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We demonstrate, for the first time, the multiplexed determination of microbial species from whole blood using the paper‐folding technique of origami to enable the sequential steps of DNA extraction, loop‐mediated isothermal amplification (LAMP), and array‐based fluorescence detection. A low‐cost handheld flashlight reveals the presence of the final DNA amplicon to the naked eye, providing a “sample‐to‐answer” diagnosis from a finger‐prick volume of human blood, within 45 min, with minimal user intervention. To demonstrate the method, we showed the identification of three species of Plasmodium, analyzing 80 patient samples benchmarked against the gold‐standard polymerase chain reaction (PCR) assay in an operator‐blinded study. We also show that the test retains its diagnostic accuracy when using stored or fixed reference samples.

This study is uniquely based on the Research Interest Score (RGRI) and not on other existing bibliometric criteria for evaluation of published biological inventory products (articles and monographs). RGRI is a ResearchGate.net score that measures scientists’ interest in the publication and is based on its citations, recommendations, and reads. Our data revealed that high RGRI scores of publications were generally not determined by the journal’s Impact Factor (IF) or high quartiles (Q). However, open access to publications undoubtedly creates the strongest preconditions for the rise of RGRI. The importance and popularity of a publications can also be affected by its various other characteristics, for example, international collaboration of authors, ecological issues such as plant-insect interactions, and even the wording of the publication title., Žurnalo citavimo rodiklis (IF) arba aukštos kvartilės (Q1, Q2) nebūtinai nulemia aukštesnes „Research Gate Research Interest“ (RGRI) rodiklio reikšmes, publikacijų populiarumą ir svarbą. Tačiau laisva publikacijų prieiga neabejotinai sudaro didžiausias prielaidas RGRI balo kilimui, t. y. publikacijų skaito-mumui ir cituojamumui. Be atviros prieigos, RGRI rodiklio dydžiui turi įtakos ir įvairios kitos straips-nių charakteristikos. Autorių duomenimis, didžiau-sią reikšmę gali turėti tokie veiksniai kaip tarp-tautinis bendradarbiavimas, ekologinių klausimų nagrinėjimas (pvz., augalų ir vabzdžių sąveika) bei pati straipsnio pavadinimo formuluotė. RGRI ro-diklio reikšmės kinta laike, tačiau nepaisant skirtin-gų starto pozicijų, jų didėjimo tempas nepriklauso nuo to, ar publikacija yra paskelbta aukšto citavimo (IF) žurnale ar nereitinguotame mokslo leidinyje; šiek tiek spartesnis publikacijų RGRI rodiklio didė-jimas pastebimas tarp tų publikacijų, kurios yra lais-vos prieigos. Šiais komercializuoto ir formalizuoto mokslo laikais profesionalūs tyrėjai netiesiogiai, ta-čiau sistemiškai yra skatinami nepublikuoti straips-nių nereitinguotuose žurnaluose arba žurnaluose, neturinčiuose aukštų kvartilių. Straipsnio autoriai mano, kad tai nė kiek neskatina svarbiausio tikslo siekimo – greitesnės biologinės įvairovės inventori-zacijos, netgi tikriausiai daro priešingą poveikį.v

Objective: This study evaluated the bibliometric data of the most common publication types of the neuroimaging literature. Methods: PubMed was searched to identify all published papers with “neuroimaging” as their MeSH Major Topics, and they were further searched by the following publication types: case report, clinical trial, comparative study, editorial, evaluation study, guideline, meta-analysis, multicenter study, randomized controlled trial, review, technical report, and validation study. The proportion of papers belonging to each publication type published in neuroimaging journals was calculated. Year-adjusted mean citation counts for each publication type were computed using data from Web of Science. Publication trend and its correlation with citation performance were assessed. Results: Review and comparative study were the most common publication types. Publication types with the highest proportion in neuroimaging journals were guideline, validation study, and technical reports. Since the year 2000, multicenter study, review, and meta-analysis showed the strongest linear increase in annual publication count. These publication types also had the highest year-adjusted citation counts (4.7–10.0). Publication types with the lowest year-adjusted citation counts were editorial and case report (0.5–1.0). It was estimated that 12.5% of the publications labeled as case reports were incorrectly labeled. Conclusions: Neuroimaging literature has been expanding with papers of higher levels of evidence, such as meta-analyses, multicenter studies, and randomized controlled trials.

Listeria monocytogenes is the third most deadly foodborne pathogen in the United States. The bacterium is found in soil and water, contaminating raw food products and the processing environment, where it can persist for an extended period. Currently, testing of food contact and non-food contact surfaces is performed using an array of sampling devices and endpoint technologies, offering various levels of sensitivity, cost, user skill, and time to detection. Paper-based microfluidic devices (µPADs) are a rapid detection platform amenable to low-cost, user-friendly, and portable diagnostics. In this study, we developed and evaluated a µPAD platform specific for the colorimetric detection of the Listeria genus following recovery from food contact and non-food contact surfaces. For detection, four colorimetric substrates specific for the detection of β-glucosidase, two broths selective for the detection of Listeria spp., and a nonselective broth were evaluated to facilitate detection of Listeria spp. The limit of detection and time to detection were determined by using pure bacterial cultures. After 8 h enrichment, L. monocytogenes (102 Colony Forming Units (CFU)/coupon) was detected on every surface. After 18 h enrichment, L. monocytogenes (102 CFU/coupon) was detected on all surfaces with all swabbing devices. This study demonstrated the ability of the µPAD-based method to detect potentially stressed cells at low levels of environmental contamination.

We introduce Synthetic Microfluidic Paper, a novel porous material for microfluidic applications that consists of an OSTE polymer that is photostructured in a well-controlled geometry of slanted and interlocked micropillars. We demonstrate the distinct benefits of Synthetic Microfluidic Paper over other porous microfluidic materials, such as nitrocellulose, traditional paper and straight micropillar arrays: in contrast to straight micropillar arrays, the geometry of Synthetic Microfluidic Paper was miniaturized without suffering capillary collapse during manufacturing and fluidic operation, resulting in a six-fold increased internal surface area and a three-fold increased porous fraction. Compared to commercial nitrocellulose materials for capillary assays, Synthetic Microfluidic Paper shows a wider range of capillary pumping speed and four times lower device-to-device variation. Compared to the surfaces of the other porous microfluidic materials that are modified by adsorption, Synthetic Microfluidic Paper contains free thiol groups and has been shown to be suitable for covalent surface chemistry, demonstrated here for increasing the material hydrophilicity. These results illustrate the potential of Synthetic Microfluidic Paper as a porous microfluidic material with improved performance characteristics, especially for bioassay applications such as diagnostic tests. QC 20160115

We have amplified an 84 bp fragment from the insertion sequence IS6110 of Mycobacterium tuberculosis (MTB) DNA on a paper substrate in 10 min. Tuberculosis (TB) is a highly infectious disease with a high mortality rate in the developing countries. Cheap and rapid screening assays are needed at the point of care for timely intervention and treatment. While PCR-based detection of TB is faster than bacterial culture and more specific compared to microscopy, PCR is not viable as a routine screening test in many low-income countries where TB is endemic due to the requirement of an expensive thermocycler. Disease screening based on isothermal amplification techniques is, therefore, being explored as an alternative to PCR. Successful isothermal amplification of DNA from HIV, H1N1, Chlamydia trachomatis, E. coli, etc. on cheap and disposable paper substrates has been reported in the literature. Isothermal amplification of MTB DNA on paper has been challenging due to the high GC content (65%) of MTB genome. Here we report helicase dependent amplification of a fragment of MTB DNA on a paper substrate in 10 min starting from 100 copies of the template using inexpensive heat sources, such as hot plates and hand warmers. The enzyme mix used to amplify DNA can be spotted and stored dry on paper at ambient temperatures for more than a month. The DNA amplified on paper can be detected by incorporating a suitable fluorescence marker in the reaction mixture or by directly loading the paper in a standard gel electrophoresis set up. Finally, as a surrogate to real clinical sputum samples, MTB DNA was successfully amplified on paper in the viscous environment of artificial sputum.

The prey range of the invasive Asian paper wasp, Polistes chinensis antennalis, was studied using molecular diagnostics. Nests of paper wasps were collected from urban residential and salt marsh habitats, larvae were removed and dissected, and DNA in the gut of the paper wasp larvae was amplified and sequenced with cytochrome c oxidase subunit I (COI). Seventy percent of samples (211/299) yielded medium-to high-quality sequences, and prey identification was achieved using BLAST searches in BOLD. A total of 42 taxa were identified from 211 samples. Lepidoptera were the majority of prey, with 39 taxa from 91% of samples. Diptera was a relatively small component of prey (three taxa, 19 samples). Conclusive species-level identification of prey was possible for 67% of samples, and genus-level identification, for another 12% of samples. The composition of prey taken was different between the two habitats, with 2.5× more native prey species being taken in salt marsh compared with urban habitats. The results greatly extend the prey range of this invasive species. The technique is a more effective and efficient approach than relying on the collection of “prey balls”, or morphological identification of prey, for the study of paper wasps.

Control of human African trypanosomiasis (HAT) in the Democratic Republic of Congo is based on mass population screening by mobile teams; a costly and labor-intensive approach. We hypothesized that blood samples collected on filter paper by village health workers and processed in a central laboratory might be a cost-effective alternative. We estimated sensitivity and specificity of micro-card agglutination test for trypanosomiasis (micro-CATT) and enzyme-linked immunosorbent assay (ELISA)/T.b. gambiense on filter paper samples compared with parasitology-based case classification and used the results in a Monte Carlo simulation of a lot quality assurance sampling (LQAS) approach. Micro-CATT and ELISA/T.b. gambiense showed acceptable sensitivity (92.7% [95% CI 87.4-98.0%] and 82.2% [95% CI 75.3-90.4%]) and very high specificity (99.4% [95% CI 99.0-99.9%] and 99.8% [95% CI 99.5-100%]), respectively. Conditional on high sample size per lot (> or = 60%), both tests could reliably distinguish a 2% from a zero prevalence at village level. Alternatively, these tests could be used to identify individual HAT suspects for subsequent confirmation.

Research in paper-based analytical devices has been increasing rapidly in recent years. Manyof these devices are used as low-cost alternatives for diagnostics and biosensing. In this work,two novel paper-based technologies were developed. The first paper-based technology achieved was measuring streaming potential on paper-based microfluidic devices. The streaming potential measurements were able to detect the presence of adsorbed polyvinylamine or potassium polyvinylsulfate in paper-based microfluidic channels. The measured streaming potential ranged from -80 mV to 80 mV and the polarity was sensitive to the adsorbed polymer. Furthermore, the measured streaming potential on paper treated with BSA showed a polarity switch when the pH was changed from below the pKa to above the pKa of BSA. Lastly, streaming potential measurements may provide an electronic interface for paperbased sensors. The second technology developed was a paper-based chromatographic pre-concentration device for biological and chemical applications. The device successfully concentrated a protein, streptavidin, via biotinylated microgels immobilized onto a selected area of the filter paper. The device was able to process a large volume of fluid with the incorporation of a passive pump made of superabsorbent polymer. The concentration factor achieved by the device was over 3000-fold. The flow dynamics through the paper was modeled using Darcy’s law. This technology could be an excellent low-cost alternative for biochemical analysis for samples thatrequire preconcentration, especially for the analysis of trace compounds in wastewater and drinking water. Master of Applied Science (MASc)

The first part of paper deals with the base information about diagnostics of power transformers. In this part are presented differently insulating methods, for example method of recovery voltage method, method of polarization and depolarization currents and chromatographic analysis.The second part of paper deals use of method of frequency domain spectroscopy for oil power transformers. This method is used in analysis insulating condition of power transformer with system of oil-paper. It was found, that the results of these tests are highly impacted by the operating temperature during the experimental measurement. Moisture and conductivity between insulating paper and oil in an insulating system are highly dependent from temperature.In the other part, the paper presents experimental results of the frequency diagnostic measurement for a real single-phase traction transformer 110/27 kV at different operating temperatures and states (with oil and without).Finally in the last part, the paper presents comparing frequency insulating measurements among several the same single-phase transformers 110/27 kV.

Ethnic minority females are less likely to receive a diagnosis or treatment for an eating disorder (ED). This study captured the experiences of therapists who have worked with ethnic minority females to improve outcomes for this group. Twelve therapists in the United Kingdom, London were recruited for semi-structured interviews and thematic analysis was used to analyse the data. Shame was cited as a barrier to accessing help. This influenced therapeutic work such as not challenging shame or linking this to a negative interpretation of parents. Emotional and interpersonal factors were thought to be more common risk factors for the ED. The minimising of weight and shape concern and non-fat-phobic anorexia was thought to lead to a complex and delayed route to accessing ED services. Therapists felt restricted by service management who they felt required them to deliver a narrow range of therapies that had not necessarily demonstrated therapeutic outcomes in ethnic minority females. Nevertheless, therapists reported using curiosity to guide their cultural adaptations when feeling uncertain. When working with ethnic minority females, therapists face challenges from the therapeutic and diagnostic framework that services are aligned to. Creative solutions to address this include adapting the patient care pathway, referral guides, cultural reflective practice, and the use of cultural genograms and scripts in therapeutic work to address unmet needs.

This study reports the development of a sensitive magnetic bead-based enzyme-linked immunoassay (MELISA) for the pan-reactive detection of the Influenza A virus. The assay combines immunomagnetic beads and biotin-nanoparticle-based detection to quantify a highly conserved viral nucleoprotein in virus lysates. At the capture step, monoclonal antibody-coated magnetic microbeads were used to bind and concentrate the nucleoprotein in samples. The colorimetric detection signal was amplified using biotinylated silica nanoparticles (NP). These nanoparticles were functionalized on the surface with short DNA spacers bearing biotin groups by an automated supported synthesis method performed on nano-on-micro assemblies with a DNA/RNA synthesizer. A biotin-nanoparticle and immunomagnetic bead-based assay was developed. We succeeded in detecting Influenza A viruses directly in the lysis buffer supplemented with 10% saliva to simulate the clinical context. The biotin-nanoparticle amplification step enabled detection limits as low as 3 × 103 PFU mL−1 and 4 × 104 PFU mL−1 to be achieved for the H1N1 and H3N2 strains respectively. In contrast, a classical ELISA test based on the same antibody sandwich showed detection limit of 1.2 × 107 PFU mL−1 for H1N1. The new enhanced MELISA proved to be specific, as no cross-reactivity was found with a porcine respiratory virus (PRRSV). Graphical abstract Supplementary Information The online version contains supplementary material available at 10.1007/s00216-020-03081-x.

Purpose In many cases, the diagnosis of a periprosthetic joint infection (PJI) consisting of the clinical appearance, laboratory tests, and other diagnostic tools remains a difficult task. Single serum biomarkers are easy to collect, are suitable for periodical assessment, and are a crucial tool in PJI diagnosis, but limited sensitivity or specificity is reported in literature. The aim of this study was to combine the best-performing single serum biomarkers into a multi-biomarker model aiming to improve the diagnostic properties. Methods Within a 27-month period, 124 surgical procedures (aseptic or septic revision total knee arthroplasty (TKA) or total hip arthroplasty (THA)) were prospectively included. The serum leukocyte count, C-reactive protein (CRP), interleukin-6, procalcitonin, interferon alpha, and fibrinogen were assessed 1 day prior to surgery. Logistic regression with lasso-regularization was used for the biomarkers and all their ratios. After randomly splitting the data into a training (75%) and a test set (25%), the multi-biomarker model was calculated and validated in a cross-validation approach. Results CRP (AUC 0.91, specificity 0.67, sensitivity 0.90, p value 0.03) and fibrinogen (AUC 0.93, specificity 0.73, sensitivity 0.94, p value 0.02) had the best single-biomarker performances. The multi-biomarker model including fibrinogen, CRP, the ratio of fibrinogen to CRP, and the ratio of serum thrombocytes to CRP showed a similar performance (AUC 0.95, specificity 0.91, sensitivity 0.72, p value 0.01). Conclusion In this study, multiple biomarkers were tested for their diagnostic performance, with CRP and fibrinogen showing the best results regarding the AUC, accuracy, sensitivity, and specificity. It was not possible to further increase the diagnostic accuracy by combining multiple biomarkers using sophisticated statistical methods.

Background Fungal infections of the skin, hair, and nails are the largest and most widespread group of all mycoses. Nannizzia nana is a relatively rare etiological factor of dermatomycosis in humans, as it usually affects animals, e.g. pigs and boars. In addition to the zoophilic nature, there are also reports of the geophilic reservoir of this dermatophyte species. Objective In this study, we present symptomatic infections with N. nana aetiology in humans reported recently in Poland. Interestingly, these cases had a non-specific clinical picture and occurred as skin lesions on the neck and foot as well as onychomycosis of the toenails. From the medical history, the patients had no contact with pigs. Methods Diagnostics of these infections was performed with a combination of classical phenotypic and molecular genomic methods. The genomic diversity of the isolates was determined using the MP-PCR method. In vitro antifungal susceptibility tests against itraconazole, ketoconazole, terbinafine and naftifine hydrochloride were also performed. Results Nannizzia nana has been identified as an etiological factor of dermatomycosis. Moreover, heterogeneity of the genomes was revealed for the obtained strains. In vitro activities of antifungal agents showed that isolates were susceptible to all tested drugs. The patients were treated with oral terbinafine and topical ketoconazole cream, which led to a complete recovery. Conclusions In conclusion, the cases studied by us may indicate that the infrequency of N. nana infections may not necessarily be related to the low infectivity of this fungal agent, but they are rather associated with misdiagnosis. Furthermore, N. nana reservoirs should also be sought in soil.

Frontal fibrosing alopecia (FFA) is recognized to represent a generalized process of inflammatory scarring alopecia. Apart from the classic form affecting the frontal hairline, there are a range of disease manifestations involving loss of eyebrows and of eyelashes, loss of peripheral body hair, fibrosing alopecia in a pattern distribution, facial and extrafacial skin, mucous membrane, and nail involvement. Classic linear, diffuse “zigzag”, pseudo “fringe sign”, androgenetic alopecia-like, cockade-like, ophiasis-like and incomplete patterns are distinguished. The aetiology of FFA remains obscure, but a number of pathogenetic hypotheses and treatments to halt disease progression have been proposed.

We introduce a novel diagnostic approach for the uncovering of hidden content in layered documents, based on a photoacoustic imaging setup with optimized acoustic detection parameters. By exploiting the laser-induced ultrasound following the intense absorption of pulsed optical radiation by printed ink, this traditional biomedical imaging method is proven capable of revealing text characters buried under several layers of stacked paper sheets at high spatial resolution and excellent contrast levels. Such a remarkable performance benefits from the effective combination of strong light scattering by paper fibrils and the virtually unobstructed propagation of the generated ultrasonic waves through multiple layers, enabling thus the accurate recording of text, even in double-sided prints, with minimum shadowing artifacts. The imaging effectiveness, the implementation simplicity, the robustness and the relatively low cost features offered by the proposed photoacoustic modality, are anticipated to contribute in its wide adoption by the preservation of cultural heritage community, as a powerful diagnostic tool for the in-depth investigation of complex multi-layered objects.

Echinococcus granulosus sensu lato is a globally prevalent zoonotic parasitic cestode leading to cystic echinococcosis (CE) in both humans and sheep with both medical and financial impacts, whose reduction requires the application of a One Health approach to its control. Regarding the animal health component of this approach, lack of accurate and practical diagnostics in livestock impedes the assessment of disease burden and the implementation and evaluation of control strategies. We use of a Bayesian Latent Class Analysis (LCA) model to estimate ovine CE prevalence in sheep samples from the Río Negro province of Argentina accounting for uncertainty in the diagnostics. We use model outputs to evaluate the performance of a novel recombinant B8/2 antigen B subunit (rEgAgB8/2) indirect enzyme-linked immunosorbent assay (ELISA) for detecting E. granulosus in sheep. Necropsy (as a partial gold standard), western blot (WB) and ELISA diagnostic data were collected from 79 sheep within two Río Negro slaughterhouses, and used to estimate individual infection status (assigned as a latent variable within the model). Using the model outputs, the performance of the novel ELISA at both individual and flock levels was evaluated, respectively, using a receiver operating characteristic (ROC) curve, and simulating a range of sample sizes and prevalence levels within hypothetical flocks. The estimated (mean) prevalence of ovine CE was 27.5% (95%Bayesian credible interval (95%BCI): 13.8%–58.9%) within the sample population. At the individual level, the ELISA had a mean sensitivity and specificity of 55% (95%BCI: 46%–68%) and 68% (95%BCI: 63%–92%), respectively, at an optimal optical density (OD) threshold of 0.378. At the flock level, the ELISA had an 80% probability of correctly classifying infection at an optimal cut-off threshold of 0.496. These results suggest that the novel ELISA could play a useful role as a flock-level diagnostic for CE surveillance in the region, supplementing surveillance activities in the human population and thus strengthening a One Health approach. Importantly, selection of ELISA cut-off threshold values must be tailored according to the epidemiological situation., Highlights • Development of a Bayesian LCA model for ovine cystic echinococcosis (CE) in Argentina. • Evaluation of diagnostic performance for a novel rEgAgB8/2 indirect ELISA. • Identification of optimal ELISA thresholds for use as a flock-level surveillance tool. • Improvement of CE surveillance in sheep will help strengthen the One Health approach.

Significance: Optical scattering signals obtained from tissue constituents contain a wealth of structural information. Conventional intensity features, however, are mostly dictated by the overall morphology and mean refractive index of these constituents, making it very difficult to exclusively sense internal refractive index fluctuations. Aim: We perform a systematic analysis to elucidate how changes in internal refractive index profile of cell nuclei can best be detected via optical scattering. Approach: We construct stochastically inhomogeneous nuclear models and numerically simulate their azimuth-resolved scattering patterns. We then process these two-dimensional patterns with the goal of identifying features that directly point to subnuclear structure. Results: Azimuth-dependent intensity variations over the side scattering range provide significant insights into subnuclear refractive index profile. A particular feature we refer to as contrast ratio is observed to be highly sensitive to the length scale and extent of refractive index fluctuations; further, this feature is not susceptible to changes in the overall size and mean refractive index of nuclei, thereby allowing for selective tracking of subnuclear structure that can be linked to chromatin distribution. Conclusions: Our analysis will potentially pave the way for scattering-based assessment of chromatin reorganization that is considered to be a key hallmark of precancer progression.

Purpose The aim of this prospective study was to identify the tumour characteristics that are associated with invasiveness and those that are relevant for disease-specific survival (DSS) in upper tract urothelial carcinoma, UTUC. Methods From a prospective consecutive cohort of patients with suspicion of UTUC, those who were diagnosed with UTUC using URS prior to rNU between 2005 and 2012 were included. Tumour characteristics were analysed for prediction of invasiveness and association with DSS. Stages were categorised as superficial (pTa-1 and CIS only) or invasive (≥ pT2). Tumours were graded according to WHO 1999 classification. DSS was analysed regarding possible association with stage, grade, size, multifocality, location, ploidy and rate of proliferation. Associations were tested using Fisher’s exact test, Pearson Chi-square or Cox’s regression. Kaplan–Meier survival curves were constructed. Results Forty-five consecutive patients were included, and 43 of them were included in the final analyses because their rNU specimens were available for reassessment. The only tumour characteristics that were significantly associated with stage were tumour grade (P

Accurate methods for determining the duration of HIV infection at the individual level are valuable in many settings, including many critical research studies and in clinical practice (especially for acute infection). Since first published in 2003, the ‘Fiebig staging system’ has been used as the primary way of classifying early HIV infection into five sequential stages based on HIV test result patterns in newly diagnosed individuals. However, Fiebig stages can only be assigned to individuals who produce both a negative and a positive test result on the same day, on specific pairs of tests of varying ‘sensitivity’. Further, in the past 16 years HIV-testing technology has evolved substantially, and three of the five key assays used to define Fiebig stages are no longer widely used. To address these limitations, we developed an improved and more general framework for estimating the duration of HIV infection by interpreting any combination of diagnostic test results, whether obtained on single or multiple days, into an estimated date of detectable infection, or EDDI. A key advantage of the EDDI method over Fiebig staging is that it allows for the generation of a point estimate, as well as an associated credibility interval for the date of first detectable infection, for any person who has at least one positive and one negative HIV test of any kind. The tests do not have to be run on the same day; they do not have to be run during the acute phase of infection and the method does not rely on any special pairing of tests to define ‘stages’ of infection. The size of the interval surrounding the EDDI (and therefore the precision of the estimate itself) depends largely on the length of time between negative and positive tests. The EDDI approach is also flexible, seamlessly incorporating any assay for which there is a reasonable diagnostic delay estimate. An open-source, free online tool includes a user-updatable curated database of published diagnostic delays. HIV diagnostics have evolved tremendously since that original publication more than 15 years ago, and it is time to similarly evolve the methods used to estimate timing of infection. The EDDI method is a flexible and rigorous way to estimate the timing of HIV infection in a continuously evolving diagnostic landscape.

A proof-of-concept aptamer-based optical assay is described for the determination of the immuno signalling molecule interleukin-6 (IL-6), a key marker of acute inflammation. The optical assay is based on the aggregation of gold nanoparticles (AuNP) coated in two complimentary “sandwich-style” aptamers, each with different IL-6 target moieties. IL-6 will recognise the complimentary aptamer pair and bind to it, thereby causing the aggregation of the corresponding functionalised nanoparticles. The aggregation of the AuNPs after exposure to IL-6 induces a visible colour change from red to pink, with a corresponding change in the absorption maximum from 520 to 540 nm. The change in the absorption maximum can be monitored visually, or by using a spectrophotometer or a plate reader. The optimal size and functionalisation of aptamer-coated AuNPs, and the potential assay formats were investigated using UV-vis spectrophotometry, transmission electron microscopy, and dynamic light scattering. The optical assay was applied for detecting mouse IL-6 in a mixed protein solution as a representative biological sample. The assay works in the 3.3 to 125 μg·mL−1 IL-6 concentration range, and the detection limit (at S/N = 3) is 1.95 μg·mL−1. This study was performed as a proof-of-concept demonstration of this versatile assay design, with a view to developing a similar assay for use in clinical samples in future. Graphical abstractSchematic representation of the aggregation of aptamer-functionalised nanoparticles in the presence of interleukin-6 (IL-6). The presence of mouse IL-6 in a mixed protein solution leads to a visible colour change, and a change in the absorption spectrum of the nanoparticles. Electronic supplementary material The online version of this article (10.1007/s00604-019-3975-7) contains supplementary material, which is available to authorized users.

Purpose: It is challenging to deliver the full-length dysferlin gene or protein to restore cellular functions of dysferlin-deficient (DYSF-/-) myofibres in dysferlinopathy, a disease caused by the absence of dysferlin, which is currently without effective treatment. Exosomes, efficient membranous nanoscale carriers of biological cargoes, could be useful. Experimental design: Myotube- and human serum-derived exosomes were investigated for their capabilities of restoring dysferlin protein and cellular functions in murine and human DYSF-/- cells. Moreover, dysferlinopathic patient serum- and urine-derived exosomes were assessed for their abilities as diagnostic tools for dysferlinopathy. Results: Here we show that exosomes from dysferlin-expressing myotubes carry abundant dysferlin and enable transfer of full-length dysferlin protein to DYSF-/- myotubes. Exogenous dysferlin correctly localizes on DYSF-/- myotube membranes, enabling membrane resealing in response to injury. Human serum exosomes also carry dysferlin protein and improve membrane repair capabilities of human DYSF-/- myotubes irrespective of mutations. Lack of dysferlin in dysferlinopathic patient serum and urine exosomes enables differentiation between healthy controls and dysferlinopathic patients. Conclusions: Our findings provide evidence that exosomes are efficient carriers of dysferlin and can be employed for the treatment and non-invasive diagnosis of dysferlinopathy.

The worldwide emergence of multidrug‐resistant (MDR) bacteria is associated with significant morbidity, mortality, and healthcare costs. Rapid and accurate diagnostic methods to detect antibiotic resistance are critical for antibiotic stewardship and infection control measurements. Here a cantilever nanosensor‐based diagnostic assay is shown to detect single nucleotide polymorphisms (SNPs) and genes associated with antibiotic resistance in Gram negative (Pseudomonas aeruginosa) and positive (Enterococcus faecium) bacteria, representing frequent causes for MDR infections. Highly specific RNA capture probes for SNPs (ampRD135G or ampRG154R) or resistance genes (vanA, vanB, and vanD) allow to detect the binding of bacterial RNA within less than 5 min. Serial dilutions of bacterial RNA indicate an unprecedented sensitivity of 10 fg µL−1 total RNA corresponding to less than ten bacterial cells for SNPs and 1 fg µL−1 total RNA for vanD detection equivalent to single bacterial cell sensitivity., A new nanosensor array method is presented to investigate antibiotic resistances in bacteria. Single nucleotide polymorphisms in ampicillin resistance markers from gram negative Pseudomonas aeruginosa and vancomycin resistance genes in gram positive Enterococcus faecium are studied. The method shows unprecedented sensitivity and specificity (1 bacterial cell) as well as drastically reduced times to obtain a diagnosis, compared to standard procedures.

Background Telestroke has developed rapidly as an assessment tool for patients eligible for reperfusion therapy. Objective To investigate whether vascular neurologists can diagnose intracranial large vessel occlusion (LVO) as quickly and accurately using a smartphone application compared to a hospital-based desktop PC monitor. Methods We retrospectively enrolled 108 consecutive patients with acute ischemic stroke in the middle cerebral artery territory who underwent magnetic resonance imaging (MRI) within 24 hours of their stroke onset. Two vascular neurologists, blinded to all clinical information, independently evaluated magnetic resonance angiography and fluid-attenuated inversion recovery images for the presence or absence of LVO in the internal carotid artery and middle cerebral artery (M1, M2, or M3) on both a smartphone application (Smartphone-LVO) and a hospital-based desktop PC monitor (PC-LVO). To evaluate the accuracy of an arterial occlusion diagnosis, interdevice variability between Smartphone-LVO and PC-LVO was analyzed using κ statistics, and image interpretation time was compared between Smartphone-LVO and PC-LVO. Results There was broad agreement between Smartphone-LVO and PC-LVO evaluations regarding the presence or absence of arterial occlusion (Reader 1: κ=0.94; P Conclusions The results indicate the evaluation of neuroimages using a smartphone application can provide an accurate and timely diagnosis of anterior intracranial arterial occlusion that can be shared immediately with members of the stroke team to support the management of patients with hyperacute ischemic stroke.

Endometrial cancer is one of the most common cancers experienced by women throughout the world. It is also the most common malignancy within the female reproductive system, representing 37.7% of all disorders. The incidence increases with age, and is diagnosed most frequently in women between 45 and 65 years old. In the last few years, numerous studies have been performed to identify tumour biomarkers. Biomarkers include not only protein routinely used as tumour markers but also genes and chromosomes. The limiting factor in the use of markers in the diagnosis of endometrial cancer is their lack of specificity. However, specific markers for endometrial cancer are the subject of much research attention. Although moderately elevated levels of markers are present in a number of inflammatory or non-malignant diseases, significantly increased levels of markers indicate the development of cancer. Recently, research has been focused on the identification of molecular changes leading to different histological subtypes of endometrial cancer. In this paper the authors reviewed several currently investigated markers. Progress in these investigations is very important in the diagnostics and treatment of endometrial cancer. In particular, the identification of novel mutations and molecular profiles should enhance our ability to personalise adjuvant treatment with genome-guided targeted therapy.

// Zenglin Pei 1 , Song-Mei Liu 2 , Jing-Tao Huang 2 , Xuan Zhang 1 , Dong Yan 3 , Qianlin Xia 1 , Chunxia Ji 1 , Weiping Chen 4 , Xiaoyan Zhang 1 , Jianqing Xu 1 , Jin Wang 1 1 Scientific Research Center, Shanghai Public Health Clinical Center, Fudan University, Jinshan District, Shanghai 201508, P.R. China 2 Center for Gene Diagnosis, Zhongnan Hospital of Wuhan University, Wuhan, Hubei 430071, P.R. China 3 Department of Medical Oncology, Beijing Chaoyang Hospital affiliated to Capital Medical University, Beijing, China 4 Genomics Core, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA Correspondence to: Jin Wang, email: wangjin@shaphc.org Keywords: pancreatic cancer, meta-analysis, diagnostics, multiple miRNA, SROC Received: October 24, 2016 Accepted: January 25, 2017 Published: February 07, 2017 ABSTRACT Background: Pancreatic cancer (PaCa) is the most lethal gastrointestinal (GI) tumor. Although many studies on differentially expressed miRNAs as candidate biomarkers of pancreatic cancer have been published, reliability of these findings generated from investigations performed in single laboratory settings remain unclear. Results: There were 29 articles with a total of 2,225 patients and 1,618 controls included in this meta-analysis. The pooled sensitivity was 82% (95% CI, 79–85%); the specificity was 85% (95% CI, 79–89%); and area under the curve (AUC) was 0.89 (95% CI, 0.86–0.92). Subgroup analyses indicated that there were significant divergences between Caucasian and Asian subgroups for circulating miRNA analysis. Materials And Methods: To comprehensively investigate the potential utility of miRNAs as biomarkers of the disease, we searched publications diagnosing PaCa using miRNAs from PubMed, Medline, Embase, Google Scholar and Chinese National Knowledge Infrastructure (CNKI) databases. The sensitivity (SEN), specificity (SPE), and summary receiver operating characteristic (SROC) curve were used to examine the overall test performance, and heterogeneity was analyzed with the I 2 test. Conclusions: Our analysis demonstrated that multiple miRNAs (SEN: 85%; SPE: 89%; AUC: 0.93) were more accurate for diagnosing PaCa than a single miRNA (SEN: 78%; SPE: 79%; AUC: 0.84), and future studies are still needed to confirm the diagnostic value of these pooled miRNAs for PaCa.

INTRODUCTION: This study aimed to establish whether an alcoholic antiseptic, wiped or not before venipuncture, may jeopardize alcohol testing with a commercial enzymatic assay and a reference head-space gas chromatography (GC) technique. MATERIALS AND METHODS: Venous blood was collected from 23 healthy volunteers, with two sequential procedures. In the first blood collection, 2 mL of alcoholic antiseptic (0.5% chlorhexidine, 70% ethanol) were place on a gauge pad, the venipuncture site of right arm was cleaned but the antiseptic was not let to dry before phlebotomy. In the second blood collection, 2 mL of the same alcoholic antiseptic were placed on another gauge pad, the venipuncture site of left harm was cleaned and the antiseptic was accurately cleansed before phlebotomy. Ethanol was measured with a reference GC technique in whole blood and EDTA plasma, and a commercial enzymatic assay in EDTA plasma. RESULTS: No subject complained about feeling a particular itchy sensation when the alcohol was not wiped before puncturing the vein. The concentration of alcohol in all EDTA plasma samples was always lower than the limit of detection of the enzymatic assay (i.e., 2.2 mmol/L ; 0.1 g/L). Similarly, alcohol concentration was also undetectable using a reference GC technique (i.e., < 0.22 mmol/L ; 0.01 g/L) in EDTA plasma and whole blood. CONCLUSION: It seems reasonable to conclude that using ethanol-containing antiseptics before venipuncture may not be causes of spurious or false positive results of alcohol measurement at least when ideal venipunctures can be performed.

Contact lens is a ubiquitous technology used for vision correction and cosmetics. Sensing in contact lenses has emerged as a potential platform for minimally invasive point‐of‐care diagnostics. Here, a microlithography method is developed to fabricate microconcavities and microchannels in a hydrogel‐based contact lens via a combination of laser patterning and embedded templating. Optical microlithography parameters influencing the formation of microconcavities including ablation power (4.3 W) and beam speed (50 mm s−1) are optimized to control the microconcavity depth (100 µm) and diameter (1.5 mm). The fiber templating method allows the production of microchannels having a diameter range of 100–150 µm. Leak‐proof microchannel and microconcavity connections in contact lenses are validated through flow testing of artificial tear containing fluorescent microbeads (Ø = 1–2 µm). The microconcavities of contact lenses are functionalized with multiplexed fluorophores (2 µL) to demonstrate optical excitation and emission capability within the visible spectrum. The fabricated microfluidic contact lenses may have applications in ophthalmic monitoring of metabolic disorders at point‐of‐care settings and controlled drug release for therapeutics.

Circulating anodic antigen (CAA) testing is a powerful, increasingly-used tool for diagnosis of active schistosome infection. We sought to determine the feasibility and reliability of measuring CAA in blood spots collected on Whatman 903 protein saver cards, which are the predominant filter papers used worldwide for dried blood spot (DBS) research and clinical care. CAA was eluted from blood spots collected from 19 individuals onto Whatman 903 cards in Mwanza, Tanzania, and the assay was optimized to achieve CAA ratios comparable to those obtained from the spots' corresponding serum samples. The optimized assay was then used to determine the correlation of serum samples (n=16) with DBS from cards that had been stored for 8 years at ambient temperature. Using a DBS volume equivalent to approximately four times the quantity of serum, CAA testing in DBS had a sensitivity of 76% and a specificity of 79% compared to CAA testing in serum. CAA testing was reliable in samples eluted from Whatman 903 cards that had been stored for 8 years at ambient temperature. The overall kappa coefficient was 0.53 (standard error 0.17, p

Background Diagnostic services are an essential component of high-quality surgical, anesthesia and obstetric (SAO) care. Efforts to scale up SAO care in Latin America have often overlooked diagnostics capacity. This study aims to analyze the capacity of diagnostic services, including radiology, pathology, and laboratory medicine, in hospitals providing SAO care in the states of Chiapas, Mexico and Amazonas, Brazil. Methods A stratified cross-sectional evaluation of diagnostic capacity in hospitals performing surgery in Chiapas and Amazonas was performed using the Surgical Assessment Tool (SAT). National data sources were queried for indicators of diagnostics capacity in terms of workforce, infrastructure and diagnosis utilization. Fisher's exact tests and chi-square tests were used to compare categorical variables between the private and public sector in Chiapas while descriptive statistics are used to compare Amazonas and Chiapas. Findings In Chiapas, 53% (n = 17) of public and 34% (n = 20) of private hospitals providing SAO care were assessed. More private hospitals than public hospitals could always provide x-rays (35% vs 23.5%) and ultrasound (85% vs 47.1%). However neither sector could consistently perform basic laboratory testing such as complete blood counts (70.6% public, 65% private). In Amazonas, 30% (n = 18) of rural hospitals were surveyed. Most had functioning x-ray machine (77.8%) and ultrasound (55.6%). The majority of hospitals could provide complete blood count (66.7%) but only one hospital (5.6%) could always perform an infectious panel. Both Chiapas and Amazonas had dramatically fewer diagnostic practitioners per capita in each state compared to the national average capacity. Interpretation Facilities providing SAO care in low-resource states in Mexico and Brazil often lack functioning diagnostics services and workforce. Scale-up of diagnostic services is essential to improve SAO care and should occur with emphasis on equitable and adequate resource allocation.

Point-of-care devices that are inexpensive, disposable, and environmentally friendly are becoming increasingly predominant in the field of biosensing and biodiagnostics. Here, microfluidics is a suitable option to endow portability and minimal reagent and material consumption. Nanocellulose is introduced to manufacture microfluidic channels and as a storage and immobilization compartment of glucose oxidase. Improved enzymatic activity retention is demonstrated in a simple and disposable point-of-care diagnostic unit that is able to detect glucose from fluid matrices at 0.1 mM concentration and in less than 10 min. It is concluded that the patterning and fluidic technologies that are possible with nanocellulose enable easily scalable multianalyte designs. publishedVersion

Introduction: Hartnup disorder is caused by a deficiency of the sodium dependent B0 AT1 neutral amino acid transporter in the proximal kidney tubules and jejunum. Biochemically, Hartnup disorder is diagnosed via amino acid excretion patterns. However, these patterns can closely resemble amino acid excretion patterns of generalized aminoaciduria, which may induce a risk for misdiagnosis and preclusion from treatment. Here we explore whether calculating a diagnostic ratio could facilitate correct discrimination of Hartnup disorder from other aminoacidurias. Methods: 27 amino acid excretion patterns from 11 patients with genetically confirmed Hartnup disorder were compared to 68 samples of 16 patients with other aminoacidurias. Amino acid fold changes were calculated by dividing the quantified excretion values over the upper limit of the age-adjusted reference value. Results: Increased excretion of amino acids is not restricted to amino acids classically related to Hartnup disorder (“Hartnup amino acids”, HAA), but also includes many other amino acids, not classically related to Hartnup disorder (“other amino acids”, OAA). The fold change ratio of HAA over OAA was 6.1 (range: 2.4–9.6) in the Hartnup cohort, versus 0.2 (range: 0.0–1.6) in the aminoaciduria cohort (p

A fluorescence intensity monitor has been developed for the non-invasive, pulse-by-pulse normalization of LCLS-II experiments. The linearity, noise and position sensitivity of the detectors have been characterized., For LCLS-II, a fluorescence intensity monitor for the non-invasive, pulse-by-pulse normalization of experiments has been developed. A prototype diagnostic was constructed with a microchannel plate assembly and two photodiodes. The diagnostic was then installed in the LCLS SXR instrument Kirkpatrick–Baez mirror chamber with the detectors located above the vertically reflecting mirror. The linearity, noise and position sensitivity of the detectors have been characterized. The photodiode responsivity is suitable for high pulse energies. The microchannel plate detector shows sufficient responsivity over a wide range of pulse energies. The relative signal from the two photodiodes provides a sensitive measure of the X-ray beam position. The fluorescence intensity monitor provides intensity normalization while being compatible with high incident power, a 0.93 MHz repetition rate and ultra-high vacuum.

Options for diagnosing Fasciola hepatica infection in groups of cattle are limited. Increasing the opportunities for herd-level diagnosis is important for disease monitoring, making informed treatment decisions and for flukicide efficacy testing. The sensitivity of a simple sedimentation method based on composite faecal samples for the detection of fluke eggs in cattle was assessed through a combination of experimental and statistical modelling techniques. Initially, a composite sample method previously developed for sheep was used to investigate the sensitivity of composite sample testing compared with individual counts on the same samples in cattle. Following this, an optimised, validated, qualitative (presence-absence) composite sample field test was developed for cattle. Results showed that fluke egg counts obtained from a composite sample are representative of those expected from individual counts. The optimal sampling strategy was determined to be 10 individual 10 g samples (100 g composite sample) from which a 10 g subsample is taken for sedimentation. This method yielded a diagnostic sensitivity of 0.69 (95 per cent CI 0.5 to 0.85). These results demonstrate the validity and usefulness of a composite faecal egg sedimentation method for use in the diagnosis and control of F. hepatica in groups of cattle, with the caveat that a negative test should be followed up with a second test due to limitations relating to test sensitivity.

Introduction Renal cell carcinoma (RCC) accounts for 3% of adult malignancies and more than 90% of kidney neoplasms. High rates of undiagnostic percutaneous kidney biopsies and difficulties in reliable pre-operative differentiation between malignant and benign renal tumors using contemporary imaging techniques result in large numbers of redundant surgeries. Absence of specific biomarkers for early detection and monitoring complicates on-time diagnosis of the disease and relapse. For the patients followed up after having a nephrectomy, a noninvasive and sensitive biomarker enabling early detection of disease relapse would be extremely useful. Material and methods The study is a review of recent knowledge regarding potential clinical applications of microRNAs (miRNAs) as biomarkers of RCC. Results MicroRNAs are essential regulators of various processes such as cell proliferation, differentiation, development and death; they have been implicated in diverse biological and pathological processes in RCC. There is a class of miRNAs that promote RCC development (oncomirs) and a class of miRNAs that negatively regulate oncogenes, suppress tumor growth and invasion, and thus could be considered treatment agents (anti-oncomirs). Separate miRNAs and specific miRNAs expression profiles have been identified, enabling early detection of the disease, prediction of response to systemic therapy, or prognostication of biological behavior of the disease. Conclusions The miRNA network analysis and gene profiling may help to identify the most sensible molecular signatures of RCC that can be used for diagnostic purposes, as well as poor prognosis signatures and poor therapeutic response signatures in patients who undergo systemic therapy.

Both acute and chronic inflammation of the pancreas often lead to complications that nowadays can be resolved using endoscopic and surgical procedures. In many cases, intraoperative ultrasound examination (IOUS) enables correct assessment of the extent of the lesion, and allows for safe surgery, while also shortening its length.At the authors' clinic, intraoperative ultrasound is performed in daily practice. In this paper, we try to share our experiences in the application of this particular imaging technique.Intraoperative examination conducted by a surgeon who has assessed the patient prior to surgery, which enabled the surgeon to verify the initial diagnosis. The material presented in this paper includes 145 IOUS procedures performed during laparotomy due to lesions of the pancreas, 57 of which were carried out in cases of inflammatory process.IOUS is a reliable examination tool in the evaluation of acute inflammatory lesions in the pancreas, especially during the surgery of chronic, symptomatic inflammation of the organ. The procedure allows for a correct determination of the necessary scope of the planned surgery. The examination allows for the differentiation between cystic lesions and tumors of cystic nature, dictates the correct strategy for draining, as well as validates the indications for the lesion's surgical removal. IOUS also allows the estimation of place and scope of drainage procedures in cases of overpressure in the pancreatic ducts caused by calcification of the parenchyma or choledocholitiasis in chronic pancreatitis. In pancreatic cancer, IOUS provides a verification of the local extent of tumor-like lesions, allowing for the assessment of pancreatic and lymph nodes metastasis, and indicating the presence of distant and local metastases, including the liver. IOUS significantly improves the effectiveness of intraoperative BAC aspiration or drainage of fluid reservoirs.Zarówno ostre, jak i przewlekłe stany zapalne trzustki prowadzą często do powstania powikłań, które współcześnie rozwiązuje się na drodze zabiegów endoskopowych oraz operacyjnych. Śródoperacyjna ultrasonografia (IOUS) w wielu przypadkach umożliwia prawidłowe określenie rozległości zmian chorobowych, pozwala na bezpieczne przeprowadzenie zabiegu operacyjnego oraz skrócenie czasu jego trwania.W klinice, w której pracują autorzy, śródoperacyjne badanie ultrasonograficzne jest wykonywane w codziennej praktyce. W niniejszej pracy pragniemy podzielić się własnymi doświadczeniami w zastosowaniu tej techniki obrazowania.Badania śródoperacyjne przeprowadzane były przez chirurga, który dokonywał oceny przed zabiegiem, co pozwoliło na weryfikację rozpoznań. Przytoczony materiał dotyczy 145 procedur IOUS wykonywanych podczas laparotomii z powodu zmian chorobowych trzustki, w tym 57 zastosowanych w przypadku procesu zapalnego.IOUS jest wiarygodnym badaniem w ocenie ostrych zmian zapalnych w trzustce, głównie w trakcie operacji przewlekłego, objawowego zapalenia tego narządu. Prawidłowo wyznacza zakres planowanej operacji. Badanie różnicuje zmiany torbielowate, guzy o torbielowatym charakterze, określa prawidłowy sposób drenażu lub weryfikuje wskazania do wycięcia zmiany chorobowej. Ocenia miejsce i zakres operacji drenażowych w przypadku nadciśnienia w drogach trzustkowych spowodowanego zwapnieniami miąższu lub kamicą przewodową w przebiegu przewlekłego zapalenia trzustki. W raku trzustki – weryfikuje stan miejscowego zaawansowania zmian guzowatych, pozwala na ocenę zajęcia naczyń okołotrzustkowych, węzłów chłonnych oraz wykazuje obecność przerzutów miejscowych i odległych, w tym w obrębie wątroby. IOUS znacznie poprawia skuteczność śródoperacyjnej BACC oraz aspiracji lub drenażu zbiorników płynowych.

Many common and inexpensive materials, such as, paper, fabric and thread are currently being explored as alternate substrates for low-cost and field-based sensing applications. While these substrates have primarily been utilized to develop low-cost diagnostic devices, it is important to study whether these materials are suitable for more general chemical sensing applications. Here, we report chemical syntheses (inorganic and organic) and sensing using threads as microchannels. We first characterized the liquid transport through thread channels and then measured typical sample losses due to adsorption and evaporation. Next, we demonstrated synthesis of ferric hydroxide in a Y-geometry thread reactor and detected the product by colorimetry. Our low-cost device gave a product yield of 84% compared to a test tube-based synthesis. We also synthesized (with similar to 72% yield) and detected colorless 2,4-dichloro-N-(2-morpholinoethyl) benzamide in an attempt to demonstrate an organic reaction in a thread device. Finally, we successfully detected bovine serum albumin and the glucose present in blood plasma as examples of detection of chemical substances relevant in diagnostics. Our results indicate that cotton and silk threads are indeed suitable materials for incorporation into field-deployable disposable chemical reaction and sensing systems. (C) 2013 Elsevier B.V. All rights reserved.

Biomarkers for active tuberculosis (TB) are urgently needed to improve rapid TB diagnosis. The objective of this study was to identify serum protein expression changes associated with TB but not latent Mycobacterium tuberculosis infection (LTBI), uninfected states, or respiratory diseases other than TB (ORD). Serum samples from 209 HIV uninfected (HIV−) and co-infected (HIV+) individuals were studied. In the discovery phase samples were analyzed via liquid chromatography and mass spectrometry, and in the verification phase biologically independent samples were analyzed via a multiplex multiple reaction monitoring mass spectrometry (MRM-MS) assay. Compared to LTBI and ORD, host proteins were significantly differentially expressed in TB, and involved in the immune response, tissue repair, and lipid metabolism. Biomarker panels whose composition differed according to HIV status, and consisted of 8 host proteins in HIV− individuals (CD14, SEPP1, SELL, TNXB, LUM, PEPD, QSOX1, COMP, APOC1), or 10 host proteins in HIV+ individuals (CD14, SEPP1, PGLYRP2, PFN1, VASN, CPN2, TAGLN2, IGFBP6), respectively, distinguished TB from ORD with excellent accuracy (AUC = 0.96 for HIV− TB, 0.95 for HIV+ TB). These results warrant validation in larger studies but provide promise that host protein biomarkers could be the basis for a rapid, blood-based test for TB., Highlights • Active tuberculosis leads to the differential expression of serum proteins involved in associated host processes. • Serum protein expression changes in tuberculosis involve the immune response, tissue repair, and lipid metabolism. • Panels of 8–10 host proteins can distinguish active tuberculosis from latent infection, and other respiratory diseases. Accurate biomarkers for active tuberculosis (TB) are urgently needed to improve rapid diagnosis. Current diagnostics for TB rely on microbiologic or molecular confirmation of M. tuberculosis, and are therefore dependent on a specimen from the site of disease which is not always accessible. This study demonstrates that human host proteins are differentially expressed in TB compared to latent M. tuberculosis infection, or respiratory diseases other than TB. Our data thus provide promise that host proteins have the potential to become the basis of rapid blood tests that do not require a sample from the site of disease.

The past decade has seen human leukocyte antigen (HLA) typing emerge as a remarkably popular test for the diagnostic work-up of coeliac disease with high patient acceptance. Although limited in its positive predictive value for coeliac disease, the strong disease association with specific HLA genes imparts exceptional negative predictive value to HLA typing, enabling a negative result to exclude coeliac disease confidently. In response to mounting evidence that the clinical use and interpretation of HLA typing often deviates from best practice, this article outlines an evidence-based approach to guide clinically appropriate use of HLA typing, and establishes a reporting template for pathology providers to improve communication of results.

Introduction Atopic dermatitis (AD) and psoriasis are chronic inflammatory skin diseases frequently accompanied by itching. The exact pathogenesis of dermatological pruritus remains unknown, but it is believed that altered skin innervation may play a role. Aim The assessment of the sensory threshold in AD and psoriasis in relation to pruritus experienced by patients. Material and methods A total of 18 subjects with AD, 20 with psoriasis and 49 healthy controls were exposed to alternating current generated by the current source. A selected preset of current frequencies (ranging from 5 Hz to 2000 Hz) allowed a selective stimulation of different nerve endings (Aβ, Aδ and C-type). Pruritus severity was measured with visual analogue scale (VAS) and an itch questionnaire developed in house. All results were analyzed statistically. Results Sensory thresholds within the uninvolved skin of AD or psoriasis patients were significantly higher than in healthy volunteers (p < 0.001), and no significant differences were found between AD and psoriasis (p > 0.05). Similarly, sensory thresholds within the diseased skin of AD or psoriasis were significantly higher than in the normal skin (p < 0.01), and patients with psoriasis had also a significantly higher threshold than AD individuals (p < 0.05). The sensory threshold inversely correlated with pruritus severity in AD and psoriasis and the highest correlation was found for 5 Hz frequency predominantly stimulating C fibers (VAS: R = –0.32, p < 0.05; pruritus questionnaire: R = 0.54, p < 0.001). Conclusions Evaluation of the sensory threshold may be a valuable tool for pruritus assessment, but further studies are still warranted.

Cystic fibrosis-associated liver disease (CFLD) affects ca. 30% of patients. The CFLD is now considered the third cause of death, after lung disease and transplantation complications, in CF patients. Diagnostics, clinical assessment and treatment of CFLD have become a real challenge since a striking increase of life expectancy in CF patients has recently been observed. There is no elaborated "gold standard" in the diagnostic process of CFLD; clinical evaluation, laboratory tests, ultrasonography and liver biopsy are used. Clinical forms of CFLD are elevation of serum liver enzymes, hepatic steatosis, focal biliary cirrhosis, multilobular biliary cirrhosis, neonatal cholestasis, cholelithiasis, cholecystitis and micro-gallbladder. In children, CFLD symptoms mostly occur in puberty. Clinical symptoms appear late, when damage of the hepatobiliary system is already advanced. The CFLD is more common in patients with severe mutations of CFTR gene, in whom a complete loss of CFTR protein function is observed. CFLD, together with exocrine pancreatic insufficiency and meconium ileus, is considered a component of the severe CF phenotype. Treatment of CFLD should be complex and conducted by a multispecialist team (gastroenterologist, hepatologist, dietician, radiologist, surgeon). The main aim of the treatment is to prevent liver damage and complications associated with portal hypertension and liver cirrhosis. Ursodeoxycholic acid is used in the treatment of CFLD. There is no treatment of proven long-term efficacy in CFLD. Liver transplantation is a treatment of choice in end-stage liver disease.

// Saber Imani 1, 2 , Xianqin Zhang 1 , Hossein Hosseinifard 3 , Shangyi Fu 4 , Junjiang Fu 1 1 Key Laboratory of Epigenetics and Oncology, The Research Center for Precision Medicine, Southwest Medical University, Luzhou, Sichuan, PR China 2 Chemical Injuries Research Center, Baqiyatallah Medical Sciences University (BMSU), Tehran, Iran 3 Research Center for Evidence Based Medicine (RCEBM), Tabriz University of Medical Sciences, Tabriz, Iran 4 Honors College, University of Houston, Houston, TX, USA Correspondence to: Junjiang Fu, email: fujunjiang@swmu.edu.cn , fujunjiang@hotmail.com Keywords: breast cancer, microRNA-34a, biomarker, diagnostics, meta-analysis Received: November 18, 2016 Accepted: January 30, 2017 Published: February 20, 2017 ABSTRACT Background: MicroRNA-34a (miR-34a) is a master regulator of tumor suppression in breast cancer (BC). This systematic review aims to analyze the diagnostic accuracy of miR-34a in the detection of BC as a biomarker. Results: A total of 1858 BC cases and 494 controls from thirteen eligible studies reported in 9 publications were included. The overall pooled sensitivity, specificity, negative likelihood ratio (NLR), positive likelihood ratio (PLR), and diagnostic odds ratio (DOR) were 85.50% (95% CI: 83.80-87.00%), 70.00% (95% CI: 65.80–74.10%), 0.29 (95% CI: 0.19–0.43), 2.58 (95% CI: 1.91–3.43), and 9.39 (95% CI: 5.47–16.12), respectively. Similarly, the overall area under the curve (AUC) of the summary receiver operating characteristic (SROC) was 0.80, indicating the high conservation of miR-34a as a biomarker. Furthermore, subgroup analysis suggested that the use of miR-34a as a biomarker is more accurate in tissue-based sample of invasive BC. We also indicated that miR-34a is a capable biomarker in diagnosing BC in people of Caucasian descent. Materials and Methods: A systematic search was conducted for eligible publications that address miR-34a expression level in BC cases and noncancerous controls. Diagnostic capacity of miR-34a for BC was assessed using pooled sensitivity and specificity, DOR, and AUC of SROC. PLR and NLR were verified to estimate the miR-34a diagnostic accuracy in clinical level. The quality of the included studies was assessed by QUADAS-2. Conclusions: These findings suggest miR-34a is a promising non-invasive biomarker in diagnosing BC. Well-designed cohort studies should be implemented to warrant the diagnostic value of miR-34a in clinical purposes.

Background: Obtaining a diagnosis of neuropsychiatric disorders such as autism requires long waiting times that can exceed a year and can be prohibitively expensive. Crowdsourcing approaches may provide a scalable alternative that can accelerate general access to care and permit underserved populations to obtain an accurate diagnosis. Objective: We aimed to perform a series of studies to explore whether paid crowd workers on Amazon Mechanical Turk (AMT) and citizen crowd workers on a public website shared on social media can provide accurate online detection of autism, conducted via crowdsourced ratings of short home video clips. Methods: Three online studies were performed: (1) a paid crowdsourcing task on AMT (N=54) where crowd workers were asked to classify 10 short video clips of children as “Autism” or “Not autism,” (2) a more complex paid crowdsourcing task (N=27) with only those raters who correctly rated ≥8 of the 10 videos during the first study, and (3) a public unpaid study (N=115) identical to the first study. Results: For Study 1, the mean score of the participants who completed all questions was 7.50/10 (SD 1.46). When only analyzing the workers who scored ≥8/10 (n=27/54), there was a weak negative correlation between the time spent rating the videos and the sensitivity (ρ=–0.44, P=.02). For Study 2, the mean score of the participants rating new videos was 6.76/10 (SD 0.59). The average deviation between the crowdsourced answers and gold standard ratings provided by two expert clinical research coordinators was 0.56, with an SD of 0.51 (maximum possible SD is 3). All paid crowd workers who scored 8/10 in Study 1 either expressed enjoyment in performing the task in Study 2 or provided no negative comments. For Study 3, the mean score of the participants who completed all questions was 6.67/10 (SD 1.61). There were weak correlations between age and score (r=0.22, P=.014), age and sensitivity (r=–0.19, P=.04), number of family members with autism and sensitivity (r=–0.195, P=.04), and number of family members with autism and precision (r=–0.203, P=.03). A two-tailed t test between the scores of the paid workers in Study 1 and the unpaid workers in Study 3 showed a significant difference (P

Background: Clinical implementation of Next-Generation Sequencing (NGS) is challenged by poor control for stochastic sampling, library preparation biases and qualitative sequencing error. To address these challenges we developed and tested two hypotheses. Methods: Hypothesis 1: Analytical variation in quantification is predicted by stochastic sampling effects at input of (a) amplifiable nucleic acid target molecules into the library preparation, (b) amplicons from library into sequencer, or (c) both. We derived equations using Monte Carlo simulation to predict assay coefficient of variation (CV) based on these three working models and tested them against NGS data from specimens with well characterized molecule inputs and sequence counts prepared using competitive multiplex-PCR amplicon-based NGS library preparation method comprising synthetic internal standards (IS). Hypothesis 2: Frequencies of technically-derived qualitative sequencing errors (i.e., base substitution, insertion and deletion) observed at each base position in each target native template (NT) are concordant with those observed in respective competitive synthetic IS present in the same reaction. We measured error frequencies at each base position within amplicons from each of 30 target NT, then tested whether they correspond to those within the 30 respective IS. Results: For hypothesis 1, the Monte Carlo model derived from both sampling events best predicted CV and explained 74% of observed assay variance. For hypothesis 2, observed frequency and type of sequence variation at each base position within each IS was concordant with that observed in respective NTs (R2 = 0.93). Conclusion: In targeted NGS, synthetic competitive IS control for stochastic sampling at input of both target into library preparation and of target library product into sequencer, and control for qualitative errors generated during library preparation and sequencing. These controls enable accurate clinical diagnostic reporting of confidence limits and limit of detection for copy number measurement, and of frequency for each actionable mutation.

Daniel J. O’Shannessy 1 , Elizabeth B. Somers 1 , Earl Albone 1 , Xin Cheng 1 , Young Chul Park 1 , Brian E. Tomkowicz 2 , Yoshitomo Hamuro 3 , Thomas O. Kohl 4 , Tracy M. Forsyth 4 , Robert Smale 5 , Yao-Shi Fu 5 , Nicholas C. Nicolaides 1 1 Morphotek Inc., 210 Welsh Pool Road, Exton, PA 19342, USA 2 Centocor Inc. Malvern, PA, USA 3 ExSAR Corporation, Monmouth Junction, NJ, USA 4 Rockland Immunochemicals, Inc., Gilbertsville, PA, USA 5 Laboratory Corporation of America, Los Angeles, CA, USA Received: December 15, 2011; Accepted: December 27, 2011; Published: December 27, 2011; Keywords: Folate receptor alpha, cancer, diagnostics, monoclonal antibody, protein structure, 26B3 Correspondence: Daniel J. O’Shannessy, email: // // Abstract Folate receptor alpha (FRA) is a cell surface protein whose aberrant expression in malignant cells has resulted in its pursuit as a therapeutic target and marker for diagnosis of cancer. The development of immune-based reagents that can reproducibly detect FRA from patient tissue processed by varying methods has been difficult due to the complex post-translational structure of the protein whereby most reagents developed to date are highly structure-sensitive and have resulted in equivocal expression results across independent studies. The aim of the present study was to generate novel monoclonal antibodies (mAbs) using modified full length FRA protein as immunogen in order to develop a panel of mAbs to various, non-overlapping epitopes that may serve as diagnostic reagents able to robustly detect FRA-positive disease. Here we report the development of a panel of FRA-specific mAbs that are able to specifically detect FRA using an array of diagnostic platforms and methods. In addition, the methods used to develop these mAbs and their diverse binding properties provide additional information on the three dimensional structure of FRA in its native cell surface configuration.

The ligation of active pharmaceutical ingredients (API) for working with image processing systems in diagnostics (MRT) attracts increasing notice and scientific interest. The Diels-Alder ligation Reaction with inverse electron demand (DAR(inv)) turns out to be an appropriate candidate. The DAR(inv) is characterized by a specific distribution of electrons of the diene and the corresponding dienophile counterpart. Whereas the reactants in the classical Diels-Alder Reaction feature electron-rich diene and electron-poor dienophile compounds, the DAR(inv) exhibits exactly the opposite distribution of electrons. Substituents with pushing electrones increase and, with pulling electrons reduce the electron density of the dienes as used in the DAR(inv).We report here that the DAR(inv) is an efficient route for coupling of multifunctional molecules like active peptides, re-formulated drugs or small molecules like the alkyalting agent temozolomide (TMZ). This is an example of our contribution to the "Click chemistry" technology. In this case TMZ is ligated by DAR(inv) as a cargo to transporter molecules facilitating the passage across the cell membranes into cells and subsequently into subcellular components like the cell nucleus by using address molecules. With such constructs we achieved high local concentrations at the desired target site of pharmacological action. The DAR(inv) ligation was carried out using the combination of several technologies, namely: the organic chemistry and the solid phase peptide synthesis which can produce 'tailored' solutions for questions not solely restricted to the medical diagnostics or therapy, but also result in functionalizations of various surfaces qualified amongst others also for array development.We like to acquaint you with the DAR(inv) and we like to exemplify that all ligation products were generated after a rapid and complete reaction in organic solutions at room temperature, in high purity, but also, hurdles and difficulties on the way to the TMZ-BioShuttle conjugate should be mentioned.With this report we would like to stimulate scientists working with the focus on "Click chemistry" to intensify research with this expanding DAR(inv )able to open the door for new solutions inconceivable so far.

// R. Jonas A. Nilsson 1,2,3,* , Niki Karachaliou 4,* , Jordi Berenguer 1 , Ana Gimenez-Capitan 5 , Pepijn Schellen 1,3 , Cristina Teixido 5 , Jihane Tannous 6 , Justine L. Kuiper 7 , Esther Drees 1 , Magda Grabowska 1 , Marte van Keulen 6 , Danielle A. M. Heideman 8 , Erik Thunnissen 8 , Anne-Marie C. Dingemans 9 , Santiago Viteri 4 , Bakhos A. Tannous 6 , Ana Drozdowskyj 10 , Rafael Rosell 4,5,11,12,** , Egbert F. Smit 7,** and Thomas Wurdinger 1,3,6,** 1 Cancer Center Amsterdam, Department of Neurosurgery, VU University Medical Center, Amsterdam, The Netherlands 2 Department of Radiation Sciences, Oncology, Umea University, Umea, Sweden 3 ThromboDx B.V., Amsterdam, The Netherlands 4 Translational Research Unit, Dr, Rosell Oncology Institute, Quiron Dexeus University Hospital, Barcelona, Spain 5 Pangaea Biotech SL, Barcelona, Spain 6 Department of Neurology, Massachusetts General Hospital and Neuroscience Program, Harvard Medical School, Boston, MA, USA 7 Cancer Center Amsterdam, Department of Pulmonary Diseases, VU University Medical Center, Amsterdam, The Netherlands 8 Cancer Center Amsterdam, Department of Pathology, VU University Medical Center, Amsterdam, The Netherlands 9 Department of Pulmonary Diseases, Maastricht University Medical Center, Maastricht, The Netherlands 10 Pivotal, Madrid, Spain 11 Catalan Institute of Oncology, Hospital Germans Trias i Pujol, Barcelona, Spain 12 Molecular Oncology Research (MORe) Foundation, Barcelona, Spain * These two authors are co-first authors of the manuscript ** These three authors are co-senior authors of the manuscript Correspondence to: Thomas Wurdinger, email: // Keywords : diagnostics, NSCLC, liquid biopsies, platelets, EML4-ALK Received : August 23, 2015 Accepted : October 06, 2015 Published : November 02, 2015 Abstract Purpose: Non-small-cell lung cancers harboring EML4-ALK rearrangements are sensitive to crizotinib. However, despite initial response, most patients will eventually relapse, and monitoring EML4-ALK rearrangements over the course of treatment may help identify these patients. However, challenges associated with serial tumor biopsies have highlighted the need for blood-based assays for the monitoring of biomarkers. Platelets can sequester RNA released by tumor cells and are thus an attractive source for the non-invasive assessment of biomarkers. Methods: EML4-ALK rearrangements were analyzed by RT-PCR in platelets and plasma isolated from blood obtained from 77 patients with non-small-cell lung cancer, 38 of whom had EML4-ALK-rearranged tumors. In a subset of 29 patients with EML4-ALK-rearranged tumors who were treated with crizotinib, EML4-ALK rearrangements in platelets were correlated with progression-free and overall survival. Results: RT-PCR demonstrated 65% sensitivity and 100% specificity for the detection of EML4-ALK rearrangements in platelets. In the subset of 29 patients treated with crizotinib, progression-free survival was 3.7 months for patients with EML4-ALK+ platelets and 16 months for those with EML4-ALK− platelets (hazard ratio, 3.5; P = 0.02). Monitoring of EML4-ALK rearrangements in the platelets of one patient over a period of 30 months revealed crizotinib resistance two months prior to radiographic disease progression. Conclusions: Platelets are a valuable source for the non-invasive detection of EML4-ALK rearrangements and may prove useful for predicting and monitoring outcome to crizotinib, thereby improving clinical decisions based on radiographic imaging alone.

Objective: To investigate glial and neuronal biomarkers in cerebrospinal fluid (CSF) samples from patients with relapsing–remitting multiple sclerosis (RRMS) and clinically isolated syndrome (CIS) suggestive of multiple sclerosis (MS), and to evaluate their ability to predict conversion from CIS to clinically definite MS (CDMS) and also disability progression in MS. Methods: CSF levels of neurofilament light protein (NFL), t-tau, p-tau, glial fibrillary acidic protein (GFAP), S-100B, human chitinase 3-like 1 protein (YKL-40), monocyte chemoattractant protein-1 (MCP-1), α-sAPP and β-sAPP; and Aβ38, Aβ40 and Aβ42, were analyzed in 109 CIS patients and 192 RRMS patients. The mean follow-up time of these 301 patients was 11.7 ± 6.4 years. Results: High levels of NFL were associated with early conversion from CIS to CDMS (hazard ratio (HR) with 95% confidence interval (CI): 2.69 (1.75 – 4.15); p < 0.0001). High levels of YKL-40 and GFAP were associated with earlier progression in the Expanded Disability Status Scale (EDSS), score 3: YKL-40 (HR (95% CI): 2.78 (1.48 – 5.23); p = 0.001) and GFAP (HR (95% CI): 1.83 (1.01 – 3.35); p = 0.04). High levels of YKL-40 were associated with earlier progression to EDSS 6 (HR (95% CI): 4.57 (1.01 – 20.83); p = 0.05). Conclusions: CSF levels of NFL in CIS patients are an independent prognostic marker for conversion to CDMS. Whereas, CSF levels of YKL-40 and GFAP are independent prognostic markers for disability progression in MS.

In the present communication, we report the fabrication of a unique core-shell inorganic nanomaterial with potential therapeutic and diagnostic functions. It contains an iron–cobalt (FeCo) core that demonstrates magnetic resonance imaging (MRI) contrast property and a thin nanoshell of gold that inhibits the function of a pro-angiogenic growth factor, VEGF165. Au(FeCo) core-shell nanomaterials are fabricated in the gas phase and characterized using transmission electron microscopy, energy dispersive spectrum, inductively coupled plasma analysis, and MRI. Inhibition of VEGF165 function by Au(FeCo) is demonstrated against VEGF165/VPF-induced signaling cascades and proliferation of human umbilical vein endothelial cells (HUVECs). The self-contrast property of Au(FeCo) is determined in vitro by MRI after incubating HUVECs with Au(FeCo), demonstrating intrinsic contrast property of this potentially therapeutic nanomaterial. In brief, we report here the successful fabrication of an inorganic core-shell nanomaterial with potential therapeutic and diagnostic functions. It inhibits the function of VEGF165 and functions as a MRI contrast agent.