BACKGROUND: Preventive laparoscopic closure of the epiploic foramen (EF) has previously been described, but methods for EF closure during laparotomy for treatment of colic are lacking. OBJECTIVES: To describe the foramen epiploicum mesh closure (FEMC) technique through a ventral midline laparotomy in horses under general anaesthesia and its outcome. STUDY DESIGN: Terminal surgical experiment and in vivo study. METHODS: In a pilot study, a standard ventral midline laparotomy was performed under general anaesthesia in 10 experimental horses and a three-dimensional expandable diabolo-shaped mesh construct was manually introduced through the EF into the omental vestibule. The laparotomy incision was routinely closed after which the horses were subjected to euthanasia. Subsequently, the cadavers were extensively manipulated and finally mesh position was verified during necropsy. Thereafter, a ventral midline laparotomy, with simulation of abdominal manipulations during colic surgery was performed on six experimental horses. A three-dimensional expandable diabolo-shaped mesh with a radiological marker was introduced as in the pilot study. Clinical and laboratory parameters were recorded during the post-operative period. Four weeks after the intervention, abdominal radiography and laparoscopy were performed in all six horses. RESULTS: Insertion of all 16 diabolo-shaped mesh constructs during laparotomy was uneventful. The 10 mesh constructs of the pilot study were positioned correctly at necropsy. No post-operative clinical or laboratory parameter alterations were noticed in the other six horses, apart from a transient leucocytosis and an increase in serum amyloid A. Abdominal radiography at 4 weeks revealed consistent positioning of the mesh marker. Laparoscopy confirmed that all six EFs were closed, without intestinal adhesions. MAIN LIMITATIONS: Mesh insertion in clinical cases and during clean contaminated surgery was not investigated. CONCLUSION: The FEMC technique via laparotomy provides a fast, simple and reliable procedure to obliterate the EF and may be useful during EFE surgery to prevent recurrence of the disease, avoiding a subsequent laparoscopic procedure.

An atlas of bloodstream-accessible bone marrow proteins for site-directed therapy of acute myeloid leukemia.

The concept of arming antibodies with bioactive payloads for a site-specific therapy of cancer has gained considerable interest in recent years. However, a successful antibody-based targeting approach critically relies on the availability of a tumor-associated target that is not only preferentially expressed in the tumor tissue but is also easily accessible for antibody therapeutics coming from the bloodstream. Here, we perfused the vasculature of healthy and acute myeloid leukemia (AML)-bearing rats with a reactive ester derivative of biotin and subsequently quantified the biotinylated proteins to identify AML-associated bone marrow (BM) antigens accessible from the bloodstream. In total, >1400 proteins were identified. Overall, 181 proteins were >100-fold overexpressed in AML as compared with normal BM. Eleven of the most differentially expressed proteins were further validated by immunohistochemistry and confocal microscopic analyses, including novel antigens highly expressed in AML cells (for example, adaptor-related protein complex 3 ß2) and in the leukemia-modified extracellular matrix (ECM) (for example, collagen-VI-α-1). The presented atlas of targetable AML-associated BM proteins provides a valuable basis for the development of monoclonal antibodies that could be used as carriers for a site-specific pharmacodelivery of cytotoxic drugs, cytokines or radionuclides to the BM in AML.

A quantitative swab is a good non-invasive alternative to a quantitative biopsy for quantifying bacterial load in wounds healing by second intention in horses.

The aim of this study was to evaluate different techniques for diagnosing wound infection in wounds healing by second intention in horses and to assess the effect of a vortex and sonication protocol on quantitative bacteriology in specimens with a histologically confirmed biofilm. In 50 wounds healing by second intention, a clinical assessment, a quantitative swab, a semi-quantitative swab, and a swab for cytology were compared to a quantitative tissue biopsy (reference standard). Part of the biopsy specimen was examined histologically for evidence of a biofilm. There was a significant, high correlation (P<0.001; r=0.747) between the outcome of the quantitative swabs and the quantitative biopsies. The semi-quantitative swabs showed a significant, moderate correlation with the quantitative biopsies (P<0.001; ρ=0.524). Higher white blood cell counts for cytology were significantly associated with lower log10 colony-forming units (CFU) in the wounds (P=0.02). Wounds with black granulation tissue showed significantly higher log10 CFU (P=0.003). Specimens with biofilms did not yield higher bacteriological counts after a vortex and sonication protocol was performed to release bacteria from the biofilm. Based on these findings, a quantitative swab is an acceptable non-invasive alternative to a quantitative biopsy for quantifying bacterial load in equine wounds healing by second intention.

Anomalies in the Charge Yields of Fission Fragments from the

Fast-neutron-induced fission of ^{238}U at an energy just above the fission threshold is studied with a novel technique which involves the coupling of a high-efficiency γ-ray spectrometer (MINIBALL) to an inverse-kinematics neutron source (LICORNE) to extract charge yields of fission fragments via γ-γ coincidence spectroscopy. Experimental data and fission models are compared and found to be in reasonable agreement for many nuclei; however, significant discrepancies of up to 600% are observed, particularly for isotopes of Sn and Mo. This indicates that these models significantly overestimate the standard 1 fission mode and suggests that spherical shell effects in the nascent fission fragments are less important for low-energy fast-neutron-induced fission than for thermal neutron-induced fission. This has consequences for understanding and modeling the fission process, for experimental nuclear structure studies of the most neutron-rich nuclei, for future energy applications (e.g., Generation IV reactors which use fast-neutron spectra), and for the reactor antineutrino anomaly.

S-shaped non-paraxial corrections to general astigmatic beams.

Non-paraxial perturbation wave equations are solved for general astigmatic Gaussian beams for the first time, to the best of our knowledge, in the angular spectrum representation by taking into account generic boundary conditions. Expressions for second-order corrections are derived and exemplified with an optical cavity made of two cylindrical mirrors. Non-paraxial corrections can lead, depending on the choice of boundary conditions, to a transverse S-shaped beam mode, which has been qualitatively been observed in a highly divergent non-planar four-mirror cavity.