Ubiquitination of proteins is a post-translational modification process with many different cellular functions, including protein stability, immune signaling, antiviral functions and virus replication. While ubiquitination of viral proteins can be used by the host as a defense mechanism by destroying the incoming pathogen, viruses have adapted to take advantage of this cellular process. The ubiquitin system can be hijacked by viruses to enhance various steps of the replication cycle and increase pathogenesis. Emerging viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), flaviviruses like Zika and dengue, as well as highly pathogenic viruses like Ebola and Nipah, have the ability to directly use the ubiquitination process to enhance their viral-replication cycle, and evade immune responses. Some of these mechanisms are conserved among different virus families, especially early during virus entry, providing an opportunity to develop broad-spectrum antivirals. Here, we discuss the mechanisms used by emergent viruses to exploit the host ubiquitin system, with the main focus on the role of ubiquitin in enhancing virus replication. Ubiquitin is a 76-amino acid protein well known for its function in marking proteins for degradation by the proteasome, although multiple non-degradative functions are well established. Covalent ubiquitination depends on a series of enzymes, including the E1 ubiquitin-activating enzyme, an E2 ubiquitin-conjugating enzyme, and an E3 ubiquitin ligase . UbiquitViruses have evolved to antagonize the host immune response by interfering with host ubiquitin-dependent signaling pathways, or by hijacking the cellular ubiquitination machinery, to promote viral replication and pathogenesis. With the ongoing pandemic caused by the novel strain of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it is becoming critical to identify common steps used by different viruses to replicate in host cells that could potentially be targeted with broad-spectrum antivirals. Specifically, ubiquitination of viral proteins is emerging as an important host cellular process that is utilized by multiple viruses to promote their replication. In this review, we explore the various ways that emergent viruses exploit the host ubiquitin system, and we focus on recent literature highlighting the role of ubiquitin in enhancing virus replication .Viruses gain entry to the host cell through highly specific interactions with cellular surface receptors, although some viruses can also use less-specific mechanisms of entry via interactions with lipids or carbohydrates on the surface of the cell membrane . FollowiFurther evidence that flaviviruses in general may use a non-degradative ubiquitination step for uncoating of the virus genome comes from experiments with yellow fever virus (YFV). Similar to DENV, inhibition of the E1-activating enzyme also inhibited the initial round of YFV translation, and this required the function of the valosin-containing protein VCP/p97 , which i−/−Trim7 animals. A proportion of infectious viral particles released during replication contained ubiquitinated E, and ubiquitination on the E-K38 residue provided the virion the ability to interact with at least one potential cellular receptor, TIM-1, enhancing virus entry, replication and pathogenesis. In this case, ubiquitination of E not only functions in the early steps of virus entry, but also provides a mechanism of tissue tropism . Th. Th82]. The host E3-ubiquitin ligase TRIM6, along with the E2-ubiquitin conjugase Ube2K, synthesize unanchored K48-linked polyubiquitin chains that enhance the activation and kinase activity of IKKε . The unaOther henipaviral proteins block other steps of the IFN-I induction and signaling pathways. Similar to IAV and SARS- and MERS-CoVs, the accessory protein V (NiV-V) of NiV and other paramyxoviruses is capable of inhibiting TRIM25-dependent activation of RIG-I [To successfully replicate, viruses depend on specific factors expressed by the host cell. Many viruses use the ubiquitin system at different stages of their replication cycle, highlighting the importance of this cellular machinery for viral replication and spread. In this review, we provided an overview of the complexity of the interaction between emerging and highly pathogenic viruses such as SARS-CoV-2 and the ubiquitin system. Importantly, ubiquitination of structural proteins that are important in the process of virus entry is developing as a common non-degradative mechanism used by emergent viruses from different families, and could provide targets for broad-spectrum antivirals. One aspect that needs further investigation is how and when ubiquitinated proteins, or in some cases unanchored ubiquitin, are packaged in the infectious virions. Future studies should focus on identifying the specific steps and the subcellular compartments in which ubiquitination occurs to promote viral replication, and whether these processes are conserved through the different families of viruses. Identifying the specific factors of the host ubiquitin system that are commonly required for replication of different viruses could provide targets for broad-spectrum antivirals.


Growing evidence suggests that travelling waves are functionally relevant for cognitive operations in the brain. Several electroencephalography (EEG) studies report on a perceptual alpha-echo, representing the brain response to a random visual flicker, propagating as a travelling wave across the cortical surface. In this study, we ask if the propagating activity of the alpha-echo is best explained by a set of discrete sources mixing at the sensor level rather than a cortical travelling wave. To this end, we presented participants with gratings modulated by random noise and simultaneously acquired the ongoing MEG. The perceptual alpha-echo was estimated using the temporal response function linking the visual input to the brain response. At the group level, we observed a spatial decay of the amplitude of the alpha-echo with respect to the sensor where the alpha-echo was the largest. Importantly, the propagation latencies consistently increased with the distance. Interestingly, the propagation of the alpha-echoes was predominantly centro-lateral, while EEG studies reported mainly posterior-frontal propagation. Moreover, the propagation speed of the alpha-echoes derived from the MEG data was around 10 m/s, which is higher compared to the 2 m/s reported in EEG studies. Using source modelling, we found an early component in the primary visual cortex and a phase-lagged late component in the parietal cortex, which may underlie the travelling alpha-echoes at the sensor level. We then simulated the alpha-echoes using realistic EEG and MEG forward models by placing two sources in the parietal and occipital cortices in accordance with our empirical findings. The two-source model could account for both the direction and speed of the observed alpha-echoes in the EEG and MEG data. Our results demonstrate that the propagation of the perceptual echoes observed in EEG and MEG data can be explained by two sources mixing at the scalp level equally well as by a cortical travelling wave. This conclusion however does not put into question continuous travelling waves reported in intracranial recordings. Cortical travelling waves has been proposed as a potential mechanism supporting communication and computations in the brain . MathemaTravelling waves have been identified using both invasive and non-invasive EEG recordings. While invasive recordings provide a high signal-to-noise ratio, the intracortical electrodes cover a relatively small area of the brain. In contrast, non-invasive EEG provides whole-head coverage, but the signals are strongly affected by spatial mixing and noise. These discrepancies between the techniques introduce inconsistency across studies. First, the reported velocity of the travelling waves varies within a wide range, 0.1 – 10 m/s , suggestMany studies have reported travelling waves associated with endogenous brain rhythms that are not dependent on external stimulation . HoweverIn this study, we applied broadband visual stimulation to participants while we recorded the ongoing MEG. The perceptual echoes were estimated from the temporal response functions derived from the visual broadband input and the MEG sensors. We further applied source modelling as well as forward simulation to understand the neuronal sources explaining the propagation of the perceptual alpha-echoes.Twenty-four participants  with no history of neurological disorders partook in the study. Six participants were excluded from the analysis because the perceptual echo was at the noise level. The study was approved by the local ethics committee  and written informed consent was acquired before enrolment in the study. All participants conformed to standard inclusion criteria for MEG experiments. Participants had normal or corrected-to-normal vision.Two grating stimuli, 5.8 degrees of visual angle, were presented bilaterally on a projector screen, placed at 1.4 meters from the participants’ eyes . After 1The luminance of the stimuli was modulated using a broadband (i.e. noise with uniform distribution) flickering signal . The broPrior to the measurement, the head position of the participant was digitized using the Polhemus Fastrack electromagnetic digitiser system . The MEG data were acquired using a 306-sensor TRIUX Elekta Neuromag system , while the participant was sitting in an upright position (60°) in a dim light magnetically shielded room. The magnetic signals were bandpass filtered within 0.1 – 330 Hz with embedded anti-aliasing filters and sampled at 1000 Hz. Simultaneously with the MEG, we also acquired the coordinates of the pupil and the pupil diameter using an EyeLink eye-tracker .The broadband flickering (or tagging) signals were recorded using a custom-made photodetector  that was connected to a miscellaneous channel of the MEG system. This allowed us to acquire the tagging signal with the same temporal precision as the MEG data.Additionally, a high-resolution T1-weighted anatomical image  was acquired using 3-Tesla Phillips Achieva scanner.Since magnetometer signals are strongly affected by the noise, additional pre-processing methods are needed to attenuate external interference and sensor artifacts. To this end, we used MNE-Python tools . MNE-PytTo further suppress artifacts caused by the cardiac signal and blinks, we applied independent component analysis . Prior The artifact-free MEG time series were segmented into 6 s epochs; −2 to 4 s relative to the onset of the stimulus motion. Although ICA allowed removing blinks from the MEG signal, we additionally analysed the X-axis and Y-axis channels of the eye-tracker, to exclude trials containing blinks and saccades. The eye blinks were detected in the eye-tracker channels by z-scoring the time series and applying a threshold of 5 SD was applied so that any deflection above the threshold was classified as a blink. The saccades were detected using a scatter diagram of X-axis and Y-axis time series of the eye-tracker for each trial. An event was classified as a saccade if the gaze shifted away from the fixation point by 2° and its duration was more than 500 ms. Trials contaminated by blinks and saccades were removed from further analysis. We also rejected trials containing large-amplitude events (above 5 SD) in MEG which are mainly associated with motion and muscle artefacts. As a result, the number of trials that remained after exclusion was 304 ± 12 (mean ± SD) per participant. For each participant, the number of trials per condition was equalized by randomly selecting the same number of trials.Prior to computing TRF, the magnetometer signals were bandpass filtered within 1 – 99 Hz.i is the time lag ranging from - 0.25 to 0.75 s with a step of 0.001 s; t is the vector of length 1.5 seconds; r is Person correlation; TRF denotes response function.The TRFs were computed between the broadband flickering signal and the MEG gradiometers for each trial. To this end, we applied the cross-correlation approach , TRF(i)=It should be noted that this approach is essentially equivalent to more sophisticated approaches such as ridge regression  in case The TRFs computed for each trial were then averaged across the trials.The alpha-echoes were computed in the source space using the linearly constrained minimum variance (LCMV) beamformer , as implTo build a forward model, we first segmented the MRI images using Fieldtrip, and then manually aligned the MRI images to the head shape digitization points acquired with the Polhemus Fastrak. Finally, a single shell head model was prepared using surface spherical harmonics fitting the brain surface .The individual anatomy was wrapped to the standard MNI template using non-linear normalisation . The temThe covariance matrix was estimated in the time interval  s relatively to the onset of stimuli motion. Since, the SSS method  reduces the rank of data, we estimated the rank after SSS and initialise the regularisation parameter kappa using the estimated rank. We computed the unit-noise-gain or weight-normalized beamformer  to avoidAs suggested elsewhere , the traIn this study, we presented participants with grating stimuli while modulating the luminance of the stimuli by random noise, and simultaneously acquiring MEG. Temporally uncorrelated flickering stimuli were shown in the left and right hemifield.The brain response to the randomly flickering stimuli was estimated using temporal response functions (TRFs). The TFR reflects the kernels best explaining the brain response when convolved with the input stimuli. The individual temporal responses were averaged across the participants for both the right  and leftp < 10-12). We then used LCMV beamformers to assess the power of TRFs in source space. To this end, the sensor-space MEG times series were projected into source space, and then the TRFs were computed for each dipole (or grid point). The maximum power of the TRF was localised in the parietal regions near the midline  for both the right and left stimulation sides , predominantly in the centro-lateral direction  propagation delay of perceptual echoes across the scalp. Using source modelling, we found that the perceptual alpha-echoes were well explained by two generators in the primary visual and parietal cortices. This is consistent with the notion that travelling waves at the scalp level may occur as a result of a summation of neuronal activity generated by two phase-lagged dipoles. To further clarify this, we used a realistic forward model to estimate the activity generated by two sources, in occipital and parietal areas, in the MEG sensors. These stimulations explained the MEG findings well. Finally, we used an EEG forward model to project the two-dipole activity to EEG sensors. The sensor-level EEG travelling waves closely resembled those reported in previous EEG studies. Thus, our simulation results show that the sensor-level travelling waves reported in MEG and EEG can be explained by a common two-dipole model.i) delayed excitation from a single oscillator, (ii) propagating pulses in an excitable network, and (iii) phase-locked weakly coupled oscillators. Recently, Alamia and VanRullen (Three mathematical models for generating travelling waves in the brain have been proposed . These manRullen  proposedIn contrast to EEG studies, our MEG results revealed a more focal propagation of the perceptual alpha-echoes, essentially restricted by the occipital and parietal areas. Interestingly, EEG studies reveal predominantly occipital-frontal propagation of the travelling waves for either a single flickering stimulus presented centrally  or two iA growing body of evidence suggests that travelling waves are functionally relevant for memory and attention . AlthougIn our recent study , we foun


Encepur Adults, GSK), administered approximately 3 years post-primary vaccination according to 1 of 3 licensed vaccination schedules in adults and adolescents, resulted in antibody persistence for 10 years post-boosting. We used different power-law models (PLMs) to predict long-term persistence of anti-TBE virus neutralization test (NT) antibody titers over a period of 20 years post-booster dose, based on individual antibody NT titers measured for 10 years post-booster vaccination. The PLMs were fitted on pooled data for all vaccine schedules. A mean NT titer of 261 , considerably above the accepted threshold of protection (NT titers ≥10), was predicted 20 years post-booster vaccination with TBE vaccine. Our modeled data suggest that the intervals of booster doses could be increased without compromising protection against TBE.In tick-borne encephalitis (TBE)-endemic regions, long-term vaccination programs are efficient in preventing the disease. A booster dose of a polygeline-free inactivated TBE vaccine ( Flavivirus genus, transmitted mainly through Ixodes ticks. In the European Union, TBE is notifiable since 2012; notification rates were the same in 2014 and 2015 ,1 although emerging data from 2016 illustrate an increase in the number of cases for eight countries.2Tick-borne encephalitis (TBE), an infectious disease of the central nervous system, is caused by the TBE virus (TBEV) of the 3 Case-fatality rates differ according to the TBEV subtype, but can reach up to 40% for the Far-Eastern subtype, while being significantly lower for the Siberian (6–8%), and European (≤1.4%) variation.4 Several vaccines, based on different strains of TBEV, are available and considered effective in preventing TBE.4The non-specificity of clinical symptoms in the initial viremic phase and the lack of effective treatment make TBE very difficult to manage. This can lead to long-term sequelae in more than 30% of cases.Encepur Adults, GSK), targeting the European subtype, is licensed in Europe for use in adults and adolescents ≥12 years of age and has been shown to be immunogenic and well tolerated,6 with persistently high antibody titers induced up to 10 years following a first booster vaccination.8 The TBE vaccine has also been reported to induce antibodies against various Asian TBEV isolates.9The polygeline-free inactivated TBE vaccine , for which anti-TBEV NT titers ≥2 and ≥10 were chosen as thresholds indicative of seropositivity and of a clinically meaningful antibody response, respectively.11Although there is no established correlate of protection against TBE, the World Health Organization supports the use of a surrogate marker of protection represented by the presence of circulating antibodies against TBEV at or above a locally agreed concentration considered as clinically meaningful.4 Several TBE-endemic European countries recommend routine vaccination as a three-dose primary series according to one out of three different approved schedules, followed by repeated booster doses.12 However, the optimal intervals for booster vaccination necessary to ensure long-term protection against TBE are still being debated, as only a few studies reported on long-term immunogenicity of available vaccines.13 A recent study demonstrated that ≥90% of individuals vaccinated with the TBE vaccine maintained NT titers ≥10 up to 10 years after a first booster dose, regardless of age and primary vaccination schedule received.8 Moreover, no TBE infections occurred during this period.8The World Health Organization recommends vaccination against TBE for individuals of all ages living in highly endemic areas and to targeted cohorts (>50–60-year-olds) in low-to-moderate-endemic regions, as well as to travelers planning outdoor activities in TBE-endemic areas.11 We used several power-law models (PLMs), based on antibody levels measured in two extension studies in participants followed 57 and 10 years8 post-booster dose to predict the development of antibody levels beyond the time period covered by clinical trials.With the aim of estimating the evolution of the immune response over time beyond available clinical data, we modeled anti-TBEV NT antibody levels up to 20 years post-first boosting following primary vaccination with one of the 3 licensed schedules.A summary contextualizing the results, the potential clinical research relevance and the impact of our study is displayed in Encepur) according to a licensed  or an unlicensed  vaccination schedule.11 In the first extension study (NCT00387634), 283 participants received a booster dose at study entry, 3 years post-primary vaccination. Additionally, 40 participants who had received a booster dose 12–18 months post-primary were enrolled; all were followed up for 5 years.7 Of these, 201 individuals who had received one of the licensed vaccination schedules were followed up in the second extension study (NCT01562444) from year 6 to 10 post-booster dose.8In the primary study, a randomized trial conducted in Czechia, adolescents and adults ≥12 years old received three primary doses of the TBE vaccine . The PPLM combines multiple PL functions, each fitted to different, consecutive intervals of data, which allows for more flexible estimation of the evolution of antibody levels over the entire period of time. The monotone PPLM (MPPLM) further restrained these functions from increasing in value to exclude a potentially biologically not meaningful increase in antibody level over time. Finally, we considered the extended PLM (EPLM) proposed by Fraser et al. This model is an extension of the PLM which accounts for two populations of B-cells involved in a long-term antibody plateau: activated and memory B-cells.The models were compared using the classic model selection based on the Akaike information criterion (AIC). Moreover, models were compared based on prediction error. To that end, a separate calculation was done in which the two last years of measured data were not considered. A sensitivity analysis excluding outliers was also performed, with outliers identified by the standard interquartile range rule (1.5 × IQR).The values predicted by the models were then compared with the measured values. All predicted values were reported as geometric means.All statistical analyses were performed using the PROC NLMIXED subroutine in Statistical Analysis Systems (SAS). Formulas, SAS syntax, and estimated parameters for each model are provided in the Supplementary Material.For all three primary vaccination schedules , a similar yearly distribution of anti-TBEV antibody mean NT titers was observed following the booster dose . Only 14p-value = 0.11). Therefore, all models were applied on pooled data from participants receiving any of the three vaccination schedules and considering the timing of the booster dose  as the starting time point for the prediction .To our knowledge, this is the first study modeling long-term antibody persistence after vaccination against TBE, providing modeled data beyond the 10 years post-first booster period investigated to date. Twenty years post-booster vaccination with the TBE vaccine, different models predict a mean NT titer considerably above the established threshold of 10 for more than 95% of individuals who were included in these analyses. Therefore, sufficient protection against TBE is predicted for at least 20 years after booster vaccination with the TBE vaccine, when any of the licensed primary three-dose schedules is completed and a booster dose is administered at 1 or 3 years post-primary vaccination, depending on the primary schedule.7 while at 10 years post-booster, they were 166–245 in the total-vaccinated cohort and 260–307 in the per-protocol set.8 Of note, over 10 years of follow-up post-boosting, no consistent decline in geometric mean NT titers was observed within the same age groups, although smaller overall values were observed for participants receiving the booster dose at 50–59 and ≥60 years of age than at 15–49 years of age.8 However, the comparison was hindered by a relatively small sample size for the ≥60-year age group, which also applied for our analysis. Therefore, analyses were not performed by age stratum.When considering the follow-up of adolescents and adults who received the full primary vaccination series and a booster dose, no consistent decline in antibody NT titers was measured over a period of 10 years. For instance, at 5 years post-booster dose, the observed geometric mean titers ranged between 300 and 429 across groups,17 An exposure to TBEV itself appears unlikely, since in the case of infection a more sustained increase in antibody levels would be expected. However, the predicted values as well as the prediction interval indicate that participants not showing elevated results due to potential cross-reactivity or high baseline levels would still have NT titers consistently above threshold levels after 20 years post-boosting.According to the AIC, the best model for the provided data was the PPLM, which predicted an increase of antibody levels over the timeframe considered. This increase simulates the slight increase in antibodies observed in the measured data obtained during the trials. One possibility is that exposure to or vaccination against other flaviviruses  could result in antibody formation with a certain cross-reactivity within the assay; although this is unlikely for NT, which was shown to distinguish between pathogens belonging to the TBEV family.12 may be increased without compromising the expected protection, thus potentially lowering associated costs from both the societal and patient perspective. However, this remains to be confirmed by further empirical evidence and results from future studies following up vaccinated individuals will provide data against which the predicted rate of antibody titer decline could potentially be validated. Evaluation up to 15 years post-booster dose is currently ongoing as part of a third extension study (NCT03294135).Our modeled data suggest that the interval between booster doses currently recommended for immunization against TBE (every 3–5 years)While generally, results of a statistical analysis may vary depending on the model used, a strength of this analysis is the use of several different models leading to similar results demonstrating the robustness of the prediction. However, the analysis has also several potential limitations. Among those, one of the most important is the assumption about the dynamics and decline of B- and T-cells, as the statistical analysis cannot be the sole driver for the conclusions. Another potential limitation is the fact that a drop in NT titers was not predicted by any of the models, as the observed data used as basis for this modeling exercise showed a steady trend beyond one year post-booster dose, without reaching the turning point where antibodies started to fall, which made it impossible to estimate the antibody half-life. Due to the relatively small sample size for the older age range, in particular, the ≥60-year age group, the analyses were not carried out by age strata; potential extrapolation of results from the pooled age groups to individuals ≥60 years of age should therefore be interpreted with caution. Moreover, these results are specific to the TBE vaccine used, and cannot be generalized to other vaccines.In conclusion, the predicted antibody persistence was considerably above the surrogate marker of protection (NT titers ≥10) up to 20 years post-booster vaccination with the TBE vaccine. Our results suggest that booster schedules could be increased in the future, although our modeled estimates still need to be confirmed by field data.Click here for additional data file.


This cohort study analyzes the implications of pandemic-related stay-at-home restrictions for the positivity rates of influenza and other seasonal respiratory viruses in Northern California. Was the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–related shelter-in-place order associated with lower rates of common respiratory viruses in a community?In this cohort study using interrupted time analysis of 46 128 tests for viral respiratory pathogens, statistically significant lower rates of influenza and rhinovirus or enterovirus infections were found after the institution of a shelter-in-place order to prevent SARS-CoV-2 transmission.Results of this study showed that public health initiatives, such as shelter-in-place orders during a pandemic, appeared to be associated with lower transmission rates of other respiratory viruses in a community. Public health initiatives that include shelter-in-place orders are expensive and unpopular. Demonstrating the success of these initiatives is essential to justify their systemic or individual cost.To examine the association of a shelter-in-place order with lower rates of seasonal respiratory viral activity.This cohort study with interrupted time series analysis obtained monthly counts of respiratory virus testing results at UC Davis Health from August 1, 2014, to July 31, 2020. Patients of all ages underwent testing conducted by the laboratory at UC Davis Health, a referral center for a 65 000-square-mile area that includes 33 counties and more than 6 million Northern California residents.A statewide shelter-in-place order was instituted on March 19, 2020, restricting residents to their homes except for traveling for essential activities. Large social gatherings were prohibited, schools were closed, and nonessential personnel worked remotely. Those who had to leave their homes were mandated to wear face masks, engage in frequent handwashing, and maintain physical distancing.Positivity rates of common respiratory viruses within the community served by UC Davis Health.A total of 46 128 tests for viral respiratory pathogens over a 6-year period were included in the analysis. For the postexposure period (March 25-July 31), approximately 168 positive test results occurred for the studied organisms in the 2020 virus year, a positivity rate of 9.88 positive results per 100 tests that was much lower than the positivity rate of 29.90 positive results per 100 tests observed for this date range in the previous 5 virus years. In contrast, the positivity rates were similar for the preexposure time frame (August 1-March 24) in the 2020 virus year and for the same time periods in the 5 previous years (30.40 vs 33.68 positive results per 100 tests). In the regression analyses, statistically significant decreases in viral activity were observed in the postexposure period for influenza  and for rhinovirus or enterovirus  infections. Lower rates of postexposure viral activity were seen for respiratory syncytial virus, parainfluenzavirus, coronaviruses, and adenoviruses; however, these associations were not statistically significant.Using interrupted time series analysis of testing for viral respiratory pathogens, this study found that statistically significant lower rates of common community respiratory viruses appeared to be associated with a shelter-in-place order during the coronavirus disease 2019 pandemic. As part of this directive, nonessential personnel worked remotely, large social gatherings and worship services were canceled, and California schools were closed. Essential workers and consumers leaving their homes were mandated to wear face masks, engage in frequent handwashing, and maintain a 6-foot distance from people outside their household.1 The March 19 stay-at-home order remained in effect through the summer and fall of 2020, with travel permissible only for health care needs; food procurement; outdoor exercise or recreation; outdoor museum visits; and activities at open, low-risk businesses and workplaces for which telework was not possible, including retail, manufacturing, and offices.The 2019-2020 respiratory virus season was like no other in human history, with the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its circulation alongside common respiratory viruses. In response to the coronavirus disease 2019 (COVID-19) pandemic, federal, state, and local governments placed restrictions on travel and instituted physical distancing protocols to curb the spread of this novel virus. On March 19, 2020, California governor Gavin Newsom issued a statewide shelter-in-place order.The end of the 2019-2020 respiratory virus season presented a unique opportunity to evaluate the outcome of public health interventions in response to SARS-CoV-2 for the epidemiologic characteristics of other circulating respiratory viruses. In this cohort study, we examined the association of the shelter-in-place order with lower rates of seasonal respiratory viral activity using the incidence of common respiratory viruses in a Northern California health system during the statewide shelter-in-place order. Public health initiatives that include shelter-in-place orders are expensive and unpopular. Demonstrating their success is essential to justify their systemic or individual cost.TREND) reporting guideline.All data used in this study were deidentified, so the study did not qualify as human research according to the policies of the US Department of Health and Human Services or the US Food and Drug Administration; therefore, institutional review board approval was not required. We followed the Transparent Reporting of Evaluations With Nonrandomized Designs , which tests for influenza A subtypes H1, H1 2019, and H3; influenza B; respiratory syncytial virus (RSV) A and B; parainfluenzavirus 1 to 4; human coronaviruses 229E, HKU1, NL63, and OC43; human metapneumovirus; rhinovirus or enterovirus; and adenovirus. Respiratory syncytial virus and coronaviruses were added to UC Davis Health’s current respiratory pathogen panel in July 2016, limiting our evaluation of these 2 viruses to 5 rather than 7 years. Human metapneumovirus rates during the study period were substantially lower than the rates of other viruses; thus, this organism was not included in the analysis.Results of point-of-care real-time polymerase chain reaction test for influenza A and B and RSV  were also included in this study. Point-of-care RSV testing was available at UC Davis Health starting in 2018. Respiratory pathogen panel and point-of-care testing data were extracted from clinical encounters in multiple settings, including the hospital, emergency department, and more than 40 outpatient clinics.Our interest was in ascertaining whether the seasonally adjusted incidence rate of respiratory virus detection during the shelter-in-place period differed from the expected rates based on previous years. The units of analysis were monthly counts of positive and negative test results. In this study, the shelter-in-place order divided the current virus season into preexposure  and postexposure  periods. This calendar time division was used for the 5 previous virus years to permit the comparison of similar calendar periods between virus years while accounting for between-year variation. In the unadjusted analysis of virus incidence, we proportionally allocated the March data in a 24:7 ratio to the preexposure and postexposure periods.2 (SAS Institute Inc) to fit marginal regression models for overdispersed Poisson data with autocorrelated errors for each organism’s monthly time series of positive test result counts. This model included a variable coded 0 for all months before or 1 for all months after March 2020, and coded with the value 7/31 for March 2020, to adjust for the shelter-in-place order affecting the last week of March and fully affecting all weeks subsequent to that. The estimated regression coefficient for this variable was used to estimate the seasonally adjusted incidence rate ratio (IRR) associated with the shelter-in-place order, which was also expressed as a percentage reduction (100 × [1 – IRR]). The Wald test for this regression coefficient was evaluated for statistical significance with a 2-sided α = 5%. All models included additional covariates to adjust for year-to-year differences and cosinor analysis terms to account for sinusoidal seasonal effects.3 For each organism, we fit and compared 6 models that varied in how they controlled for historical patterns  and how they controlled for seasonal effects ,4 and we used the Akaike information criterion to select the best model.5For analyses adjusting for between-year and seasonal effects, we used the GLIMMIX procedure in SAS/STAT softwareOffset terms based on the monthly number of definitive test results were included in the model so that that the incidence rates refer to incidence proportions. The small number of indeterminate test results (<0.14%) were excluded from the analysis. In sensitivity analyses, we changed the offset terms to refer to number of days in the month so that incidence rates refer to number of positive test results per day , approximately 168 positive test results occurred in the 2020 virus year, a positivity rate of 9.88 positive results per 100 tests, which was much lower than the positivity rate of 29.90 positive results per 100 tests observed for this date range in the previous 5 virus years. In contrast, the positivity rates were similar at the preexposure time frame (August 1-March 24) for the 2020 virus year and for the previous 5 virus years (30.40 vs 33.68 positive results per 100 tests) .We observed lower rates of all incident viral respiratory infections after the institution of the statewide shelter-in-place order compared with the previous 5 to 7 years (2014-2019). Specifically, a statistically significant decrease of 93%  was found in 2020 for laboratory-confirmed cases of influenza and 81%  for laboratory-confirmed cases of rhinovirus or enterovirus. Lower rates of viral activity ranging from 63% to 91%  were also observed in all other viral pathogens, but these rates were not statistically significant because of smaller incidence rates, higher seasonal variability, and thus wider CIs .6 indicated that in Japan, seasonal influenza activity was lower in the 2019-2020 respiratory season compared with the 5 previous years. Studies from Taiwan, Singapore, Korea, and Thailand have also shown decreased influenza incidence after countrywide public health measures were instituted to reduce the transmission of COVID-19.10 To our knowledge, this study is the first in the United States to examine the implications of SARS-CoV-2 public health measures for influenza rates as well as the first in the world to examine the implications of these public initiatives for other seasonal respiratory viruses.We hypothesized that the statewide shelter-in-place order that was instituted to curb the spread of SARS-CoV-2 would be associated with lower rates of common viral infections circulating in the community. In April 2020, a report by Sakamoto et alThis study has several limitations. First, although UC Davis Health serves as a referral center for more than 6 million Northern California residents, this is a study of a single health system. Second, although the statistical analysis rigorously estimated the seasonally adjusted outcome of a shelter-in-place order as an IRR, the order was issued toward the end of the respiratory viral season in California. The implication of this public health intervention for the 2020-2021 respiratory viral season is unknown and could be altered by favorable  or unfavorable  factors. Third, health care avoidance because of concerns about COVID-19, particularly by those with milder infections, was a possible confounder in the number of laboratory-confirmed cases. Increased use of telehealth and limits on testing capabilities are other possible confounders. We used the rate of positive cases to control for fewer tests attributed to changes in health-seeking behavior. In sensitivity analyses that included daily rates of positive cases, the estimated IRRs were similar, with IRRs for parainfluenzavirus and adenovirus becoming statistically significant (eTable in the 11 However, this cohort study shows that public health strategies have been associated with lower rates of respiratory viruses in the community served by UC Davis Health.The restrictions on travel, mandated mask wearing, and physical distancing that are embedded in the California shelter-in-place order have been controversial because of their substantial social and economic costs.


The filamentous actin (F-actin) cytoskeleton is progressively damaged after status epilepticus (SE), which is related to delayed neuronal death, aberrant recurrent circuits and epileptogenesis. Glucocorticoids regulate dendritic spine remodeling by acting on glucocorticoid receptors and the dynamics of the F-actin cytoskeleton. Our previous study showed that administration of dexamethasone (DEX) in the latent period of the pilocarpine epileptic model reduces damage to the hippocampal filamentous actin cytoskeleton and the loss of hippocampal neurons and aids in maintaining the synaptic structures, but it is not sufficient to stop epileptogenesis. In this work, we focused on the role of glucocorticoids in regulating the hippocampal F-actin cytoskeleton during SE. We examined the abundance of synaptic F-actin, analyzed the hippocampal F-actin/G-actin (F/G) ratio and pCofilin, and evaluated the number of hippocampal neurons and pre/postsynaptic markers in pilocarpine-induced status epilepticus mice with or without administration of dexamethasone (DEX). We found that the latency of Stage 3 seizures increased, the mortality decreased, the damage to the synaptic F-actin cytoskeleton in the hippocampal subfields was significantly attenuated, and a greater number of postsynaptic structures were retained in the hippocampal subfields after treatment with DEX. These results indicate that treatment with dexamethasone stabilizes the synaptic F-actin cytoskeleton and reduces the damage to the brain due to SE. This approach is expected to be beneficial in alleviating delayed neuron damage and the process of epileptogenesis. SE is manifested by the continuous and persistent onset of seizures, which is a life-threatening neurological condition. One of the consequences of SE is extensive brain damage and secondary temporal lobe epilepsy with recurrent spontaneous seizures and hippocampal-dependent cognitive impairment in vivo: mineralocorticoid receptors (MRs) of high affinity and glucocorticoid receptors (GRs) of low affinity Glucocorticoids have proven to be helpful in treatment of epilepsy in numerous clinical and experimental studies The study protocol for animals was approved by the Research Ethics Committee of the First Hospital of Jilin University, China (reference number 2014-044). The experiments were designed using the principle of "The Three Rs". All experiments were performed on ICR adult male mice weighing 22-24 g. Mice were purchased from Changsheng Biotechnology Co., Ltd. . Male animals were chosen to prevent differences in epileptic susceptibility and basal hormone levels due to gender. Mice were housed in quiet rooms with temperature controlled at 22-26 °C with a 12/12 light and dark cycle and were allowed to freely access food and water. Animals were acclimated for 3 days before performing any experiments. To reduce the influence of circadian rhythm on seizures, all pilocarpine-induced SE models were performed between 8 AM and 10 AM.In the morning of the experimental day, mice were injected intraperitoneally with 1 mg/kg methylscopolamine (0.1 mg/ml) , followed by 300 mg/kg pilocarpine (30 mg/ml)  30 minutes later. After pilocarpine administration, the behaviors of the animals were monitored for 2 hours, and the onset of seizures was evaluated by Racine scale To observe the effects of glucocorticoids on remodeling of the hippocampal F-actin cytoskeleton during pilocarpine-induced SE, animals were randomly divided into three groups: control, pilocarpine, and pilocarpine with dexamethasone treatment, which were identified as Control, PILO and PILO+DEX groups respectively. For animals in the PILO+DEX group, a dose of 10 mg/kg dexamethasone   was injected intraperitoneally together with pilocarpine.To verify whether glucocorticoids modulate F-actin dynamics through GRs, pretreatment of RU486, an antagonist of GRs, was administered. In this assay, animals were randomly divided into four groups: control, PILO, PILO+DEX and RU486. For animals in the RU486 group, RU486 (2 mg/ml)  was pretreated at a dose of 20 mg/kg for 4 consecutive days intragastrically prior to pilocarpine and dexamethasone administration. The last dose of RU486 was given 1 hour before the pilocarpine and dexamethasone injection. The other three groups were given equal volumes of normal saline intragastrically.Preparation of samples was performed according to a previously published method As mentioned previously As described in previous studies, hippocampal pyramidal cells were detected by Anti-NeuN antibody To test the effect of DEX treatment on presynaptic and postsynaptic components in SE mice, we labeled the presynaptic and postsynaptic structures with synapsin I and PSD95, respectively. Protocols for standard immunofluorescence staining were performed for synapsin I and PSD95 staining. The primary antibodies used were rabbit anti-synaptic polyclonal antibody  and PSD-95/SAP90 . For staining of PSD95, pretreatment with pepsin was used to improve the detection The ratio of F-actin to G-actin reflects the dynamic changes in the actin cytoskeleton in the brain. To measure the F/G ratio, the hippocampi were homogenized, and a standard protocol according to the manufacturer was followed using an Assay KIT .The actin-severing protein cofilin is inactivated by Ser3 phosphorylation, thereby allowing further elongation of the polymerizing actin filaments and maintaining the filamentous structures For confocal scanning and statistical analysis, we randomly selected 7 animals survived from SE in the PILO and PILO+DEX groups and 7 animals from the control group, respectively. A Zeiss 710 confocal laser scanning microscope was used in imaging of F-actin, NeuN, PSD95 and synapse I. Each experiment used the same parameter settings to avoid deviation. Hippocampal slices were randomly selected from the-1.28 to -2.12 mm coordinates of anterior-posterior Bregma. The labeling density of F-actin was detected by Image-Pro Plus analysis software . The fluorescence intensity of PSD95 and synapsin I was measured using ImageJ software . We measured at least 3 fields for each hippocampal subregion in each animal. The numbers of pyramidal cells in areas CA1 and CA3 were counted as in the previous description The raw data were input to GraphPad Prism 8  for statistical analysis and graph preparation after the normality test using SPSS25 . Data that followed a normal distribution were expressed as the mean ± SD. One-way ANOVA with the post hoc Tukey's test was performed for data with three or more groups. Data that followed a nonnormal distribution were expressed as M . For comparison between two groups, the Kolmogorov-Smirnov test was used. For conditions among three groups, the Kruskal-Wallis test was performed, followed by the post hoc multiple comparison of Dann's test. The significance set for statistical analysis was P < 0.05.After pilocarpine injection, the behavioral changes of animals in this research were similar to those of previous studies We observed the latency of Stage 3 seizures in all animals of the PILO and PILO+DEX groups, counted the number of seizures of greater than or equal to Stage 3, and calculated the average stage in the survived animals. The average latencies of Stage 3 seizures were 10.60  minutes and 16.99  minutes in the PILO and PILO+DEX groups, respectively. Compared with the PILO group, dexamethasone treatment significantly increased the latency of Stage 3 seizures (P<0.01)  Fig. B and CA1Numerous studies have focused on the effects of glucocorticoids in epilepsy and convulsions, but the conclusions are controversial so far MRs and GRs are two types of glucocorticoid receptors in the brain. High-dose glucocorticoids activate GRs, and low-dose glucocorticoids activate MRs. However, the activation of these two types of glucocorticoid receptors often plays opposite roles in many physiological activities Dexamethasone treatment showed a protective effect on the hippocampal F-actin cytoskeleton in this study. Numerous studies have shown that glucocorticoids regulate the F-actin cytoskeleton through two main pathways, i.e., transcriptional and nontranscriptional pathways. The dynamics of the actin cytoskeleton might be affected by transcriptional regulation of glucocorticoid-induced gene expression In view of the evidence presented, we inferred that DEX might promote the polymerization of actin and maintain the filamentous state by activating GRs on the dendritic spines in our study.Many reports have stated that glucocorticoids have a protective effect on neuronal loss in epilepsy models Evidence suggests that actin cytoskeleton exerts a key role in the communication network to determine cell death or survival F-actin is located in both pre- and postsynaptic structures, interacts with pre- and postsynaptic proteins and plays a key role in synaptic remodeling F-actin depolymerization after status epilepticus has been widely known and might be related to subsequent remodeling of synapses and delayed neuronal death. Therefore, protecting the F-actin cytoskeleton from depolymerization during SE might play an important role in blocking the process of epileptogenesis.In summary, we examined the remodeling of actin dynamics in the SE model after glucocorticoid treatment. Our results demonstrate that administration of 10 mg/kg dexamethasone ameliorates the damage to the hippocampal filamentous actin cytoskeleton, contributes to the maintenance of synaptic structures, and might play a role in blocking the process of epileptogenesis.


Objectives: We aimed to assess the clinical pharmacist-initiated telephone-based patient education and self-management support for patients with cardiovascular disease during the nationwide lockdown during COVID-19 pandemic.Methods: A prospective single-center telephone-based cross-sectional study was conducted among patients at the Cardiology Department and its speciality clinic at a 1,800-bed tertiary care hospital in Southern India. A validated 8-item clinical pharmacist aided on-call questionnaire with two Domains was administered during and after lockdown (15 March and 8 June 2020). Clinical pharmacist-provided educational assistance on self-management practices were in accordance with the guidelines of Indian Council of Medical Research (ICMR) and World Health Organization. Comparisons was performed using sign test and association of responses were analyzed using the Goodman and Kruskal's gamma test. All the tests were two-tailed, p < 0.05 was considered to be statistically significant.Results: Of the 1,080 patients, 907 consented with a response rate of (83.9%) and 574 (96.36%) patients were analyzed post-intervention. Majority of the patients were male (54.7%) and had Acute Coronary Syndrome . The majority of subjects had at least two co-morbid conditions  and were rural population (82.5%) as self-employed (43.1%) with a middle-class economy (31.6%). In the Domain-1 of checklist the awareness toward complications caused by COVID-19 in cardiovascular diseases  and the importance of universal safety precautions enhanced after clinical pharmacist assistance [ and ]. In Domain-II of checklist there was a significant improvement in patients awareness toward fatal complications caused by COVID-19 , maintenance of self-hygiene , practice of universal safety precautions  and self-isolation . The results of our study population varied from baseline evaluation  to post-intervention  based on Literacy, employment status and economic status.Conclusions: The proactive role of clinical pharmacists in providing instructional services in collaboration with cardiologist during the pandemic circumstances increased patients understanding and mitigated infection exposure among patients, health care professionals and also assuring the continuity of care in patients with established cardiovascular diseases. In the last two decades, clustering and incidence of severe acute respiratory infections are one of the major threats to public health. Coronavirus disease COVID-19) was first recorded in Wuhan, China, by the end of December 2019. Since then, COVID-19 has rapidly spread around the world. The COVID-19 was declared as a global pandemic on 11th March 2020 by the World Health Organization. COVID-19 has a major impact on public health and has a direct or indirect impact on social and economic activities. The exponential increase in the number of patients with COVID-19 in the past 6 months has overwhelmed health-care systems across the world. This is due to an inadequate understanding of the dynamic interplay of shifting epidemiology, publicity, pandemic prevention strategies, risk identification, and public health behavior .The primary outcome of the study is to identify the impact of the clinical pharmacist-initiated educational guidance on COVID-19 pandemic among patients with established cardiovascular disease. The secondary outcome is to ensure continuity of care and compliance with the prescribed drugs.p < 0.05 was considered to be statistically significant.Data were entered in MS Office Excel 2019 and analyzed using the IBM SPSS Statistics Version 25. Continuous variables were presented as mean ± standard deviation (SD). Categorical variables were presented as absolute numbers and percentages. Comparisons between baseline and post assistance scores among the individuals were performed using sign test, Association of Responses with socio-demographic variables were analyzed using the Goodman and Kruskal's gamma test. All tests were two-tailed, p = 0.000) and their current condition (p = 0.000). However, majority of them were not aware of the complications caused by COVID-19 among patients with cardiovascular diseases  and the importance of universal safety precautions, their awareness enhanced after clinical pharmacist assistance [ and ]. In Domain-II regarding knowledge aspect majority of the patient's knowledge improved regarding fatal complications caused by COVID 19 , the process of self-isolation, maintenance of self-hygiene , the importance of universal safety precaution  and regarding self-quarantine  depicted in Of the 1,080 patients contacted by telephone, the response rate at the baseline was 907 (83.9%) and 574 (63.28%) post-intervention. The majority (54.7%) of the study population were male and had at least two co-morbid conditions (44.56%) in the age group (61–80 years) . The patPandemics and epidemics are a widespread problem then and now as COVID-19. During such periods, people in the community face several challenges. Lack of awareness and consciousness often leads to an uneasy attitude which could adversely affect the patients with established cardiovascular complications. Different stakeholders in their respective countries are working together to “flatten the curve” by joint prevention initiatives led by the WHO. With a practically sufficient global lockdown, Pharmacists appear to be the first contact point for meeting the health requirements of the public .We studied the role of clinical pharmacists' assistance for patients with established cardiovascular diseases during the COVID-19. The principal findings in our study at initial assessment were (1) Most of the patients were aware of their medical condition (CVD and comorbidities), (2) Most of the patients were aware of SARS-CoV-2 (COVID-19) infection, (3) majority of the patients were unaware of fatal complications caused by COVID-19 and association of COVID severity with CVD and comorbidities, (4) most of them were unaware of the importance of universal safety precautions, (5) majority of them don't know that self-quarantine is a procedure followed by people who are at risk during the epidemic.Pharmacists continue to play their role in promoting continuity of pharmaceutical care, as well as supporting governments for disseminating information on precautions related to COVID-19 spread . PharmacThe other measure which has been mentioned a lot in recent weeks is hand hygiene. The World Health Organization (WHO) regards handwashing with soap and water and friction with hydroalcoholic gel as the most effective measures for the prevention of infections and antimicrobial resistance . ResearcAs a consequence of COVID-19, the need for social distancing forced us to use all the resources in our toolbox, and telehealth is one of them that accelerated its adoption globally . TeleheaWhat was wonderful about it was during this pandemic when our patients are generally considered to be at greater risk of having more severe COVID-19 disease and when they have been asked to stay at home, through virtual visits, we are still able to maintain their continuity of care, evaluate their COVID-19 awareness and knowledge, and provide instructional assistance and mitigate their exposure to infection. Overall, the results reflect what might develop into a new standard of future health care, particularly during contagious outbreaks . In thisDuring the times of stretched clinical resources due to COVID-19, our research results helped to add new ways to reduce COVID-19 spread in patients with established cardiovascular diseases. Although our study is a single-center study involving a clinical pharmacist, despite its limitations, the results in our study suggest that the extended role of the clinical pharmacist may also be beneficial to other clinical groups. In addition, more awareness amongst the study patients could also be attributed to govt initiated awareness programmes on COVID-19 . The resThe clinical pharmacist may, however, play a pro-active role in promoting patient-specific treatment decisions by serving as a resource for physicians and other health care professionals to mitigate adverse events caused by SARS nCoV-2 infection in patients with established cardiovascular disease. The enhanced role of clinical pharmacists in providing instructional services should mitigate infection transmission during the COVID-19 pandemic.The original contributions presented in the study are included in the article/supplementary materials, further inquiries can be directed to the corresponding author/s.The institutional ethics committee has approved this study and participants have been informed of the purpose of the study before participating and voluntary consent is obtained virtually. All procedures performed in this study involving human participants were consistent with the Declaration of Helsinki 1964 and its subsequent amendments or comparable ethical standards.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.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.


The induction of synaptic plasticity requires the presence of temporally patterned neural activity. Numerous cellular studies in animals and brain slices have demonstrated that long-term potentiation (LTP) enhances synaptic transmission, which can be evoked by high-frequency intermittent stimulation. In humans, plasticity processes underlying perceptual learning can be reliably induced by repetitive, LTP-like sensory stimulation. These protocols lead to improvement of perceptual abilities parallel to widespread remodeling of cortical processing. However, whether maintained rhythmic cortical activation induced by the LTP-like stimulation is also present during human perceptual learning experiments, remains elusive. To address this question, we here applied a 20 Hz intermittent stimulation protocol for 40 min to the index-, middle- and ring-fingers of the right hand, while continuously recording EEG over the hand representation in primary somatosensory cortex in young adult participants. We find that each train of stimulation initiates a transient series of sensory-evoked potentials which accumulate after about 500 ms into a 20 Hz steady-state response persisting over the entire period of the 2-s-train. During the inter-train interval, no consistent evoked activity can be detected. This response behavior is maintained over the whole 40 min of stimulation without any indication of habituation. However, the early stimulation evoked potentials (SEPs) and the event-related desynchronization (ERD) during the steady-state response change over the 40 min of stimulation. In a second experiment, we demonstrate in a separate cohort of participants that the here-applied pneumatic type of stimulation results in improvement of tactile acuity as typically observed for electrically applied 20 Hz intermittent stimulation. Our data demonstrate that repetitive stimulation using a 20 Hz protocol drives rhythmic activation in the hand representation of somatosensory cortex, which is sustained during the entire stimulation period. At the same time, cortical excitability increases as indicated by altered ERD and SEP amplitudes. Our results, together with previous data underlining the dependence of repetitive sensory stimulation effects on NMDA-receptor activation, support the view that repetitive sensory stimulation elicits LTP-like processes in the cortex, thereby facilitating perceptual learning processes. Under everyday-live conditions, humans learn largely through practicing and repetition. In the laboratory, however, by exploiting adequate timing of pulse trains, learning can be induced merely through electrical stimulation of brain cells and synapses. The approach of repetitive sensory stimulation closes the apparent gap between these extremes by translating protocols that induce plasticity at a cellular level into sensory stimulation protocols applicable in humans.For example, high-frequency stimulation is used to induce long-term potentiation (LTP), whereas low-frequency stimulation evokes long-term depression (LTD) . ApplyinWhile these forms of cortical reorganization imply the presence of plastic processes, up to now it remained unclear whether LTP-like protocols led to a robust and maintained repetitive activation of the somatosensory cortex, which would be the prerequisite for the induction of plastic LTP-like processes. We therefore performed continuous EEG recordings over the hand representation of the somatosensory cortex during a period of 40 min of 20 Hz intermittent stimulation of the index finger. To avoid contamination of the EEG signal with artifacts arising from electrical stimulation, we used a pneumatic device allowing reproducible air-puff stimulation of the skin . We founThe study consists of two separate experiments, performed with two independent groups of healthy, right-handed participants . Handedness of participants was confirmed using the Edinburg Handedness Inventory . Afterward, another two baseline measures were assessed: one on the right (pre2-right) and on the left index finger (pre-left), in randomized order. Subsequently, repetitive sensory stimulation was applied for 30 min during which participants watched an animal documentary. Finally, a post measure of tactile acuity was assessed on each index finger, in randomized order (post-right and post-left).®; see ®), which were placed in an adjacent room to reduce operating noise. The airflow was transmitted to the participant using plastic standard tubes (Festo®), which inflated the circular membrane attached to the skin . The latency between the Master 8 trigger output and the air-puff arriving at the membrane was 25 ms. This latency was subtracted in the presented data, so that the latency of 0 reflects the arrival of the stimulus on the skin. For a detailed description see In both experiments, repetitive sensory stimulation was applied to the index-, middle- and ring-fingers of the right hand using an airflow-driven membrane  with a sampling rate of 512 Hz. Participants sat in a comfortable chair inside of a Faraday cage. Before electrode placement, the skin was cleaned with alcohol and prepared with SkinPure preparation gel. The Ag/AgCl electrodes were placed with Elefix conduction gel and arranged according to the international 10–20 system . Additionally, four ocular electrodes were applied. Baseline measures alternated between two eyes-open (each lasting 1 min) and eyes-closed conditions (each lasting 30 s). Both eyes-open conditions were combined to serve as the baseline measure.EEG-recordings were performed using a 13-channel DC-EEG amplifier  was band pass filtered between 1 and 250 Hz, notch filtered around 47 and 53 Hz with a linear finite impulse response (FIR) filter, and segmented into 7 s epochs. Ocular artifacts were removed from the EEG signals using least mean squares regression . The corTime-frequency analyses were performed using Morlet wavelet convolution . Resulting power values were normalized with the following formula:where the average time interval of 700–400 ms before stimulation onset over all trials and conditions constituted the baseline.Intertrial phase coherence values (ITPC) were derived by transforming phase angles of the complex wavelet transformation with the following formula:T-test to each data point and corrected for multiple comparisons using a cluster-based procedure. In this approach, the 1% largest clusters, that are adjacent data points reaching significance, were identified for each iteration. Only clusters the size or larger than the 1% largest of these clusters, were considered significant. To analyze possible changes of evoked responses over 40 min of stimulation, we compared the first and last 10 min of stimulation . To this end, we used a similar procedure but shuffled the condition instead of the temporal domain in each iteration. Furthermore, condition differences were tested with Welch’s Test appropriate for within subject designs. The same multiple comparison correction was applied. To analyze signal processing dynamics over time more closely, we divided 40 min of stimulation into four segments of 10 min each. Frequencies and time periods of interest were analyzed using repeated measures ANOVA, correcting for multiple comparisons with Tukey-Kramer post-hoc tests. Finally, tactile acuity changes were analyzed using paired T-Tests. All statistical calculations were performed in MATLAB R2019a.Stimulus evoked power- and phase-changes from baseline were analyzed using non-parametrical permutation testing . The timGrand average sensory evoked potentials recorded during 40 min repetitive intermittent 20 Hz stimulation of the fingertips, display clearly discernible components  analysis over all participants revealed a large synchronous activation right after stimulation onset, which comprises all measured frequencies from 3 to 60 Hz. For the 3 Hz band, this synchronization lasts up to 850 ms, while it is reduced for higher oscillatory frequencies to around 200 ms duration see . Most noTo further investigate the stability of the 20 Hz cortical responses induced by repetitive sensory stimulation over time, we compared the first and the last 10 min of the 40-min stimulation period. No significant power differences could be observed between time points overall see . The iniAdditionally, to monitor the time course of power and phase changes more closely, we performed repeated measures ANOVAS comparing four 10-min bins of stimulation. The ERD present from 500 to 2500 ms after stimulation onset between 10 and 13 Hz declined significantly over time see . ConversTo provide evidence that the early SEP components are subject to stimulation-induced changes we analyzed separately the cortical responses evoked during the first and the last 10 min of stimulation . SignifiT(9) = 3.18; p = 0.011; dz = 1.01], indicating improved tactile acuity (see T(9) = 0.80; p = 0.447].To examine, whether pneumatic air-puff stimulation induces similar effects on tactile acuity as the typically applied electrical stimulation , we compuity see . In contOur results demonstrate that 20 Hz repetitive sensory stimulation delivered through air-puffs onto the fingertips induces a 20 Hz steady-state response in the hand representation of somatosensory cortex, which is maintained over a period of 40 min with no indication of habituation. In a second experiment, we could show that this type of stimulation increases tactile acuity of the fingertips to an extent comparable to electrical stimulation .Temporal processing induced by cutaneous stimulation has been studied extensively. Touch stimuli are transmitted by mechanoreceptors in the glabrous skin of the fingertip. It is likely that FA-I (fast-adapting type I) Meissner endings, sensitive to high frequency dynamic skin deformation, are involved in transmitting the 20 Hz repetitive sensory stimulation. Another type of receptors possibly contributing as well, are the FA-II (fast-adapting type II) Pacini endings, which are highly sensitive to mechanical transients and high-frequency vibrations . Touch iWhen using electrical stimulation of the median nerve, there is general agreement that the N20 component originates mainly in the granular layer (layer IV) of Brodmann’s area (BA) 3b, which occupies the posterior bank of the rolandic fissure . The oriCompared to electrical nerve stimulation, air-puff stimulation applied to the fingers will not alter the potential origins of the different components but will markedly modify latencies and peak properties due to less synchronized afferent activation. As illustrated in About 500 ms after stimulation onset, the evoked potentials accumulate into a steady-state response characterized by a peak at 20 Hz. Steady-state evoked potentials (SSEPs) are recorded using repetitive stimuli in a range between 5 and 50 Hz, and are analyzed using a frequency domain analysis. They elicit cortical responses, which reach maximal amplitudes in certain frequency ranges, specific for each sensory modality. In the visual system, the amplitudes of steady-state EPs are largest close to 10 and 18 Hz for unpatterned flash stimuli and at somewhat lower frequencies for patterned stimuli . In the Using the mean phase coherence as a statistical measure for phase synchronization, we further analyzed the response properties of the evoked potentials during the intertrial phase. A first broad phase coherence covering all frequency bands most likely reflects the initial SEP components, while the following steady-state phase is reflected in the subsequent 20 Hz coherence. A less distinct, but still significant phase coherence is also apparent in the 40 Hz frequency band for the whole duration of the stimulation train, which most likely reflects the first harmonics. Repeated measures comparisons of the first 10 and the last 10 min of stimulation did not reveal significant differences in coherence, suggesting a maintained cortical steady state response without signs of habituation.A significant event-related synchronization in the low-frequency ranges during the first 500 ms can be assumed to reflect the transient sensory evoked potentials, while the synchronization in the 20 Hz frequency range probably illustrates the steady-state response. The prominent event-related desynchronization (ERD) covering the beta and low gamma range evoked after onset and offset of the stimulation train most likely reflects the transients of the sensory evoked potentials see . During The strength of event-related cortical responses and harmonics differed between participants. It is conceivable that differences in cortical responses during repetitive sensory stimulation are markers for effective plasticity and learning processes in the brain and only in the right combination, optimal learning conditions can be established. Therefore, future studies should analyze the effects of pneumatic repetitive sensory stimulation and perceptual learning in the same participants.Although the ERD in the alpha band slightly declined over time, both phase coherence and stimulus evoked power in the 20 Hz range did not change over the course of stimulation. This is an important observation as it indicates that prolonged periods of identical types of stimulation will not lead to habituation of the response. The diminished ERD can be interpreted as a signature of cortical plasticity processes indicating enhanced excitability. In this case, over the period of stimulation, less and less alpha ERD is required to maintain the same level of stimulus processing. To further support this claim, we could show that the N1, N2, and P2 component of the event-related potential responses to the stimulation train was significantly altered after 20 min . The obsIn cellular research, long-term potentiation (LTP) and long-term depression (LTD) of synaptic transmission comprise persistent forms of activity-dependent changes in synaptic strength . TypicalSome limitations of our study need to be addressed. Monitoring of potential finger flexor/extensor muscle activity could improve the signal-to-noise ratio of tactile acuity measurements, as the possibility of involuntary movement could be taken into account.Furthermore, aiming to increase the cortical activation elicited by air-puff stimulation, we stimulated three fingers simultaneously , leading to a mix of median nerve (digit 2 and 3) as well as ulnar and radial nerve (digit 4) mediated activation. As such, comparison to more specific stimulation approaches are limited.We have already discussed the differences between SEP characteristics obtained by median nerve and by cutaneous finger stimulation. Direct comparison is hampered by a number of factors, most critical probably the fact that median nerve stimulation is applied to the wrist, thereby activating different somatic structures as well as motor fibers, which is not the case when using finger stimulation.Finally, it would be of great interest to combine experiment 1 and 2 in the same cohort of participants, offering the opportunity to correlate psychophysical and electrophysiological data, a direction that should be explored in the future.Our data demonstrate that repetitive stimulation using a 20 Hz protocol drives rhythmic activation in the hand representation of somatosensory cortex, which is sustained during the entire stimulation period. At the same time, cortical excitability increases as indicated by altered ERD and SEP amplitudes, while on a perceptual level, tactile acuity is improved. Our results, together with previous data underlining the dependence of repetitive sensory stimulation effects on NMDA-receptor activation, support the view that repetitive sensory stimulation elicits LTP-like processes in the cortex, thereby facilitating perceptual learning processes.The datasets generated for this study are available on request to the corresponding author.The studies involving human participants were reviewed and approved by the Ethics Committee of the Ruhr-University Bochum. The patients/participants provided their written informed consent to participate in this study.MB and MS carried out the experiment, performed the data acquisition, and analyzed the data. MB, MK, and HD designed the study, interpreted the data, and drafted the manuscript. All authors contributed to the article and approved the submitted version.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.


New variants of the severe acute respiratory distress syndrome coronavirus 2 (SARS-CoV-2) are emerging around the world and causing widespread concern regarding their ability to escape natural or vaccine-induced immunity and available therapeutics. Here, we will briefly review the potential clinical implications of these new variants. Since its emergence in late 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has given rise to a global pandemic that remains uncontrolled, in part related to the virus’ ability to adapt. Through genomic surveillance of SARS-CoV-2, a number of variants have recently been identified. Although the emergence of viral variants is anticipated with ribonucleic acid (RNA) viruses such as SARS-CoV-2, it is critically important to determine the impact of these variants on pandemic control and, in particular, on therapeutics and vaccines. This primer is not intended as an exhaustive review of all SARS-CoV-2 variants. Instead, here, we use some of the known variants to illustrate the potential challenges arising from their emergence.RNA viruses, including coronaviruses, acquire mutations quickly during viral replication in the host cytoplasm. Despite the presence of RNA proofreading activity that yields high replication fidelity, coronaviruses have several features that provide opportunities to acquire mutations compared with DNA viruses . The terSince it was first identified in Wuhan, China, the SARS-CoV-2 genome has developed several mutations. Of particular clinical relevance are those mutations in the S protein. The S protein is responsible for binding the virus and is the target for neutralizing antibodies during infection as well as antibody-containing treatments and vaccines. Neutralizing antibodies bind to the S protein through the receptor-binding domain (RBD), and by doing so, these antibodies prevent the virus from attaching to the angiotensin-converting enzyme 2 (ACE2) receptor on human cells  emerged and became the dominant circulating variant globally by June 2020 4G emerge. AlthougDespite sharing some of the same mutations, the 3 main variants emerged independently of each other. In September 2020, the B.1.1.7 variant was first detected in the United Kingdom and has rapidly spread around the world  (3).The B.1.1.7 variant has different biological properties compared with previous variants. It has accounted for approximately 28% of SARS-CoV-2 cases in England, with population genetic models suggesting that it spreads 56% more quickly than other variants . HeighteFurthermore, the P.1 variant has been linked to reinfection. It was detected in 42% of cases during a devastating surge in Manaus, Brazil, where it was estimated that 76% of the population had already been infected with SARS-CoV-2 . These rThe possibility of worse outcomes due to these variants is currently being studied carefully. At the time of this writing, the data are inconclusive. Preliminary studies disclosed that the B.1.1.7 strain was not associated with increased hospitalization, 28-day case fatality, or reinfection ; howeverAnother potential consequence of emerging variants is their ability to evade detection by diagnostic tests. Most available reverse-transcription polymerase chain reaction– based tests have been designed to detect specific RNA sequences found in the viral genome. These molecular tests are highly sensitive, as they identify multiple SARS-CoV-2 genetic targets, making them less vulnerable to potential pitfalls. However, no test is perfect. False negative results are possible if mutations occur in the parts of the virus’ genome assessed by that specific test. For example, a mutation present in the B.1.1.7 variant (69/70 deletion) has been found to affect the performance of some diagnostic molecular assays . ReportiOne of the greatest concerns with emerging SARS-CoV-2 variants is the potential to escape natural or vaccine-induced immunity. A number of the S protein mutations have raised concern for escape from neutralizing antibodies produced by convalescent sera or monoclonal antibodies and potential decrease in protection by vaccines. This may also mean that patients with a prior SARS-CoV-2 infection could become reinfected with emerging new variants. The effectiveness of other existing therapies such as remdesivir should not be altered, as the target for this drug has not been associated to the known mutations.In vitro studies have shown that neutralizing antibodies in convalescent sera have a lack of potency against the B.1.351 variant. Cocktails of monoclonal antibodies binding different areas in the S protein appear to retain activity against some of these emerging variants, although they may be less effective against B.1.351 . With reIn the current primer, we have described the main emerging variants of SARS-CoV-2 and the potential impact on existing therapies. It is expected that SARS-CoV-2 will continue to acquire new mutations that will give rise to new variants. For the present, there is cause for optimism with respect to the efficacy of the current generation of vaccines against some of the emerging variants of SARS-CoV-2. However, it should be emphasized that we do not have detailed knowledge about the efficacy of the vaccines with many of the emerging variants. Given that each newly infected person represents an opportunity to acquire new mutations and develop novel variants, slowing the spread of the virus through continued public health interventions not only is critical to prevent further morbidity and mortality, but also can potentially impact the emergence of viral variants.Both authors have reported that they have no relationships relevant to the contents of this paper to disclose.


Understanding the role of children in the transmission of SARS-CoV-2 is urgently required given its policy implications in relation to the reopening of schools and intergenerational contacts.We conducted a rapid review of studies that investigated the role of children in the transmission of SARS-CoV-2. We synthesized evidence for four categories: 1) studies reporting documented cases of SARS-CoV-2 transmission by infected children; 2) studies presenting indirect evidence on the potential of SARS-CoV-2 transmission by (both symptomatic and asymptomatic) children; 3) studies reporting cluster outbreaks of COVID-19 in schools; 4) studies estimating the proportions of children infected by SARS-CoV-2, and reported results narratively.A total of 16 unique studies were included for narrative synthesis. There is limited evidence detailing transmission of SARS-CoV-2 from infected children. We found two studies that reported a 3-month-old whose parents developed symptomatic COVID-19 seven days after caring for the infant and two children who may have contracted COVID-19 from the initial cases at a school in New South Wales. In addition, we identified six studies presenting indirect evidence on the potential for SARS-CoV-2 transmission by children, three of which found prolonged virus shedding in stools. There is little data on the transmission of SARS-CoV-2 in schools. We identified only two studies reporting outbreaks of COVID-19 in school settings and one case report of a child attending classes but not infecting any other pupils or staff. Lastly, we identified six studies estimating the proportion of children infected; data from population-based studies in Iceland, Italy, South Korea, Netherlands, California and a hospital-based study in the UK suggest children may be less likely to be infected.Preliminary results from population-based and school-based studies suggest that children may be less frequently infected or infect others, however current evidence is limited. Prolonged faecal shedding observed in studies highlights the potentially increased risk of faeco-oral transmission in children. Further seroprevalence studies (powered adequately for the paediatric population) are urgently required to establish whether children are in fact less likely to be infected compared to adults.https://www.ed.ac.uk/usher/uncover/completed-uncover-reviews.We plan to update this rapid review as new data becomes available. These updates are available at  COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was declared a pandemic on 11 March 2020 by the World Health Organization. Children are less likely to develop severe disease from COVID-19 compared to adults, although the reasons for this remain unclear . UnderstWe searched PubMed, medRxiv and the WHO COVID-19 database on 30 April 2020 with entry date limits from late 2019  and checked by a senior epidemiologist (ET). We synthesized evidence thematically and reported results narratively.A total of 993 publications were retrieved and 16 unique studies were finally included for narrative synthesis  from New South Wales traced the close contacts (735 students and 128 staff) of 18 initial COVID-19 cases (nine students and nine staff) to see who they subsequently infected. They found that two children (secondary cases) may have contracted COVID-19 from the initial cases at their schools [We identified six studies ,6-10 preOne study assessed environmental contamination in the isolation room of an infected infant . AlthougAt least three studies have shown that children show prolonged faecal shedding compared to adults (and in some cases longer than four weeks) -9. NotabIt is estimated that the proportion of infected children with latent asymptomatic or with mild symptoms of respiratory illness is higher than in adults, which highlights the possibility that children and young adolescents may be potential sources of undetected community transmission . A natioSpecific evidence related to the transmission of SARS-CoV-2 in schools is lacking, which is probably due to the early closure of schools at the start of the pandemic in many countries. We identified only two studies reporting a cluster outbreak of COVID-19 in school settings ,11 and oOne of these studies retrospectively followed up 661 pupils and staff in a French high school affected by an outbreak of COVID-19 . They foA school-based study from New South Wales, identified 18 individuals (nine pupils and nine staff) from 15 schools (10 high schools and five primary schools) that were confirmed COVID-19 cases . A totalOne case report describes a paediatric patient who was infected during a holiday in France . DespiteWe next sought to identify studies which estimated the proportion of children infected in the community. We included five studies -17 estimTargeted (n = 9199) and random population screening n = 13 607) from Iceland found that children under 10 were less likely to test positive than those 10 and over   607 from. A demogThis rapid evidence review summarises the currently available evidence on the role of children in SARS-CoV-2 transmission. The key messages based on the findings of this review are: (i) Case series and outbreak reports detailing transmission in children are few. Those studies which are available demonstrate that transmission by children is possible but do not quantify the likelihood of transmission in children compared to adults. (ii) There is theoretical evidence that alternative transmission routes such as faeco-oral transmission may pose a higher risk in children. Further investigation is required to confirm whether longer viral shedding times in stools in children pose a higher risk of transmission. (iii) There is little evidence on transmission dynamics in school settings. Given the current paucity of data, further investigation and close monitoring will be essential where schools have re-opened and in settings where schools have remained open. (iv) Children are infected, but perhaps less frequently than adults. However, these studies are limited by small numbers and additional community seroprevalence surveys are needed to confirm the proportion of children infected.Direct evidence showing children as a source of transmission is scarce and largely based on small studies or studies investigating few paediatric cases. We identified two studies indicating that children can transmit COVID-19 to other children or adults ,5, howevWhile there is lack of direct evidence on the frequency that children transmit the disease, indirect evidence illustrates the potential risk of viral transmission by children. First, there is evidence showing that an infected infant contaminated (with SARS-CoV-2) the surfaces they were in contact with, thus rendering the risk of transmitting the virus to caregivers through indirect contact with fomites . This fiFew studies have described SARS-CoV-2 transmission in schools. There are two studies that investigated school outbreaks of COVID-19 ,11 and oUnderstanding the proportion of SARS-CoV-2 infection in children will help to determine children’s relative contribution to SARS-CoV-2 transmission in the community. Based on the infection rates reported from Iceland, Italy, South Korea, Netherlands, and US -17, it shttps://www.ed.ac.uk/usher/uncover).There is insufficient evidence to robustly answer the key study question. The absence of conclusive evidence on children’s role in transmission increases the uncertainty in relation to policy decisions on school closures and re-openings. During the COVID-19 pandemic, many countries have taken precautionary measures by closing schools to slow transmission among children. As the rate of infection subsides in some areas, some countries have started re-opening schools, and this further highlights the urgency of obtaining robust data on transmission in children and monitoring the impact re-opening schools have on country-level epidemics. A phased re-introduction of face-to-face learning in schools has started in Taiwan (February), Denmark (April), Norway (April), China (May), France (May) and Germany (May). We are committed to reviewing the literature on this topic regularly as new data emerge and to re-evaluate the conclusions of this review given the rapid pace of ongoing research on COVID-19. These updates will be available online , are urgently required to establish whether children are in fact less likely to be infected compared to adults. Faeco-oral transmission appears to pose a higher risk of onward transmission in infected children compared to adults, given the longer faecal shedding time observed in several studies. This may have substantial implications for community spread in day-care centres, schools, and homes and the need for a high level of basic hygiene measures in these settings to limit spread.