Multiplexed biosensor for point-of-care COVID-19 monitoring: CRISPR-powered unamplified RNA diagnostics and protein-based therapeutic drug management.

In late 2019 SARS-CoV-2 rapidly spread to become a global pandemic, therefore, measures to attenuate chains of infection, such as high-throughput screenings and isolation of carriers were taken. Prerequisite for a reasonable and democratic implementation of such measures, however, is the availability of sufficient testing opportunities (beyond reverse transcription PCR, the current gold standard). We, therefore, propose an electrochemical, microfluidic multiplexed polymer-based biosensor in combination with CRISPR/Cas-powered assays for low-cost and accessible point-of-care nucleic acid testing. In this study, we simultaneously screen for and identify SARS-CoV-2 infections (Omicron-variant) in clinical specimens (Sample-to-result time: ∼30 min), employing LbuCas13a, whilst bypassing reverse transcription as well as target amplification of the viral RNA (LODs of 2,000 and 7,520 copies/µl for the E and RdRP genes, respectively, and 50 copies/ml for combined targets), both of which are necessary for detection via PCR and other isothermal methods. In addition, we demonstrate the feasibility of combining synthetic biology-driven assays based on different classes of biomolecules, in this case protein-based ß-lactam antibiotic detection, on the same device. The programmability of the effector and multiplexing capacity (up to six analytes) of our platform, in combination with a miniaturized measurement setup, including a credit card sized near field communication (NFC) potentiostat and a microperistaltic pump, provide a promising on-site tool for identifying individuals infected with variants of concern and monitoring their disease progression alongside other potential biomarkers or medication clearance.

Designing electrochemical microfluidic multiplexed biosensors for on-site applications.

Clinical assessment based on a single biomarker is in many circumstances not sufficient for adequate diagnosis of a disease or for monitoring its therapy. Multiplexing, the measurement of multiple analytes from one sample and/or of the same target from different samples simultaneously, could enhance the accuracy of the diagnosis of diseases and their therapy success. Thus, there is a great and urgent demand for multiplexed biosensors allowing a low-cost, easy-to-use, and rapid on-site testing. In this work, we present a simple, flexible, and highly scalable strategy for implementing microfluidic multiplexed electrochemical biosensors (BiosensorX). Our technology is able to detect 4, 6, or 8 (different) analytes or samples simultaneously using a sequential design concept: multiple immobilization areas, where the assay components are adsorbed, followed by their individual electrochemical cells, where the amperometric signal readout takes place, within a single microfluidic channel. Here, first we compare vertical and horizontal designs of BiosensorX chips using a model assay. Owing to its easier handling and superior fluidic behavior, the vertical format is chosen as the final multiplexed chip design. Consequently, the feasibility of the BiosensorX for multiplexed on-site testing is successfully demonstrated by measuring meropenem antibiotics via an antibody-free ß-lactam assay. The multiplexed biosensor platform introduced can be further extended for the simultaneous detection of other anti-infective agents and/or biomarkers (such as renal or inflammation biomarkers) as well as different (invasive and non-invasive) sample types, which would be a major step towards sepsis management and beyond.

Occurrence, loadings and removal of EU-priority polycyclic aromatic hydrocarbons (PAHs) in wastewater and sludge by advanced biological treatment, stabilization pond and constructed wetland.

Eight polycyclic aromatic hydrocarbon (PAH) compounds which have been accepted as priority micropollutants by European Union (EU) were analyzed both in wastewater and sludge lines throughout three full scale (located in city, sub-province and village) WWTPs during 12-month sampling period. Investigated WWTPs have different treatment types including advanced biological treatment, stabilization pond (SP) and constructed wetland (CW). Removal efficiencies for total PAH compounds varied from 48% in CW to 85% in advanced biological treatment plant. The maximum concentrations of 360-2282 ng/L observed for naphthalene in raw wastewater were decreased to 103-370 ng/L by treatment processes. Minimum concentration were detected for benzo(k)fluoranthene (B[k]F) and benzo(g,h,i)perylene (B[g,h,i]P) ranged between 8 and 12 ng/L and 19-33 ng/L, respectively. While minimum removal efficiencies were obtained for B[k]F and B[g,h,i]P maximum removal efficiencies were obtained for naphthalene in all WWTPs. PAHs present in minimum and maximum levels in the sludge samples were detected as 54 and 6826 ng/g for the B[g,h,I]P and naphthalene, respectively. Considering the removal mechanisms, PAHs have been determined to be removed by biodegradation or vaporization up to 84% and by settling (adsorption onto sludge) up to 2%. The greatest portion (99%) of naphthalene and anthracene were determined to be biodegraded or vaporized in biological treatment due to their low molecular weights. On the other hand, mechanism of adsorption onto sludge was determined as negligible for these two compounds. In addition, approximately 14% of PAHs were discharged to the receiving environment. Among the different WWTP types investigated, advanced biological treatment was found to be the most efficient plant for the removal of PAH compounds.

Removal of PAHs from leachate using a combination of chemical precipitation and Fenton and ozone oxidation.

In this study, six emerging pollutants, belonging to the polycyclic aromatic hydrocarbons (PAHs) group, found in landfill leachate were investigated for their removal by sequential treatment processes including chemical precipitation (CP), Fenton oxidation (FO) and ozone oxidation (OO). Each treatment process was run under different conditions using an experimental design program. Optimization of both CP and FO processes was designed based on the measured values of the residual chemical oxygen demand (COD) of the samples analyzed. The analysis of variance test was applied to the obtained results for determination of statistical significance of the model. Removal efficiencies of micropollutants were determined in the optimal conditions both for CP and FO processes. Samples obtained after these processes were treated with different pH and ozonation times for observing the performances of ozonation on micropollutant removal under different operating conditions. In this study, the removal of acenaphthylene, acenaphthene, fluorene, phenanthrene, fluoranthene and pyrene micropollutants was investigated. The values obtained for PAHs in leachate were determined to be above 10 ppb. In the CP process, the removal efficiencies for PAHs were ranged between 6% and 40% except for pyrene. Removal efficiencies of all micropollutants with FO were over 70% except for fluorene (55%). The removal efficiencies of the investigated micropollutants were 80-100% as a result of consecutive treatment processes including CP, FO and OO respectively.

Choice of marker for assessment of RV dysfunction in acute pulmonary embolism : NT-proBNP, pulmonary artery systolic pressure, mean arterial pressure, or blood pressure index.

BACKGROUND: We aimed to examine the value of NT-proBNP, pulmonary artery systolic pressure (PASP), blood pressure index (BPI), and mean arterial pressure (MAP) in the determination of right ventricular dysfunction (RVD) in patients with acute pulmonary embolism (APE). PATIENTS AND METHODS: A total of 547 patients diagnosed with APE were included in the study. Demographic characteristics and comorbid conditions of patients were recorded in patient files. For blood pressure measurement, a calibrated digital blood pressure monitor was used at regular intervals. Blood samples were taken from patients at the time of admission for hemogram, biochemical, and hemostasis blood tests. Echocardiography was performed on all patients to detect RVD and evaluate pulmonary artery pressure. RESULTS: PASP (p < 0.001), MAP (p < 0.001), diastolic blood pressure (p < 0.001), D­dimer (p = 0.001), NT-proBNP (p = 0.001), white blood cell (p < 0.001), and platelet (p = 0.001) counts were higher in APE patients with RVD compared with those without RVD, whereas the mean BPI level (p < 0.001) was lower. BPI had a negative correlation with PASP, NT-proBNP, platelet count, and triglyceride levels in patients with RVD. In regression analysis, BPI and PASP were found to be independent predictors of RVD. In receiver operating characteristic curve analysis, BPI (AUC ± SE = 0.975 ± 0.006; p < 0.001) was found to be the best predictor of RVD with a higher sensitivity (92.8%) and specificity (100%). CONCLUSION: We found that BPI had a better diagnostic discrimination for RVD compared with PASP and NT-proBNP.

Unraveling the impact of therapeutic drug monitoring via machine learning for patients with sepsis.

Clinical studies investigating the benefits of beta-lactam therapeutic drug monitoring (TDM) among critically ill patients are hindered by small patient groups, variability between studies, patient heterogeneity, and inadequate use of TDM. Accordingly, definitive conclusions regarding the efficacy of TDM remain elusive. To address these challenges, we propose an innovative approach that leverages data-driven methods to unveil the concealed connections between therapy effectiveness and patient data, collected through a randomized controlled trial (DRKS00011159; 10th October 2016). Our findings reveal that machine learning algorithms can successfully identify informative features that distinguish between healthy and sick states. These hold promise as potential markers for disease classification and severity stratification, as well as offering a continuous and data-driven "multidimensional" Sequential Organ Failure Assessment (SOFA) score. The positive impact of TDM on patient recovery rates is demonstrated by unraveling the intricate connections between therapy effectiveness and clinically relevant data via machine learning.

Occurrence, distribution, and fate evaluation of endocrine disrupting compounds in three wastewater treatment plants with different treatment technologies in Türkiye.

Nowadays, two of the endocrine disrupting compounds (EDCs) in the group of alkylphenols (APs), nonylphenol (4-NP) and octylphenol (4-t-OP), have attracted great scientific and regulatory attention mainly due to concerns about their aquatic toxicity and endocrine disrupting activity. This paper investigated the occurrence, distribution behavior, fate, and removal of 4-NP and 4-t-OP in liquid and solid phases of three full-scale wastewater treatment plants (WWTPs) with different treatment technologies comparatively. In this context, (i) advanced biological WWTP, (ii) wastewater stabilization pond (WSP), and (iii) constructed wetland (CW) were utilized. In all three investigated WWTPs, the concentrations of 4-NP (219.9-19,354.4 ng/L) in raw wastewater were higher than those of 4-t-OP (13.9-2822.4 ng/L). Within the scope of annual average removal efficiencies, 4-NP was treated highly in advanced biological WWTP (93.5 %), while it was almost not treated in WSP (3.1 %) and treated with negative removal (<0 %) in CW. While 4-t-OP was treated at a similar removal rate (93.5 %) to 4-NP in advanced biological WWTP, it was treated moderately in WSP (52.5 %) and very poorly in CW (12.4 %). It has been determined that the most important removal mechanism of both 4-NP and 4-t-OP in WWTPs is biodegradation, followed by sorption onto sewage sludge. According to the mass balance performed in advanced biological WWTP, the biodegradation rates for 4-NP and 4-t-OP were found to be 70.4 % and 86.6 %, respectively, while the sorption onto sewage sludge were determined to be 23.3 % and 6.8 %. One of the critical findings obtained within the scope of the study is that while the concentrations of both metabolites, especially 4-NP, in wastewater and sewage sludge, decreased considerably under aerobic conditions, on the contrary, their concentrations increased significantly under anaerobic conditions. Both compounds were detected at higher concentrations in primary sludge compared to secondary sludge in advanced biological WWTP, while in WSP, they were determined at higher concentrations in anaerobic stabilization pond sludge compared to facultative stabilization pond sludge. Besides, it was also determined that the sorption behavior of these alkylphenols is much more dominant than desorption.

A three-level model for therapeutic drug monitoring of antimicrobials at the site of infection.

The silent pandemic of bacterial antimicrobial resistance is a leading cause of death worldwide, prolonging hospital stays and raising health-care costs. Poor incentives to develop novel pharmacological compounds and the misuse of antibiotics contribute to the bacterial antimicrobial resistance crisis. Therapeutic drug monitoring (TDM) based on blood analysis can help alleviate the emergence of bacterial antimicrobial resistance and effectively decreases the risk of toxic drug concentrations in patients' blood. Antibiotic tissue penetration can vary in patients who are critically or chronically ill and can potentially lead to treatment failure. Antibiotics such as ß-lactams and glycopeptides are detectable in non-invasively collectable biofluids, such as sweat and exhaled breath. The emergence of wearable sensors enables easy access to these non-invasive biofluids, and thus a laboratory-independent analysis of various disease-associated biomarkers and drugs. In this Personal View, we introduce a three-level model for TDM of antibiotics to describe concentrations at the site of infection (SOI) by use of wearable sensors. Our model links blood-based drug measurement with the analysis of drug concentrations in non-invasively collectable biofluids stemming from the SOI to characterise drug concentrations at the SOI. Finally, we outline the necessary clinical and technical steps for the development of wearable sensing platforms for SOI applications.

End-to-end design of wearable sensors.

Wearable devices provide an alternative pathway to clinical diagnostics by exploiting various physical, chemical and biological sensors to mine physiological (biophysical and/or biochemical) information in real time (preferably, continuously) and in a non-invasive or minimally invasive manner. These sensors can be worn in the form of glasses, jewellery, face masks, wristwatches, fitness bands, tattoo-like devices, bandages or other patches, and textiles. Wearables such as smartwatches have already proved their capability for the early detection and monitoring of the progression and treatment of various diseases, such as COVID-19 and Parkinson disease, through biophysical signals. Next-generation wearable sensors that enable the multimodal and/or multiplexed measurement of physical parameters and biochemical markers in real time and continuously could be a transformative technology for diagnostics, allowing for high-resolution and time-resolved historical recording of the health status of an individual. In this Review, we examine the building blocks of such wearable sensors, including the substrate materials, sensing mechanisms, power modules and decision-making units, by reflecting on the recent developments in the materials, engineering and data science of these components. Finally, we synthesize current trends in the field to provide predictions for the future trajectory of wearable sensors.

Evaluation of occurrence, fate and removal of priority phthalate esters (PAEs) in wastewater and sewage sludge by advanced biological treatment, waste stabilization pond and constructed wetland.

Phthalate Esters (PAEs), detected in high concentrations generally in treated wastewater discharged from wastewater treatment plants (WWTPs), are important pollutants that restrict the reuse of wastewater. Investigating the fate of these endocrine-disrupting chemicals in WWTPs is crucial in order to protect both receiving environments and ecosystems. For this purpose, di(2-ethylhexyl) phthalate (DEHP), di-n-octyl phthalate (DNOP) and benzyl butyl phthalate (BBP) in the group of PAEs were monitored in simultaneously both in wastewater and sludge lines of selected two nature-based WWTPs and one advanced biological WWTP. Although it was frequently stated that phthalates were significantly removed in WWTPs in many studies found in literature, negative removal efficiencies of selected phthalates in investigated WWTPs during the sampling period were observed generally in this study. One of the reasons for this concentration increase could be releasing of phthalates from microplastics in wastewater during the treatment process or the desorption of PAEs from treatment sludge. DNOP was the compound with the highest concentration increase at almost each treatment unit of the three WWTPs. On the other hand, total PAEs load was 1997 g d-1 in advanced biological WWTP and adsorption onto sludge of PAEs were determined as 90%. The side-stream total load returned from the decanter supernatant was 0.02% of the total PAEs load coming to advanced biological WWTP from the sewer system. As a result of detailed statistical analysis, the correlation between raw wastewater and primary clarifier (PC) effluent was determined as an increasing linear relation for DEHP and DNOP. On the other hand, moderate and strong correlations were observed both between septic tank and constructed wetland (CW) processes with raw wastewater. In the waste stabilization pond (WSP), while a significant correlation was not found between the sludge line data, homogeneous variance, strong and moderate correlations were obtained in the wastewater line data. However, while mean differences for all investigated PAEs were not significant (p > 0.05) in the wastewater line, mean differences of DEHP (p < 0.05) were significant in the sludge line according to ANOVA analysis.

Biosensor-Enabled Multiplexed On-Site Therapeutic Drug Monitoring of Antibiotics.

Personalized antibiotherapy ensures that the antibiotic concentration remains in the optimal therapeutic window to maximize efficacy, minimize side effects, and avoid the emergence of drug resistance due to insufficient dosing. However, such individualized schemes need frequent sampling to tailor the blood antibiotic concentrations. To optimally integrate therapeutic drug monitoring (TDM) into the clinical workflow, antibiotic levels can either be measured in blood using point-of-care testing (POCT), or can rely on noninvasive sampling. Here, a versatile biosensor with an antibody-free assay for on-site TDM is presented. The platform is evaluated with an animal study, where antibiotic concentrations are quantified in different matrices including whole blood, plasma, urine, saliva, and exhaled breath condensate (EBC). The clearance and the temporal evaluation of antibiotic levels in EBC and plasma are demonstrated. Influence of matrix effects on measured drug concentrations is determined by comparing the plasma levels with those in noninvasive samples. The system's potential for blood-based POCT is further illustrated by tracking ß-lactam concentrations in untreated blood samples. Finally, multiplexing capabilities are explored successfully for multianalyte/sample analysis. By enabling a rapid, low-cost, sample-independent, and multiplexed on-site TDM, this system can shift the paradigm of "one-size-fits-all" strategy.

Low-Cost Optical Assays for Point-of-Care Diagnosis in Resource-Limited Settings.

Readily deployable, low-cost point-of-care medical devices such as lateral flow assays (LFAs), microfluidic paper-based analytical devices (µPADs), and microfluidic thread-based analytical devices (µTADs) are urgently needed in resource-poor settings. Governed by the ASSURED criteria (affordable, sensitive, specific, user-friendly, rapid and robust, equipment-free, and deliverability) set by the World Health Organization, these reliable platforms can screen a myriad of chemical and biological analytes including viruses, bacteria, proteins, electrolytes, and narcotics. The Ebola epidemic in 2014 and the ongoing pandemic of SARS-CoV-2 have exemplified the ever-increasing importance of timely diagnostics to limit the spread of diseases. This review provides a comprehensive survey of LFAs, µPADs, and µTADs that can be deployed in resource-limited settings. The subsequent commercialization of these technologies will benefit the public health, especially in areas where access to healthcare is limited.

On-Site Therapeutic Drug Monitoring.

Recent technological advances have stimulated efforts to bring personalized medicine into practice. Yet, traditional application fields like therapeutic drug monitoring (TDM) have remained rather under-appreciated. Owing to clear dose-response relationships, TDM could improve patient outcomes and reduce healthcare costs. While chromatography-based routine practices are restricted due to high costs and turnaround times, biosensors overcome these limitations by offering on-site analysis. Nevertheless, sensor-based approaches have yet to break through for clinical TDM applications, due to the gap between scientific and clinical communities. We provide a critical overview of current TDM practices, followed by a TDM guideline to establish a common ground across disciplines. Finally, we discuss how the translation of sensor systems for TDM can be facilitated, by highlighting the challenges and opportunities.

Evaluation of chromatographic descriptors for the prediction of gastro-intestinal absorption of drugs.

The use of chromatographic descriptors in QSAR was evaluated. Therefore, retentions were measured on an immobilized artificial membrane system, 2 micellar liquid chromatography systems and 17 orthogonal or disimilar reversed-phase liquid chromatographic systems. It was investigated whether it was possible to model gastro-intestinal absorption as a function of chromatographic retentions applying two linear and one non-linear multivariate modeling technique. In a second step it was evaluated if models built with theoretical descriptors could be improved by adding the measured retention factors to the data set of descriptive variables. It was seen that gastro-intestinal absorption could be modelled in function of chromatographic retention using the non-linear modeling technique multivariate adaptive regression splines (MARS). The best models were obtained using a combination of theoretical and chromatographic descriptors with MARS as modeling technique.

Unusual metastases of lung cancer: bulbus oculi and maxillary sinus.

Lung adenocarcinoma often makes metastasis to the brain, liver, kidneys, bone, bone marrow and adrenal glands. It can also make metastasis to other parts of the body rarely for example eye, nose, parotid gland and paranasal sinus. We did not encounter with combined ocular bulbus and the maxillary sinus metastases of lung cancer in the accessible literature. In this case report, a patient who was combined ocular bulbus and the maxillary sinus metastases of lung adenocarcinoma will be discussed.

A simple "lasso" for intraocular foreign bodies.

Three patients had foreign bodies in their anterior chambers following penetrating ocular injuries. These foreign bodies were removed by a closed chamber technique using a simple loop. The loop was created by a 22-gauge intravenous cannula and a 7-0 polypropylene suture. Retained cilia in one patient and metallic foreign bodies in two patients were removed using this intraocular "lasso." Sutures were not placed at the incision sites at the end of the surgery. This is an inexpensive and easy to prepare technique that introduces minimal surgical trauma. In addition, two hands are not needed for loop manipulation. This technique may be an excellent alternative for removal of small intraocular foreign bodies.

Effect of Sialic Acid on Platelet Cryopreservation.

Sialic acid is a molecule which is responsible for the net negative surface charge of platelets. We investigated the effect of sialic acid on fresh and cryopreserved platelets. Platelet samples were obtained by platelet apheresis from 8 healthy donors. Platelet suspensions with different sialic acid concentrations (0, 1, 2 and 4 mg/mL) were studied for ADP and ristocetin induced platelet aggregation, basal and ADP induced P-selectin and glycoprotein- Ib/IX expression. Then platelet samples were cryopreserved in 5% DMSO with or without 4 mg/mL sialic acid. After thawing, P-selectin expression was compared with the control group. Six samples were also washed after thawing and P-selectin expression was again compared to unwashed samples. Sialic acid suppressed ADP induced platelet aggregation and P-selectin expression in a dose dependent manner. In cryopreserved samples, P-selectin expression of 4 mg/mL sialic acid containing group was found significantly higher than the control group (p< 0.001). In cryopreserved control group, P-selectin expression of thawedwashed group was significantly higher than thawed-unwashed group (p< 0.05). Our results indicate that sialic acid is not a good cryoprotective agent. Washing procedure after thawing to eliminate DMSO causes significant platelet activation.

The Role of Interleukin-1 Inhibitors on Acute Myeloblastic Leukemia Blast Proliferation; Future Potential for Biotherapy.

UNLABELLED: The effect of interleukin-1 (IL-1) as an autocrine growth factor on the proliferation of the acute myeloblastic leukemia (AML) blasts was studied. Bone marrow specimens were obtained from nine patients with different subgroups of AML. IL-1 receptor antagonist (IL-1RA) and IL-1 ß neutralizing antibody (IL-1ß NA) alone or in combination were added to the culture mediums of the AML blast cultures for the detection of their inhibitory effect on AML blast cell proliferation and colony formation. Average colony numbers in the IL-RA, IL-ßNA, and IL-IRA plus IL-IßNA included culture flasks, were 63.7 ± 21.5 %, 69.5 ± 19 %, 53.4 ± 23.7 %, respectively, as compared to those of the control (p < 0.01). Inhibition of colony formation by IL-IRA plus IL-IßNA was more prominent than by IL-IßNA alone (p < 0.01). No correlation between the inhibition of AML blast colony ormation and FAB AML subgroups was seen. RESULT: Both IL-1RA or IL-IßNA or in combination induced varying degrees of inhibition on blast colony formation. IL-I inhibitory molecules could be considered as an alternative therapy for AML in patients whose blast cells are sensitive to IL-1 inhibition.