Combinatorial mixtures of organic solutes for improved liquid/liquid extraction of ions.

Biological systems routinely extract and organize ions in complex yet highly ordered and active systems. Much of this function is attributed to proteins, although recent evidence indicates aggregates of lipids are also capable of molecular recognition. Here we tested the hypothesis that combinatorial mixtures of organic solutes might lead to enhanced liquid/liquid extraction. We started with liquid oleic acid as an organic phase extracting copper ions from water and added a library of additives. By using Bayesian optimization to autonomously direct the combinatorial formulation, we discovered mixtures that enhanced the extraction performance. The main additive that improved the system was octylphosphonic acid. Interestingly, the optimal mixture has a significant improvement compared to this additive alone. This suggests that the combinations of organic solutes are better than using pure components in liquid/liquid extraction. Furthermore, we found that precipitation occurs in the samples showing better extraction efficiency, which has interesting material properties and potential for new types of supramolecular biosensors.

Helium atom scattering from potassium tantalate/niobate(001) surfaces.

The results of He atom scattering experiments on KTa0.48Nb0.52O3(001) surfaces are presented and compared with similar experiments on mixed potassium tantalate/niobate perovskites with lower concentrations of Nb. The results are puzzling, unique and intriguing. Angular distributions of the He scattering intensities (the He surface diffraction pattern) are found to be very similar to those obtained from targets with lower Nb fractions. However, drift spectra (the intensity of the He specular reflection as a function of the He wave vector) are not. Whereas the drift spectra in the 〈10〉 azimuth do resemble those of the 0, 6 and 10% Nb fraction targets, in the 〈11〉 azimuth they are more similar to the strange drift spectra found from the 30% Nb-doped targets. Most intriguing are the surface temperature scans (the He specular intensity as a function of the target surface temperature); for they are quite distinct from those with lower Nb fractions. Finally, the inelastic He scattering experiments provide phonon dispersion results that are similar in most respects to those found for the lower Nb level samples. In particular, the most prominent feature is an Einstein-like mode at about 13.5 meV which spans the entire surface Brillouin zone in both 〈10〉 and 〈11〉 azimuths.

Quantification of Protein-Induced Membrane Remodeling Kinetics In Vitro with Lipid Multilayer Gratings.

The dynamic self-organization of lipids in biological systems is a highly regulated process that enables the compartmentalization of living systems at micro- and nanoscopic scales. Consequently, quantitative methods for assaying the kinetics of supramolecular remodeling such as vesicle formation from planar lipid bilayers or multilayers are needed to understand cellular self-organization. Here, a new nanotechnology-based method for quantitative measurements of lipid-protein interactions is presented and its suitability for quantifying the membrane binding, inflation, and budding activity of the membrane-remodeling protein Sar1 is demonstrated. Lipid multilayer gratings are printed onto surfaces using nanointaglio and exposed to Sar1, resulting in the inflation of lipid multilayers into unilamellar structures, which can be observed in a label-free manner by monitoring the diffracted light. Local variations in lipid multilayer volume on the surface is used to vary substrate availability in a microarray format. A quantitative model is developed that allows quantification of binding affinity (K D ) and kinetics (kon and koff ). Importantly, this assay is uniquely capable of quantifying membrane remodeling. Upon Sar1-induced inflation of single bilayers from surface supported multilayers, the semicylindrical grating lines are observed to remodel into semispherical buds when a critical radius of curvature is reached.

Natural history and quality of life in patients with cystine urolithiasis: a single centre study.

OBJECTIVE: To describe the natural history and quality of life (QoL) in patients with cystine urolithiasis. PATIENTS AND METHODS: A cohort study was carried out involving participants recruited from a single surgeon's case mix. Patients with cystinuria and related urolithiasis were invited to complete a questionnaire involving demographic information, use of medical treatment, surgical interventions and the 36-item short-form 36-item short-form health survey (SF-36). RESULTS: In all, 14 patients completed the survey. The SF-36 survey showed lower QoL than the general public in seven of eight domains. The mean interventional rate in patients with cystinuria was 10.6 procedures per patient. Most patients reported previous use of d-penicillamine and urinary alkalinisation medications, with most ceasing due to side-effects or lack of perceived efficacy. CONCLUSION: Cystinuria is associated with a high rate of surgical intervention and lower QoL than the general public. Individuals with this condition report that medical management is either ineffective or poorly tolerated. There is a need for further improvements in medical management of cystinuria, to reduce the rate of operative intervention.

Lipid Multilayer Grating Arrays Integrated by Nanointaglio for Vapor Sensing by an Optical Nose.

Lipid multilayer gratings are recently invented nanomechanical sensor elements that are capable of transducing molecular binding to fluid lipid multilayers into optical signals in a label free manner due to shape changes in the lipid nanostructures. Here, we show that nanointaglio is suitable for the integration of chemically different lipid multilayer gratings into a sensor array capable of distinguishing vapors by means of an optical nose. Sensor arrays composed of six different lipid formulations are integrated onto a surface and their optical response to three different vapors (water, ethanol and acetone) in air as well as pH under water is monitored as a function of time. Principal component analysis of the array response results in distinct clustering indicating the suitability of the arrays for distinguishing these analytes. Importantly, the nanointaglio process used here is capable of producing lipid gratings out of different materials with sufficiently uniform heights for the fabrication of an optical nose.

Lipid-Based Catalysis Demonstrated by Bilayer-Enabled Ester Hydrolysis.

Lipids have not traditionally been considered likely candidates for catalyzing reactions in biological systems. However, there is significant evidence that aggregates of amphiphilic compounds are capable of catalyzing reactions in synthetic organic chemistry. Here, we demonstrate the potential for the hydrophobic region of a lipid bilayer to provide an environment suitable for catalysis by means of a lipid aggregate capable of speeding up a chemical reaction. By bringing organic molecules into the nonpolar or hydrophobic region of a lipid bilayer, reactions can be catalyzed by individual or collections of small, nonpolar, or amphiphilic molecules. We demonstrate this concept by the ester hydrolysis of calcein-AM to produce a fluorescent product, which is a widely used assay for esterase activity in cells. The reaction was first carried out in a two-phase octanol-water system, with the organic phase containing the cationic amphiphiles cetyltrimethylammonium bromide (CTAB) or octadecylamine. The octanol phase was then replaced with phospholipid vesicles in water, where the reaction was also found to be carried out. The reaction was monitored using quantitative fluorescence, which revealed catalytic turnover numbers on a scale of 10-7 to 10-8 s-1 for each system, which is much slower than enzymatic catalysis. The reaction product was characterized by 1H-NMR measurements, which were consistent with ester hydrolysis. The implications of thinking about lipids and lipid aggregates as catalytic entities are discussed in the context of biochemistry, pharmacology, and synthetic biology.

Scalable lipid droplet microarray fabrication, validation, and screening.

High throughput screening of small molecules and natural products is costly, requiring significant amounts of time, reagents, and operating space. Although microarrays have proven effective in the miniaturization of screening for certain biochemical assays, such as nucleic acid hybridization or antibody binding, they are not widely used for drug discovery in cell culture due to the need for cells to internalize lipophilic drug candidates. Lipid droplet microarrays are a promising solution to this problem as they are capable of delivering lipophilic drugs to cells at dosages comparable to solution delivery. However, the scalablility of the array fabrication, assay validation, and screening steps has limited the utility of this approach. Here we take several new steps to scale up the process for lipid droplet array fabrication, assay validation in cell culture, and drug screening. A nanointaglio printing process has been adapted for use with a printing press. The arrays are stabilized for immersion into aqueous solution using a vapor coating process. In addition to delivery of lipophilic compounds, we found that we are also able to encapsulate and deliver a water-soluble compound in this way. The arrays can be functionalized by extracellular matrix proteins such as collagen prior to cell culture as the mechanism for uptake is based on direct contact with the lipid delivery vehicles rather than diffusion of the drug out of the microarray spots. We demonstrate this method for delivery to 3 different cell types and the screening of 92 natural product extracts on a microarray covering an area of less than 0.1 cm2. The arrays are suitable for miniaturized screening, for instance in high biosafety level facilities where space is limited and for applications where cell numbers are limited, such as in functional precision medicine.

The N-terminal region of wheat streak mosaic virus coat protein is a host- and strain-specific long-distance transport factor.

Understanding the genetics underlying host range differences among plant virus strains can provide valuable insights into viral gene functions and virus-host interactions. In this study, we examined viral determinants and mechanisms of differential infection of Zea mays inbred line SDp2 by Wheat streak mosaic virus (WSMV) isolates. WSMV isolates Sidney 81 (WSMV-S81) and Type (WSMV-T) share 98.7% polyprotein sequence identity but differentially infect SDp2: WSMV-S81 induces a systemic infection, but WSMV-T does not. Coinoculation and sequential inoculation of SDp2 with WSMV-T and/or WSMV-S81 did not affect systemic infection by WSMV-S81, suggesting that WSMV-T does not induce a restrictive defense response but that virus-encoded proteins may be involved in differential infection of SDp2. The viral determinant responsible for strain-specific host range was mapped to the N terminus of coat protein (CP) by systematic exchanges of WSMV-S81 sequences with those of WSMV-T and by reciprocal exchanges of CP or CP codons 1 to 74. Green fluorescent protein (GFP)-tagged WSMV-S81 with CP or CP residues 1 to 74 from WSMV-T produced similar numbers of infection foci and genomic RNAs and formed virions in inoculated leaves as those produced with WSMV-S81, indicating that failure to infect SDp2 systemically is not due to defects in replication, cell-to-cell movement, or virion assembly. However, these GFP-tagged hybrids showed profound defects in long-distance transport of virus through the phloem. Furthermore, we found that four of the five differing amino acids in the N terminus of CP between the WSMV-S81 and WSMV-T isolates were collectively involved in systemic infection of SDp2. Taken together, these results demonstrate that the N-terminal region of tritimoviral CP functions in host- and strain-specific long-distance movement.

Genital premalignant and malignant diseases: a retrospective study of male genital skin biopsies.

BACKGROUND: Genital skin malignancies are a rare entity encountered by clinicians that may result in significant morbidity and mortality. Lack of familiarity or expertise in this area among clinicians can delay appropriate management of these conditions and may result in disease progression. METHODS: We performed a retrospective descriptive cohort study of male patients who received a genital skin biopsy reported by one major dermatopathology laboratory between January 2017 and December 2018 with a histological diagnosis of a premalignant or malignant condition. Patient age, type of clinician, clinical notes, genital site, type of biopsy performed, and histopathological diagnosis were evaluated. RESULTS: Of the 1525 male genital skin biopsies available for analysis, 5% (74/1525) were premalignant or malignant diseases. These included penile intraepithelial neoplasia (PeIN) (42/74, 57%), followed by invasive squamous cell carcinoma (SCC) (11/74, 15%), Bowenoid papulosis (BP) (9/74, 12%), basal cell carcinoma (8/74, 11%), malignant melanoma (2/74, 2.7%), extramammary Paget's (1/74, 1.4%), and metastatic cutaneous deposits (1/74, 1.4%). PeIN and BP most commonly affected the penile shaft ([18/42] 43% and [4/9] 44%, respectively), invasive SCCs most commonly affected the glans penis (4/11, 36%), and all BCCs (8/8, 100%) were located on the scrotum. Invasive SCCs were most biopsied by urologists, BP was most biopsied by dermatologists, and PeINs and BCCs were most biopsied by general practitioners. CONCLUSIONS: A variety of genital malignancy types were found in males. A greater understanding of male genital premalignant and malignant conditions may help guide education and further research in this area.

Penile squamous cell carcinoma following childhood hypospadias repair.

We report a case of a 46-year-old man who presented with recurrent lower urinary tract infections and obstructive voiding symptoms on the background of a proximal hypospadias repair at 4 years of age. A mass was later identified at his distal urethra, which on biopsy was found to be a penile squamous cell carcinoma. The lesion was excised en bloc, and at 10 months post-excision, the patient has no evidence of local or regional recurrence.

Ureaplasma urealyticum infection presenting as pyelonephritis and perinephric abscess in an immunocompromised patient.

We report a case of an immunosuppressed 67-year-old woman who presented with fever of unknown origin. Further investigation revealed multiple left renal and perinephric abscesses. These were managed with percutaneous drainage and broad-spectrum antibiotics; however, no clinical improvement resulted. No organism was identifiable on standard microscopy and culture of all drain, blood and urine samples taken. Left nephrectomy with right urinary diversion was performed for sepsis management and to protect the patient's right kidney. Eventually, Ureaplasma species' RNA was detected in the patient's drain fluid using PCR and 16S ribosomal RNA gene sequencing. The patient was treated successfully with targeted antibiotic therapy and underwent extensive rehabilitation following this. Histopathology of the nephrectomy specimen suggested xanthogranulomatous pyelonephritis.

Organic Composomes as Supramolecular Aptamers.

Information contained in the sequences of biological polymers such as DNA and protein is crucial to determining their function. Lipids are not generally thought of as information-containing molecules. However, from a supramolecular perspective, the number of possible combinations of lipids in a mixture is comparable to the complexity of DNA or proteins. Here, we test the idea that an organic composome can exhibit molecular recognition. We use water/octanol as a model two-phase system and investigate the effect of organic solutes in different combinations in the organic phase on selective partitioning of two water-soluble dyes (Brilliant Blue FCF and Allura Red AC) from the aqueous phase into the organic phase. We found that variation in the concentration of the surfactant cetyltrimethylamonium bromide (CTAB) in the octanol phase alone was sufficient to cause a switch in selectivity, with low CTAB concentrations being selective for the red dye and high CTAB concentrations being selective for the blue dye. Other organic components were added to the organic phase to introduce molecular diversity into the composome and directed evolution was used to optimize the relative concentrations of the solutes. An improvement of selective partitioning in the heterogeneous system over the pure CTAB solution was observed. The results indicate that supramolecular composomes are sufficient for molecular recognition processes in a way analogous to nucleic acid aptamers.

Aptamer Functionalized Lipid Multilayer Gratings for Label-Free Analyte Detection.

Lipid multilayer gratings are promising optical biosensor elements that are capable of transducing analyte binding events into changes in an optical signal. Unlike solid state transducers, reagents related to molecular recognition and signal amplification can be incorporated into the lipid grating ink volume prior to fabrication. Here we describe a strategy for functionalizing lipid multilayer gratings with a DNA aptamer for the protein thrombin that allows label-free analyte detection. A double cholesterol-tagged, double-stranded DNA linker was used to attach the aptamer to the lipid gratings. This approach was found to be sufficient for binding fluorescently labeled thrombin to lipid multilayers with micrometer-scale thickness. In order to achieve label-free detection with the sub-100 nm-thick lipid multilayer grating lines, the binding affinity was improved by varying the lipid composition. A colorimetric image analysis of the light diffracted from the gratings using a color camera was then used to identify the grating nanostructures that lead to an optimal signal. Lipid composition and multilayer thickness were found to be critical parameters for the signal transduction from the aptamer functionalized lipid multilayer gratings.

Urethral syringocele: unseen but existing.

BACKGROUND: Bulbourethral syringocele is an uncommon and under-diagnosed condition most commonly seen in the paediatric population, although there is increasing recognition in adults. Due to the difficulty in diagnosis, we report our experience of urethral syringocele in a quaternary paediatric hospital, with differing presentations, diagnosis and treatment. METHODS: This is a retrospective review of seven cases of children over a period of 14 years, including their presentations, diagnosis, treatment and follow-up. A review of the current literature is presented. RESULTS: The median age of these seven cases at presentation was 11 years (6 days to 16 years). Clinical features varied with age, with obstructive uropathy in a neonate, urinary tract infection in an infant, scrotal abscess in two children and lower urinary tract obstructive symptoms in three teenagers. Diagnostic voiding cystogram diagnosed the majority of syringoceles and two were seen on magnetic resonance imaging. Five boys underwent endoscopic transurethral deroofing and two children required transperineal marsupialization. Long-term follow-up showed all had complete resolution of symptoms. CONCLUSION: Urethral syringocele presents from the neonatal period to late adolescence, with the presenting features reflective of age. Surgical management can be performed endoscopically or by open approach. Awareness of this condition and inclusion in the differential diagnosis, particularly in the setting of an atypical or recurrent scrotal abscess, could avoid a prolonged therapeutic course.

Characterization of a U.S. Isolate of Beet black scorch virus.

ABSTRACT The first reported U.S. isolate of Beet black scorch necrovirus (BBSV) was obtained and characterized. Host range of the virus for localized and occasionally systemic infection included the Chenopodiaceae and Tetragonia expansa; Nicotiana benthamiana supported symptomless systemic infection by the virus. The complete nucleotide sequence of the genomic RNA of the virus, designated BBSV-Co, exhibits 93% similarity to the genome of the 'Ningxia' isolate of BBSV from China. Amino acid sequence similarity in predicted genes ranged from 95% in the p4 gene to 97% in the p82 and coat protein genes. A potential additional gene exists within the U.S. isolate of BBSV that is absent from Chinese isolates of BBSV due to nucleotide differences between these isolates within the coat protein gene. Coat protein analysis by isoelectric focusing and by mass spectroscopy indicated the presence of phosphorylated residues. Using primer extension analysis of the 5' end of the genome and site-directed mutants of genomic clones of BBSV-Co from which infectious RNA was produced, the native 5' end of the BBSV-Co genome was determined to be 5'-GAAACCTAACC...3', lacking the two terminal adenosine nucleotides in the published sequences of BBSV from China.

Nonmonotonic DNA-length-dependent mobility in pluronic gels.

Two-dimensional electrophoresis was used to analyze the mobility of DNA fragments in micellar gels of pluronic F127 (EO_{100}PO_{70}EO_{100}) and pluronic P123 (EO_{20}PO_{70}EO_{20}). The 20-3500 base pair DNA fragments were separated by size first in agarose gels, and then in pluronic gels at room temperature. In agarose gels, the DNA mobility decreases monotonically with increasing DNA length. In pluronic gels, however, the mobility varies nonmonotonically according to fragment lengths that are strongly correlated with the diameter of the spherical micelles. Brownian dynamics (BD) simulations with short-ranged intra-DNA hydrodynamic interactions were performed to numerically calculate the length-dependent mobility in pluronic lattices. The rising and falling trends, as well as the oscillations of mobility, were captured by the coarse-grained BD simulations. Molecular dynamics simulations in pluronic F127, with explicitly modeled micelle coronas, justified that the hydrodynamic interactions mediated by the complex fluid of hydrated poly(ethylene oxide) are a possible reason for the initial rise of mobility with DNA length.

Variation in growth and competitive ability between sexually and clonally produced hydroids.

In controlled laboratory experiments, colonies of Podocoryna carnea typically overgrow and kill colonies of Hydractinia symbiolongicarpus. Generally, these experiments have used colonies grown from tissue explants (clonal replicates) surgically removed from mature colonies taken from natural populations. In contrast, experiments involving interspecific bouts between small, sexually produced colonies reveal that both the characteristics and outcomes of competition differ from previous studies. During competition between small sexually produced colonies, H. symbiolongicarpus exhibits directional growth toward P. carnea and produces nematocyte-rich hyperplastic stolons more readily than P. carnea does. Nevertheless, P. carnea can still overcome H. symbiolongicarpus if it initially grows away from the contact zone and subsequently flanks H. symbiolongicarpus. Overall, sexually produced colonies of H. symbiolongicarpus destroyed their P. carnea counterparts in more than 35% of competitive bouts, whereas P. carnea dominated their H. symbiolongicarpus counterparts in all similar encounters between clonally produced colonies. In natural populations, competition between small sexually produced colonies of H. symbiolongicarpus may predominate, and these results support the hypothesis that this species is adapted to competition early in colony development. More generally, studies of competition between sexually produced colonies should complement similar studies of clonally produced colonies.

Lower urinary tract symptoms: thinking beyond the urinary tract.

We present a case of a 54-year-old man with progressive lower urinary tract symptoms over 12 months. Physical examination, urinalysis, serum biochemistry and ultrasound of the renal tract were all unremarkable. Flexible cystoscopy was normal. Urodynamic assessment revealed an overactive bladder of unknown aetiology. The patient went on to have an MRI of the lumbosacral spine which showed a spinal cord tumour of the conus medullaris. The patient underwent a laminectomy and resection of the tumour. Histology showed myxopapillary ependymoma of the spinal cord. This case highlights the need to consider the full spectrum of causes, urological and non-urological, in assessing a patient with voiding dysfunction.

Single molecule observation of DNA electrophoresis in pluronic f127.

Single molecule fluorescence microscopy is used to follow the motion of long DNA molecules undergoing electrophoresis in Pluronic gels. We find that for low fields most DNA molecules follow tortuous paths through the gels, at an angle up to 90 degrees from the field direction, while some molecules find paths along the field lines. In high fields, virtually all of the DNA molecules follow the field lines. In many cases, the molecules travel as compact coils with optically discernible radii smaller than in free solution. In other cases, the molecules extend and contract or travel in an extended configuration.