Prevalence and correlates of Trichomonas vaginalis infection among female US federal prison inmates

BACKGROUND: Previous studies have observed high prevalences of Trichomonas vaginalis infection among women entering US jails and state prisons (22%–47%). We sought to determine the prevalence among women incarcerated in 2 US female-only federal prisons. METHODS: Female inmates were recruited at 2 prisons (n = 624). Participants completed a self-administered questionnaire and provided self-collected first-catch urine and vaginal swab specimens. Specimens were tested for T. vaginalis DNA. RESULTS: Approximately 8.5% of participants at the first prison, and 8.3% at the second prison had a positive urine result, vaginal swab result or both, for a combined prevalence of 8.5%. Using positivity in either specimen as the reference standard, urine polymerase chain reaction had a sensitivity of 66.7% and vaginal swab polymerase chain reaction had a sensitivity of 84.4%. The only significant positive correlate of T. vaginalis infection was lower household income before arrest. Other variables nonsignificantly positively correlated with T. vaginalis were being employed at the time of arrest, having experienced sexual, physical, or emotional abuse by a family member, having a parent who had not had a drug or alcohol addiction, never exchanging sex for money or drugs, ever being pregnant, having abnormal vaginal bleeding/spotting, and having concurrent chlamydia or gonorrhea. CONCLUSIONS: Although not as high as in other studies of women entering US jails and state prisons, our observed T. vaginalis prevalence of 8.5% was much higher than in the general US population. Therefore, screening for T. vaginalis infection may be warranted at federal prison entry, as well as sexual health education during prison stay

A paperfluidic platform to detect Neisseria gonorrhoeae in clinical samples

Globally, the microbe Neisseria gonorrhoeae (NG) causes 106 million newly documented sexually transmitted infections each year. Once appropriately diagnosed, NG infections can be readily treated with antibiotics, but high-risk patients often do not return to the clinic for treatment if results are not provided at the point of care. A rapid, sensitive molecular diagnostic would help increase NG treatment and reduce the prevalence of this sexually transmitted disease. Here, we report on the design and development of a rapid, highly sensitive, paperfluidic device for point-of-care diagnosis of NG. The device integrates patient swab sample lysis, nucleic acid extraction, thermophilic helicase-dependent amplification (tHDA), an internal amplification control (NGIC), and visual lateral flow detection within an 80 min run time. Limits of NG detection for the NG/NGIC multiplex tHDA assay were determined within the device, and clinical performance was validated retroactively against qPCR-quantified patient samples in a proof-of-concept study. This paperfluidic diagnostic has a clinically relevant limit of detection of 500 NG cells per device with analytical sensitivity down to 10 NG cells per device. In triplicate testing of 40 total urethral and vaginal swab samples, the device had 95% overall sensitivity and 100% specificity, approaching current laboratory-based molecular NG diagnostics. This diagnostic platform could increase access to accurate NG diagnoses to those most in need.This work was funded by the National Institute of Health National Institute of Allergy and Infectious Diseases award number R01 AI113927 to Boston University and the NIH National Institute of Biomedical and Bioengineering award number U54 EB007958 to Johns Hopkins University. (R01 AI113927 - National Institute of Health National Institute of Allergy and Infectious Diseases; U54 EB007958 - NIH National Institute of Biomedical and Bioengineering)Accepted manuscrip

Quantification of long-range power law correlations among healthy and pathologic subjects using detrended fluctuation analysis and multifractal detrended fluctuation analysis

The healthy heartbeat is traditionally thought to be regulated according to the classical principle of homeostasis whereby physiologic systems operate to reduce variability and achieve an equilibrium-like state. However recent studies reveal that under normal conditions, beat-to-beat fluctuation in heart rate display the kind of long-range correlations typically exhibited by the dynamical system far away from equilibrium. In contrast, heart rate time series from patients with severe congestive heart failure show a breakdown of this long-range correlation behavior. Two different non-linear dynamic methods namely Detrended Fluctuation Analysis (DFA) and Multifractal (MF) DFA are used for the quantification of this correlation property in non-stationary physiological time series and it revealed the presence of long-range power law correlation for the group of healthy subjects while breakdown in the long-range power law correlation for the group of subjects with cardiac heart failure. Application of DFA analysis shows evidence for a crossover phenomenon associated with a change in short(αl) and long(α2) range scaling exponents. For healthy subjects, calculated value of αl and α2 (mean value ± S.D.) are 1.31 ± 0.17 and 1.00 ± 0.07 respectively. For subjects with cardiac heart failure calculated value of ctl and a2 is 0.71 ± 0.20 and 1.24 ± 0.07 respectively i.e. only one scaling exponent is not sufficient to characterize the entire heart-rate time series which resulted into MF-DFA approach. This suggested that there is more than one exponent values needed to characterize the heart rate time series. Multifractal DFA is based on generalization of DFA and a MATLAB code is developed to implement the MF-DFA algorithm and to identify whether the given time series under analysis exhibits multifractality or not by generating more than one exponent values for multifractal signal. The value of a for q\u3eO for healthy is 1.04 ± 0.02 and for CHF is found to be 1.32 ± 0.02 and the value of a for q\u3c0 for healthy subjects is 3.01 ±0.26 and for CHF subjects is found to be 3.53 ± 0.14 (mean value ± S.D.) The student\u27s ttest suggests that p-value is 0.00001 which is less than 0.05 thus the value of a for q \u3c0 and q\u3e0 among healthy subjects and CHF subjects are statistically different. Value of a for q\u3e0 is less than that for q\u3c0. And for q =2 MF-DFA retains monofractal DFA. Thus, MF-DFA is clearly able to discriminate among the healthy and CHF for q\u3c0 as well for q\u3e0. MF-DFA also determines which fluctuations i.e. (small or large) dominate for the given interbeat interval time series because for q\u3c0 the slow fluctuations dominate whereas for q\u3e0 large fluctuations dominate. DFA and MF-DFA were able to discriminate 23 Healthy subjects out of 26 Healthy subjects data sets i.e. true positive specificity is 0.89 and false negative specificity is 0.12 and 9 CHF subjects out of 11 CHF subjects data sets i.e. true positive specificity is 0.82 and false negative specificity is 0.19. These methods may be of use in distinguishing healthy from pathologic data sets based on the difference in the scaling properties

Nanofiber fabrication in a temperature and humidity controlled environment for improved fibre consistency

To fabricate nanofibers with reproducible characteristics, an important demand for many applications, the effect of controlled atmospheric conditions on resulting electrospun cellulose acetate (CA) nanofibers was evaluated for temperature ranging 17.5 - 35°C and relative humidity ranging 20% - 70%. With the potential application of nanofibers in many industries, especially membrane and filter fabrication, their reproducible production must be established to ensure commercially viability.
Cellulose acetate (CA) solution (0.2 g/ml) in a solvent mixture of acetone/DMF/ethanol (2:2:1) was electrospun into nonwoven fibre mesh with the fibre diameter ranging from 150nm to 1µm.
The resulting nanofibers were observed and analyzed by scanning electron microscopy (SEM), showing a correlation of reducing average fibre diameter with increasing atmospheric temperature. A less pronounced correlation was seen with changes in relative humidity regarding fibre diameter, though it was shown that increased humidity reduced the effect of fibre beading yielding a more consistent, and therefore better quality of fibre fabrication.
Differential scanning calorimetry (DSC) studies observed lower melt enthalpies for finer CA nanofibers in the first heating cycle confirming the results gained from SEM analysis. From the conditions that were explored in this study the temperature and humidity that gave the most suitable fibre mats for a membrane purpose were 25.0°C and 50%RH due to the highest level of fibre diameter uniformity, the lowest level of beading while maintaining a low fibre diameter for increased surface area and increased pore size homogeneity. This study has highlighted the requirement to control the atmospheric conditions during the electrospinning process in order to fabricate reproducible fibre mats

Single-use primary capture technology with the promise to deliver new standards for the economics, convenience and reliability of mAb bioprocessing

Product capture chromatography has been crucial in the process development and manufacture of mAb therapeutics over the past 20 years, and in particular Protein A affinity. Chromatography is a step that has had a lot of process development time attributed to getting packed beds to perform to their maximum capability and most of the optimisation stems from limitations of the inherent media and its pack form, such as mass transfer & pressure drop limitations, channeling and bead-wall support effects. The limited throughput that this media offers has prevented this unit operation from being economically accessible in a single-use format. Here we present 3 case studies of work using this a novel nanofibre adsorbent which takes the well-developed performance characteristics of chromatography utilising the same chemical base materials and process infrastructure and delivers a productivity improvement of 50-fold while maintaining product CQAs. This huge throughput advantage enables the drug manufacture to choose whether they want to reduce this unit process size such that the chromatography cartridge’s lifetime (in terms of cycles) can be exhausted over a single batch, with the aim of making in single-use operation economically feasible, or whether they what to trade that off with operating their unit operation in a significantly reduced time period. The goal of this is not just to reduce COGs associated with chromatography, but to enable new overall processing strategies to be employed giving drug manufacturers greater flexibility in their choice of operations and thereby maximising the productivity of new and existing drug manufacturing facilities. The work presented here explores the physical properties that enable this high productivity operation and discusses the resulting product characterisation and process considerations. The industrial suitability of the nanofibre technology has been tested across a 1,000x increase in scale. Initial development work focused on high-throughput screening studies on the Tecan liquid handling system (10-50μL) and lab scale (125μL-1mL nanofibre volume). Run times of less than 3 minutes allowed the impact of key process parameters on quality attributes such as host cell protein and product concentration to be quickly optimised. This work was then scaled to a pilot purification of a 50L CHO cell culture in a single batch. In this initial feasibility study, using a 10mL prototype housing unit, a productivity of 460g/L/h was achieved, with a recovery yield over 90%. A 3-log reduction in host cell proteins was also maintained over 200 cycles, and the Protein A leaching was less than 7ppm. Please click Additional Files below to see the full abstract

Abschlussbericht für ein FuEuI-Vorhaben des im Rahmen der Bekanntmachung des BMFTR "Innovationsräume Bioökonomie" geforderten Innovationsraums "NewFoodSystems - Neue Lebensmittelsysteme"

Ziel dieses Vorhabens war es ein in Bezug auf die Aminosäurezusammensetzung bedarfsgerechtes Starterfutter für Larven der Schwarzen Soldatenfliege (Herme ia illucens) zu formulieren. Das erste Larvenstadium ist das sensibelste Stadium während der Aufzucht von Herme ia illucens. Dafür wurden Versuchsansätze von jeweils 10.000 Neonates durch Wiegung aufgeteilt. Um ausschließlich lebende Neonates auszuwählen, fand vor der Wiegung eine Aussiebung abgestorbener Neonates statt. Ziel der Versuche war es durch Limitierung und Supplementierung einzelner Aminosäuren im Vergleich zu einer Kontrolldiät (Junghennenfutter) und einer Positiv-Kontrolle (Basisfutter-Diät, in der alle Aminosäuren auf das Niveau des Junghennenfutters ergänzt wurden) den Aminosäurenbedarf abzuleiten. Ein proteinreduziertes Basisfutter aus einer Getreidemischung (Zentrakorn, 40%), Junghennenfutter (33%), Stroh (gemahlen, 20%) und Zucker (7%) diente als Negativkontrolle. Im Versuch wurden die Aminosäuren Methionin, Lysin, Threonin, Tryptophan, Isoleucin und Arginin untersucht. Die Bewertung fand anhand der folgenden Untersuchungsparameter statt: Mortalität/Überlebensrate, Dynamik der Larvengewichte, Biomassekonversionsrate, sonstige relevante Eigenschaften und Beobachtungen (Feuchtegehalt des Restsubstrats, Schimmelbildung etc.). Es erfolgte eine Futtergabe von 50g je Versuchsansatz pro 10.000 Neonates am Versuchsbeginn sowie eine zweite Futtergabe mit gleicher Menge sobald die erste Ration vollständig gefressen wurde (visuelle Beurteilung). Das Versuchsende erfolgte nach vollständigem Verzehr der zweiten Ration

Struktur-basiertes Design, Synthese und pharmakokinetische Charakterisierung von kovalenten Inhibitoren zur Adressierung von Krebs-relevanten Proteinkinasen

Die vorliegende Arbeit beinhaltet zwei Abschnitte: (i) die pharmakokinetische Charakterisierung von Inhibitoren mit dem Fokus auf der Analyse der metabolischen Stabilität in Anwesenheit von Lebermikrosomen, die einen Phase I Metabolismus nachbilden. Hierzu wurde ein entsprechendes Assaysystem in der Arbeitsgruppe etabliert. (ii) Das Struktur basierte Wirkstoffdesign, die organische Synthese von niedermolekularen Verbindungen und deren Charakterisierung, mit dem Ziel selektive und potente Inhibitoren für Krebs relevante Proteinkinasen zu identifizieren. Die Kombination aus biochemischem Aktivitätsassay und zellulärem Viabilitätsassay erlaubte eine Bewertung der generierten Substanzbibliotheken. Massenspektrometrische Studien ermöglichten die Analyse der kovalenten Adressierung des Zielproteins, sowie Komplexkristallstrukturen verwendet wurden, welche tiefere Einblicke in die Bindungsmodi der Inhibitoren erlaubten. Im Rahmen dieser Arbeit wurde ein Phase I Metabolismus Assays etabliert, der einen wichtigen Beitrag zur Optimierung und Weiterentwicklung der Inhibitoren leistet. Der Assay wurde in Gegenwart von humanen und murinen Lebermikrosomen durchgeführt, welche die Cytochrom P450 Enzyme beinhalten, durch die die Mehrheit der auf dem Markt zugelassenen Wirkstoffe abgebaut werden. Untersucht wurde die metabolische Stabilität von 200 Verbindungen über die Zeit, sodass Parameter wie die Halbwertszeit (t1/2) und die intrinsische Clearance (CLint) für eine Vielzahl an Verbindungen aus unterschiedlichen Projekten bestimmt und mit Referenzverbindungen verglichen werden konnten. Die Proteinkinase MKK7 ist Teil des JNK Signalwegs, dessen Fehlregulierung in neurodegenerativen Erkrankungen sowie bei Herzinfarkten mit darauffolgenden ischämischen Reperfusionsschäden identifiziert wurde. Unzureichende Selektivitätsprofile und mangelnde Wirksamkeit wurden bislang für JNK Inhibitoren beschrieben, weshalb die Adressierung der JNK aktivierenden Kinase MKK7 eine gute Alternative darstellt die genannten Selektivitätsmängel zu umgehen. Der literaturbeschriebene, potente Pyrazolopyrimidin-basierte kovalente MKK7 Inhibitor 24 sollte in komplexen zelluläre in vitro und ersten in vivo Modellsystemen getestet werden, weshalb zunächst ein Gramm der Verbindung hergestellt wurde. Die Identifizierung einer geeigneten Formulierung für die Substanz sowie die Bestimmung der maximal tolerierbaren Dosis bildeten die Grundlage für erste in vivo Mausstudien. Aus diesen ging bei 600 mg pro kg Körpergewicht der Maus durch IV Applikation eine maximale Plasmakonzentration von 5 µM hervor, sowie eine gute Verträglichkeit der Substanz. Die nur mäßige Löslichkeit der Verbindung stellte sich als problematisch dar, weshalb verschiedene Salzformen dargestellt wurden, von denen das HCl Salz 34 eine verbesserte Löslichkeit zeigte. Weiterhin wurde der Einfluss von 24 auf Reperfusionsschäden infolge eines Herzinfarktes in Mausmodellen untersucht. Hierbei wurden Herzinfarkte induziert und durch die präventive Vorab Gabe der entstandene ischämische Reperfusionsschaden im Vergleich zu unbehandelten Mäusen analysiert. Basierend auf den bislang erzielten Ergebnissen lässt sich ein geringerer ischämischer Reperfusionsschaden bei vorheriger Verabreichung von 24 vermuten. Die Behandlung von Krebs wurde in den letzten Jahren durch den Ansatz der Präzisionsmedizin revolutioniert. Hierbei können durch die Identifizierung von prädiktiven Biomarkern große Patientenpopulationen in Subgruppen unterteilt werden, die somit eine individuelle und maßgeschneiderte Therapie erhalten. Die zielgerichtete Adressierung mit TKIs wurde erfolgreich für die Proteinkinasen EGFR, KIT und PDGFR beschrieben. Her2, ein Mitglied der ErbB Rezeptorfamilie, wurde bei etwa 4 % aller Patienten mit NSCLC als Treiber identifiziert, wovon ca. 80 % Insertionsmutationen in Exon20 entsprechen. Geringe Überlebens und Ansprechraten von < 40 % auf die bislang angewendeten Therapien mit TKIs verdeutlichen den dringenden Bedarf an wirksamen Inhibitoren für Krebspatienten mit positivem Her2 Mutationsstatus. In dieser Arbeit wurden kovalente, Pyrrolopyrimidin basierte Typ II Inhibitoren mit dem Ziel weiterentwickelt, die Löslichkeit der Inhibitoren durch Einführung von Löslichkeitsgruppen zu steigern und somit eine verbesserte zelluläre Aktivität erzielen zu können. Durch Einbringung von polaren Alkohol und Amin-funktionalitäten in ortho Position zum Michael Akzeptor wurde eine gesteigerte biochemische, aber gleichbleibende zelluläre Aktivität festgestellt. Kristallisationsstudien verifizierten den angenommenen Bindungsmodus und die pharmakokinetische Charakterisierung zeigte bspw. Optimierungsbedarf bzgl. der Stabilität in murinem Blutplasma sowie in Lebermikrosomen. Mutationen in den Proteinkinasen KIT und PDGFR wurden bei der Mehrheit von Patienten mit gastrointestinalen Stromatumoren identifiziert. Die derzeitig zugelassenen Wirkstoffe für die Behandlung von GIST sind reversible, ATP kompetitive Inhibitoren, von denen lediglich Avapritinib und Ripretinib speziell für die Therapie von GIST entwickelt wurden. Im Rahmen dieser Arbeit wurde basierend auf der Ponatinibstruktur eine Substanzbibliothek von Typ II bzw. Typ III Inhibitoren synthetisiert. Diese wurden mit einer reaktiven Gruppe dekoriert, um eine kovalente Bindung mit Cys788 oder Cys809 in KIT bzw. Cys814 oder Cys835 in PDGFR auszubilden. In MS und MS/MS Experimenten konnte die spezifische Bindung der synthetisierten Inhibitoren an Cys788 in KIT nicht nachgewiesen werden. Ferner wurden einzigartige, Patienten abgeleitete PDGFR mutierte Zelllinien entwickelt und in einem HTS eingesetzt. Die Durchmusterung von fokussierten Kinaseinhibitor-bibliotheken sollte zukünftig neue Leitstrukturen identifizieren, um spezifische Inhibitoren für PDGFR mutierte gastrointestinale Stromatumore designen und synthetisieren zu können

A comparative study of fragment screening methods on the p38α kinase: new methods, new insights

The stress-activated kinase p38α was used to evaluate a fragment-based drug discovery approach using the BioFocus fragment library. Compounds were screened by surface plasmon resonance (SPR) on a Biacore(™) T100 against p38α and two selectivity targets. A sub-set of our library was the focus of detailed follow-up analyses that included hit confirmation, affinity determination on 24 confirmed, selective hits and competition assays of these hits with respect to a known ATP binding site inhibitor. In addition, functional activity against p38α was assessed in a biochemical assay using a mobility shift platform (LC3000, Caliper LifeSciences). A selection of fragments was also evaluated using fluorescence lifetime (FLEXYTE(™)) and microscale thermophoresis (Nanotemper) technologies. A good correlation between the data for the different assays was found. Crystal structures were solved for four of the small molecules complexed to p38α. Interestingly, as determined both by X-ray analysis and SPR competition experiments, three of the complexes involved the fragment at the ATP binding site, while the fourth compound bound in a distal site that may offer potential as a novel drug target site. A first round of optimization around the remotely bound fragment has led to the identification of a series of triazole-containing compounds. This approach could form the basis for developing novel and active p38α inhibitors. More broadly, it illustrates the power of combining a range of biophysical and biochemical techniques to the discovery of fragments that facilitate the development of novel modulators of kinase and other drug targets. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10822-011-9454-9) contains supplementary material, which is available to authorized users

Why nanofibers are a good adsorptive surface – fundamental understanding and industrial applications for mAb bioprocessing

Over the years, chromatography has proven to be a powerful and versatile technique for the purification of high value biotherapeutics. Yet, today’s preparative chromatography of biologics still, in principle, looks the same as it did several decades ago. Any improvements made have been incremental; constrained by the stationary phase format (porous beads), associated column size (bed height and pressure drop), and historical modes of operation. To address future manufacturing challenges such as high cost of goods, diversity in product portfolios, market dynamics and manufacturing flexibility, new, more radical approaches to the development of chromatography materials and towards associated modes of operations are needed. With the biotechnology industry maturing, wide spread adoption of new high tech tools/products such as high throughput analytics, automated process control, single use materials and real time data analysis is already taking place, which in turn will lead towards revisiting and a subsequent improvement of how chromatography will be operated in the future. Examples of such improvements that are already considered include high productivity operations such as simulated moving bed and rapid, or extreme, cycling regimes. Please click Additional Files below to see the full abstract