Self-collected oral, nasal and saliva samples yield sensitivity comparable to professionally collected oro-nasopharyngeal swabs in SARS-CoV-2 diagnosis among symptomatic outpatients.

INTRODUCTION: Containing COVID-19 requires broad-scale testing. However, sample collection requires qualified personnel and protective equipment and may cause transmission. We assessed the sensitivity of SARS-CoV-2-rtPCR applying three self-sampling techniques as compared to professionally collected oro-nasopharyngeal samples (cOP/NP). METHODS: From 62 COVID-19 outpatients, we obtained: (i) multi-swab, MS; (ii) saliva sponge combined with nasal vestibula, SN; (iii) gargled water, GW; (iv) professionally collected cOP/NP (standard). We compared ct-values for E-gene and ORF1ab and analysed variables reducing sensitivity of self-collecting procedures. RESULTS: The median ct-values for E-gene and ORF1ab obtained in cOP/NP samples were 20.7 and 20.2, in MS samples 22.6 and 21.8, in SN samples 23.3 and 22.3, and in GW samples 30.3 and 29.8, respectively. MS and SN samples showed sensitivities of 95.2% (95%CI, 86.5-99.0) and GW samples of 88.7% (78.1-95.3). Sensitivity was inversely correlated with ct-values, and became <90% for samples obtained more than 8 days after symptom onset. For MS and SN samples, false negativity was associated with language problems, sampling errors, and symptom duration. CONCLUSION: Conclusions from this study are limited to the sensitivity of self-sampling in mildly to moderately symptomatic patients. Still, self-collected oral/nasal/saliva samples can facilitate up-scaling of testing in early symptomatic COVID-19 patients if operational errors are minimized.

Characterization of ferucarbotran-loaded RBCs as long circulating magnetic contrast agents.

AIM: The biomedical application of contrast agents based on superparamagnetic iron oxide nanoparticles is still limited because of their short intravascular half-life. The potential of red blood cells (RBCs) loaded with new ferucarbotran nanoparticles as magnetic contrast agents with longer blood retention time has been investigated. MATERIALS & METHODS: Ferucarbotran was loaded into RBCs by a procedure of hypotonic dialysis and isotonic resealing. Ferucarbotran amounts encapsulated in RBCs were determined by NMR. The survival of ferucarbotran-loaded RBCs and bulk ferucarbotran was evaluated in the mouse bloodstream. RESULTS: Blood retention time of these RBC constructs is longer (∼14 days) than the bulk ferucarbotran (∼1 h) with a slower Fe clearance from liver and spleen. CONCLUSION: Ferucarbotran-loaded RBCs could be used as potential contrasting agents for diagnostic applications in MRI/magnetic particle imaging.

Current limitations of molecular magnetic resonance imaging for tumors as evaluated with high-relaxivity CD105-specific iron oxide nanoparticles.

OBJECTIVE: Tumor imaging via molecular magnetic resonance imaging (MRI) that uses specific superparamagnetic iron oxide particles (SPIOs) has been addressed in the literature several times in the last 20 years. To our knowledge, none of the reported approaches is currently used for routine clinical diagnostic evaluation, nor are any in clinical development. This raises questions as to whether SPIO-enhanced molecular MRI is sensitive and specific enough for use in clinical practice. The aim of our preclinical study was to investigate the minimum requirements for obtaining sensitive molecular MRI for use in tumor evaluations under optimal conditions. The well-vascularized F9 teratocarcinoma tumor model, which exhibits high levels of the highly accessible target CD105 (endoglin), was used to compare the accumulation and visualization of target-specific SPIOs by MRI. MATERIAL AND METHODS: Superparamagnetic iron oxide particles were optimized in the following ways: (a) proton relaxivity was increased for higher imaging sensitivity, (b) a coating material was used for optimal loading density of the αCD105 antibody, and (c) binding activity to the target CD105 was increased. Binding activity and specificity were confirmed in vitro using enzyme-linked immunosorbent assay and in vivo using pharmacokinetic and biodistribution studies of 11 F9 teratoma-bearing mice together with micro-autoradiography. CD105 target expression was determined using immunohistochemistry and quantitative enzyme-linked immunosorbent assay. The transverse relaxation rate R2* was quantified by 3.0-T MRI in the tumors, kidneys, and muscles before and up to 60 minutes after injection in 11 mice. The use of [Fe]-labeled SPIOs for all in vivo experiments allowed for the direct correlation of the imaging results with SPIO accumulation. RESULTS: High-relaxivity αCD105-polyacrylic acid-SPIOs (r2 up to 440 L mmol Fe s) with strong binding activity accumulated specifically in tumors (1.4% injected dose/g) and kidneys (4.1% injected dose/g) in a manner dependent on the target concentration. The accumulation occurred within the first 3 minutes after injection. Visualization of specific SPIOs was accomplished with MRI. In contrast to the successful use of MRI in all examined kidneys (mean ± SEM ΔR2*, 61 ± 11 s), only 6 of 11 tumors (mean ± SEM ΔR2*, 15 ± 7 s) showed a clear signal when compared with the control even though optimal conditions were used. CONCLUSION: The accumulation of CD105-specific SPIOs in F9 mouse teratomas was robust. However, visualization of the specifically accumulated SPIOs by MRI was not reliable because of its limited signal detection sensitivity. We postulate that it will be challenging to improve the imaging properties of targeted SPIOs further. Therefore, molecular MRI by targeted SPIOs is currently not suitable for clinical tumor imaging using routinely applicable sequences and field strength.

Synergistic effects of miconazole and polymyxin B on microbial pathogens.

The therapeutic value of antibiotics depends on the susceptibility of the infecting microorganism and the pharmacological profile of the drugs. To assess the value of an antibiotic combination of polymyxin B and miconazole this study examined the in vitro synergistic potential of the two drugs on Gram-negative and Gram-positive bacteria and yeast. Antifungal and antibacterial activity was tested by minimum inhibitory concentration (MIC) of broth macrodilution and urea broth microdilution, by fluorescence microscopy and flow cytometry. Synergism was calculated using the fractional inhibitory concentration index (FICi). With Staphylococcus intermedius as target we found up to an eightfold reduction of the individual MICs when both drugs were combined. However, the FICi was 0.63 suggesting no real interaction between the two drugs. With Escherichia coli, Pseudomonas aeruginosa, and Malassezia pachydermatis as targets the antimicrobial drug combination reduced the MICs of polymyxin B and miconazole from fourfold to hundredfold resulting in FICi between 0.06 and 0.5 which defines a synergistic action. Thus, if polymyxin B and miconazole are combined their effect is greater than the sum of the effects observed with polymyxin B and miconazole independently, revealing bactericidal and fungicidal synergism. Our results indicate a strong therapeutic value for the combination of these antimicrobial agents against Gram-negative bacteria and yeast and a weaker value against Gram positive bacteria for clinical situations where these pathogens are involved.

One-pot synthesis of pegylated ultrasmall iron-oxide nanoparticles and their in vivo evaluation as magnetic resonance imaging contrast agents.

A well-defined copolymer poly(oligo(ethylene glycol) methacrylate-co-methacrylic acid) P(OEGMA-co-MAA) was studied as a novel water-soluble biocompatible coating for superparamagnetic iron oxide nanoparticles. This copolymer was prepared via a two-step procedure: a well-defined precursor poly(oligo(ethylene glycol) methacrylate-co-tert-butyl methacrylate), P(OEGMA-co-tBMA) (M(n) = 17300 g mol(-1); M(w)/M(n) = 1.22), was first synthesized by atom-transfer radical polymerization in the presence of the catalyst system copper(I) chloride/2,2'-bipyridyl and subsequently selectively hydrolyzed in acidic conditions. The resulting P(OEGMA-co-MAA) was directly utilized as a polymeric stabilizer in the nanoparticle synthesis. Four batches of ultrasmall PEGylated magnetite nanoparticles (i.e., with an average diameter below 30 nm) were prepared via aqueous coprecipitation of iron salts in the presence of variable amounts of P(OEGMA-co-MAA). The diameter of the nanoparticles could be easily tuned in the range 10-25 nm by varying the initial copolymer concentration. Moreover, the formed PEGylated ferrofluids exhibited a long-term colloidal stability in physiological buffer and could therefore be studied in vivo by magnetic resonance (MR) imaging. Intravenous injection into rats showed no detectable signal in the liver within the first 2 h. Maximum liver accumulation was found after 6 h, suggesting a prolongated circulation of the nanoparticles in the bloodstream as compared to conventional MR imaging contrast agents.

Nanoparticle-based diagnosis and therapy.

Nanoparticles are at the leading edge of the rapidly developing field of material science in nanotechnology with many potential applications in clinical medicine and research. Due to their unique size-dependent properties nanoparticles offer the possibility to develop both new therapeutic and diagnostic tools. The ability to incorporate drugs into nanosystems displays a new paradigm in pharmacotherapy that could be used for cell-targeted drug delivery. Nontargeted nanosystems such as nanocarriers that are coated with polymers or albumin and solid lipid particles have been used to transport a large number of compounds. However, nowadays drugs can be coupled to nanocarriers that are specific for cells and/or organs. Thus, drugs that are either trapped within the carriers or deposited in subsurface oil layers could be specifically delivered to organs, tumors and cells. These strategies can be used to concentrate drugs in selected target tissues thus minimizing systemic side effects and toxicity. In addition to these therapeutic options, nanoparticle-based "molecular" imaging displays a field in which this new technology has set the stage for an evolutionary leap in diagnostic imaging. Based on the recent progress in nanobiotechnology, nanoparticles have the potential to become useful tools as therapeutic and diagnostic tools in the near future.

Maghemite nanoparticles protectively coated with poly(ethylene imine) and poly(ethylene oxide)-block-poly(glutamic acid).

Superparamagnetic iron oxide particles (SPIO) of maghemite were prepared in aqueous solution and subsequently stabilized with polymers in two layer-by-layer deposition steps. The first layer around the maghemite core is formed by poly(ethylene imine) (PEI), and the second one is formed by poly(ethylene oxide)-block-poly(glutamic acid) (PEO-PGA). The hydrodynamic diameter of the particles increases stepwise from D(h) = 25 nm (parent) via 35 nm (PEI) to 46 nm (PEI plus PEO-PGA) due to stabilization. This is accompanied by a switching of their zeta-potentials from moderately positive (+28 mV) to highly positive (+50 mV) and finally slightly negative (-3 mV). By contrast, the polydispersity indexes of the particles remain constant (ca. 0.15). Mössbauer spectroscopy revealed that the iron oxide, which forms the core of the particles, is only present as Fe(III) in the form of superparamagnetic maghemite nanocrystals. The magnetic domains and the maghemite crystallites were found to be identical with a size of 12.0 +/- 0.5 nm. The coated maghemite nanoparticles were tested to be stable in water and in physiological salt solution for longer than 6 months. In contrast to novel methods for magnetic nanoparticle production, where organic solvents are necessary, the procedure proposed here can dispense with organic solvents. Magnetic resonance imaging (MRI) experiments on living rats indicate that the nanoparticles are useful as an MRI contrast agent.

Nanomedicine for respiratory diseases.

Nanotechnology provides new materials in the nanometer range with many potential applications in clinical medicine and research. Due to their unique size-dependent properties nanomaterial such as nanoparticles offer the possibility to develop both new therapeutic and diagnostic tools. Thus, applied nanotechnology to medical problems--nanomedicine--can offer new concepts that are reviewed. The ability to incorporate drugs into nanosystems displays a new paradigm in pharmacotherapy that could be used for cell-targeted drug delivery. Nontargeted nanosystems such as nanocarriers that are coated with polymers or albumin and solid lipid particles have been used as transporter in vivo. However, nowadays drugs can be coupled to nanocarriers that are specific for cells and/or organs. Thus, drugs that are either trapped within the carriers or deposited in subsurface oil layers could be specifically delivered to organs, tumors and cells. These strategies can be used to concentrate drugs in selected target tissues thus minimizing systemic side effects and toxicity. In addition to these therapeutic options, nanoparticle-based "molecular" imaging displays a field in which this new technology has set the stage for an evolutionary leap in diagnostic imaging. Based on the recent progress in nanobiotechnology there is potential for nanoparticles and -systems to become useful tools as therapeutic and diagnostic tools in the near future.

Evaluation of the -26G>A CC16 polymorphism in acute respiratory distress syndrome.

OBJECTIVE: Different risk factors are presumably involved in the pathogenesis of acute respiratory distress syndrome (ARDS) including genetic factors. Clara cell protein 16 (CC16) is a potential candidate gene for ARDS susceptibility because reduced levels of the anti-inflammatory CC16 have been observed in bronchoalveolar lavage fluids or serum of patients with different inflammatory lung diseases. Furthermore, CC16 potently inhibits phospholipase A2, which plays a major role in ARDS pathophysiology. A functional polymorphism (-26G>A) was previously identified and related to decreased CC16 levels, asthma, and asthma severity. DESIGN: Observational study. SETTINGS: Adults with ARDS were recruited from intensive care units in two university medical centers. SUBJECTS: We evaluated the role of this genetic variant in 117 German patients with ARDS and 373 German controls. MEASUREMENTS: The CC16 -26G>A polymorphism was analyzed by melting-curve analysis using a pair of fluorescence resonance energy transfer probes. MAIN RESULTS: CC16 genotype frequencies in ARDS patients did not differ from those seen in controls. Also, the allele frequencies were identical in patients compared with controls (0.66 and 0.34). Moreover, only one of the patients who died (n = 27) was homozygous for the -26A allele. CONCLUSIONS: The CC16 -26G>A polymorphism does not affect the susceptibility to and the outcome of ARDS.

DCL-Hb for trauma patients with severe hemorrhagic shock: the European "On-Scene" multicenter study.

OBJECTIVE: A major cause of death in patients with severe hemorrhagic shock following trauma is the subsequent occurrence of multiple organ failure due to tissue hypoxia. Early administration of an oxygen carrier may reduce the occurrence of organ failures and improve survival. It may also reduce the need of blood products. DESIGN AND SETTING: Prospective multicenter study in a university clinic. PATIENTS: 121 patients with severe hemorrhagic shock. INTERVENTIONS: Patients were randomly assigned "on-scene" to receive either up to 1000 ml of a 10% diaspirin cross-linked hemoglobin (DCLHb) solution or the study center's standard therapy. MEASUREMENTS AND RESULTS: Demographic and physiological characteristics of the two treatment groups at baseline were comparable. Organ failures and survival rates until day 5 and day 28 showed no significant differences. The sponsor therefore terminated this trial prematurely after an interim evaluation of the data indicated no evidence of efficacy to offset concerns raised about the safety of DCLHb. Median volumes of cumulative blood products administered on 1 (1595 vs. 3716 ml) and 7 days (3139 vs. 4746 ml) after admission were lower in the DCLHb group. CONCLUSIONS: The early application of an oxygen carrier (DCLHb) to patients with severe hemorrhagic shock following trauma had no significant effect on the occurrence of organ failure or on 5- and 28-day survival in this abbreviated trial. However, early infusion of up to 1000 ml DCLHb reduces the need for blood products without changing morbidity or survival.