Images are presented of the change in conductivity distribution during the REG/cardiac cycle, at 50 fps, showing maximum local conductivity change of approx. 1% in grey/white matter. A total of 17 tests were performed during short  carotid artery occlusions on 5 volunteers, monitored by Transcranial Doppler ultrasound. From EIT measurements averaged over complete REG/cardiac cycles, 13 occlusion tests showed consistently decreased conductivity of cerebral regions on the occluded side, and increased conductivity on the opposite side. The maximum local conductivity change during occlusion was approx. 20%. The simplicity of the carotid artery intervention provides a striking validation of the scalp-mounted measurement system in imaging cerebral hemodynamics, and the REG images indicate its unique combination of sensitivity and temporal resolution. An Electrical Impedance Tomography (EIT) system has been developed for dynamic three-dimensional imaging of changes in conductivity distribution in the human head, using scalp-mounted electrodes. We attribute these images to changes in cerebral perfusion. At 100 frames per second (fps), voltage measurement is achieved with full-scale signal-to-noise ratio of 105 dB and common-mode rejection ratio > 90 dB. A novel nonlinear method is presented for 3-D imaging of the difference in conductivity distribution in the head, relative to a reference time. The method achieves much reduced modelling error. It successfully localizes conductivity inclusions in experimental and simulation tests, where previous methods fail. For > 50 human volunteers, the rheoencephalography (REG) waveform is observed in EIT voltage measurements for every volunteer, with peak-to-peak amplitudes up to approx. 50  Ifunctional Electrical Impedance Tomography of Evoked Responses, first described in . App. App4]. ribed in . The sysIn the case of human head studies, convincing validation of EIT sensitivity to dynamic variation in cerebral perfusion status is an important step in its potential uptake for clinical use. However, such a demonstration is non-trivial in several respects, e.g. the wide variety of entirely normal phenomena that can influence the trans-impedance EIT measurement, and the safety of volunteer subjects. In developing the fast EIT system reported here, it was found that the trans-impedances recorded on the scalp undergo significant cyclical variation in synchronism with cardiac activity. This bio-impedance phenomenon, referred to as rheoencephalography (REG), has been widely studied in non-imaging mode , 10] an an10] anet al. [Carotid artery occlusion is known to have a gross impact on cerebral perfusion –14]. It. It14]. et al.  have preIIAThe EIT measurements in this paper were made using two variants of our fEITER instrument. The original version of the system (as used in ) is termpk–pk) is applied via any two selected electrodes by a pair of mirror-image Voltage-Controlled Current Sources (VCCSs) based on an enhanced Howland design . T. T13]. TThe THR process was carried out 17 times on five volunteer subjects from within the research team. In each THR test, a 5-second (approx.) occlusion was applied to the left or the right MCA whilst recordings were made of data from EIT (fEITER Mk. I), TCD  and ECG (Mindray PM8000E Monitor).For EIT image reconstruction by the above method of difference imaging, the reference frame of voltage measurement values was set to the average values for a complete REG (i.e. cardiac) cycle immediately prior to the start of the THR occlusion.IIIASuccessful imaging within the human head using fEITER with scalp-mounted electrodes is critically dependent upon two attributes of the methodology described in 1)ε was large in this case.A head-shaped tank was filled with a saline solution of conductivity 2.7 S/m for tests as described in , one of A cylindrical piece of carrot  was inserted into the saline. With its long axis parallel to the vertical axis of the tank, it was placed to the left of center, then to the right, and then withdrawn. To track the movement of the carrot, difference imaging was applied, using a reference frame that was recorded prior to insertion. The traditional image reconstruction method  and the The traditional method failed to produce images that resembled the physical distribution in any respect. The proposed new method yielded images that successfully localized the carrot and its movement, as illustrated in 2)The system performance was assessed by simulating the inclusion of a 20 mm-diameter sphere of conductivity 0.65 S/m (i.e. blood) at four different locations inside the head, as shown in In the simulations, the current patterns listed in The images  resulting from the traditional reconstruction method of  are showMany other simulations were performed with the above inclusions and with different combinations of forward and inverse models that present larger modelling errors than the case discussed above. In all cases, the proposed new method of  achievedBThe raw EIT signals in μV.The use of the Mk. II system allows measurement of REG in lower trans-impedance situations  that wouDuring the REG cycle, the peak local magnitude of the conductivity change of the grey/white matter (relative to the reference period) within the 3-dimensional images, is of the order of 1%. Data taken on different days with the above subject were found to be highly repeatable and reconstructed images show clearly the same general behaviour. For each of the other two subjects measured with Mk. II, only a single trial session was conducted; in each case, the measurement data showed that several electrodes were poorly attached , thus reducing the quality of the reconstructed images. Nevertheless, for the many well-attached electrodes, similar behavior of EIT voltages, as described for the first subject above, was observed during the REG cycle.CDuring the left-side occlusion, the EIT measurements near the left ear , show a Using EIT data averaged over complete REG cycles  THR trials reported in et al. [During long-term (60s) common carotid artery occlusion on human subjects, Shi et al.  report 2VThe fEITER system reported here achieves a unique combination of high imaging speed (100 fps) and measurement sensitivity . Using a novel nonlinear 3D reconstruction method, EIT imaging of human cerebral hemodynamics using scalp-mounted electrodes has been demonstrated. The reconstruction method does not use a subject-specific model. Simulations show that it is resilient against modelling error and provides good localization of included conductivity contrasts  for measurement SNR of 70 dB or better.For > 50 resting human volunteers, trans-impedance measurements using fEITER Mk. I display the REG waveform, synchronized with the cardiac cycle. For a resting human volunteer measured by Mk. II, EIT imaging of the REG process reveals ~1% tidal variation in cerebral conductivity. This variation shows considerable left-right symmetry and cycle-to-cycle repeatability. We believe that the resulting reconstructed images of conductivity difference in the white and grey matter during the REG cycle represent the first time-resolved images of cerebral perfusion due to the normal cardiac cycle. The intervals over which significant changes occur in these images are short, suggesting a minimum EIT frame rate of 25 fps for such studies.EIT measurements during THR are highly correlated with TCD data. Using data from fEITER Mk. I, EIT images during the THR process (for measurements averaged over the REG cycle) show large changes in conductivity (~20%), far exceeding the steady-state variation. The images show strong left-right asymmetry, reflecting the application of the carotid artery occlusion. Similar behaviour was observed in 13 out of 17 trials, with the remaining 4 traceable to a single experimental error. We believe that these are the first 3D images to be observed during the THR procedure.Taken together, these results provide compelling evidence that both steady-state and evoked variations in cerebral hemodynamics are observable using state-of-the-art EIT instruments and algorithms. The challenge now is to demonstrate the collection of clinically useful data.


Transfemoral transcatheter aortic valve replacement (TAVR) is currently thestandard catheter-based treatment of severe aortic stenosis patients. Being thetransfemoral route not feasible, other access sites could be chosen. TransaorticTAVR via either a J mini-sternotomy or a right anterolateral mini-thoracotomy isa good option for patients having tricky thoracoabdominal aorta. Some tips andtricks may help in getting a fast and safe transaortic procedure. As indications forTAVR are getting broader due to current guidelines recommendations to be an efficient route when direct cannulation of the commonfemoral artery or navigation through the aortic arch and thoracoabdominal aorta istricky or unsafe, when atransapical (TA) approach is not judged the optimal strategy, and when other routes are notindicated.Transaortic (TAo) TAVR has demonstratedA proper hybrid operating room (HOR) setting is mandatory to perform the procedurefluently and securely. Our standard setting is depicted in Briefly, the patient is lying supine, in a standard fashion. The fluoroscopic C-armis set as per the TF route, with the machine on the patients’ left side. The screensare obliquely or transversally put at the patient’s feet. The first operator is onthe right side in front of the C-arm, and the assistant is on the left side, betweenthe C-arm and the transesophageal machine (if required). At the head of the patient,the anesthetist and the ventilator are set. We normally use three tables. The smallone (Mayo) is used at the beginning and at the end and supports all the surgicalinstruments. The second table is used for the device preparation, and the third -put behind and to the left of the first operator - carries all the transcathetermaterials and harbor the delivery catheter and valve before the TAo insertion, thusproviding less distance and curve before their insertion within the delivery sheath.A preassembled and deaired cardiopulmonary bypass machine is always running justoutside the HOR.We generally get percutaneous access of the common femoral vein, with introduction ofa 6-Fr short sheath and a multipolar right ventricular pacing catheter for treatingpotential bradyarrhythmias at incision; at this stage a half-dose of heparin isgiven.nd or 3rd rightintercostal space or a right anterolateral mini-thoracotomy at the 2ndintercostal space is chosen according to the length and depth of the aorta, itsanatomical relationship to the sternum, and to the presence or absence of horizontalaorta (> 48°), respectively. An extension to the 4th intercostal spacemay be needed in case of deep aorta location (> 6 cm). The pericardial sac isincised and suspended, and the remaining dose of heparin is administered.A manubrotomy (J-mini-sternotomy) to the 2We normally manage both the 5-F pigtail and the device delivery sheath from theascending aorta.Even though the non-coronary sinus is located between four and six o’clock from theoperator’s view, we found easier to engage it with the pigtail catheter from theanterolateral surface of the aorta (towards the inner curvature), on which a double4-0 proline purse-string suture is performed. A 6-Fr short sheath istranscutaneously introduced in the ascending aorta via a Seldinger technique tostabilize the catheter-sheath unit during the deployment.e.g., a Teflonfelt), if present, and at least 5.5 cm away from the aortic annular plane:Because of the typical tissue frailty of the elderly patients normally referred forTAVR, we generally perform three large, concentric, purse-string sutures on theouter curvature of the ascending aorta , ~ 2-3 cThe inner purse-string is made of 3-0 braided stitch facing the assistantThe intermediate is made of 4-0 proline, facing the operator (180° from thefirst suture)The outer is a U-shape 4-0 proline stitch facing the assistant with doubleTeflon felt supportA 7-Fr short sheath is then placed as previously described. We do not exchange ituntil a stiff pre-curved shape wire is within the left ventricle. Through thissheath, the valve is crossed as per operator preference; in challenging horizontalaortas, we found useful for this purpose a 3.5 to 5-Fr Judkins right catheter. Wethen exchange the short sheath with the delivery one  to deploRegarding the complications of the TAo procedures, they could be addressed as generalTAVR-related complications  and TAo specific ones. The latterencompasses access site complications  and general surgical complications .All our TAo-TAVR cases are performed in a HOR with the capability for bothcardiothoracic surgical and interventional procedures, in general anesthesia andorotracheal intubation. We obtained informed consent from the patient, while ourInstitutional Review Board waived the need of publication consent due to theretrospective nature of this report.In our practice, we have progressively shifted from the TA to the Tao-TAVR as analternative access whenever the ascending aorta is suitable for harboring theintroducer sheath.nd postoperative day.Since 2016, we have adopted this route in case of unsuitable femoral access. Sofar, we have successfully treated 20 patients, with one operative death due tomassive ischemic stroke on the 2We currently reserve the TA access mainly for cases of short or heavilyatherosclerotic ascending aorta. For a proper indication of Tao-TAVR, wegenerally rely on high-resolution multi-detector computed tomography of thethorax. In this manner, we are able to properly estimate the ascending aortalength from the virtual basal ring and the wall’s atherosclerotic burden. Sometips and tricks on this access management could be useful for a fast procedureand favorable outcomes.. Also, a direct transiliac approachthrough a small abdominal incision has been described in highly selectedpatients.Currently, other than TF-TAVRs account only for ≤ 15% of all thecatheter-based aortic valve replacements. The main alternative routes described inthe literature are: transcarotid, transaxillary, TA, and transcavalUsing the ascending aorta to perform the procedure is a familiar circumstance for acardiac surgeon, thus increasing self-confidence on the procedure. In the nearfuture, the number of TAVR procedures will further expand; choosing the therapy witha multidisciplinary team will be more and more worthy, and different possibilitieswill lead to a tailored therapy.Several observational studies have reported a comparison between TF and Tao-TAVRsresults. reported similar 30-day and 1-yearoutcomes, but with a trend in favor of TF-TAVR. Nevertheless, the TAo approachtended to present better outcomes than the TA one.The first direct comparison of these two accesses made by Arai et al.The TAo approach have several advantages over the others. The first is no need foraortic arch crossing; in case of severe arch pathology, this may result in a reducedhazard of embolic neurological events due to debris dislodgement during wires andcatheters exchanges.Secondly, there will be only one site of operation directly controlled, with nofurther risk of secondary access complications. Finally, the short distance from theannulus may help in challenging situations requiring fine deploymentadjustments., and these possibilities enrich the surgicalarmamentarium.Modifications of the classical TAo technique have been made in search for a lessinvasive direct aortic approach. In our experience, we find them useful in thecontext of minimally invasive cardiac surgery and redo or multi-valvular operations.In general, once a patient is considered at moderate or higher risk for surgery andis more than 75 years old, we would treat him by TAVR.In recent years, the introduction of the so-called rapid deployment bioprosthesis haswidened the possibilities to treat the aortic stenosisIn conclusion, as per our experience, handling the TAo approach could be of worth forthose surgeons performing TAVR, and some simple rules and tricks help in getting theprocedure safe and effective.


The COVID-19 pandemic prompted widespread implementation of telehealth, including in the inpatient setting, with the goals to reduce potential pathogen exposure events and personal protective equipment (PPE) utilization. Nursing workflow adaptations in these novel environments are of particular interest given the association between nursing time at the bedside and patient safety. Understanding the frequency and duration of nurse-patient encounters following the introduction of a novel telehealth platform in the context of COVID-19 may therefore provide insight into downstream impacts on patient safety, pathogen exposure, and PPE utilization.The aim of this study was to evaluate changes in nursing workflow relative to prepandemic levels using a real-time locating system (RTLS) following the deployment of inpatient telehealth on a COVID-19 unit.In March 2020, telehealth was installed in patient rooms in a COVID-19 unit and on movable carts in 3 comparison units. The existing RTLS captured nurse movement during 1 pre- and 5 postpandemic stages (January-December 2020). Change in direct nurse-patient encounters, time spent in patient rooms per encounter, and total time spent with patients per shift relative to baseline were calculated. Generalized linear models assessed difference-in-differences in outcomes between COVID-19 and comparison units. Telehealth adoption was captured and reported at the unit level.P<.001). Frequency of encounters decreased (difference-in-differences range –6.6 to –14.1 encounters) and duration of encounters increased (difference-in-differences range 1.8 to 6.2 minutes) from baseline to a greater extent in the COVID-19 units relative to the comparison units. At most stages of the pandemic, the change in total time nurses spent in patient rooms per patient per shift from baseline did not differ between the COVID-19 and comparison units . The primary COVID-19 unit quickly adopted telehealth technology during the observation period, initiating 15,088 encounters that averaged 6.6 minutes (SD 13.6) each.Change in frequency of encounters and time spent per encounter from baseline differed between the COVID-19 and comparison units at all stages of the pandemic (all RTLS movement data suggest that total nursing time at the bedside remained unchanged following the deployment of inpatient telehealth in a COVID-19 unit. Compared to other units with shared mobile telehealth units, the frequency of nurse-patient in-person encounters decreased and the duration lengthened on a COVID-19 unit with in-room telehealth availability, indicating “batched” redistribution of work to maintain total time at bedside relative to prepandemic periods. The simultaneous adoption of telehealth suggests that virtual care was a complement to, rather than a replacement for, in-person care. However, study limitations preclude our ability to draw a causal link between nursing workflow change and telehealth adoption. Thus, further evaluation is needed to determine potential downstream implications on disease transmission, PPE utilization, and patient safety. The COVID-19 pandemic prompted the widespread implementation of telehealth throughout the health care sector to protect patients and health care workers by reducing the risk of infection . In the An RTLS captures the time spent in specific locations, providing a direct measure of workflow for health care professionals as well as a proxy for outcomes associated with staff movement, such the use of PPE upon each entrance into a room -17. In t. Understanding direct nurse-patient care following the introduction of a novel telehealth platform in the context of COVID-19 may provide insight into the downstream impacts on patient safety, pathogen exposure, and PPE utilization.The impact of telehealth on nursing workflows is of particular interest, as nurses spend approximately 6-fold more time at the bedsides of hospitalized patients than attending physicians . More tiThus, we aimed to evaluate changes in nursing workflow relative to prepandemic levels using an RTLS following the deployment of inpatient telehealth on a COVID-19 unit. Given isolation precautions and ready availability of telehealth equipment on the COVID-19 unit, we hypothesized a reduction in the frequency and duration of in-person nurse-patient encounters in the COVID-19 unit relative to prepandemic comparator units.Telehealth was implemented throughout an acute care academic hospital in response to the COVID-19 pandemic in March of 2020. The setup of telehealth differed between the hospital’s primary COVID-19 unit and comparison units. Our primary aim was to explore changes in nursing workflows in these novel circumstances through a retrospective, observational evaluation using RTLS data to capture the frequency of direct nurse-patient encounters at the bedside, time nurses spent in patient rooms per encounter, and total time nurses spent with each patient in the patient room per shift. For each outcome, the change from the prepandemic level was calculated, and difference-in-differences analyses were used to determine if changes in nurse movement differed between the COVID-19 unit and comparison units. The simultaneous adoption of telehealth in terms of video call frequency and duration was captured and is reported at the unit level.The academic acute care hospital is in a major metropolitan area in the western United States and serves a diverse population. The four inpatient units in this evaluation were identical in size and layout with 22-bed single-room capacity. Prepandemic, these four units focused on inpatient general medicine populations. Other units serving primarily surgical, oncological, and intensive care patients were excluded. At the beginning of the pandemic, one of the units became the primary COVID-19 unit for the hospital and was therefore compared to the other three units. Hospital administration reported that standard registered nurse-to-patient ratio on the four units ranged from 1:4 to 1:3, varying with disease acuity based on state law . The regThe first patient tested positive for SARS-CoV-2 in the hospital’s emergency department on March 2, 2020, and local stay-at-home orders were announced on March 16, 2020 . For theIn response to the pandemic, telehealth was rapidly implemented throughout inpatient settings in mid-March 2020, as previously reported  and descAll clinical team members, including hospitalist and specialist physicians, nurses, respiratory therapists, trainees, and clinical researchers, received instructions on and were encouraged to incorporate telehealth into their clinical or research activities. Patients received incoming calls passively given the default automatic turn-on feature of the video tablet, but received no other instruction or guidance on its use .The primary data source for this evaluation was extracted from the existing RTLS platform . The RTLS captured movement of nurses into and out of patient rooms on the selected units. These data were used to calculate the following 3 outcomes related to nurses’ movement and direct patient care: (1) number of nurse-patient direct encounters within patient rooms, (2) time nurses spent in patient room per encounter, and (3) total time nurses spent in the patient room with each patient per shift.The key components of the RTLS are infrared and radiofrequency sensors installed in each room and staff badges worn alongside their name badge ,29. LineSince the direct line of sight between the room sensor and nurse badge could be briefly interrupted , a single nurse-patient encounter could appear as several short, consecutive events. Thus, to identify unique direct nurse-patient encounters and calculate the time nurses spent in a patient room, multiple successive RTLS events that occurred within 30 seconds of another and were associated with an individual nurse in a single patient room were collapsed into a single direct nurse-patient encounter. The duration of individual encounters was summed to calculate the total time nurses spent with each patient per shift using the first and last timestamp of each encounter associated with a unique nurse in a specific patient room. Since nursing needs and thus workflows may differ between shifts , movemenStaff RTLS badges were linked within the system to an employee identifier and role. Nurses categorized as “nurse” or “float nurse” within the RTLS were included in the analysis. Hospital administration and managers encourage all nurses to wear the badges to utilize the system’s beneficial features, including automatic silencing of patient room alarms when a nurse enters a room and a discreet button to call security. Compliance with badge wearing is near universal among nursing staff compared to other members of the clinical team  . HoweverData were extracted from the EHR  to determine the presence of a patient in each hospital room in the unit at midnight, and the results of the patient’s most recent COVID-19 test result (positive or negative) within the prior 14 days or from hospital admission. Besides this information, no other patient-level data such as identifiers or clinical characteristics were obtained. The RTLS and EHR data were then merged by patient room and time to identify direct encounters between nurses and patients.Data were extracted from Zoom video conferencing software and included the unit associated with the host (originating hardware) user ID, call start time, call end time, and number of participants. Only calls lasting between 30 seconds and 2 hours with 2 or more participants were included in analysis. Patients were not instructed or encouraged to initiate calls to clinical staff or family members themselves during the observation period, although anecdotal reports suggest that nurses occasionally set up calls between patients and their families. This data platform did not link a telehealth encounter to individual physicians, staff members (nurses), or patients. Consequently, telehealth use described all possible use cases at the unit level.Descriptive statistics were generated to describe patient census by COVID-19 status and telehealth utilization in the COVID-19 and comparison units prepandemic during the 5 stages of the pandemic. Telehealth utilization is expressed as the number of telehealth calls per patient (using the midnight patient census) per unit for all case uses such as clinical encounters, research activities, and patient-family connections. Since telehealth events could not be linked to individual users , no additional analysis was performed with these data.To investigate differences in the three primary outcomes based on RTLS data between the COVID-19 and comparison units, a difference-in-differences approach was applied. Change in each outcome was calculated for each of the 5 stages of the pandemic, as defined by local case rates, relative to prepandemic levels. To determine if the change from the prepandemic stage differed between the COVID-19 and comparison units, difference-in-differences was determined using a generalized linear model in SAS  for each of the three outcomes.P<.05 was considered statistically significant. When multiple comparisons were made, P values were adjusted using an adaptive, two-stage linear setup procedure to control the false discovery rate [Means, standard deviations, and ranges are reported where appropriate. For all models, ery rate .Data were obtained from multiple sources, including RTLS movement data at the individual nurse level, EHR data at the patient and thus room level, and telehealth log data at the unit level. All data were deidentified and data points that did not occur in patient rooms were removed before being sent to the analytics team to protect the anonymity of the workforce. The evaluation was exempt per the Stanford University's institutional review board (protocol 55927).The COVID-19 unit was the main care location for hospitalized patients diagnosed with COVID-19, but it was also the care location for some patients without COVID-19  for the primary COVID-19 unit and comparison units is shown in P<.001). Nurses in the COVID-19 unit entered patient rooms less frequently during each surge period when compared to the nonsurge period just before; this pattern was seen for both the morning and night shifts .For each stage of the pandemic, change in number of times nurses entered patient rooms relative to that prepandemic are shown in The daily median time nurses spent in patient rooms per shift (morning and night) for the primary COVID-19 unit and comparison units is shown in P=.79) and night (P=.81) shifts (P<.001). Nurses in the COVID-19 unit spent more time in patient rooms per entry during each surge period when compared to that in the nonsurge period just before; this pattern was seen for both the morning and night shifts .Time nurses spent in patient rooms per entry did not differ between the COVID-19 unit and comparison units during the prepandemic stages for both the morning  shifts . For eac) shifts . In the ) shifts . At all P<.001) but not the night shift (P=.57), as shown in The daily median total time nurses spent in patient rooms per patient per shift (morning and night) for the primary COVID-19 unit and comparison units is shown in P>.17), with the exception of the telehealth rollout phase.In each stage of the pandemic, the change in the total time nurses were in a patient room per patient per shift relative to baseline is shown in Evaluating changes in the frequency and duration of direct nurse-patient encounters using an RTLS following an inpatient telehealth deployment during the COVID-19 pandemic was feasible and provided novel insights into nursing workflow redistribution in this setting. Relative to the prepandemic stage, nurses in a COVID-19 unit with in-room ready access to telehealth decreased the frequency of entries into patient rooms to a greater extent than that for nurses in other units with shared mobile telehealth units. Counter to our hypothesis, the average in-person encounter length increased proportionally, such that the total in-person time nurses spent with patients on the COVID-19 unit did not significantly differ from that in prepandemic comparator units. The simultaneous adoption of telehealth, presented at the unit level, suggests it was used as a complement to, rather than a replacement for, in-person care.To put the above findings into context, the average decrease in encounters weighted by time period in the COVID-19 unit relative to other units was 11.25 and 9.13 encounters per patient per morning and night shift, respectively. Assuming full capacity on the 22-bed COVID-19 unit and PPE use for a quarter of such encounters (as isolation precautions were not required for every patient on the unit), these data suggest workflow adaptations saved approximately 785 PPE units over the course of a week. Similarly, given the increased time burden required in caring for isolated patients—approximately 4 minutes to don and 3 minutes to doff PPE outside the patient room —workflowThe change in total time spent at patient bedside per shift from baseline did not differ significantly between the COVID-19 and comparison units, given the increased duration of each encounter. Past work suggests that this “batching” or “clustering” of bedside work includes performing physical assessments, administering medications, and delivering a food tray all in one bedside encounter ,32. The Notably, The COVID-19 unit and comparison units had important differences that likely influenced the adoption of telehealth. In the COVID-19 unit, hardware was readily available in each patient room, whereas the comparison units had a limited number of carts in a central location on the unit; the hardware had to be retrieved and then set up in the patient rooms prior to use. In addition to the ready availability of technology, the increased threat of pathogen exposure on the COVID-19 unit likely further promoted rapid staff uptake relative to comparison units, which did not adopt telehealth until late in the observation period (surge #2) when the number of COVID-19 patients on those units increased. These factors appeared to help health care workers overcome typical barriers to telehealth adoption and integrate the technology into their regular clinical routine ,34.Beyond the introduction of telehealth, the observed changes in nurse-patient encounters could also be due to unobserved differences between the primary COVID-19 unit and comparator units during this real-world health crisis. Aspects of nursing care were changing, in tandem with standards for the management of COVID-19, PPE availability, and infection control recommendations ,35,36. TThis retrospective, observational study utilized readily available data in a real-world setting and thus certain limitations exist. RTLS badges were worn consistently among nurses; however, limited compliance among other health care professionals such as physicians preclude analysis of other roles . Only nuDespite these limitations, an RTLS offers an alternative to other high-burden data sources of interest, such as ethnographic observation, to provide novel insights that may not otherwise be available. The strength of this evaluation is that movement data are available for nurses and linked with individual patient encounters in this novel clinical context.Assessment of nursing workflow change following the deployment of inpatient telehealth in the context of the COVID-9 pandemic was feasible utilizing RTLS data in combination with EHR data. Compared with those of other units with shared mobile health units, direct nurse-patient encounters on a COVID-19 unit with in-room ready access to telehealth decreased in frequency and increased in duration, leading to a redistribution of work that did not impact total time at the bedside relative to prepandemic periods. The simultaneous adoption of telehealth suggests virtual care complemented, rather than replaced, in-person care in this setting. Study limitations, including the lack of telehealth utilization data at the nurse or patient level and multiple differences between the COVID-19 and comparator units , preclude our ability to draw a causal link between nursing workflow change and telehealth adoption. Further evaluation is needed to determine potential downstream implications on unmeasured outcomes such as disease transmission, PPE utilization, and patient safety.


Mycobacterium tuberculosis by inducing non-specific immune responses, was found to play an essential role against COVID-19 in various ecological, analytical, and animal studies (The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has inflicted an unprecedented and significant toll on global public health and the economy. During the fight against the pandemic, the century-old tuberculosis (TB) vaccine Bacille Calmette Guérin (BCG), which has been reported to protect against infections of various respiratory pathogens other than  studies .Our previous studies have reported that BCG vaccination could induce a non-specific immune response to fight against unrelated pathogens , 4–7. ThEarly in the COVID-19 outbreak, the hypothesis that BCG could prevent COVID-19 infection was raised. Isabel N. Kantor was the first to publicly discuss whether BCG has preventive and protective effects against COVID-19 and how strong the association is between the “more BCG vaccination” and the “ less COVID-19 infection” . Then, tAlthough this hypothesis offered a glimmer of hope to the panicked and helpless people in the early days of the COVID-19 pandemic, it still needed a lot of evidence to prove it. In the early months of the COVID-19 pandemic, findings from the ecological and analytical studies indicated that countries with low BCG coverage had significantly higher rates of COVID-19 morbidity and mortality than countries with high BCG coverage –40. On teClinicalMedicine and attracted wide attention , which aimed to evaluate the efficacy of BCG vaccination in delivering protection against COVID-19 in healthcare workers (HCWs) in South Africa, were published in the journal ttention . The stuM. tuberculosis-infected mice were resistant to secondary SARS-CoV-2 infection  and the Chinese PLA General Hospital (Grant No. QNC19047) to WG.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Proteins are direct products of the genome and metabolites are functional products of interactions between the host and other factors such as environment, disease state, clinical information, etc. Omics data, including proteins and metabolites, are useful in characterizing biological processes underlying COVID-19 along with patient data and clinical information, yet few methods are available to effectively analyze such diverse and unstructured data. Using an integrated approach that combines proteomics and metabolomics data, we investigated the changes in metabolites and proteins in relation to patient characteristics  and clinical information . We found significant enrichment of biological indicators of lung, liver, and gastrointestinal dysfunction associated with disease severity using publicly available metabolite and protein profiles. Our analyses specifically identified enriched proteins that play a critical role in responses to injury or infection within these anatomical sites, but may contribute to excessive systemic inflammation within the context of COVID-19. Furthermore, we have used this information in conjunction with machine learning algorithms to predict the health status of patients presenting symptoms of COVID-19. This work provides a roadmap for understanding the biochemical pathways and molecular mechanisms that drive disease severity, progression, and treatment of COVID-19. A number of observational studies have established a relationship between severe COVID-19 and pre-existing conditions such as Type 2 Diabetes and obesity3. Utilizing a multi-omic approach to investigate how comorbidities may contribute to both sides of the virus-host interaction will allow for a molecular-level understanding of the infection and may lead to improved preventive or therapeutic interventions. Omics data sets are highly complex, often containing a high degree of dimensionality and zero-inflation which can complicate analyses that rely on conventional statistical testing. Therefore, as new computational tools are developed to handle omics data, reanalysis of publicly available COVID-19 data may reveal novel findings. Our group, alongside our collaborators, has recently generated several innovative methods for identifying key clusters4, molecules5, and biological processes6 from omics data.The COVID-19 pandemic continues to make an impact globally as communities reshape their activities due to the spread of various emerging SARS-CoV-2 strains. Despite the generation of multiple efficacious vaccines, our understanding of the factors that contribute to disease severity remains limited7. SARS-CoV-2 genome variation over time has introduced new strains with various functional characteristics such as a set of mutations in spike protein, the primary vaccine antigen8, and folding conformations in the virus variants and related functions9. Host response to infection can be measured by profiling small molecules (metabolites) and large molecules (proteins). For example, 3′-Deoxy-3′,4′-didehydro-cytidine (ddhC), a human antiviral metabolite, was significantly increased in COVID-19 patients10, and Gamma aminobutyric acid (GABA) metabolite was suggested as a potential signaling molecule by activating B cells and plasma cells11. We aimed to investigate metabolite and protein profiles to develop a comprehensive snapshot of host response and identify potential molecular biomarkers associated with COVID-19 disease severity.We previously have shown that SARS-CoV-2 genomic variation is independent of host characteristics 12. The measurements included a complete blood cell count panel, a comprehensive metabolic test panel, and quantification of 941 metabolites and 894 proteins from serum samples. Metabolite profiling and protein profiling were performed using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and stable isotope-labeled proteomics strategy TMTpro (16plex)13.We investigated the factors that influence COVID-19 disease severity by reanalyzing a previously published integrated study of metabolite and protein profiles, epidemiological data, and clinical data14. This is especially the case for severe COVID-19, which results in prolonged systemic inflammation that can damage multiple organ systems16. The mechanism behind these events can be investigated through the interrogation of soluble factors present within the circulation during infection. This includes proteins which are the downstream products of gene expression and metabolites which are products of biological reactions. Quantification and joint analysis of these molecules may identify the molecular determinants of the aberrant inflammation observed in COVID-19 cases leading to improved diagnostic and therapeutic methods . Pairwise comparisons (Dunn’s test with Benjamini–Hochberg adjustment) revealed that the age distribution was significantly different between the severe and healthy groups   , and health outcome and BMI   and also by various health outcomes. However, sex seems to have the lowest effect on overall metabolites and protein profiles compared to other clinical variables  Fig. a, and thWe identified important proteins Fig. a and metp-value = 9.7E-18) groups. This finding is consistent with a similar study analyzing metabolite profiles of COVID-19 patients, which found cytosine to be the distinguishing feature that determined infection status17. It is hypothesized that changes in levels of cytosine are critically involved with RNA virus evolution, including SARS-CoV-218. Notably, the underrepresentation of cytosine within the SARS-CoV-2 genome suggests an alternative role for this metabolite beyond the synthesis of viral RNA. While it is unclear why cytosine levels are higher in COVID-19 patients, this finding points towards cytosine as an effective biomarker for COVID-19 infection.Viruses have adapted over time to hijack cellular machinery and resources for their own replication. Consequently, homeostatic synthesis and recycling of nucleobases may be disrupted in favor of producing new copies of the viral genome. We found that cytosine levels are elevated in the COVID-19 groups as compared to the non-COVID-19 Fig. b and hea19. Circulating uridine levels are typically high in healthy individuals but tend to undergo short term fluctuations in response to diet. Changes in synthesis are tightly regulated by the liver along with the adipose tissue21. Abnormal liver function appears to be a commonality between SARS-CoV23 and SARS-CoV-2 infection29. Therefore, liver impairment may mediate the lower uridine levels observed in COVID-19 patients. The functional consequences of this reduction are also unclear. In an animal model of pulmonary fibrosis, it was shown that uridine supplementation has anti-inflammatory and anti-fibrotic effects30. While additional studies are needed, a similar therapeutic strategy may mitigate prolonged inflammation within the lungs that leads to eventual injury and disruption of the epithelium31.Our results indicate that uridine levels are lower in the COVID-19 groups as compared to the healthy and non-COVID-19 groups. Uridine is a biologically dynamic metabolite that is critical to the synthesis of RNA and glycogen32, is depleted in the severe  and the non-severe COVID-19 groups  compared to the healthy group . BCAAs play a critical role in protein anabolism38, lowered BCAA levels are often observed in various conditions, including liver cirrhosis39, urea cycle disorders40, chronic renal failure42, and impaired renal function43. Thus, our finding is congruent with the impaired renal function observed in severe COVID-19 cases44.Levels of branched-chain amino acids (BCAAs) leucine, isoleucine, and valine are lower in the COVID-19 groups Fig. d as compcoefficient = 4.5, p-value = 8.5E-40) and non-COVID-19  groups compared to the healthy group  in the gastrointestinal tract68 and in mice models of liver injury and dysfunction71. The lower levels of PCYOX1 protein seen in the COVID-19 groups may contribute to the liver dysfunction associated with COVID-1972.Prenylcysteine oxidase 1  levels are heavily depleted in the severe COVID-19 group as compared to the other groups. HRG is a plasma protein that is involved in many biological processes, including immune system regulation, cell adhesion, angiogenesis, and coagulation94. It is associated with inflammation and can display either pro-inflammatory or anti-inflammatory effects95. However, the anti-inflammatory effects of 15-HETE are more well-studied and include inhibition of leukotriene B4 action on polymorphonuclear neutrophils (PMN)96 and the migration of PMN in response to cytokines97. We found severe depletion of 15-HETE levels in the COVID-19 groups compared to the non-COVID-19 and healthy groups. The depletion of 15-HETE and subsequent loss of anti-inflammatory signals could contribute to the heightened inflammation seen during COVID-19 infection98. Interestingly, 15-HETE also plays a role in promoting pulmonary vascular remodeling during hypoxia by exerting pro-angiogenic effects100. A reduction in serum 15-HETE levels would suggest that this metabolite does not directly contribute to the cardiovascular dysfunction observed in severe COVID-19 cases.15-HETE is a metabolite produced when arachidonic acid is oxygenated by arachidonate 15-lipoxygenase102. This has primarily been attributed to prolonged overactivation of platelets and high levels of neutrophil degranulation, both of which represent significantly enriched pathways in our analysis of this group. Increases in platelet mediated prothrombosis are typical in response to many invading pathogens. However, SARS-CoV-2 infection in particular triggers significant changes in platelet gene expression and aggregation. Furthermore, excessive platelet activation from COVID-19 leads to alterations in innate immune responses that contribute to thrombotic events. This includes the accumulation of platelet-monocyte complexes, which directly express high tissue factor levels, which directly increases the risk of clotting103. High plasma levels of platelet factors also contribute to dysregulated neutrophil responses, such as the excessive formation of neutrophil extracellular traps and degranulation105. In addition to their direct detrimental role to cardiovascular health, these factors may also exacerbate systemic inflammation leading to damage within the tissues.To date, severe COVID-19 has been associated with an increase in the immediate and long-term risk of thrombosis and coagulation abnormalities106. Sustained levels of extracellular ATP trigger the expression of ectoenzymes on the surface of immune cells that convert ATP to adenosine. Binding of adenosine to cognate purinergic receptors tempers inflammation by reducing neutrophil chemotaxis and platelet aggregation in addition to promoting wound healing via the release of vascular endothelial growth factor (VEGF) by macrophages and dendritic cells107. Under hypoxic conditions, hypoxia inducible factors HIF-α and HIF-β can alter adenosine metabolism in order to protect tissue from further damage brought about by prolonged inflammation108. Acute lung injury that occurs during severe cases of COVID-19 may trigger such pathways leading to the observed enrichment in purine metabolism.Pathway level analysis may also be beneficial for identifying overarching mechanisms that have a role in regulating rampant inflammation. Collectively, the purine metabolism pathway is significantly enriched within the COVID-19 groups compared to the healthy control Fig. b. Cell-f109. In mice models, it has been found that lumican is critical for host immune innate response111, and its deficiency has been associated with cardiomyocyte hypertrophy112. In a study of Nepalese children, lumican levels were negatively associated with levels of α-1-acid glycoprotein113, an acute phase protein that increases during inflammation, infection, or injury to tissues114. We found that lumican levels were depleted in the COVID-19 groups compared to the non-COVID-19 and healthy groups and is also associated with some glycerophospholipids in block-wise association testing. Thus, lower lumican levels may act as an additional biomarker of rampant inflammation in the COVID-19 groups. Alternatively, lower lumican may lead to disruption of the collagen and fibril assembly pathways as a consequence of infection.Lumican (protein ID P51884) regulates fibril assembly and stromal collagen matrix assembly116. Identification of additional biomarkers could facilitate diagnosing the severity of lung injury prior to the induction of respiratory failure. Our analysis identified significant alterations in processes that maintain cell or tissue structure, including enrichment of proteins involved in the keratinization pathway  matching previous studies125. While the direct impact of COVID-19 infection on glucose levels remains to be elucidated, inflammation may be responsible for the differences in glucose levels observed between the groups. Okin and Medzhitov found that sustained inflammation can lead to elevated glucose levels in the plasma126. Alternatively, IFN-gamma production in response to viral infection has been shown to induce insulin resistance127 and subsequent higher glucose levels.128 and increased levels of citrulline in the plasma is associated with a reduction in glucose production129. Regardless of the biological significance, the correlation of uridine and citrulline with glucose points towards these metabolites being modest candidates for COVID-19 severity biomarkers.In addition to its association with COVID-19 severity, we also found some correlations between glucose and metabolites such as citrulline and uridine. Both of these metabolites were severely depleted in the COVID-19 groups compared to the non-COVID-19 group. While it is unclear if there is a biological connection to these relationships, the metabolism of glucose has been linked with the metabolism of uridine and citrulline. Specifically, uridine has been shown to induce glucose uptake by skeletal musclesC-Reactive Protein (CRP): We found CRP levels to be lower in the non-severe COVID-19 group compared to the non-COVID-19 group , whereas an opposite trend appears when compared to the severe COVID-19 group. This reinforces what has been established in previous studies133. CRP plays a critical role in inflammation and response to infection via the complement pathway and cytokine production136. Thus, our finding of increased CRP levels in the severe group is in agreement with previous studies that suggest elevated inflammatory markers including procalcitonin, D-dimer, and lactate dehydrogenase137 are associated with COVID-19 disease severity.139. Additionally, within COVID-19, kynurenine has been found to be positively correlated with proinflammatory cytokines141, and activation of the kynurenine pathway has been observed in COVID-19 patients142.Similar to glucose, CRP has some metabolic and protein correlates which may be able to serve as novel biomarkers for COVID-19 severity. Specifically, CRP is positively correlated with kynurenine and lipopolysaccharide-binding protein (LBP) SFig. . Kynuren143. Additionally, previous studies have shown that LBP is associated with inflammation markers, including CRP, in patients who have undergone hemodialysis144 and in patients with conditions such as acute respiratory distress syndrome and inflammatory bowel disease146.The correlation between CRP and LBP is likely a product of these molecules’ role in inflammation. LBP has been shown to be increased in patients with inflammatory conditions like systemic inflammatory response syndromeMonocyte: We found monocyte levels to be significantly decreased in the severe COVID-19 group . Based on a meta-analysis of COVID-19 studies involving severe and non-severe patients, lower monocyte counts have been observed as part of a larger trend of immune dysregulation147.Salicylate: When investigating associations across all groups, we observed a positive relationship between Monocyte counts and Salicylate . Salicylate is commonly found in non-steroidal anti-inflammatory drugs, including aspirin. Sodium salicylate has been shown to have potent effects on limiting monocyte migration, expression of inflammatory cytokines, and preventing proliferation150. Whether or not there is a prophylactic benefit to administering aspirin to limit COVID-19 infection is currently the subject of debate153. A positive association between salicylate and monocyte counts seems to indicate that other factors, either directly related to infection or other forms of treatment, are responsible for the decrease in monocytes observed in patients with severe COVID-19.Sphingomyelin: An additional positive correlation was found between monocyte counts and sphingomyelin . Sphingolipids are a class of membrane-associated molecules that play an important role in cell-to-cell interactions and intracellular signaling within the immune system154. Sphingomyelin is cleaved by sphingomyelinases to produce ceramide which induces a signaling cascade that can lead to the differentiation, proliferation, apoptosis, or cytokine secretion by select immune cell populations. It was recently shown that neutral sphingomyelinase 2 can cause monocyte migration and secretion of inflammatory cytokines in response to soluble TNF-α 155. Elevated TNF-α has been associated with both obesity and severe cases of COVID-19156. Most of the COVID-19 biomarkers and their related pathways reported in our study are novel, and some have already been discussed previously157 . Block 20 in particular shows that 5-methyluridine, citrulline, choline, and uridine are jointly associated with the C4b-binding protein chains  and Vitamin-K dependent protein S. Inflammation may explain the observed association between the proteins and most of the metabolites in block 20. The C4b-binding protein is involved with the inhibition of the classical and lectin complement system pathwaysWe used four machine learning (ML) approaches for disease severity prediction, including deep neural networks (DNN), k-nearest neighbors (KNN), Random Forest (RF), and Logistic Regression (LR) SFig. . We haveDNN outperformed all other methods with an accuracy score of 81.78% when trained using the metabolomics data and clinical data . The other ML processes that were used had accuracy in the range of 60–70%. A higher F-1 score suggests better efficiency of models, which in turn means that the number of false positives is less, thus better prediction.Integrative analysis of multi-omics enables a more accurate and comprehensive understanding of biological activities at the molecular level. Utilizing omics data can provide finer resolution for identifying the specific molecules or processes that distinguish severe cases of COVID-19. This analysis holds the potential to not only improve our understanding of COVID-19 pathogenesis, but may also lead to improved diagnostic and therapeutic avenues. In this study, we used protein and metabolite profiles from a cohort of donors with varying COVID-19 and health statuses to measure the changes across groups after appropriate adjustment for data properties such as zero inflation and confounding factors. We followed up with pathway enrichment analysis to provide context at a functional level and then investigated associations between significantly different omics features  and clinical information (metabolic panel and complete blood count). Lastly, we utilized our findings to investigate the predictive potential of several machine learning algorithms benchmarked against conventional logistic regression to determine which model is best suited for diagnosing the status of COVID-19 infections.162.Phenotype association testing revealed significantly altered proteins and metabolites based on health status  by searching for the Project IDs: IPX0002106000 and IPX0002171000. Before feeding the publicly available dataset12 into machine learning algorithms, various data cleaning steps were taken. The steps included removing any columns of the 942 metabolites and 640 proteins with missing values across samples and imputing missing BMIs for healthy individuals based on the optimal BMI of Chinese people167. We removed other clinical information like platelet counts, etc., since they were only present for the unhealthy patients and not for the healthy subjects. Removing missing values resulted in a dataset that included 404 metabolites and 374 proteins. We also normalized the dataset based on the min–max function available in the sklearn package168. The cleaned and normalized dataset was split into training (80% of data) and testing (20% of data) subsets to train and test various prediction algorithms, including Logistic Regression, Random Forest, K-Nearest Neighbor, Decision Tree, and Deep Neural Network.The study’s aim was to predict health outcome status and discover important features using proteomics and metabolomics data and clinical information. Metabolomic and proteomic data from the original study were downloaded from ProteomeXchange Consortium , also provides an enrichment score for each metadata to measure the potential influence of each metadata on detected structure (clusters). omeClust first discretizes metadata; and then calculates enrichment score as normalized mutual information between cluster labels and discretized metadata.We applied We used multivariate association testing with considering noisy, sparse (zero-inflated), high-dimensional, and extremely non-normal data.omePath package6. omePath assigns an importance score  to each omics feature  and performs statistical tests (Wilcoxon sum rank) between rank of feature score in a given pathway against all ranks to calculate a p-value for the null hypothesis. There is no difference between the distribution of score of features with the pathways of interest vs. all other features in the study. We used an alpha level of 0.05 for significance. omePath is an open-source software implemented as an R package with code, tutorials, and documentation at https://github.com/omicsEye/omePath. The result for each association contains the identified pathway, members of the pathways, number of observed members in the study (n), and the total number of pathway members in the database (HMDB database for metabolite pathways120 and Reactome pathways database121 for proteins), used for the analysis, p-value, q-value from Benjamini–Hochberg FDR correction (q = 0.25).Enrichment analyses were performed using the 171. Many studies have been targeted to build and execute model-agnostic interpretability tools174. We use the term feature importance to explain how important a feature is to the model’s predictive performance. The most well-known approach is using permutation importance which was introduced by Breiman175. Using permutation importance, we have quantified the importance score of features for predicting health outcomes. The features consist of metabolomics, proteomics, and clinical info for all the patients considered in the study. The samples were labeled into four labels based on the health status; Healthy, non-COVID-19, COVID-19, and severe-COVID-19. We predicted the health status of a patient based on the various proteomics and metabolomics data and found the importance of each feature inside the prediction model such as beta-alanine and 15-HETE metabolites which both were ranked as top influential features in RF and DNN models  algorithms like neural networks are often considered to be a black box because of their inability to provide a simple and straightforward explanation of their predictions. Nevertheless, this prediction model generally exceeds simple linear models or decision trees and random forest predictions. Yet, such simple models are still preferred in the field of medical science due to their simplicity and interpretabilitysklearn package.Data from a real-world scenario are never flawless. The medical records and the clinical information for the patients affected by COVID-19 are no exception. The data in this study contained many values which were lost due to machine or human error. The data thus was cleaned of all the unknown values and also the null values by dropping the instance with the missing feature value. Serum panel data tends to have a significant difference between the maximum and minimum value, to rectify this issue, we used normalization. Normalization is a scaling-down transformation in data where the difference between min and max values is significantly big. The data were normalized using a min–max scaler function which is present in the Decision Tree follows a flow-chart-like structure where the nodes are the features, the branches are the decision rules, and the leaves are the outcomes. Decision Tree is a supervised learning method that utilizes a divide and conquer approach; it selects the best attribute using Information Gain and then divides the dataset into a subset. This division is performed repeatedly until the method reaches a child node which satisfies the condition of no remaining attributes or no more remaining instances.176. Then, the mode of the labels of K- nearest neighbors using the sorted table is given as an output.K-Nearest Neighbor (KNN) is a supervised machine learning technique that is dependent on the training dataset. The K, in KNN, stands for a user-defined number. This algorithm assumes that data points with similar features reside in close proximity to each other. Proximity is generally calculated in the form of euclidean distances among points. In a classification problem such as ours, the distances between the test data points and the training data points are calculated, sorted, and stored in a tableRandom Forest is a supervised algorithm that randomly selects a subset of the training dataset and creates a decision tree on the subset; it then carries out a vote to predict the class of the test data points.177. Logistic regression uses a functional approach to estimate the probability of binary response based on input features. LR finds the best-fit parameters to a nonlinear function called sigmoid178. Logistic regression models probability for a binary class, however, our health outcome variable has more than two classes. To address the binary class limitation of logistic regression, we used a ‘newton-cg’ solver. In our study, we use logistic regression as a baseline for the other methods.A predominant part of published propensity results uses Logistic Regression (LR). Logistic regression is a very sought after technique because of its mathematical ability to produce probability in the range 179. The inputs of DNN are fed in the input layers, which are passed through one or more hidden layers, which consist of neurons, where they are analyzed and processed to determine the output of the next layer. DNN uses a learning rule which correctly decides the weight and the bias of each neuron in the hidden layer and output layer. The power of DNN to determine and adapt the weight and bias dynamically makes it a powerful tool to capture the various complex and non-linear relationships among the various features, which in turn facilitates classification and prediction of correct labels, thus increasing the accuracy and efficiency of the model181.Deep neural network (DNN) is a type of machine learning architecture that mimics the working of the neurons located in the brain, and how they transfer information to learn new problems for the purpose of solving themWe have incorporated the full extent of the data since some were discarded due to missing values. Including a more extensive set of data and features in deep learning, the model brings out a more comprehensive hidden complex relationships among all the proteins and metabolites. This enables a more accurate prediction and prioritization of metabolites and proteins for further studies to show how they affect a patient's health status. The importance scores for metabolites and proteins generated by the model are based on their degree of influence on the result.The only ML model which was used in the above said paper was Random Forest (RF). Still, in our evaluation, we used decision tree (DT), k- nearest neighbors (KNN), random forest (RF), logistic regression (LR), and deep neural network (DNN). We have made a thorough comparison of all the methods using various metrics like accuracy, precision, recall, and F-1 score to evaluate the performance of each model. The accuracy of DNN comes higher than all other methods that were considered. We then used the DNN model for importance evaluation that leads to the discovery of numerous metabolites and proteins which when done, a thorough study shows a relationship with covid and health status.The different evaluation metrics used in machine learning section are as follows:Supplementary Information 1.Supplementary Information 2.Supplementary Information 3.Supplementary Information 4.


Can untrained persons correctly perform and interpret the results of SARS-CoV-2 rapid antigen detection tests (RADT), and can performance be optimized?In this cross-sectional study of 278 participants self-performing SARS-CoV-2 RADT in an intended-use setting, the accuracy of RADT interpretation was poor when the manufacturer’s instructions were used. A modified quick reference guide was associated with significantly better user performance.These findings suggest that longitudinal mass RADT testing for SARS-CoV-2 could be accurately self-performed in an intended-use setting but there are potential interventions to optimize performance. This cohort study assesses the execution and interpretation of SARS-CoV-2 rapid antigen detection tests when performed by untrained persons an intended-use setting. Longitudinal mass testing using rapid antigen detection tests (RADT) for serial screening of asymptomatic persons has been proposed for preventing SARS-CoV-2 community transmission. The feasibility of this strategy relies on accurate self-testing.To quantify the adequacy of serial self-performed SARS-CoV-2 RADT testing in the workplace, in terms of the frequency of correct execution of procedural steps and accurate interpretation of the range of possible RADT results.This prospective repeated cross-sectional study was performed from July to October 2021 at businesses with at least 2 active cases of SARS-CoV-2 infection in Montreal, Canada. Participants included untrained persons in their workplace, not meeting Public Health quarantine criteria . Interpretation and performance were compared between participants who received instructions provided by the manufacturer vs those who received modified instructions that were informed by the most frequent or most critical errors we observed. Data were analyzed from October to November 2021.RADT testing using a modified quick reference guide compared with the original manufacturer’s instructions.The main outcome was the difference in correctly interpreted RADT results. Secondary outcomes included difference in proportions of correctly performed procedural steps. Additional analyses, assessed among participants with 2 self-testing visits, compared the second self-test visit with the first self-test visit using the same measures.Overall, 1892 tests were performed among 647 participants, of whom 278 participants  had at least 1 self-testing visit. For self-test visit 1, significantly better accuracy in test interpretation was observed among participants using the modified quick reference guide than those using the manufacturer’s instructions for reading results that were weak positive , positive , strong positive , and invalid . Use of the modified guide was associated with improvements on self-test visit 2 for results that were weak positive , positive , and invalid . For procedural steps identified as critical for test validity, adherence to procedural testing steps did not differ meaningfully according to instructions provided or reader experience.In this cross-sectional study of self-performed SARS-CoV-2 RADT in an intended-use setting, a modified quick reference guide was associated with significantly improved accuracy in RADT interpretations. However, several factors impede this deployment. These include regulatory requirements that RADT be performed by trained health care workers in some countriesIn this study, we report the results of a prospective field evaluation of serial self-performed RADT for SARS-CoV-2 among untrained, asymptomatic persons in their workplaces. The primary objective was to quantify the adequacy of self-testing, in terms of the frequency of correct execution of procedural steps and the accuracy of interpretation of the range of possible RADT results. We also aimed to compare results using the instructions provided by the manufacturer with results using modified instructions that were informed by the most frequent or most critical errors we observed.STROBE) reporting guideline for cross-sectional studies.This cross-sectional study was approved by the Research Ethics Board of the Research Institute of the McGill University Health Centre. All participants provided written informed consent. The study followed the Strengthening the Reporting of Observational Studies in Epidemiology . Study data were collected and managed using REDCap.11In this prospective, repeated, cross-sectional study, businesses in Montreal, Canada, with at least 2 active cases of SARS-CoV-2 infection within a 14-day period were identified by the Montreal Department of Public Health. Businesses with newly identified outbreaks were contacted by the study team by telephone and offered participation in the study via the deployment of 1 of 4 mobile teams on-site. Among outbreak businesses, priority was given to contacting those with greater than 50 employees to increase the probability of identifying participants with unsuspected SARS-CoV-2 infection.Any worker in outbreak businesses who could provide written informed consent was considered eligible to participate. Participants were not compelled by the study team or by their employer to attend all 4 study visits and could withdraw from further testing if they no longer wished to participate.Data on self-reported participant age, gender, race and ethnicity, language preference (English or French), COVID-19 symptoms, exposures, and vaccination were collected. Race and ethnicity were self reported and categorized as Asian, Black, Hispanic, Indigenous, mixed, White, and other . Race and ethnicity and other sociodemographic measures were recorded to account for the heterogeneity of SARS-CoV-2 community transmission. Additionally, we assessed testing results, adequacy of performance of test procedures, and result interpretation.The RADT used was the Panbio COVID-19 Ag Rapid Test Device (Abbott Laboratories). At each visit, trained study personnel instructed participants on procedures to perform self-collected nasal midturbinate swab specimens. On visits 1 and 3, all postspecimen collection steps and result interpretations were performed by study personnel. On visits 2 (first self-testing visit) and 4 (second self-testing visit), these steps were performed by the participant. During self-testing, study personnel assessed 13 RADT procedural steps performed by the participant as correct or incorrect. If any of 6 steps deemed to be critical to testing integrity were assessed as incorrectly performed, the participant was informed at the end of the procedure and testing was redone by study personnel. Participants with a positive RADT result were offered confirmatory laboratory-based polymerase chain reaction (PCR) testing at a local testing center.A proficiency panel of 7 RADT test results was created using serial dilutions of the positive control reagent and aimed to evaluate participants’ ability to interpret the range of possible signal intensities of the test line, including results classified as weak positive, positive, strong positive, invalid, and negative  proceeded until the overall weighted mean of the accurate readings could be estimated with 6.0% precision at the 95% confidence level. A modified quick reference guide informed by the analysis of phase 1 data was expected to yield at least a 12.0–percentage point improvement in RADT reading accuracy vs the manufacturer's instructions. Based on an expected overall weighted mean of the accurate readings of 60.0%  in phase 1, the number of readings required to achieve an accuracy of 72.0% with 6.0% precision was 216 readings per result category . At the rate of 2 readings per person per visit, this corresponds to a recruitment of 108 people using the modified instructions.sensitivity to refer to the proportion of accurately identified RADT result interpretations of positive and of invalid results; specificity referred to accurate RADT result interpretation of negative results.Baseline characteristics were summarized according to self-testing visit and reference guide type. Estimates of 95% CIs around a proportion or around the difference of 2 proportions were calculated according to a binomial distribution using the Wilson Score method. We used the term Generalized linear mixed-effects models were fit to evaluate the association of the use of the modified quick reference guide with accurate RADT proficiency panel result interpretation. Outbreak business and participant were modeled as a random intercepts; participant age (in years), gender, RADT result type  and an interaction term between RADT result type and instruction type were included as covariates (fixed effects).Data were analyzed using R statistical software version 3.5.2 . Because of the small number of variables to be analyzed and because missingness was no greater than 5%, a complete case analysis was performed for the modeling. Statistical significance was assessed by using 2-tailed tests, with α = .05. Data were analyzed from October to November 2021.From July 7, to October 8, 2021, 236 outbreak businesses were identified ; 168 busAt the first self-test visit, significantly better accuracy in interpreting proficiency panel RADT results was observed among participants using the modified quick reference guide compared with those using the manufacturer’s instructions for results that were weak positive , positive , strong positive , and invalid . Use of the modified guide, compared with using the manufacturer’s instructions, was associated with statistically significant improvements on self-test visit 2 for results that were weak positive , positive , and invalid  .In multivariable models accounting for outbreak business and participant age and gender, the modified quick reference guide was associated with higher accuracy in result interpretation for self-test visit 1  but not for self-test visit 2 . For procedural steps identified as critical for the validity of test results, adherence did not differ meaningfully according to instructions provided or reader experience .13 but was not associated with improving the proper execution of procedural steps. Repeated self-testing was associated with improved signal interpretation, although the difference was less apparent with modified instructions, as initial accuracy was already high. These results complement recent data on how consumers act on the results of at-home COVID-19 self-test kits.14In this prospective cross-sectional study of self-performed SARS-CoV-2 RADT in an intended-use setting, the accuracy of signal interpretation was proportional to test-band intensity and was extremely low for tests with weak bands. An evidence-based modified quick reference guide improved signal interpretation by untrained workers to a level of accuracy similar to that of professional technologists for other RADTs15 In several studies of malaria RADT, weak positive test bands consistently have correlated with lower sensitivity.15 Our study agrees with these findings for SARS-CoV-2 RADT, and provides evidence for a simple and effective potential corrective intervention.Self-testing using RADT has been studied for other infections. Among users attending a pretravel clinic, only 70.6% of all malaria RADT strips were correctly interpreted by untrained users.It is notable that during the study period, Health Canada had not yet authorized self-performance of RADT by nonprofessionals. Because of this, the instructions for use of the RADT used in this test and all other RADT sold in Canada during this period stated for professional use only. These instructions may differ from those for the same products sold in other countries. Also, while the RADT used in this study is not available in the United States, several comparable products are sold on the US market.Study strengths include the recruitment of participants from diverse racial and ethnic, socioeconomic, and educational backgrounds. The modified quick reference guide was informed by the most frequent or most critical errors observed and validated with different participants. Further, we assessed the independent outcomes associated with modified instructions and repeated self-testing.This study has some limitations. One limitation was the small proportion of eligible businesses that entered the study. This proportion depended on several factors other than the agreement of the businesses to participate, such as our team’s capacity  and the decision not to attend businesses that agreed to participate but where the last recorded COVID-19 case was more than 14 days earlier than the day our teams could visit. Additionally, a single RADT was assessed in this study, with uncertain generalizability to the full diversity of available RADTs.17 This use case may become more pertinent with the emergence of new SARS-CoV-2 variants of concern.This cross-sectional study found that longitudinal mass SARS-CoV-2 RADT testing could be accurately self-performed and provides evidence for a potential intervention to optimize performance. Self-administered RADTs are inexpensive and can be decentralized and implemented at scale for serial mass testing of asymptomatic persons to break chains of transmission and reduce SARS-CoV-2 incidence.


When exposed to UV light, this compound can undergo a photochemically induced trans/cis isomerization and a 6π electrochemical cyclization with the subsequent formation of 2,4,6-trihydroxyphenanthrene (THP). THP is a potentially harmful compound which can exert genotoxic effects. In this work we improved the chromatographic separation and determination of the two resveratrol isomers and of THP by using a non-commercial pentafluorophenyl stationary phase. We assessed the effect of natural deep eutectic solvents (NaDES) as possible photo-protective agents by evaluating cis-resveratrol isomer and THP formation under different UV-light exposure conditions with the aim of enhancing resveratrol photostability and inhibiting THP production. Our results demonstrate a marked photoprotective effect exerted by glycerol-containing NaDES, and in particular by proline/glycerol NaDES, which exerts a strong inhibitory effect on the photochemical isomerization of resveratrol and significantly limits the formation of the toxic derivative THP. Considering the presence of resveratrol in various commercial products, these results are of note in view of the potential genotoxic risk associated with its photochemical degradation products and in view of the need for the development of green, eco-sustainable and biocompatible resveratrol photo-stable formulations. The trans- form can be converted into the cis- form by exposure to UV-light irradiation or direct sunlight [trans-resveratrol, further reactions may occur other than the trans/cis isomerization, and different chemical species can be formed that can have a different biological activity than trans-resveratrol. As a matter of fact, the trans-resveratrol photodegradation pathway includes quinonoid and reactive radical species, whose identification is often elusive to spectroscopic and spectrometric investigations [cis- form can undergo a 6π photochemical electrocyclization with subsequent oxidation to form a phenanthrenoid product previously characterized in our laboratory as 2,4,6-trihydroxyphenanthrene -electron conrotatory pericyclic ring closure with subsequent oxidation and generation of the fully aromatic tricyclic system of THP [Resveratrol  demonstrated that THP can induce DNA damage through a pro-oxidant mechanism, indicating a genotoxic risk even at very low micromolar concentrations . trans-resveratrol by using different formulations and stabilizing agents [trans-resveratrol in terms of degradation and concomitant THP formation after UV-light exposure [trans-resveratrol photostability by applying an innovative green chemistry approach via the use of natural deep eutectic solvents (NaDES) and to inhibit the conversion into the cis- isomer and subsequent THP formation.Several strategies have been used in the past years to improve the stability of g agents ,12. To texposure ,17,18,19trans-resveratrol subjected to UV-light irradiation  and a hydrogen bond donor  in a solid (or solid/liquid) state that associates by hydrogen bonding ,21,22. Tadiation . trans-resveratrol in different biological and pharmaceutical matrices are present in the literature [cis-resveratrol is more complicated due to its low stability and the lack of an analytical standard. However, it is possible to quantify this compound by the induced photo-generation of the cis-standard in situ, starting by irradiating the trans-isomer and analyzing the sample in the timeframe in which only the isomerization process occurs [trans-resveratrol, cis-resveratrol and THP by using a non-commercial sub-3 µm pentafluorophenyl (PFP) silica-based stationary phase. This method was applied to assess the capacity of NaDES to act as trans-resveratrol protecting agents against photo-chemical induced degradation, with a particular focus on the inhibition of THP formation.The determination of resveratrol and its photoconversion products was improved by the use of a novel chromatographic method, employing for the first time a non-commercial reverse phase column. Several chromatographic methods for the quantification of terature ,26,27,28s occurs ,29,30,31s occurs . In the trans-resveratrol, cis-resveratrol and THP) using the PFP stationary phase is shown in trans-Resveratrol standard solution was analyzed before and after one hour of direct sunlight exposure.A representative chromatographic separation of the three compounds of interest  present the same pseudo-molecular ion in their mass spectrum at 227 m/z [The black chromatogram represents the chromatographic profile of a  227 m/z ,31 .THP is formed via photo-induced trans-isomer UV-light irradiation. The use of the pentafluorophenyl stationary phase offers better results with respect to common C18 reverse-phase columns reported in the literature [cis-resveratrol from the phenanthrenoid species THP (α = 1.32). The better selectivity of pentafluorophenyl stationary phase towards these two compounds could arise from the stronger π–π interaction that the tricyclic THP system could exert with the aromatic moiety of the perfluorinated phenilic stationary phase. This optimized chromatographic method offers a complete resolution of the three photochemical products derived from the terature . One of 3CN) and avoiding column washing and re-equilibration.In our previous work, we described the HPLC analytical determination and semi-preparative purification of THP on a C18 reverse phase column . In thistrans-resveratrol UV-light-induced photochemical conversion.trans/cis photo-induced isomerization process is strongly inhibited when trans-resveratrol is irradiated in the presence of glycerol containing NaDES. In particular, ProGlc and BetGlc were revealed to be the best solvents in terms of isomerization inhibition. The control solvent (DMSO) displays a similar behavior to that of propylene glycol containing NaDES with a rapid conversion of trans- into cis- isomer within the first 30 min of analysis. ProGlc and BetGlc showed a pronounced effect also on THP production over two hours of UV-light exposure. The photo-induced production of THP turned out to be less than 5% in the presence of ProGlc NaDES, while it gradually increased in the other NaDES and more than 20% in the control solvent after two hours of UV-light exposure. The trans-resveratrol dissolved in two glycerol-containing NaDES (ProGlc and CholGlc) and in the control solvent (DMSO).Given the results obtained, three representative samples were exposed to direct sunlight in order to verify the photo-conversion trend in a system that was much closer to environmental conditions. trans-Resveratrol exposure to direct sunlight radiation results in a rapid almost complete isomerization into the cis-form within one hour of irradiation and up to 30% THP conversion after two hours. In the presence of both NaDES, these values are significantly reduced, particularly when trans-resveratrol is dissolved in ProGlc eutectic solvent. Notably, after two hours of exposure in ProGlc, the cis/trans isomerization was remarkably lower than in the control sample and THP formation was around 2%, i.e., more than ten times lower than control in which was around 30%. The CholGlc solvent also showed an important photoprotective effect, with only a 60% trans-resveratrol conversion after two hours of exposure and less than 15% of THP production.The results are in good agreement with the previous data obtained by the UV-lamp irradiation. trans-resveratrol and to correlate it with the photoisomerization and cyclization trend, the absolute amount of resveratrol (mg/mL) in the two different conditions after 2 h irradiation was calculated. In trans-resveratrol in DMSO and ProGlc is reported. As shown in the table, the absolute amount of trans-resveratrol present under the two conditions is significantly higher in the presence of ProGlc NaDES.Photochemical isomerization and electrocyclization/oxidation are not the only processes that can occur during resveratrol irradiation. To verify the protective effect on trans/cis-resveratrol and THP) was assessed to be ≥90% of the starting amount of irradiated material. Our data confirmed the photo-protective effect of ProGlc NaDES on trans-isomer photo-chemical degradation and evidenced that isomerization and electrocyclization processes represent one the main photochemical degradation pathways under irradiation.In all experimental conditions, the total amount of the three compounds  and more probably to hydrogen bond donor effect of resveratrol in the eutectic system with respect to common solvents. It is interesting to note how glycerol and propylene glycol differ only for one hydroxyl moiety. The photoconversion kinetics of resveratrol in common organic solvents including alcohols such as ethanol or propanol are very similar to those observed in DMSO. The observed photo-stabilizing effect of glycerol-containing NaDES could be unrelated to the polarity of the solvent or to the solvation of the molecule itself, instead being related to the ability of the resveratrol to make hydrogen bonds in the eutectic solvent and in this way to maintain a more rigid conformation that increases the isomerization energy barrier [The inversion of isomerization equilibrium could be probably unrelated to the physico-chemical properties of the solvents  but in part may be due to solvation . All other chemicals and solvents were of analytical grade and were from Sigma-Aldrich . THP was photochemically synthesized and purified in our laboratory as previously described .NaDES were prepared by mixing two components at 70 °C in a water bath under magnetic stirring for 1 h. Six different NaDES were prepared with specific molar ratios after empirical experimental optimization. Choline/glycerol (1:1.5 molar ratio) (CholGlc); proline/glycerol (1:2 molar ratio) (ProGlc); proline/propylene glycol (1:4 molar ratio) (ProProp); betaine/propylene glycol (1:4 molar ratio) (BetProp); betaine/glycerol (1:2.2 molar ratio) (BetGly); choline/propylene glycol (1:3 molar ratio) (CholProp).trans-Resveratrol was dissolved at a concentration of 10 mg/mL both in DMSO and in the NaDES. All the solutions were extensively sonicated in an ultrasonicator bath to allow the complete dissolution of the molecule, and were carefully protected from direct light. After samples preparation, 0.5 mL of each solution was spread on a glass slide and exposed to UV-lamp irradiation  or to direct sunlight (solar light spectrum) for different times. At each time point, a known amount of each sample was diluted in 30% aqueous acetonitrile at a final concentration of 0.2 mg/mL for subsequent chromatographic analysis.3CN. The flow rate was 0.8 mL/min, the column temperature was set at 25 °C and the elution was performed isocratically with 30% B. Samples were diluted in the mobile phase and injected through the needle. The PDA detector was set up in the range 200 to 600 nm. Mass spectrometric detection was performed in the negative electrospray ionization mode, using nitrogen as the nebulizer gas. Analyses were performed in the Total Ion Current (TIC) mode with a mass range of 100–500 m/z. The capillary voltage was 0.8 kV, cone voltage 15 V, ion source temperature 120 °C and probe temperature 600 °C. Quantification of each compound was performed by using standard calibration curves in the range of 0.02–5 nmoles.UPLC-DAD/MS was performed on a Waters Acquity H-Class UPLC system , including a quaternary solvent manager (QSM), a sample manager with a flow through needle system (FTN), a photodiode array detector (PDA) and a single-quadruple mass detector with electrospray ionization source (ACQUITY QDa). Chromatographic analyses were performed on a non-commercial pentafluorophenyl column  developed and prepared by Ciogli and co-workers . The mobtrans-Resveratrol and THP were quantified via UPLC-PDA (254 nm) by using standard calibration curve of the pure compounds. Cis- isomer standard was generated photochemically as reported by Francioso and co-workers by exposing hydro-alcoholic solutions of trans-resveratrol to UV light . The identity of the compounds was confirmed by chromatographic retention properties, UV-visible and MS spectra. The conversion rate was estimated by differential calculation between quantified trans- isomer after the treatment with UV light . The calibration curves were obtained by plotting the peak-area ratio of each analyte versus its concentration . The relative percentage of each compound in the reaction mixture was calculated by expression as 100% of the sum of three compounds present at each time of UV-light exposure. Repeatability of the analysis for the three compounds was assessed as lower than 5% expressed (RSD) for standard solutions and irradiated samples.trans-resveratrol from its photochemically induced conversion and transformation products. In particular, the method is highly powerful for the determination of resveratrol isomers and THP phenanthrenic photoproducts. THP is a polycyclic aromatic hydrocarbon derived from trans-resveratrol photo-chemical isomerization and subsequent electrocyclization followed by oxidation. This product was isolated and identified for the first time in our laboratory, and its chemical structure was elucidated by means of spectroscopic and spectrometric techniques [This paper describes an improved liquid chromatographic method for the separation of chniques . This phchniques .trans-resveratrol exposure to UV light.Nowadays, resveratrol is widely used in cosmetic and pharmaceutical topical formulations that are often in contact with the skin and exposed to sunlight UV radiation. Despite its wide use and the many studies aimed at improving resveratrol photostability, to date there are no works in which the production of THP has been evaluated under conditions of different UV light exposure. We therefore applied our improved chromatographic method to evaluate the effect of NaDES on the photoconversion and THP production after trans/cis resveratrol isomerization process and a strong reducing effect on the production of the toxic derivative THP.Our results demonstrate a pronounced photoprotective effect exerted by some NaDES under different irradiation conditions (UV-lamp light and direct sunlight exposure). In particular, it was shown that NaDES containing glycerol as a hydrogen bond donor have a strong inhibitory effect on the photo-induced These data provide new insights for the development of green eco-friendly and biocompatible resveratrol photo-stable pharmaceutical formulations. Furthermore, this work will pave the way for future more in-depth studies on the interactions and molecular mechanisms behind the photochemical stabilization of resveratrol by natural deep eutectic solvents.