Use of Point-of-Care Ultrasound to Diagnose a Ruptured Splenic Hemangioma.

An 89-year-old female presented to the emergency department (ED) with hypotension and altered mental status. The patient had no external signs of trauma or hemorrhage and no abdominal tenderness on examination. The patient remained hypotensive after initial fluid resuscitation, and laboratory testing revealed a significant anemia. Point-of-care ultrasound (POCUS) was used to perform a rapid ultrasound in shock (RUSH) exam in an attempt to uncover the etiology of undifferentiated hypotension. The exam displayed free fluid in the right upper quadrant and the left upper quadrant exam demonstrated a large splenic lesion with mixed echogenicity. Subsequent computed tomography (CT) of the abdomen and pelvis with intravenous contrast suggested a ruptured hemorrhagic splenic cyst, and the patient underwent an emergent splenectomy for hemorrhage control. Operative pathologic examination revealed the cystic lesion to be a splenic hemangioma. This case report highlights the utility of the Rapid Ultrasound for Shock and Hypotension (RUSH) protocol when evaluating patients with undifferentiated nontraumatic shock, and a rare cause of spontaneous intra-abdominal hemorrhage.

A redox switch allows binding of Fe(II) and Fe(III) ions in the cyanobacterial iron-binding protein FutA from Prochlorococcus.

The marine cyanobacterium Prochlorococcus is a main contributor to global photosynthesis, whilst being limited by iron availability. Cyanobacterial genomes generally encode two different types of FutA iron-binding proteins: periplasmic FutA2 ABC transporter subunits bind Fe(III), while cytosolic FutA1 binds Fe(II). Owing to their small size and their economized genome Prochlorococcus ecotypes typically possess a single futA gene. How the encoded FutA protein might bind different Fe oxidation states was previously unknown. Here, we use structural biology techniques at room temperature to probe the dynamic behavior of FutA. Neutron diffraction confirmed four negatively charged tyrosinates, that together with a neutral water molecule coordinate iron in trigonal bipyramidal geometry. Positioning of the positively charged Arg103 side chain in the second coordination shell yields an overall charge-neutral Fe(III) binding state in structures determined by neutron diffraction and serial femtosecond crystallography. Conventional rotation X-ray crystallography using a home source revealed X-ray-induced photoreduction of the iron center with observation of the Fe(II) binding state; here, an additional positioning of the Arg203 side chain in the second coordination shell maintained an overall charge neutral Fe(II) binding site. Dose series using serial synchrotron crystallography and an XFEL X-ray pump-probe approach capture the transition between Fe(III) and Fe(II) states, revealing how Arg203 operates as a switch to accommodate the different iron oxidation states. This switching ability of the Prochlorococcus FutA protein may reflect ecological adaptation by genome streamlining and loss of specialized FutA proteins.

An overview of bone cement: Perioperative considerations, complications, outcomes and future implications.

Polymethyl methacrylate is commonly known as bone cement and is widely used for implant fixation in various orthopaedic arthroplasty and trauma surgery. The first bone cement use in orthopaedics is widely accredited to the famous English surgeon, John Charnley, who in 1958, used it for total hip arthroplasty. Since then, there have been many developments in cementing techniques in arthroplasty surgery. This overview aims to cover the perioperative considerations of bone cement, including cementing techniques, current outcomes and complications such as bone cement implantation syndrome. The overview will additionally consider future developments involving bone cement in orthopaedic arthroplasty.

Five historical innovations that have shaped modern orthopaedic surgery.

Throughout history, many innovations have contributed to the development of modern orthopaedic surgery, improving patient outcomes and expanding the range of treatment options available to patients. This article explores five key historical innovations that have shaped modern orthopaedic surgery: X-ray imaging, bone cement, the Thomas splint, the Pneumatic tourniquet and robotic-assisted surgery. We will review the development, impact and significance of each innovation, highlighting their contributions to the field of orthopaedic surgery and their ongoing relevance in contemporary and perioperative practice.

Identification of Potential Antimicrobial Targets of Pseudomonas aeruginosa Biofilms through a Novel Screening Approach.

Pseudomonas aeruginosa is an opportunistic pathogen of considerable medical importance, owing to its pronounced antibiotic tolerance and association with cystic fibrosis and other life-threatening diseases. The aim of this study was to highlight the genes responsible for P. aeruginosa biofilm tolerance to antibiotics and thereby identify potential new targets for the development of drugs against biofilm-related infections. By developing a novel screening approach and utilizing a public P. aeruginosa transposon insertion library, several biofilm-relevant genes were identified. The Pf phage gene (PA0720) and flagellin gene (fliC) conferred biofilm-specific tolerance to gentamicin. Compared with the reference biofilms, the biofilms formed by PA0720 and fliC mutants were completely eliminated with a 4-fold-lower gentamicin concentration. Furthermore, the mreC, pprB, coxC, and PA3785 genes were demonstrated to play major roles in enhancing biofilm tolerance to gentamicin. The analysis of biofilm-relevant genes performed in this study provides important novel insights into the understanding of P. aeruginosa antibiotic tolerance, which will facilitate the detection of antibiotic resistance and the development of antibiofilm strategies against P. aeruginosa. IMPORTANCE Pseudomonas aeruginosa is an opportunistic pathogen of high medical importance and is one of the main pathogens responsible for the mortality of patients with cystic fibrosis. In addition to inherited antibiotic resistance, P. aeruginosa can form biofilms, defined as communities of microorganisms embedded in a self-produced matrix of extracellular polymeric substances adhering to each other and/or to a surface. Biofilms protect bacteria from antibiotic treatments and represent a major reason for antibiotic failure in the treatment of chronic infections caused by cystic fibrosis. Therefore, it is crucial to develop new therapeutic strategies aimed at specifically eradicating biofilms. The aim of this study was to generalize a novel screening method for biofilm research and to identify the possible genes involved in P. aeruginosa biofilm tolerance to antibiotics, both of which could improve the understanding of biofilm-related infections and allow for the identification of relevant therapeutic targets for drug development.

Added value of mass characteristic frequency to 2-D shear wave elastography for differentiation of benign and malignant thyroid nodules.

Mass characteristic frequency (fmass) is a novel shear wave (SW) parameter that represents the ratio of the averaged minimum SW speed within the regions of interest to the largest dimension of the mass. Our study objective was to evaluate if the addition of fmass to conventional 2-D shear wave elastography (SWE) parameters would improve the differentiation of benign from malignant thyroid nodules. Our cohort comprised 107 patients with 113 thyroid nodules, of which 67 (59%) were malignant. Two-dimensional SWE data were obtained using the Supersonic Imagine Aixplorer ultrasound system equipped with a 44- to 15-MHz15-MHz linear array transducer. A receiver operating characteristic curve was generated based on a multivariable logistic regression analysis to evaluate the ability of SWE parameters with/without fmass and with/without clinical factors to discriminate benign from malignant thyroid nodules. The addition of fmass to conventional SW elasticity parameters increased the area under the curve from 0.808 to 0.871 (p = 0.02). The combination of SW elasticity parameters plus fmass plus clinical factors provided the strongest thyroid nodule malignancy probability estimate, with a sensitivity of 93.4% and specificity of 91.1% at the optimal threshold. In summary, fmass can be a valuable addition to conventional 2-D SWE parameters.

Multi-Excitation Raman Spectroscopy for Label-Free, Strain-Level Characterization of Bacterial Pathogens in Artificial Sputum Media.

The current methods for diagnosis of acute and chronic infections are complex and skill-intensive. For complex clinical biofilm infections, it can take days from collecting and processing a patient's sample to achieving a result. These aspects place a significant burden on healthcare providers, delay treatment, and can lead to adverse patient outcomes. We report the development and application of a novel multi-excitation Raman spectroscopy-based methodology for the label-free and non-invasive detection of microbial pathogens that can be used with unprocessed clinical samples directly and provide rapid data to inform diagnosis by a medical professional. The method relies on the differential excitation of non-resonant and resonant molecular components in bacterial cells to enhance the molecular finger-printing capability to obtain strain-level distinction in bacterial species. Here, we use this strategy to detect and characterize the respiratory pathogens Pseudomonas aeruginosa and Staphylococcus aureus as typical infectious agents associated with cystic fibrosis. Planktonic specimens were analyzed both in isolation and in artificial sputum media. The resonance Raman components, excited at different wavelengths, were characterized as carotenoids and porphyrins. By combining the more informative multi-excitation Raman spectra with multivariate analysis (support vector machine) the accuracy was found to be 99.75% for both species (across all strains), including 100% accuracy for drug-sensitive and drug-resistant S. aureus. The results demonstrate that our methodology based on multi-excitation Raman spectroscopy can underpin the development of a powerful platform for the rapid and reagentless detection of clinical pathogens to support diagnosis by a medical expert, in this case relevant to cystic fibrosis. Such a platform could provide translatable diagnostic solutions in a variety of disease areas and also be utilized for the rapid detection of anti-microbial resistance.

Role of the flagellar hook in the structural development and antibiotic tolerance of Pseudomonas aeruginosa biofilms.

Pseudomonas aeruginosa biofilms exhibit an intrinsic resistance to antibiotics and constitute a considerable clinical threat. In cystic fibrosis, a common feature of biofilms formed by P. aeruginosa in the airway is the occurrence of mutants deficient in flagellar motility. This study investigates the impact of flagellum deletion on the structure and antibiotic tolerance of P. aeruginosa biofilms, and highlights a role for the flagellum in adaptation and cell survival during biofilm development. Mutations in the flagellar hook protein FlgE influence greatly P. aeruginosa biofilm structuring and antibiotic tolerance. Phenotypic analysis of the flgE knockout mutant compared to the wild type (WT) reveal increased fitness under planktonic conditions, reduced initial adhesion but enhanced formation of microcolony aggregates in a microfluidic environment, and decreased expression of genes involved in exopolysaccharide formation. Biofilm cells of the flgE knock-out mutant display enhanced tolerance towards multiple antibiotics, whereas its planktonic cells show similar resistance to the WT. Confocal microscopy of biofilms demonstrates that gentamicin does not affect the viability of cells located in the inner part of the flgE knock-out mutant biofilms due to reduced penetration. These findings suggest that deficiency in flagellar proteins like FlgE in biofilms and in cystic fibrosis infections represent phenotypic and evolutionary adaptations that alter the structure of P. aeruginosa biofilms conferring increased antibiotic tolerance.

Preoperative Factors Affecting the Patient-Reported Outcome Measures following Total Knee Replacement: Socioeconomic Factors and Preoperative OKS Have a Clinically Meaningful Effect.

The Oxford Knee Score (OKS) is a patient-reported outcome questionnaire typically used to assess function and pain in patients undergoing total knee replacement (TKR). However, research is inconclusive as to which preoperative factors are important in explaining variation in outcome following TKR. The operative records of 12,709 patients who underwent primary TKR over a 9-year period were analyzed. The following variables were collected for each patient: age, sex, body mass index (BMI), Index of Multiple Deprivation decile rank, side of operation, diagnosis, the American Society of Anaesthesiologists (ASA) grade, preoperative OKS, EQ-5D index score, EuroQol visual analog scale (EQ-VAS) score, the postoperative OKS at 1 and 2 years. Generalized linear regression models were performed at 1 and 2 years to investigate the effect of the preoperative variables on the postoperative OKS. The effect of age, sex, BMI, Index of Multiple Deprivation decile rank, diagnosis, ASA grade, preoperative OKS, EuroQoL five-dimensional (EQ-5D) index score, and EQ-VAS score were all statistically significant in explaining the variation in OKS at 1 and 2 years postoperatively, with critical level of significance of 0.05 (5%). Being male aged 60 to 69 years of normal BMI, ASA grade I (fit and healthy), living in an affluent area, not reporting preoperative anxiety/depression, were associated with an enhanced mean postoperative OKS at both 1 and 2 years. When adjusted for potential confounding, age of 60-69 years, male sex, normal BMI, lower ASA grade, higher Index of Multiple Deprivation and higher pre-operative EQ-5D, EQ-VAS and OKS were identified as factors that resulted in higher post-operative OKS after primary TKR.

Comparing deep learning-based automatic segmentation of breast masses to expert interobserver variability in ultrasound imaging.

Deep learning is a powerful tool that became practical in 2008, harnessing the power of Graphic Processing Unites, and has developed rapidly in image, video, and natural language processing. There are ongoing developments in the application of deep learning to medical data for a variety of tasks across multiple imaging modalities. The reliability and repeatability of deep learning techniques are of utmost importance if deep learning can be considered a tool for assisting experts, including physicians, radiologists, and sonographers. Owing to the high costs of labeling data, deep learning models are often evaluated against one expert, and it is unknown if any errors fall within a clinically acceptable range. Ultrasound is a commonly used imaging modality for breast cancer screening processes and for visually estimating risk using the Breast Imaging Reporting and Data System score. This process is highly dependent on the skills and experience of the sonographers and radiologists, thereby leading to interobserver variability and interpretation. For these reasons, we propose an interobserver reliability study comparing the performance of a current top-performing deep learning segmentation model against three experts who manually segmented suspicious breast lesions in clinical ultrasound (US) images. We pretrained the model using a US thyroid segmentation dataset with 455 patients and 50,993 images, and trained the model using a US breast segmentation dataset with 733 patients and 29,884 images. We found a mean Fleiss kappa value of 0.78 for the performance of three experts in breast mass segmentation compared to a mean Fleiss kappa value of 0.79 for the performance of experts and the optimized deep learning model.

Quantitative assessment of ensemble coherency in contrast-free ultrasound microvasculature imaging.

PURPOSE: Contrast-free visualization of microvascular blood flow (MBF) using ultrasound can play a valuable role in diagnosis and detection of diseases. In this study, we demonstrate the importance of quantifying ensemble coherence for robust MBF imaging. We propose a novel approach to quantify ensemble coherence by estimating the local spatiotemporal correlation (LSTC) image, and evaluate its efficacy through simulation and in vivo studies. METHODS: The in vivo patient studies included three volunteers with a suspicious breast tumor, 15 volunteers with a suspicious thyroid tumor, and two healthy volunteers for renal MBF imaging. The breast data displayed negligible prior motion and were used for simulation analysis involving synthetically induced motion, to assess its impact on ensemble coherency and motion artifacts in MBF images. The in vivo thyroid data involved complex physiological motion due to its proximity to the pulsating carotid artery, which was used to assess the in vivo efficacy of the proposed technique. Further, in vivo renal MBF images demonstrated the feasibility of using the proposed ensemble coherence metric for curved array-based MBF imaging involving phase conversion. All ultrasound data were acquired at high imaging frame rates and the tissue signal was suppressed using spatiotemporal clutter filtering. Thyroid tissue motion was estimated using two-dimensional normalized cross correlation-based speckle tracking, which was subsequently used for ensemble motion correction. The coherence of the MBF image was quantified based on Casorati correlation of the Doppler ensemble. RESULTS: The simulation results demonstrated that an increase in ensemble motion corresponded with a decrease in ensemble coherency, which reciprocally degraded the MBF images. Further the data acquired from breast tumors demonstrated higher ensemble coherency than that from thyroid tumors. Motion correction improved the coherence of the thyroid MBF images, which substantially improved its visualization. The proposed coherence metrics were also useful in assessing the ensemble coherence for renal MBF imaging. The results also demonstrated that the proposed coherence metric can be reliably estimated from downsampled ensembles (by up to 90 % ), thus allowing improved computational efficiency for potential applications in real-time MBF imaging. CONCLUSIONS: This pilot study demonstrates the importance of assessing ensemble coherency in contrast-free MBF imaging. The proposed LSTC image quantified coherence of the Doppler ensemble for robust MBF imaging. The results obtained from this pilot study are promising, and warrant further development and in vivo validation.

Automatic Deep Learning Semantic Segmentation of Ultrasound Thyroid Cineclips Using Recurrent Fully Convolutional Networks.

Medical segmentation is an important but challenging task with applications in standardized report generation, remote medicine and reducing medical exam costs by assisting experts. In this paper, we exploit time sequence information using a novel spatio-temporal recurrent deep learning network to automatically segment the thyroid gland in ultrasound cineclips. We train a DeepLabv3+ based convolutional LSTM model in four stages to perform semantic segmentation by exploiting spatial context from ultrasound cineclips. The backbone DeepLabv3+ model is replicated six times and the output layers are replaced with convolutional LSTM layers in an atrous spatial pyramid pooling configuration. Our proposed model achieves mean intersection over union scores of 0.427 for cysts, 0.533 for nodules and 0.739 for thyroid. We demonstrate the potential application of convolutional LSTM models for thyroid ultrasound segmentation.

Automated Segmentation of Thyroid Nodule, Gland, and Cystic Components From Ultrasound Images Using Deep Learning.

Sonographic features associated with margins, shape, size, and volume of thyroid nodules are used to assess their risk of malignancy. Automatically segmenting nodules from normal thyroid gland would enable an automated estimation of these features. A novel multi-output convolutional neural network algorithm with dilated convolutional layers is presented to segment thyroid nodules, cystic components inside the nodules, and normal thyroid gland from clinical ultrasound B-mode scans. A prospective study was conducted, collecting data from 234 patients undergoing a thyroid ultrasound exam before biopsy. The training and validation sets encompassed 188 patients total; the testing set consisted of 48 patients. The algorithm effectively segmented thyroid anatomy into nodules, normal gland, and cystic components. The algorithm achieved a mean Dice coefficient of 0.76, a mean true positive fraction of 0.90, and a mean false positive fraction of 1.61×10-6. The values are on par with a conventional seeded algorithm. The proposed algorithm eliminates the need for a seed in the segmentation process, thus automatically detecting and segmenting the thyroid nodules and cystic components. The detection rate for thyroid nodules and cystic components was 82% and 44%, respectively. The inference time per image, per fold was 107ms. The mean error in volume estimation of thyroid nodules for five select cases was 7.47%. The algorithm can be used for detection, segmentation, size estimation, volume estimation, and generating thyroid maps for thyroid nodules. The algorithm has applications in point of care, mobile health monitoring, improving workflow, reducing localization time, and assisting sonographers with limited expertise.

Multi-parameter Sub-Hertz Analysis of Viscoelasticity With a Quality Metric for Differentiation of Breast Masses.

We applied sub-Hertz analysis of viscoelasticity (SAVE) to differentiate breast masses in pre-biopsy patients. Tissue response during external ramp-and-hold stress was ultrasonically detected. Displacements were used to acquire tissue viscoelastic parameters. The fast instantaneous response and slow creep-like deformations were modeled as the response of a linear standard solid from which viscoelastic parameters were estimated. These parameters were used in a multi-variable classification framework to differentiate malignant from benign masses identified by pathology. When employing all viscoelasticity parameters, SAVE resulted in 71.43% accuracy in differentiating lesions. When combined with ultrasound features and lesion size, accuracy was 82.24%. Adding a quality metric based on uniaxial motion increased the accuracy to 81.25%. When all three were combined with SAVE, accuracy was 91.3%. These results confirm the utility of SAVE as a robust ultrasound-based diagnostic tool for non-invasive differentiation of breast masses when used as stand-alone biomarkers or in conjunction with ultrasonic features.

Optimization of nitric oxide donors for investigating biofilm dispersal response in Pseudomonas aeruginosa clinical isolates.

Pseudomonas aeruginosa biofilms contribute heavily to chronic lung infection in cystic fibrosis patients, leading to morbidity and mortality. Nitric oxide (NO) has been shown to disperse P. aeruginosa biofilms in vitro, ex vivo and in clinical trials as a promising anti-biofilm agent. Traditional NO donors such as sodium nitroprusside (SNP) have been extensively employed in different studies. However, the dosage of SNP in different studies was not consistent, ranging from 500 nM to 500 µM. SNP is light sensitive and produces cyanide, which may lead to data misinterpretation and inaccurate predictions of dispersal responses in clinical settings. New NO donors and NO delivery methods have therefore been explored. Here we assessed 7 NO donors using P. aeruginosa PAO1 and determined that SNP and Spermine NONOate (S150) successfully reduced > 60% biomass within 24 and 2 h, respectively. While neither dosage posed toxicity towards bacterial cells, chemiluminescence assays showed that SNP only released NO upon light exposure in M9 media and S150 delivered much higher performance spontaneously. S150 was then tested on 13 different cystic fibrosis P. aeruginosa (CF-PA) isolates; most CF-PA biofilms were significantly dispersed by 250 µM S150. Our work therefore discovered a commercially available NO donor S150, which disperses CF-PA biofilms efficiently within a short period of time and without releasing cyanide, as an alternative of SNP in clinical trials in the future. KEY POINTS: • S150 performs the best in dispersing P. aeruginosa biofilms among 7 NO donors. • SNP only releases NO in the presence of light, while S150 releases NO spontaneously. • S150 successfully disperses biofilms formed by P. aeruginosa cystic fibrosis clinical isolates.

Discovery of Cephalosporin-3'-Diazeniumdiolates That Show Dual Antibacterial and Antibiofilm Effects against Pseudomonas aeruginosa Clinical Cystic Fibrosis Isolates and Efficacy in a Murine Respiratory Infection Model.

The formation of biofilms provides a formidable defense for many bacteria against antibiotics and host immune responses. As a consequence, biofilms are thought to be the root cause of most chronic infections, including those occurring on medical indwelling devices, endocarditis, urinary tract infections, diabetic and burn wounds, and bone and joint infections. In cystic fibrosis (CF), chronic Pseudomonas aeruginosa (P. aeruginosa) respiratory infections are the leading cause of morbidity and mortality in adults. Previous studies have shown that many bacteria can undergo a coordinated dispersal event in the presence of low concentrations of nitric oxide (NO), suggesting that NO could be used to initiate biofilm dispersal in chronic infections, enabling clearance of the more vulnerable planktonic cells. In this study, we describe efforts to create "all-in-one" cephalosporin-based NO donor prodrugs (cephalosporin-3'-diazeniumdiolates, C3Ds) that show both direct ß-lactam mediated antibacterial activity and antibiofilm effects. Twelve novel C3Ds were synthesized and screened against a panel of P. aeruginosa CF clinical isolates and other human pathogens. The most active compound, AMINOPIP2 ((Z)-1-(4-(2-aminoethyl)piperidin-1-yl)-2-(((6R,7R)-7-((Z)-2-(2-aminothiazol-4-yl)-2-(((2-carboxypropan-2-yl)oxy)imino)acetamido)-2-carboxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-en-3-yl)methoxy)diazene 1-oxide)-ceftazidime 12, showed higher antibacterial potency than its parent cephalosporin and front-line antipseudomonal antibiotic ceftazidime, good stability against ß-lactamases, activity against ceftazidime-resistant P. aeruginosa in vitro biofilms, and efficacy equivalent to ceftazidime in a murine P. aeruginosa respiratory infection model. The results support further evaluation of AMINOPIP2-ceftazidime 12 for P. aeruginosa lung infections in CF and a broader study of "all-in-one" C3Ds for other chronic infections.

Predictive value of comb-push ultrasound shear elastography for the differentiation of reactive and metastatic axillary lymph nodes: A preliminary investigation.

OBJECTIVES: To evaluate the predictive performance of comb-push ultrasound shear elastography for the differentiation of reactive and metastatic axillary lymph nodes. METHODS: From June 2014 through September 2018, 114 female volunteers (mean age 58.1±13.3 years; range 28-88 years) with enlarged axillary lymph nodes identified by palpation or clinical imaging were prospectively enrolled in the study. Mean, standard deviation and maximum shear wave elastography parameters from 117 lymph nodes were obtained and compared to fine needle aspiration biopsy results. Mann-Whitney U test and ROC curve analysis were performed. RESULTS: The axillary lymph nodes were classified as reactive or metastatic based on the fine needle aspiration outcomes. A statistically significant difference between reactive and metastatic axillary lymph nodes was observed based on comb-push ultrasound shear elastography (CUSE) results (p<0.0001) from mean and maximum elasticity values. Mean elasticity showed the best separation with a ROC analysis resulting in 90.5% sensitivity, 94.4% specificity, 0.97 area under the curve, 95% positive predictive value, and 89.5% negative predictive value with a 30.2-kPa threshold. CONCLUSIONS: CUSE provided a quantifiable parameter that can be used for the assessment of enlarged axillary lymph nodes to differentiate between reactive and metastatic processes.

Cephalosporin nitric oxide-donor prodrug DEA-C3D disperses biofilms formed by clinical cystic fibrosis isolates of Pseudomonas aeruginosa.

OBJECTIVES: The cephalosporin nitric oxide (NO)-donor prodrug DEA-C3D ('DiEthylAmin-Cephalosporin-3'-Diazeniumdiolate') has been shown to initiate the dispersal of biofilms formed by the Pseudomonas aeruginosa laboratory strain PAO1. In this study, we investigated whether DEA-C3D disperses biofilms formed by clinical cystic fibrosis (CF) isolates of P. aeruginosa and its effect in combination with two antipseudomonal antibiotics, tobramycin and colistin, in vitro. METHODS: ß-Lactamase-triggered release of NO from DEA-C3D was confirmed using a gas-phase chemiluminescence detector. MICs for P. aeruginosa clinical isolates were determined using the broth microdilution method. A crystal violet staining technique and confocal laser scanning microscopy were used to evaluate the effects of DEA-C3D on P. aeruginosa biofilms alone and in combination with tobramycin and colistin. RESULTS: DEA-C3D was confirmed to selectively release NO in response to contact with bacterial ß-lactamase. Despite lacking direct, cephalosporin/ß-lactam-based antibacterial activity, DEA-C3D was able to disperse biofilms formed by three P. aeruginosa clinical isolates. Confocal microscopy revealed that DEA-C3D in combination with tobramycin produces similar reductions in biofilm to DEA-C3D alone, whereas the combination with colistin causes near complete eradication of P. aeruginosa biofilms in vitro. CONCLUSIONS: DEA-C3D is effective in dispersing biofilms formed by multiple clinical isolates of P. aeruginosa and could hold promise as a new adjunctive therapy to patients with CF.

A novel application of Gini coefficient for the quantitative measurement of bacterial aggregation.

Non-surface attached bacterial aggregates are frequently found in clinical settings associated with chronic infections. Current methods quantifying the extent to which a suspended bacterial population is aggregated mainly rely on: (1) cell size distribution curves that are difficult to be compared numerically among large-scale samples; (2) the average size/proportion of aggregates in a population that do not specify the aggregation patterns. Here we introduce a novel application of Gini coefficient, herein named Aggregation Coefficient (AC), to quantify the aggregation levels of cystic fibrosis Pseudomonas aeruginosa (CF-PA) isolates in vitro using 3D micrographs, Fiji and MATLAB. Different aggregation patterns of five strains were compared statistically using the numerical AC indexes, which correlated well with the size distribution curves plotted by different biovolumes of aggregates. To test the sensitivity of AC, aggregates of the same strains were treated with nitric oxide (NO), a dispersal agent that reduces the biomass of surface attached biofilms. Strains unresponsive to NO were reflected by comparable AC indexes, while those undergoing dispersal showed a significant reduction in AC index, mirroring the changes in average aggregate sizes and proportions. Therefore, AC provides simpler and more descriptive numerical outputs for measuring different aggregation patterns compared to current approaches.

Diagnosis and treatment of biofilm infections in children.

PURPOSE OF REVIEW: Biofilm-associated infections cause difficulties in the management of childhood chronic infections and other diseases, due to the invasive nature of interventions which are often necessary for definitive management. Despite their importance, there are challenges in diagnosing biofilm infections and gaps in clinicians' understanding regarding the significance of biofilms. RECENT FINDINGS: Many chronic infections associated with biofilms remain difficult or impossible to eradicate with conventional therapy. Surgical intervention, implant removal or long-term intermittent or suppressive antimicrobial therapy may be required. There are still significant challenges in detecting biofilms which presents a barrier in clinical practice and research. Novel therapies to disrupt biofilms are currently under investigation, which may help reduce the impact of antimicrobial resistance. SUMMARY: Biofilm-associated infection should be considered wherever there is clinical concern for an infection affecting prosthetic material, where there is a predisposing condition such as suppurative lung disease; or in the setting of chronic or relapsing infections which may be culture negative. New diagnostic methods for detecting biofilms are a research priority for both clinical diagnosis and the ability to conduct high quality clinical trials of novel antibiofilm interventions.