scholarly journals Effect of host-mimicking medium and biofilm growth on the ability of colistin to kill Pseudomonas aeruginosa

2020 ◽  
Author(s):  
Esther Sweeney ◽  
Akshay Sabnis ◽  
Andrew M. Edwards ◽  
Freya Harrison
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Ex Vivo ◽  
Clinical Success ◽  
Biofilm Growth ◽  
The Matrix ◽  
Model Versus ◽  
High Doses ◽  
Fluorescent Tag

AbstractIn vivo biofilms cause recalcitrant infections with extensive and unpredictable antibiotic tolerance. Here, we demonstrate increased tolerance of colistin by Pseudomonas aeruginosa when grown in cystic fibrosis-mimicking medium versus standard medium in in vitro biofilm assays, and drastically increased tolerance when grown in an ex vivo CF model versus the in vitro assay. We used colistin conjugated to the fluorescent dye BODIPY to assess the penetration of the antibiotic into ex vivo biofilms and showed that poor penetration partly explains the high doses of drug necessary to kill bacteria in these biofilms. The ability of antibiotics to penetrate the biofilm matrix is key to their clinical success, but hard to measure. Our results demonstrate both the importance of reduced entry into the matrix in in vivo-like biofilm, and the tractability of using a fluorescent tag and benchtop fluorimeter to assess antibiotic entry into biofilms. This method could be a relatively quick, cheap and useful addition to diagnostic and R&D pipelines, allowing the assessment of drug entry into biofilms, in in vivo-like conditions, prior to more detailed tests of biofilm killing.

scholarly journals Effect of host-mimicking medium and biofilm growth on the ability of colistin to kill Pseudomonas aeruginosa

Microbiology ◽  
2020 ◽  
Vol 166 (12) ◽  
pp. 1171-1180 ◽  
Author(s):  
Esther Sweeney ◽  
Akshay Sabnis ◽  
Andrew M. Edwards ◽  
Freya Harrison
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Ex Vivo ◽  
Clinical Success ◽  
Biofilm Growth ◽  
Content Type ◽  
The Matrix ◽  
Model Versus ◽  
High Doses

In vivo biofilms cause recalcitrant infections with extensive and unpredictable antibiotic tolerance. Here, we demonstrate increased tolerance of colistin by Pseudomonas aeruginosa when grown in medium that mimics cystic fibrosis (CF) sputum versus standard medium in in vitro biofilm assays, and drastically increased tolerance when grown in an ex vivo CF model versus the in vitro assay. We used colistin conjugated to the fluorescent dye BODIPY to assess the penetration of the antibiotic into ex vivo biofilms and showed that poor penetration partly explains the high doses of drug necessary to kill bacteria in these biofilms. The ability of antibiotics to penetrate the biofilm matrix is key to their clinical success, but hard to measure. Our results demonstrate both the importance of reduced entry into the matrix in in vivo-like biofilm, and the tractability of using a fluorescent tag and benchtop fluorimeter to assess antibiotic entry into biofilms. This method could be a relatively quick, cheap and useful addition to diagnostic and drug development pipelines, allowing the assessment of drug entry into biofilms, in in vivo-like conditions, prior to more detailed tests of biofilm killing.

In Vivo Effect of Suloctidil as an Antiplatelet Agent

1979 ◽  
Vol 41 (03) ◽  
pp. 465-474 ◽  
Author(s):  
Marcia R Stelzer ◽  
Thomas S Burns ◽  
Robert N Saunders
Keyword(s):  
Ex Vivo ◽  
Antiplatelet Agent ◽  
Ratio Method ◽  
Platelet Aggregate ◽  
Permanent Effect ◽  
Platelet Aggregates ◽  
5 Ht Uptake ◽  
High Doses

SummaryThe relationship between the effects of suloctidil in vivo as an antiplatelet agent and in vitro as a modifier of platelet serotonin (5-HT) parameters was investigated. Suloctidil was found to be effective in reducing platelet aggregates formation in the retired breeder rat as determined using the platelet aggregate ratio method (PAR) with an ED50 of 16.1 mg/kg 24 hours post administration. In contrast to the hypothesis that 5-HT depletion is involved in the anti-aggregatory mechanism of suloctidil, no correlation was found between platelet 5- HT content and this antiplatelet activity. Reduction of platelet 5-HT content required multiple injections of high doses (100 mg/kg/day) of suloctidil. Suloctidil administration for 8 days at 100 mg/kg/day, which lowered platelet 5-HT content by 50%, resulted in no permanent effect on ex vivo platelet 5-HT uptake or thrombin-induced release, nor alteration in the plasma 5-HT level. However, these platelets exhibited a short-lived, significant increase in percent leakage of 5-HT after 30 minutes of incubation. Therefore, suloctidil treatment at high doses may with time result in platelet 5-HT depletion, however this effect is probably not related to the primary anti-aggregatory activity of the drug.

scholarly journals Adaptive resistance of Pseudomonas aeruginosa induced by aminoglycosides and killing kinetics in a rabbit endocarditis model.

1997 ◽  
Vol 41 (4) ◽  
pp. 823-826 ◽  
Author(s):  
Y Q Xiong ◽  
J Caillon ◽  
M F Kergueris ◽  
H Drugeon ◽  
D Baron ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Single Dose ◽  
Ex Vivo ◽  
Day Treatment ◽  
Total Daily Dose ◽  
Once Daily ◽  
Adaptive Resistance ◽  
Dosing Regimens

Adaptive resistance following the first exposure to aminoglycosides is a recently described in vitro phenomenon in Pseudomonas aeruginosa and other aerobic gram-negative bacilli. We investigated the in vivo relevance of adaptive resistance in P. aeruginosa following a single dose of amikacin in the experimental rabbit endocarditis model. Rabbits with P. aeruginosa endocarditis received either no therapy (control) or a single intravenous (i.v.) dose of amikacin (80 mg/kg of body weight) at 24 h postinfection, after which they were sacrificed at 5, 8, 12, 16, or 24 h postdose. Excised aortic vegetations were subsequently exposed ex vivo to amikacin at 2.5, 5, 10 or 20 times the MIC for 90 min. In vivo adaptive resistance was identified when amikacin-induced pseudomonal killing within excised aortic vegetations was less in animals receiving single-dose amikacin in vivo than in vegetations from control animals not receiving amikacin in vivo. Maximal adaptive resistance occurred between 8 and 16 h after the in vivo amikacin dose, with complete refractoriness to ex vivo killing by amikacin seen at 12 h postdose. By 24 h postdose, bacteria within excised vegetations had partially recovered their initial amikacin susceptibility. In a parallel treatment study, we demonstrated that amikacin given once daily (but not twice daily) at a total dose of 80 mg/kg i.v. for 1-day treatment significantly reduced pseudomonal densities within aortic vegetations versus those in untreated controls. When therapy was continued for 3 days with the same total daily dose (80 mg/kg/day), amikacin given once or twice daily significantly reduced intravegetation pseudomonal densities versus those in controls. However, amikacin given once daily was still more effective than the twice-daily regimen. These data confirm the induction of aminoglycoside adaptive resistance in vivo and further support the advantages of once-daily aminoglycoside dosing regimens in the treatment of serious pseudomonal infections.

scholarly journals The Opportunities and Use of Imaging to Measure Target Engagement

2019 ◽  
Vol 25 (2) ◽  
pp. 127-136
Author(s):  
Juliana Maynard ◽  
Philippa Hart
Keyword(s):  
Ex Vivo ◽  
Clinical Success ◽  
Receptor Occupancy ◽  
Accurate Information ◽  
Target Engagement ◽  
Drug Molecule ◽  
Drug Discovery And Development ◽  
Ex Vivo Imaging

Lack of efficacy and poor safety outcomes are deemed to be the greatest causes of clinical failure of novel therapeutics. The use of biomarkers that give accurate information on target engagement, providing confidence that pharmacological activity in the target organ is being achieved, is key in optimizing clinical success. Without a measurement of target engagement, it can be very difficult to discern the basis for any lack of efficacy of a drug molecule within the pharmaceutical industry. Target engagement can be measured in both an in vitro and in vivo setting, and in recent years imaging measurements have been used frequently in drug discovery and development to assess target engagement and receptor occupancy in both human and animal models. From this perspective, we assess and look at the advancements in both in vivo and ex vivo imaging to demonstrate the enormous potential that imaging has as an application to provide a greater understanding of target engagement with a correlative therapeutic impact.

scholarly journals Studies of Pseudomonas aeruginosa Mutants Indicate Pyoverdine as the Central Factor in Inhibition of Aspergillus fumigatus Biofilm

2017 ◽  
Vol 200 (1) ◽  
Author(s):  
Gabriele Sass ◽  
Hasan Nazik ◽  
John Penner ◽  
Hemi Shah ◽  
Shajia Rahman Ansari ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Aspergillus Fumigatus ◽  
Fungal Pathogens ◽  
Human Pathogens ◽  
Biofilm Growth ◽  
Content Type ◽  
Iron Deprivation

ABSTRACT Pseudomonas aeruginosa and Aspergillus fumigatus are common opportunistic bacterial and fungal pathogens, respectively. They often coexist in airways of immunocompromised patients and individuals with cystic fibrosis, where they form biofilms and cause acute and chronic illnesses. Hence, the interactions between them have long been of interest and it is known that P. aeruginosa can inhibit A. fumigatus in vitro. We have approached the definition of the inhibitory P. aeruginosa molecules by studying 24 P. aeruginosa mutants with various virulence genes deleted for the ability to inhibit A. fumigatus biofilms. The ability of P. aeruginosa cells or their extracellular products produced during planktonic or biofilm growth to affect A. fumigatus biofilm metabolism or planktonic A. fumigatus growth was studied in agar and liquid assays using conidia or hyphae. Four mutants, the pvdD pchE, pvdD, lasR rhlR, and lasR mutants, were shown to be defective in various assays. This suggested the P. aeruginosa siderophore pyoverdine as the key inhibitory molecule, although additional quorum sensing-regulated factors likely contribute to the deficiency of the latter two mutants. Studies of pure pyoverdine substantiated these conclusions and included the restoration of inhibition by the pyoverdine deletion mutants. A correlation between the concentration of pyoverdine produced and antifungal activity was also observed in clinical P. aeruginosa isolates derived from lungs of cystic fibrosis patients. The key inhibitory mechanism of pyoverdine was chelation of iron and denial of iron to A. fumigatus. IMPORTANCE Interactions between human pathogens found in the same body locale are of vast interest. These interactions could result in exacerbation or amelioration of diseases. The bacterium Pseudomonas aeruginosa affects the growth of the fungus Aspergillus fumigatus. Both pathogens form biofilms that are resistant to therapeutic drugs and host immunity. P. aeruginosa and A. fumigatus biofilms are found in vivo, e.g., in the lungs of cystic fibrosis patients. Studying 24 P. aeruginosa mutants, we identified pyoverdine as the major anti-A. fumigatus compound produced by P. aeruginosa. Pyoverdine captures iron from the environment, thus depriving A. fumigatus of a nutrient essential for its growth and metabolism. We show how microbes of different kingdoms compete for essential resources. Iron deprivation could be a therapeutic approach to the control of pathogen growth.

scholarly journals Rapid ratiometric biomarker detection with topically applied SERS nanoparticles

TECHNOLOGY ◽  
2014 ◽  
Vol 02 (02) ◽  
pp. 118-132 ◽  
Author(s):  
Yu "Winston" Wang ◽  
Altaz Khan ◽  
Madhura Som ◽  
Danni Wang ◽  
Ye Chen ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Clinical Success ◽  
Simultaneous Detection ◽  
Integration Time ◽  
In Vivo Experiments ◽  
Surface Enhanced Raman ◽  
Detection Probe ◽  
Multiple Cell

Multiplexed surface-enhanced Raman scattering (SERS) nanoparticles (NPs) offer the potential for rapid molecular phenotyping of tissues, thereby enabling accurate disease detection as well as patient stratification to guide personalized therapies or to monitor treatment outcomes. The clinical success of molecular diagnostics based on SERS NPs would be facilitated by the ability to accurately identify tissue biomarkers under time-constrained staining and detection conditions with a portable device. In vitro, ex vivo and in vivo experiments were performed to optimize the technology and protocols for the rapid detection (0.1-s integration time) of multiple cell-surface biomarkers with a miniature fiber-optic spectral-detection probe following a brief (5 min) topical application of SERS NPs on tissues. Furthermore, we demonstrate that the simultaneous detection and ratiometric quantification of targeted and nontargeted NPs allows for an unambiguous assessment of molecular expression that is insensitive to nonspecific variations in NP concentrations.

Optimization of Clot Formation Methodology for Assessment of Neurothrombectomy Devices in Large Animal Thrombectomy Models

2020 ◽  
Author(s):  
Cedric Jimenez ◽  
Igor Polyakov ◽  
Leigh Kleinert ◽  
André Nelson ◽  
Mark Smith
Keyword(s):  
Reliable Method ◽  
Ex Vivo ◽  
Clinical Success ◽  
Model Development ◽  
Large Animal ◽  
Soft Or ◽  
Mechanical Devices ◽  
Clot Disruption

Abstract Neurothrombectomy devices are commonly evaluated for potential clinical success in porcine models of neurothromboembolism. The majority of preclinical evaluations for these devices are performed in the vasculature of swine or dog utilizing clots created ex vivo. This investigation was conducted to develop a faster, more reliable method for creating clots ex vivo for model development. Neurothrombectomy devices are designed to perform recanalization of arterial occlusions that cause acute ischemic stroke [1]. Recanalization can be achieved via clot disruption, aspiration, or retrieval using one or more mechanical devices. In order to evaluate these devices in vivo, a fast and reliable method for creating and delivering clots to a desired artery, thereby simulating a target site for neurothrombectomy, is essential. Two types of clot analogs (soft or firm) were created using two different methods in order to compare both their mechanical properties and their ability to reliably occlude selected arteries. Utilizing both methods, pre-formed clots were qualitatively compared in vitro to evaluate elasticity, stiffness, and functionality of delivery through a catheter. These evaluations were performed prior to in vivo assessment of the effectiveness of the analogs occlusion of selected arterial vasculature.

671Assessment of irrigated-tip radiofrequency catheter ablation lesion characteristics using different irrigation fluids on ex-vivo bovine myocardium

EP Europace ◽  
2020 ◽  
Vol 22 (Supplement_1) ◽  
Author(s):  
B Candemir ◽  
E Baskovski ◽  
K Esenboga ◽  
H Yorgun ◽  
K Aytemir ◽  
...  
Keyword(s):  
Catheter Ablation ◽  
Normal Saline ◽  
Ex Vivo ◽  
Clinical Success ◽  
Ringer Lactate ◽  
Lesion Depth ◽  
Irrigation Solution ◽  
Bovine Myocardium

Abstract Introduction Certain arrythmias, particularly of ventricular origin, necessitate deep lesions in order to achieve clinical success. The introduction of irrigated-tip radiofrequency (RF) catheters has allowed creation of deeper lesions and has decreased char formation. Although normal saline has been used as a standard solution, recent observations have suggested less ionic solutions may increase lesion depth. In this in-vitro study we aimed to characterize lesions created by irrigated-tip RF catheters using standard normal saline(NS), half-normal saline (HNS), and HNS-%2.5Dextrose (HNS-DEX/2) combination solutions. METHODS Bovine myocardium was placed firstly in ringer-lactate bath. Using irrigated-tip RF catheter ablation lesions were created serially using 30, 40, 50, 60, 70Watts with NS, HNS and HNS-DEX/2 as irrigation solutions. Lesion depths, steam pops and impedance drops were measured. Subsequently the experiment was repeated in normal saline bath. RESULTS Both HNS-DEX/2 and HNS irrigation solutions increase lesion depths when compared to normal saline (Table 1, Figure 1). Steam pops were more common and earlier with HNS-DEX/2 and HNS. DISCUSSION AND CONCLUSION Our findings suggest that less ionic irrigation solutions lead to increased lesion depths using similar RF power. If confirmed in-vivo, HNS and, particularly, HNS-DEX/2 as irrigation solution may increase ablation success for intramural/deep myocardial arrhythmic foci. Additionally, studies to assess safety of this strategy are necessary as intuitively deeper lesions may lead to more complications. Table 1 Irrigation Solution/Bath solution 30W/30ml(Lesion depth mm)(Impedance drop) 40W/30ml(Lesion depth mm)(Impedance drop) 50W/30ml(Lesion depth mm)(Impedance drop) 60W/30ml(Lesion depth mm)(Impedance drop) 70W/30ml(Lesion depth mm)(Impedance drop) HNS+%2.5D / Ringer Lactate 470->61 4.583->63 5.074->62SP+(32s) 6.080->66SP+(27s) 7.577->65SP+(26s) HNS/ Ringer Lactate 3.590->65 4.580->63SP+(34s) 5.073->59SP+(33s) 5.580->56SP+(28s) 6.583->64SP+(22s) NS/ Ringer Lactate 3.077->60 472->59 4.576->61 5.081->56 674->55 SP+(30s) HNS+%2.5D/Normal saline 3.573->67 598->63 5.573->58SP+(28s) 697->76SP+(26s) 7.570->58SP (25s) HNS/Normal saline 3.563->58 4.066->52 4.567->52 5.575->52 788->60SP+(28s) NS/Normal saline 2.566->58 3.063->52 4.073->58 5.077->55 5.568->44 Lesions created by irrigated tip radiofrequency catheter with bovine myocardium Abstract Figure 1

scholarly journals Dual bioresponsive antibiotic and quorum sensing inhibitor combination nanoparticles for treatment of Pseudomonas aeruginosa biofilms in vitro and ex vivo

2019 ◽  
Vol 7 (10) ◽  
pp. 4099-4111 ◽  
Author(s):  
Nishant Singh ◽  
Manuel Romero ◽  
Alessandra Travanut ◽  
Patricia F. Monteiro ◽  
Elena Jordana-Lluch ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Ex Vivo ◽  
Biofilm Growth ◽  
Quorum Sensing Inhibitor ◽  
Quorum Sensing Inhibitors ◽  
Inhibitor Combination

Nanoparticles combining Quorum Sensing Inhibitors and anti-bacterials can eradicate biofilm growth in vitro and ex vivo.

The Thrombostat System A Useful Method to Test Antiplatelet Drugs and Diets

1995 ◽  
Vol 21 (S 02) ◽  
pp. 25-31 ◽  
Author(s):  
Michael Kratzer ◽  
Emil Negrescu ◽  
Azan Hirai ◽  
Young Yeo ◽  
Franke Petra ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Synergistic Effects ◽  
Antiplatelet Drugs ◽  
Small Samples ◽  
Fish Diet ◽  
Heart Attacks ◽  
Primary Hemostasis ◽  
High Doses

The use of platelet inhibitory drugs, like aspirin, has resulted in a significant reduction of thrombotic complications in primary and secondary prevention of heart attacks. To find more effective substances or better drug combinations, inhibition of primary hemostasis in vitro (Thrombostat system) was investigated, with different drugs and fish diet, using small samples (1 ml) of anticoagulated (Na- citrate 3.8%, 1/9) human blood. Results: 1. In the presence of 1mM aspirin, which had no effect on bleeding volume, only 0.6 nM iloprost were necessary to show a 50% inhibition, in contrast to 2.5 μM without aspirin. 2. At aspirin concentrations of 1 mM, 50% inhibition of primary hemostasis could be achieved with 20 μM SIN-I, or with 7 μM SIN-l together with iloprost (500 pM). The same effect was seen only with very high doses of SIN-l (1000 μM) alone. 3. For 50% inhibition of primary hemostasis in vitro, RGDS concentrations were reduced from 250 μM to 160 μM when blood was pretreated with 1 mM aspirin and to 75 μM when 500 pM i1oprost were added additionally. 4. Japanese fishermen (eating 270 g fish/day) demonstrated significantly longer in-vivo bleeding times and in-vitro bleeding volumes (6.49 min/224 μI), respectively, as compared to Japanese farmers (90g fish/day, 4.85 min/137 μI). 5. In Japanese subjects in-vivo bleeding times correlated with in-vitro bleeding volumes (0.69). The Thrombostat system proved to be a sensitive method to detect synergistic effects of various antiplatelet drugs in vitro and of a platelet inhibitory diet ex vivo.

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