Modeling Staphylococcus epidermidis-Induced Non-Unions: Subclinical and Clinical Evidence in Rats

S. epidermidis is one of the leading causes of orthopaedic infections associated with biofilm formation on implant devices. Open fractures are at risk of S. epidermidis transcutaneous contamination leading to higher non-union development compared to closed fractures. Although the role of infection in delaying fracture healing is well recognized, no in vivo models investigated the impact of subclinical low-grade infections on bone repair and non-union. We hypothesized that the non-union rate is directly related to the load of this commonly retrieved pathogen and that a low-grade contamination delays the fracture healing without clinically detectable infection. Rat femurs were osteotomized and stabilized with plates. Fractures were infected with a characterized clinical-derived methicillin-resistant S. epidermidis (10³, 10⁵, 10⁸ colony forming units) and compared to uninfected controls. After 56 days, bone healing and osteomyelitis were clinically assessed and further evaluated by micro-CT, microbiological and histological analyses. The biofilm formation was visualized by scanning electron microscopy. The control group showed no signs of infection and a complete bone healing. The 10³ group displayed variable response to infection with a 67% of altered bone healing and positive bacterial cultures, despite no clinical signs of infection present. The 10⁵ and 10⁸ groups showed severe signs of osteomyelitis and a non-union rate of 83–100%, respectively. The cortical bone reaction related to the periosteal elevation in the control group and the metal scattering detected by micro-CT represented limitations of this study. Our model showed that an intra-operative low-grade S. epidermidis contamination might prevent the bone healing, even in the absence of infectious signs. Our findings also pointed out a dose-dependent effect between the S. epidermidis inoculum and non-union rate. This pilot study identifies a relevant preclinical model to assess the role of subclinical infections in orthopaedic and trauma surgery and to test specifically designed diagnostic, prevention and therapeutic strategies.     

By Regulating Mitochondrial Ca2+-Uptake UCP2 Modulates Intracellular Ca2+

Introduction

The possible role of UCP2 in modulating mitochondrial Ca²⁺-uptake (mCa²⁺-uptake) via the mitochondrial calcium uniporter (MCU) is highly controversial.

Methods

Thus, we analyzed mCa²⁺-uptake in isolated cardiac mitochondria, MCU single-channel activity in cardiac mitoplasts, dual Ca²⁺-transients from mitochondrial ((Ca²⁺)m) and intracellular compartment ((Ca²⁺)c) in the whole-cell configuration in cardiomyocytes of wild-type (WT) and UCP2^-/- mice.

Results

Isolated mitochondria showed a Ru360 sensitive mCa²⁺-uptake, which was significantly decreased in UCP2^-/- (229.4±30.8 FU vs. 146.3±23.4 FU, P<0.05). Single-channel registrations confirmed a Ru360 sensitive voltage-gated Ca²⁺-channel in mitoplasts, i.e. mCa1, showing a reduced single-channel activity in UCP2^-/- (Po,total: 0.34±0.05% vs. 0.07±0.01%, P<0.05). In UCP2^-/- cardiomyocytes (Ca²⁺)m was decreased (0.050±0.009 FU vs. 0.021±0.005 FU, P<0.05) while (Ca²⁺)c was unchanged (0.032±0.002 FU vs. 0.028±0.004 FU, P>0.05) and transsarcolemmal Ca²⁺-influx was inhibited suggesting a possible compensatory mechanism. Additionally, we observed an inhibitory effect of ATP on mCa²⁺-uptake in WT mitoplasts and (Ca²⁺)m of cardiomyocytes leading to an increase of (Ca²⁺)c while no ATP dependent effect was observed in UCP2^-/-.

Conclusion

Our results indicate regulatory effects of UCP2 on mCa²⁺-uptake. Furthermore, we propose, that previously described inhibitory effects on MCU by ATP may be mediated via UCP2 resulting in changes of excitation contraction coupling.     

A monoclonal antibody specific for prophase phosphorylation of histone deacetylase 1: a readout for early mitotic cells

Histone deacetylases (HDACs) are modification enzymes that regulate a plethora of biological processes. HDAC1, a crucial epigenetic modifier, is deregulated in cancer and subjected to a variety of post-translational modifications. Here, we describe the generation of a new monoclonal antibody that specifically recognizes a novel highly dynamic prophase phosphorylation of serine 406-HDAC1, providing a powerful tool for detecting early mitotic cells.     

Who Are the High-Cost Users? A Method for Person-Centred Attribution of Health Care Spending

Objective

To develop person-centered episodes of care (PCE) for community-dwelling individuals in the top fifth percentile of Ontario health care expenditures in order to: (1) describe the main clinical groupings for spending; and (2) identify patterns of spending by health sector (e.g. acute care, home care, physician billings) within and across PCE.

Data sources

Data were drawn from population-based administrative databases for all publicly funded health care in Ontario, Canada in 2010/11.

Study design

This study is a retrospective cohort study.

Data collection/extraction methods

A total of 587,982 community-dwelling individuals were identified among those accounting for the top 5% of provincial health care expenditures between April 1, 2010 and March 31, 2011. PCE were defined as starting with an acute care admission and persisting through subsequent care settings and providers until individuals were without health system contact for 30 days. PCE were classified according to the clinical grouping for the initial admission. PCE and non-PCE costs were calculated and compared to provide a comprehensive measurement of total health system costs for the year.

Principal findings

Among this community cohort, 697,059 PCE accounted for nearly 70% ($11,815.3 million (CAD)) of total annual publicly-funded expenditures on high-cost community-dwelling individuals. The most common clinical groupings to start a PCE were Acute Planned Surgical (35.2%), Acute Unplanned Medical (21.0%) and Post-Admission Events (10.8%). Median PCE costs ranged from $3,865 (IQR = $1,712-$10,919) for Acute Planned Surgical to $20,687 ($12,207-$39,579) for Post-Admission Events. Inpatient acute ($8,194.5 million) and inpatient rehabilitation ($434.6 million) health sectors accounted for the largest proportions of allocated PCE spending over the year.

Conclusions

Our study provides a novel methodological approach to categorize high-cost health system users into meaningful person-centered episodes. This approach helps to explain how costs are attributable within individuals across sectors and has applications in episode-based payment formulas and quality monitoring.     

Antimicrobial Combinations against Pan-Resistant Acinetobacter baumannii Isolates with Different Resistance Mechanisms

The study investigated the effect of antibiotic combinations against 20 clinical isolates of A. baumannii (seven colistin-resistant and 13 colistin-susceptible) with different resistance mechanisms. Clinical data, treatment, and patient mortality were evaluated. The following methods were used: MIC, PCRs, and outer membrane protein (OMP) analysis. Synergy was investigated using the checkerboard and time-kill methods. Clonality was evaluated by PFGE. Based on clonality, the whole genome sequence of six A. baumannii isolates was analyzed. All isolates were resistant to meropenem, rifampicin, and fosfomycin. OXA-23 and OXA-143 were the most frequent carbapenemases found. Four isolates showed loss of a 43kDa OMP_. The colistin-susceptible isolates belonged to different clones and showed the highest synergistic effect with fosfomycin-amikacin. Among colistin-resistant isolates, the highest synergistic effect was observed with the combinations of colistin-rifampicin followed by colistin-vancomycin. All colistin-resistant isolates harbored bla_OXA-23-like and belonged to CC113. Clinical and demographic data were available for 18 of 20 patients. Fourteen received treatment and eight patients died during treatment. The most frequent site of infection was the blood in 13 of 14 patients. Seven patients received vancomycin plus an active drug against A. baumannii; however, mortality did not differ in this group. The synergistic effect was similar for colistin-susceptible isolates of distinct clonal origin presenting with the same resistance mechanism. Overall mortality and death during treatment was high, and despite the high synergism in vitro with vancomycin, death did not differ comparing the use or not of vancomycin plus an active drug against A. baumannii.     

Smartphone-Based Distributed Data Collection Enables Rapid Assessment of Shorebird Habitat Suitability

Understanding and managing dynamic coastal landscapes for beach-dependent species requires biological and geological data across the range of relevant environments and habitats. It is difficult to acquire such information; data often have limited focus due to resource constraints, are collected by non-specialists, or lack observational uniformity. We developed an open-source smartphone application called iPlover that addresses these difficulties in collecting biogeomorphic information at piping plover (Charadrius melodus) nest sites on coastal beaches. This paper describes iPlover development and evaluates data quality and utility following two years of collection (n = 1799 data points over 1500 km of coast between Maine and North Carolina, USA). We found strong agreement between field user and expert assessments and high model skill when data were used for habitat suitability prediction. Methods used here to develop and deploy a distributed data collection system have broad applicability to interdisciplinary environmental monitoring and modeling.     

Can Nocturnal Flight Calls of the Migrating Songbird,American Redstart,Encode Sexual Dimorphism and Individual Identity?

Bird species often use flight calls to engage in social behavior, for instance maintain group cohesion and to signal individual identity, kin or social associations, or breeding status of the caller. Additional uses also exist, in particular among migrating songbirds for communication during nocturnal migration. However, our understanding of the information that these vocalizations convey is incomplete, especially in nocturnal scenarios. To examine whether information about signaler traits could be encoded in flight calls we quantified several acoustic characteristics from calls of a nocturnally migrating songbird, the American Redstart. We recorded calls from temporarily captured wild specimens during mist-netting at the Powdermill Avian Research Center in Rector, PA. We measured call similarity among and within individuals, genders, and age groups. Calls from the same individual were significantly more similar to one another than to the calls of other individuals, and calls were significantly more similar among individuals of the same sex than between sexes. Flight calls from hatching-year and after hatching-year individuals were not significantly different. Our results suggest that American Redstart flight calls may carry identifiers of gender and individual identity. To our knowledge, this is the first evidence of individuality or sexual dimorphism in the flight calls of a migratory songbird. Furthermore, our results suggest that flight calls may have more explicit functions beyond simple group contact and cohesion. Nocturnal migration may require coordination among numerous individuals, and the use of flight calls to transmit information among intra- and conspecifics could be advantageous. Applying approaches that account for such individual and gender information may enable more advanced research using acoustic monitoring.     

Effects of Subinhibitory Concentrations of Ceftaroline on Methicillin-Resistant Staphylococcus aureus (MRSA) Biofilms

Ceftaroline (CPT) is a novel cephalosporin with in vitro activity against Staphylococcus aureus. Ceftaroline exhibits a level of binding affinity for PBPs in S. aureus including PBP2a of methicillin-resistant S. aureus (MRSA). The aims of this study were to investigate the morphological, physiological and molecular responses of MRSA clinical strains and MRSA biofilms to sub-MICs (1/4 and 1/16 MIC) of ceftaroline by using transmission, scanning and confocal microscopy. We have also used quantitative Real-Time PCR to study the effect of sub-MICs of ceftaroline on the expression of the staphylococcal icaA, agrA, sarA and sasF genes in MRSA biofilms. In one set of experiments, ceftaroline was able to inhibit biofilm formation in all strains tested at MIC, however, a strain dependent behavior in presence of sub-MICs of ceftaroline was shown. In a second set of experiments, destruction of preformed biofilms by addition of ceftaroline was evaluated. Ceftaroline was able to inhibit biofilm formation at MIC in all strains tested but not at the sub-MICs. Destruction of preformed biofilms was strain dependent because the biofilm formed by a matrix-producing strain was resistant to a challenge with ceftaroline at MIC, whereas in other strains the biofilm was sensitive. At sub-MICs, the impact of ceftaroline on expression of virulence genes was strain-dependent at 1/4 MIC and no correlation between ceftaroline-enhanced biofilm formation and gene regulation was established at 1/16 MIC. Our findings suggest that sub-MICs of ceftaroline enhance bacterial attachment and biofilm formation by some, but not all, MRSA strains and, therefore, stress the importance of maintaining effective bactericidal concentrations of ceftaroline to fight biofilm-MRSA related infections.