Amid the COVID-19 pandemic the trend points to universal use of protective face masks. The impact posed on verbal and non-verbal communication by masks is yet another challenge to be addressed in clinical care.To reflect on the consequences of face mask and covering use on communication in the clinical setting, including mental health settings.Pubmed and Google Scholar literature search using terms face mask / face covering and communication / emotion.There is a lack of literature on the impact of protective face masks and coverings on communication in clinical settings. Face masks and coverings may have a significant impact on patient-healthcare professional relationship due to disruption of verbal  and non-verbal communication  with consequences on: 1) clarity of communication with potential for misunderstanding clinical information, advice and prescriptions posing safety issues, 2) emotion perception, expression and reciprocity, 4) perception of healthcare professionals’ empathy and therefore, 3) patient satisfaction, 4) quality of care, and 5) clinical outcomes. Difficulties in communication between the patients´family or other carers and healthcare providers and between healthcare professionals are likewise challenged. People with hearing impairment, children and people with mental illness may be especially vulnerable to these difficulties in communication.Protective face masks and coverings are undoubtedly important in preventing spread of COVID-19, nonetheless mental healthcare professionals should take into account their significant impact on verbal and non-verbal communication in clinical care. Alternative strategies to enhance communication and rapport may be warranted.


In response to the COVID-19 pandemic, children’s hospitals across the country postponed elective surgery beginning in March 2020. As projective curves flattened, administrators and surgeons sought to develop strategies to safely resume non-emergent surgery. This article reviews challenges and solutions specific to a children’s hospital related to the resumption of elective pediatric surgeries. We present our tiered reentry approach for pediatric surgery as well as report early data for surgical volume and tracking COVID-19 cases during reentry.The experience of shutdown, protocol development, and early reentry of elective pediatric surgery are reported from Levine’s Children’s Hospital (LCH), a free-leaning children’s hospital in Charlotte, North Carolina. Data reported were obtained from de-identified hospital databases.Pediatric surgery experienced a dramatic decrease in case volumes at LCH during the shutdown, variable by specialty. A tiered and balanced reentry strategy was implemented with steady resumption of elective surgery following strict pre-procedural screening and testing. Early outcomes showed a steady thorough fluctuating increase in elective case volumes without evidence of a surgery-associated positive spread through periprocedural tracking.Reentry of non-emergent pediatric surgical care requires unique considerations including the impact of COVID-19 on children, each children hospital structure and resources, and preventing undue delay in intervention for age- and disease-specific pediatric conditions. A carefully balanced strategy has been critical for safe reentry following the anticipated surge. Ongoing tracking of resource utilization, operative volumes, and testing results will remain vital as community spread continues to fluctuate across the country. Since its emergence in December 2019, global spread of the novel coronavirus (SARS-CoV-2) and its associated clinical syndrome, coronavirus disease 2019 (COVID-19), has had an unprecedented impact on health and economy on a national and international level and has presented a myriad of unique challenges in the delivery of modern health care. Specifically, for the pediatric population, the COVID-19 pandemic has had a unique effect on the delivery of ongoing medical and surgical care. Although many of the early responses in health care have been documented, the specific impact on the surgical care of children and the response of children’s hospitals to the pandemic has been relatively underreported.2 The first pediatric COVID-19 case was reported in the United States on March 2, 2020, and preliminary data by the CDC reported 2572 pediatric COVID-19 cases representing 1.7% of confirmed COVID-19 cases occurring between February 12 and April 2, 2020.3 Despite the reduced burden of disease in children, the widespread shutdown of hospital-based services has had a significant impact on children’s hospitals as health care systems adapted in preparation for a potential surge of cases. The majority of children’s hospitals in the United States postponed elective and day pediatric surgery cases following The Centers for Medicare and Medicaid Services (CMS) consensus statement that recommended suspension of elective surgery in order to conserve hospital resources and reduce unnecessary exposure to patients and health care personnel.4 The Children’s Hospital Association and the American College of Surgeons (ACS) published elective case triage guidelines for surgical care during the COVID-19 pandemic, recommending only emergent and urgent pediatric surgery cases be performed.6 At the institutional level, additional modifications to normal surgical practice were reported during the shutdown such as implementing intubation protocols, restricting aerosolizing procedures such as laparoscopy and robotic surgery, and restructuring perioperative staffing using staggered teams to minimize exposure risk.7Early epidemiologic data suggested that SARS-CoV-2 affects children of all ages; however, compared to adults, children are affected less frequently with the majority being asymptomatic or reporting mild symptoms.8 Recommendations from the Children’s Hospital Association and the American Pediatric Surgical Association included considerations for expanding non-emergent operations in order to reduce the risk of hospital admission or prolonged hospital stay which could increase exposure risk and resource utilization.10 The variability in pediatric surgical guidelines likely reflects the wide range of impact across children’s hospitals and the rapidly evolving understanding of the risk of exposure vs. the delay in surgical care. In light of the complexity of this decision-making, the authors present the experience of their institution to highlight the unique considerations many children’s hospitals faced and provide a tiered approach for resumption of non-emergent essential pediatric surgical care.Just as pediatric hospitals experienced a unique impact from the shutdown during the surge phase, similar challenges were faced with developing a strategic reentry plan for surgical care within the context of their unique patient population and resource environment. While guidelines for resuming elective surgery were jointly published by the ACS, American Society of Anesthesiologists, Association of periOperative Registered Nurses, and American Hospital Association, a similar set of guidelines specific to resuming pediatric surgical cases at children’s hospitals were not clearly outlined by pediatric surgical societies, with many deferring to the adult guidelines.Levine Children’s Hospital (LCH) is a 234-bed free-leaning children’s hospital located on the campus of Atrium Health Carolinas Medical Center in Charlotte, North Carolina, part of a 40-hospital system with 900 care locations across North Carolina, South Carolina, and Georgia. Levine Children’s Hospital supports an 85-bed Level IV neonatal ICU (NICU) and 20-bed combined pediatric/cardiovascular ICU, as well as a 13-bed inpatient rehabilitation unit and provides a full spectrum of pediatric services including a Level I pediatric trauma center, extracorporeal membrane oxygenation, cardiac surgery, neurosurgery, and a dedicated pediatric emergency department. All surgical patients at LCH are managed with dedicated pediatric anesthesia. Levine Children’s Hospital shares operative resources with the adult hospital but maintains 4 general and 2 cardiac pediatric operating rooms (OR) with additional dedicated ORs in the ambulatory surgery center.As the trajectory of COVID-19 cases began to stabilize in the region surrounding the authors’ institution, the leadership of LCH along with a pediatric-specific COVID-19 Surge and Re-entry Team set about drafting a strategic plan for safely resuming electively timed, appropriately indicated essential pediatric surgery. Our institution identified a set of critical principles necessary for safe reentry, each reflecting the complex shared decision-making required to ensure a balanced approach within the context of the continually evolving pandemic response . These iThe foundation for structuring a reentry protocol was understanding the relationship of the children’s hospital within our unique health care system and region. As a free-leaning children’s hospital, decisions to resume surgical care would have ramifications on the hospital’s primary shared resources including personal protective equipment (PPE) supply and distribution, blood bank and laboratory services, diagnostic and interventional radiology support, and recalling furloughed staff members. A clear pipeline of regular communication was immediately established with leadership teams to ensure that the shared responsibility of balancing the needs of all patients and health care workers was preserved with each step toward reentry. As the rollout process began, an accruing assessment of resource supply and demand as well as divisional protocols for safe care  was maintained.13 However, due to the extension and flattening of the predicted peak in the region’s population, resumption of electively scheduled, essential surgery was determined to be necessary and safe after the rate of COVID-19 cases was approaching its anticipated peak. Adult surgical reentry was, therefore, scheduled to begin on May 11, 2020. Due to the relatively lower impact of COVID-19 cases on the pediatric population and the availability of resources in LCH as a result of the shutdown, the pediatric surgery was selected to be the torchbearer and cases were scheduled to begin 1 week ahead on May 4, 2020.As regional trends of coronavirus cases varied widely across the country, safely resuming non-emergent pediatric surgical care was implemented in alignment with local and national health authority recommendations. Timing of reentry was initially based on the ACS and CMS guidelines of a sustained reduction in the rate of new COVID-19 cases.5 Similarly, redeployment of personnel, such as anesthesiologists, pediatric critical care intensivists, and nurses, to adult intensive care units reduced pediatric surgical resources. In many cases, reallocation of resources such as ventilators, nursing staff, and PPE back toward pediatric hospitals to safely support the ramping up of pediatric surgical care was a shared decision with the adult hospital to ensure resources remained adequate across the system for the current environment and the potential for a second surge.6The redistribution of resources  from many children’s hospitals to support the anticipated surge of adult COVID-19 patients presented an additional unique challenge. The unique relationship between each children hospital and its parent health system necessarily impacted the pathway for reentry in different ways. In areas of the country greatly affected by COVID-19 known as “hotspots,” children’s hospitals mobilized resources such as ventilators and PPE to adult hospitals facing critical shortages, while others converted children’s hospital units to care for adult COVID-19 patients.Our Chief Surgical Officer (JSK) identified 8 critical decision factors that served as the pillars for how to balance the resource assessment for a surgical reentry strategy . Each reFollowing a comprehensive assessment of resources, evaluating the volume of cases suspended during the shutdown of elective surgery was the next critical step to determining the appropriate reallocation of these resources. Prior to COVID-19 restrictions, operating room utilization at LCH was at near maximum capacity with an average case volume of more than 420 cases per month in January and February 2020 immediately preceding the cessation of elective surgery on March 17, 2020. During the shutdown, pediatric surgery at LCH decreased to only 55% of the weekly case volume compared to the preceding months (January/February 2020). Similarly, compared to an identical time frame in the previous calendar year (March/April 2019), pediatric surgical case volume decreased to 53% . This re14 During the prioritization process, compiling a list of previously canceled/postponed cases and calculating the required resources and staff for each may better enable objective assessment of the backlog of surgical cases and new cases needing to be scheduled.12 At our institution, our tiered approach for reentry and resuming pre-COVID-19 surgical volume was to initially target the healthiest patients with the highest need and potential benefit from their procedure.Appropriate prioritization of surgical cases is critical to the development of a tiered approach for resuming elective surgery. For surgeries that can be performed in a non-emergent or nonurgent fashion, it is important to balance both need and benefit to the patient.10 Within the pediatric population, many operations require careful timing within a patient’s growth and development and substantial delay may hold significant consequences. Repair of congenital malformations inherently balance timing between ensuring an adequate size of the child with reestablishing function as early as possible. Musculoskeletal malformations and injuries both require careful consideration of development and healing for the timing of surgical intervention. Similarly, a delay in repair of gastrointestinal anomalies may impact a child’s nutritional support and functional ability during a critical development period.Pediatric-specific considerations that further added to the complexity of case prioritization include the impact of delaying surgery on age- and growth-dependent interventions for congenital conditions.In light of the relatively lower morbidity and mortality of COVID-19 within the pediatric population, the risks of delaying necessary surgical interventions in children was considered even more strongly. Thus, pediatric patients were selected to pilot the first phase of surgical reentry. A tiering system was developed to assist surgeons and surgical teams with case prioritization . Given tDelivering safe, uncompromised patient care while maintaining flexibility in the event of a second potential surge remained the highest priority in our coordinated effort to resume surgical cases. Redistribution of surgical cases according to the patient and procedure allowed for healthy patients with acceptable case level acuity to be preferentially directed to ambulatory surgical centers, reserving hospital outpatient and inpatient settings for patients that necessitated higher levels of care. With these considerations, the health care system initiated the reentry pilot phase with outpatient cases in the ambulatory surgery center and hospital outpatient department settings 1 week prior to resuming elective inpatient cases. A limited volume of pediatric cases was first scheduled in the hospital outpatient surgical center 1 week prior to resuming elective cases in the inpatient OR. The projected timeline was to resume 100% of pre-COVID-19 caseload volume within 6 weeks with a further ramp up to 120% until the backlog of cases has been addressed.Screening and testing practices have been essential for limiting spread and avoiding potential exposures during the surge phase of COVID-19. However, perioperative screening and testing is a constantly moving target as rates of asymptomatic or minimally symptomatic carriers continue to rise and laboratory testing technology evolves rapidly.1 Additionally, early data in the adult population demonstrated worse outcomes after surgery for COVID-positive patients.15 Consequently the decision was made to screen and test all children prior to elective surgery at LCH to promote the safety of the patients, other children, and health care providers.While children are affected by COVID-19 less frequently and less severely than adults, this may lead to a lower rate of testing in children in the community, thus underestimating the true burden of disease and raising the potential for children being asymptomatic carriers of transmission. Importantly, the severity of illness widely varies by age with 1 study of pediatric COVID-19 patients reporting severe or critical symptoms in 10.6% of infants less than 1 year of age.Based on the best available evidence and consensus within the LCH Surge and Re-entry Team, a standardized surgical COVID-19 safe screening and testing procedure was established for all elective surgery at LCH . All patOnce the reentry process was initiated, intentional tracking of surgical case volumes, pre-procedural COVID-19 screening and testing results, as well as continued trends of COVID-19 within the community were essential to ensuring the safety of providers and patients. In the first 8 weeks of reentry (May 4-June 21), 721 pediatric operations were performed at LCH. As shown in Post-procedural screening is also critical to track any incidence of potential hospital-acquired transmission of COVID-19 and identify the source of transmission as early as possible. Through the first 6 weeks of reentry, 7 patients (1.3%) have failed post-surgery screening follow-up with influenza-like illness symptoms or concern for contact with other COVID-19-positive individuals. Of these, 1 patient met criteria for COVID-19 testing which was negative. All others were instructed to quarantine and had self-limiting symptoms.In response to the evolving COVID-19 pandemic, health care facilities and surgical practices faced numerous challenges as they rapidly adapted to dramatic reductions in surgical volumes with cancellation of non-emergent/urgent procedures and disruption of perioperative resources and staff. Planning for surgical reentry presented additional challenges for children’s hospitals, particularly those incorporated within a larger health care system that may share critical perioperative resources with adult hospitals. Navigating the logistics of surgical reentry at such larger institutions can be complex and often requires balancing the needs of multiple surgical service lines and consideration of numerous factors. The described process and template, in combination with a decisive leadership team, were fundamental in the development and implementation of a tiered approach to surgical reentry at the authors’ institution. As children’s hospitals return to elective surgery and address significant backlogs, it is paramount to select the appropriate pace, patients, and location for reentry and ensure adequate resources are available. Such decisions should not be made by health care institutions in a vacuum, but rather in conjunction with regional government and public health departments in a balanced and collaborative manner, so as not to jeopardize the health of the local community. However, as community and hospital cases of COVID-19 begin to rise again following recently relaxed social distancing restrictions across the country, readiness to scale back surgical reentry in an expeditious time frame must be continually reevaluated.


JCI, Glinton et al. used state-of-the-art approaches to demonstrate that efferocytosis induced vascular endothelial growth factor C (VEGFC) in myeloid cells and stimulated lymphangiogenesis and cardiac repair. These findings provide impactful mechanistic information that can be leveraged to therapeutically target pathways in cardiac repair and ischemic heart failure.Cardiac repair following ischemic injury is indispensable for survival and requires a coordinated cellular response involving the mobilization of immune cells from the secondary lymphoid organs to the site of damage. Efferocytosis, the engulfment of cell debris and dying cells by innate immune cells, along with lymphangiogenesis, the formation of new lymphatic vessels, are emerging as central to the cardiac healing response. In this issue of the  Ischemic heart disease is estimated to affect over 120 million people worldwide. Immune and fibrotic responses work in tandem to repair the heart after ischemia by physically stabilizing the infarct and removing cell debris through efferocytosis. However, the same fibrotic scar that is critical for preventing cardiac rupture and death early on contributes to later systolic dysfunction and chronic heart failure (HF). A better understanding of the coordinated healing response is necessary to discover improved therapeutics for ischemic heart disease.The immune response to ischemic myocardial injury mobilizes innate and adaptive immune cells to the site of injury . EfferocVegfc is embryonically lethal because of the lack of lymphatics  family in mammals is composed of VEGFA, placental growth factor, VEGFB, VEGFC, and VEGFD, which signal though VEGFR1 , VEGFR2 , and VEGFR3 . Most relevant to the current discussion, genetic knockout of mphatics , and themphatics . Macroph of mice , and int of mice .JCI, Glinton et al. (–/–Cd36). They initially observed that cardiac antigen accumulation in myeloid cells in the lymph nodes after MI, concomitant with increased myocardial expression of VEGFC and the lymphatic endothelial cell marker lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1). Cell-intrinsic evidence that efferocytosis induced VEGFC was provided in vitro, as apoptotic cells added to primary cardiac macrophages resulted in elevated Vegfc mRNA levels and VEGFC secretion. –/–Cd36 mice showed reduced myeloid accumulation of cardiac antigen in the lymph nodes, a striking reduction of myocardial LYVE1 and VEGFC expression, and decreased cardiac macrophage VEGFC expression compared with control mice, further supporting the notion that VEGFC expression after MI depends on myeloid CD36. These results additionally suggest that macrophage-derived VEGFC induces lymphangiogenesis downstream of efferocytosis  in mice, they found that LYVE1 expression was increased in myocardial lymphatics at the post-I/R risk area and that macrophage Vegfc was required for improved cardiac function after reperfusion. These studies confirm a central role for myeloid-derived VEGFC in lymphangiogenesis and cardiac repair with consequences on systolic function  expression, accounting for reduced myocardial inflammation seven days after MI. Conversely, –/–Cd36 mice showed a higher fraction of myocardial macrophages expressing high levels of MHCII, supporting the notion that efferocytosis is upstream of these signals. In line with myeloid-derived VEGFC regulating adaptive immunity through myeloid MHCII expression, Glinton et al. (–/–Vegfc mice were required for post-MI cardiac repair (–/–Vegfc mice had elevated Tnfa, Il6, and Il12 gene expression that could be reduced with administration of VEGFC. In addition to the paracrine function of macrophage-produced VEGFC modulating cardiac lymphangiogenesis and immune function, these observations support an autocrine function for VEGFC through VEGFR3 to suppress excessive secretion of proinflammatory cytokines. This possibility was partially corroborated in vivo with the pharmacological inhibition of VEGFR3 using MAZ51, which reduced cardiac antigen uptake by macrophages (Vegfr3-deficient mice could determine whether this autocrine proinflammatory pathway in macrophages accounts for the observed protection of MAZ51.Glinton and colleagues considered the complexity of the immune, fibrotic, and angiogenic responses in the heart following ischemia and investigated the role of myeloid VEGFC in the immune response during cardiac repair. They combined RNA-Seq and flow cytometry of cardiac tissue to characterize the immune landscape, discovering that myeloid deficiency of n et al.  also fouc repair , 15 compGlinton et al.  provide Vegfc transcription and release remain unknown. CD36 is a fatty acid transporter, and while Glinton et al. (In addition, the specific molecular signals downstream of CD36 that lead to n et al.  used inhn et al. .Although the contribution of cardiac lymphangiogenesis to improved post-MI cardiovascular outcomes is evident  and supp


Abnormal glucose metabolism is central to neurodegeneration, and considerable evidence suggests that abnormalities in key enzymes of the tricarboxylic acid (TCA) cycle underlie the metabolic deficits. Significant recent advances in the role of metabolism in cancer provide new insight that facilitates our understanding of the role of metabolism in neurodegeneration. Research indicates that the rate-limiting step of the TCA cycle, the α-ketoglutarate dehydrogenase complex (KGDHC) and its substrate alpha ketoglutarate (KG), serve as a signaling hub that regulates multiple cellular processes: (1) is the rate-limiting step of the TCA cycle, (2) is sensitive to reactive oxygen species (ROS) and produces ROS, (3) determines whether KG is used for energy or synthesis of compounds to support growth, (4) regulates the cellular responses to hypoxia, (5) controls the post-translational modification of hundreds of cell proteins in the mitochondria, cytosol, and nucleus through succinylation, (6) controls critical aspects of transcription, (7) modulates protein signaling within cells, and (8) modulates cellular calcium. The primary focus of this review is to understand how reductions in KGDHC are translated to pathologically important changes that underlie both neurodegeneration and cancer. An understanding of each role is necessary to develop new therapeutic strategies to treat neurodegenerative disease. Exciting new research demonstrates how metabolism and mitochondria are central to processes causing neurodegeneration. Stimulating new research on metabolism in cancer suggests multiple new ways to approach the dynamic role of metabolism in neurodegenerative processes. To reveal how transformative these findings are, we first present an overview of the role of the α-ketoglutarate dehydrogenase complex (KGDHC). KGDHC regulation of the tricarboxylic acid (TCA) cycle places it central to neurodegeneration and cancer ,2,3 Sec. Decades+ to NADH, FAD+ to FADH2, and produces GTP. NADH and FADH2 pass their electrons to the protein complexes of the electron transport chain to generate ATP. However, the NADH produced from KGDHC can be directed to the enzyme nicotinamide nucleotide transhydrogenase (NNT) to transfer hydride ions to NADH forming NADPH . Seq. Seq8]. nd KGDHL .. The E3 subunit is inhibited by ATP and NADH -glucose and [U-13C] glutamate indicate that inhibition of KGDHC with specific KGDHC inhibitors in intact cerebellar granule neurons alters metabolism. The findings show that decreased KGDHC activity leads to increased KG levels and increased transamination of KG with valine, leucine, and GABA [The wide scope of KG actions raises the question of whether KGDHC controls KG levels. It has directly been demonstrated that inhibiting KGDHC increases KG levels. Measurements of KG after inhibition of KGDHC demonstrate a 50–60 percent to four-fold increase in KG levels depending on concentration and timing of inhibition ,113. Simand GABA .KGDHC and KG may also regulate cell metabolism and function through an interaction with Hypoxia Inducible Factor 1 (HIF1), a protein complex that is responsible . The α-sHypoxia inhibits KGDHC activity through SIAH2. Under hypoxia, there is no oxygen present to activate PHDs, which results in the stabilization of HIF. HIF activates the E3 ubiquitin-protein ligase SIAH2 which inhibits KGDHC. Since KGDHC is inhibited, KG cannot be oxidized into succinyl-CoA. Instead, KG will enter the reductive pathway of the TCA cycle, promoting lipid synthesis. Hypoxia Results in Accumulation of KG Which Reactivates PHD. With KGDHC inhibited, KG will begin to accumulate in the mitochondria. The KG that is produced through glutaminolysis leaves the mitochondrial and enters the cytosol. This KG activates PHD in the cytosol which then destabilizes HIF. So even under hypoxia, KGDHC regulation plays a role in response to hypoxia. Accumulation of KG prevents the activation of HIF1 by succinate, fumarate, and hypoxia. This reverses cell death and enhances glycolysis ,119. ThiThus, in cancer cells, a clear feedback loop exists between KGDHC and HIF. This circuit is necessary for the growth of cells in hypoxia and models of tumor formation but has not been explored in neurodegeneration. Not only does the inhibition of KGDHC control the activity of HIF through KG levels, but HIF controls the inhibition of KGDH through SIAH. Active HIF increases SIAH2 which ubiquitinates KGDHC and limits its activity up to 60% . Inhibit2, and NADH. KGDHC is now known to cause changes in cell function by controlling the very precise post-translational modification succinylation. Succinylation of lysine groups induces more substantial changes to a protein’s chemical properties than does lysine acetylation or methylation, as it adds a bigger structural moiety and changes the charge from a positive to a negative charge [Post-translational modifications of proteins by succinylation are regulated by KGDHC and provide an efficient and rapid biological regulatory mechanism that links metabolism to protein and cell functions. Traditionally, changes in KGDHC were thought to be transferred to cell function by changes in ATP/ADP, GTP, FADHe charge . The sece charge .+/NADH ratio. Multiple metabolic perturbations including reduced glycolysis, glutathione depletion, depressed TCA cycle activity, impairment of electron transport, ATP synthase, and uncouplers of oxidative phosphorylation dramatically decreased succinylation. In contrast, reducing the oxygen from 20% to 2.4% increased succinylation. The results demonstrate that succinylation varies with metabolic states, is not correlated to the mitochondrial NAD+/NADH ratio, and appears to coordinate the response to metabolic challenges [Multiple experiments in cells support the hypothesis that succinylation links altered metabolism to multiple cellular functions. Changes in succinylation of mitochondrial proteins following variations in metabolism were compared against the mitochondrial redox state as estimated by the mitochondrial NADallenges .KGDHC also controls succinylation in neuronal cell models ,80 Figu. ReductiThe human brain contains hundreds of succinylated proteins . A succiReductions in KGDHC activity have been reported in numerous diseases see . Since sThe changes in succinylation with AD are very specific and are closely linked to the development of the pathology . A compaOur understanding of the interactions of various post-translational modifications is in its infancy. Since both acetylation and succinylation modify lysine groups, many proteins can be modified at the same sites . The intThe signaling by KGDHC and KG through succinylation extends to the nucleus . The sucThe TCA cycle enzymes in the mitochondria are also present in the nucleus  and are Both the enzyme KGDHC and the substrate KG are involved in epigenetics. Tumorigenesis can be altered by epigenetics at the histone level as well as the DNA level . The varOne example of how KGDHC and KG regulation of gene expression may be important is in the regulation of autophagy and apoptosis by p53. In cells with low KG, activation of P53 induces ATG gene expression and apoptosis. Reduced autophagy-related gene expression contributes to P53-induced apoptosis. Addition of KG either by reducing KGDHC or adding cell permeable KG overcomes the P53 induction of apoptosis. Reduced levels of KG increase apoptosis and decrease autophagy while increased levels of KG promote apoptosis resistance and increase autophagy .KGDHC has been implicated in protein signaling through its ability to induce the release of mitochondrial proteins into the cytosol . ReductiThe capability of mitochondria to control complex cellular protein signaling with the cytosol is well documented in regard to apoptosis (for review see ). ProteiThe consequences of KGDHC inhibition on protein signaling may be mediated by alterations in surface charge on the mitochondria. Inhibition of KGDHC alters the surface charge of the mitochondria which can alter protein signaling. Increased annexin V fluorescein following treatment of cells with CESP for one hour indicates a change in surface charge which corresponds to changes in mitochondrial signaling ,149.Studies of reduced KGDHC activity inducing mitophagy support the suggestion that diminished KGDHC activity can be protective in the short-term. Hence, increased fission and mitophagy induced by mild impairment of mitochondrial function is protective in the short-term, but ultimately makes the cells more sensitive to other conditions that impair metabolism. Other studies demonstrate that a short-term reduction in KGDHC activity causes a different response than long-term or chronic reductions in both calcium levels and the response to external oxidants . For exaWhile the role of calcium in activating mitochondrial dehydrogenases such as KGDHC has been studied extensively ,150,151,A highly replicable change in AD cells is enhanced IP3-releasable calcium from the ER. This was first reported in fibroblasts from patients with non-genetic forms of AD  and was 3 receptor-evoked calcium release may involve the increase of stromal interaction molecule 1 (STIM1) expression and the decrease of STIM2 expression in the soma. These STIMs translocate to plasma membrane–ER contact sites, where they bind to Orai1 channels and open a Ca2+ influx pathway to slowly replenish ER Ca2+ stores using extracellular Ca2+ [The mechanism for the association of KGDHC to calcium is likely related to the ability of KGDHC to alter multiple protein cellular signals see . The callar Ca2+ ,158). ThKGDHC is rate limiting for energy production by the TCA cycle and new research demonstrates the exciting role KGDHC plays in the cell including the response to oxidative stress, cell signaling, post-translational modification, epigenetics, and diverse metabolic pathways as well as the surprisingly specific signal changes. These intriguing results support the hypothesis that thiamine-dependent enzymes like KGDHC are important therapeutic targets to treat diseases from neurodegeneration to cancer.


Oryza sativa), maize (Zea mays), soybean (Glycine max), and tomato (Solanum lycopersicum) are short-day plants whose photosensitivity was reduced or lost during domestication and expansion to high-latitude areas. Wheat (Triticum aestivum) and barley (Hordeum vulgare) are long-day crops whose photosensitivity is influenced by both latitude and vernalization type. Here, we summarize recent studies about where these crops were domesticated, how they adapted to photoperiodic conditions as their growing area expanded from domestication locations to modern cultivating regions, and how allelic variants of photoperiodic flowering genes were selected during this process. A deeper understanding of photoperiodic flowering in each crop will enable better molecular design and breeding of high-yielding cultivars suited to particular local environments.Flowering links vegetative growth and reproductive growth and involves the coordination of local environmental cues and plant genetic information. Appropriate timing of floral initiation and maturation in both wild and cultivated plants is important to their fitness and productivity in a given growth environment. The domestication of plants into crops, and later crop expansion and improvement, has often involved selection for early flowering. In this review, we analyze the basic rules for photoperiodic adaptation in several economically important and/or well-researched crop species. The ancestors of rice (The online version contains supplementary material available at 10.1007/s42994-021-00039-0. The history of agriculture is the process of artificial selection of numerous plants and animals. Humans began to settle down and establish agricultural societies in which they artificially domesticated wild animals and cultivated plants as crops. Natural mutations at different gene loci provided the driving force for intensive artificial selection, which can be divided into two stages: domestication and improvement. The domestication stage is characterized by continuous artificial selection by farmers of individual plants with excellent agronomic traits and further expansion of planting. In this process, landraces are developed. During the improvement stage, breeders cross varieties with different desired agronomic traits to obtain cultivars that combine multiple advantages  plants flower when exposed to light periods longer than a certain critical time; short-day (SD) plants require a long period of darkness, and thus a short day, before they flower; and day-neutral plants flower regardless of the day length  involved two stages: Triticum urartu (with an AA genome) and Aegilops peltoides (BB genome) hybridized to form emmer wheat, Triticum turgidum (AABB genome), which then hybridized with Aegilops tauschii (DD genome) to produce the hexaploid wheat Triticum aestivum (AABBDD genome)  encodes an AP1-like MADS-box transcription factor, VERNALIZATION1, a homolog of Arabidopsis AP1, and its transcription is induced by vernalization  and VRN3  to regulate their transcription .PHOTOPERIOD1 (Ppd1), which belongs to the pseudo-response regulator (PRR) gene family. The corresponding gene in barley is known as Ppd-H1 because barley contains the HH genome , which originated from wild barley, increased as barley adapted to the high latitudes of Europe. This indicates that as barley was acclimating to high latitudes with LD conditions, people chose late-heading cultivars to ensure sufficient yield , Ppd-B1 (in the BB genome), and Ppd-D1 (in the DD genome); Ppd-D1 contributes the most to the heading date of winter wheat. The two alleles of this gene, Ppd-D1a  and Ppd-D1b, can lead to a difference of 10 days in the heading time  was most likely domesticated in central China around the Huang–Huai Valley, a mid-latitude temperate region, from its wild progenitor Glycine soja 5,000 years ago  laid the foundation for adaptation to primitive cultivation of soybean and further adaptation towards northern areas  and e2 (gi) alleles contribute to flowering time adaptation in which earlier flowering and maturation is preferred  locus, on the other hand, provides advantages for soybeans in adapting to lower latitudes and increases yield , implying that they are involved in the adaptability of soybean to tropical and high-latitude regions, respectively  is domesticated from its progenitor (Oryza rufipogon). There are several subspecies of rice, including japonica and indica, and there has long been debate about their origins and domestication locations. Some genetic studies have shown that the japonica subspecies was first domesticated in the Yangtze River valley , all elite cultivars have no or very weak photoperiod response  and Rice flowering locus T1 (RFT1) , which have weak photoperiod sensitivity, are generally cultivated there. In central China, where single cropping is used, the heading time of rice can be relatively delayed to enhance yield. There, japonica cultivars (with the genotype Hd1 Ghd7 Ghd8), which have strong photoperiod sensitivity, are cultivated. In the northernmost part of China, rice types with weak photoperiod sensitivity are generally cultivated, including ecotypes with mutations in all three genes  is a member of the circadian clock, and its mutant form is distributed in both high and low latitudes  genes, encoding proteins in the CONSTANS-like (COL) family, can induce the expression of Hd3a and RFT1. Two kinds of functional nucleotide polymorphisms can lead to early flowering and improve the fitness of rice in high latitudes , has kinase activity and modifies Hd1 under LD conditions  under SD conditions, so local farmers prefer double or triple cropping in the course of a year, rather than prolonging one growth cycle, as the means to increase yield.When rice expands to lower latitudes, rice flowering is promoted as the day length becomes shorter. However, in rice, in contrast to soybean, delayed flowering time was not strongly selected for at lower latitudes to increase yield. The mutated form of Zea mays) was domesticated from teosinte (Zea mays ssp. Parviglumis), which originated in the tropical balsas river basin of southwestern Mexico and then spread to high latitudes ; the main producing countries are the United States, China, Brazil, India, and Argentina, with latitudes ranging from 40° S to 45° N , an FT homologue, was strongly selected  in Vgt1 was a major target for selection during the adaptation of maize to cool temperate and high-latitude regions  mapping in offspring of crosses between wild and cultivated species. It inhibits the expression of qLB7-1 that affects flowering time was identified. The maize allele flowers early, and DLF1 is the most likely candidate gene for this locus. DLF1 promotes floral transition by directly binding to the promoter regions of ZmMADS4 and ZmMADS67 and activating them in the shoot apex. In the evolutionary process from teosinte to maize, both DLF1 and ZmMADS67 were under selection and underwent decreases in nucleotide diversity, whereas ZmMADS4 was not under selection. DLF1 and ZmMADS67 may also play an important role in maize adaptation to higher latitudes . Through QTL mapping of recombinant inbred lines of maize and teosinte, a QTL DWARF8, ZmPRR37, ZmPHYC2, and ZmCCA1  has long been controversial, but the current consensus is that it was domesticated in the Andean region of South America and Mesoamerica, introduced to Europe by the Spanish in the sixteenth century, and then spread all over the world  is an FT homolog that inhibits flowering. In S. pimpinellifolium, it is highly expressed in LD conditions but barely expressed in SD conditions. Reduction of SP5G expression or mutation of SP5G in improved cultivars attenuates the inhibitory effect of SP5G on SFT (SINGLE FLOWER TRUSS), a florigen that directly induces flowering in tomato, under LD conditions  is also an FT homolog but is a promoter of flowering. The expression of FTL1 is much higher under SD conditions than under LD conditions and shows a stronger oscillation pattern. This gene induces the expression of ftl1 and sp5g genotypes of cherry tomato are distributed near tomato origins, and different allelic combinations are enriched in cherry tomato in the same region, indicating that they were selected independently during domestication . Some wild types carry sp5g alleles, but no wild type carries ftl1 alleles, which implies that sp5g was selected before ftl1 during the tomato domestication process Below is the link to the electronic supplementary material.


SLIT and NTRK-like protein-5 (SliTrk5) is one of the six members of SliTrk protein family, which is widely expressed in the central nervous system (CNS), regulating and participating in many essential steps of central nervous system development, including axon and dendritic growth, neuron differentiation, and synaptogenesis. SliTrk5, as a neuron transmembrane protein, contains two important conservative domains consisting of leucine repeats (LRRs) located at the amino terminal in the extracellular region and tyrosine residues (Tyr) located at the carboxyl terminal in the intracellular domains. These special structures make SliTrk5 play an important role in the pathological process of the CNS. A large number of studies have shown that SliTrk5 may be involved in the pathogenesis of CNS diseases, such as obsessive-compulsive-disorder (OCD), attention deficit/hyperactivity disorder (ADHD), glioma, autism spectrum disorders (ASDs), and Parkinson's disease (PD). Targeting SliTrk5 is expected to become a new target for the treatment of CNS diseases, promoting the functional recovery of CNS. The purpose of this article is to review the current research progression of the role of SliTrk5 in CNS and its potential mechanisms in CNS diseases. Increasing evidence has identified that SliTrk5 can modulate many important stages of central nervous system development, including axon and dendritic growth, neuron differentiation, and synaptogenesis, which may affect the pathological mechanism of many central nervous system diseases . The expExon-mapping using cDNA sequences has revealed the SliTrk5 gene is located in chromosome 13, and there are 8-50 introns in the upstream of initiation methionine, and the protein-coding region of SliTrk5 is located in a single exon. The molecular weight of SliTrk5 predicted by the deduced amino acid sequences is 108.3 kd . SliTrk5It has been confirmed that synaptogenesis is an elementary process that establishes definite connections between neurons and supports the organization of neural circuits. In the development of synaptogenesis, the membrane-anchored proteins play a crucial role in initial axon–dendrite target recognition, differentiation of presynaptic and postsynaptic specializations, which called synaptic adhesion molecules (SAMs) . The uniThe LRR domain contained in SliTrk5 plays a pivotal role in the regulation of various neuronal functions, such as neurite outgrowth, synapse formation, and dendritic morphogenesis. To execute such functions, SliTrk5 majorly employed the following two basic mechanisms. On the one hand, LRR domain works on the trans cell-cell adhesion molecule PTPRD modulating axon–dendrite adhesion , 28, whiγ isoform of phospholipase C (PLC) could be caused by the phosphorylation of Y791 in TRKA, which lead to the decrease of Ca2+ of internal stores and protein kinase C (PKC) activation, finally affecting the transmission of this signal pathway [Compared with the extracellular LRR domain, the intracellular region of SliTrk5 has a more complex architecture. There are many conserved tyrosine residues in the intracellular region of SliTrk5, which are similar to TRK both in structures and functions, including neurite outgrowth and dendritic elaboration, synapse formation, and neuronal survival , 35. One pathway –42. When pathway . TherefoThe function of SliTrk5 in regulating the process of neuronal outgrowth has been established from the beginning. The LRR domain of SliTrk5 is semblable to that of the SLIT family proteins, which are famous for modulating axon guidance and branching . NeuronsThere was a study showing that LRR-rich proteins may induce synaptic formation, and the purpose of which was to perform an expression screen for synaptogenesis proteins . SliTrk5SliTrk5 has been implicated in promoting neuronal survival. If the SliTrk5 was knocked out, the total brain volume of the mice would be reduced, which was especially obvious in the striatum . FurtherIn different types of human brain tumors, the expression of SliTrk5 was most widely in different human brain tumors, compared with other members of the SliTrks family . EspeciaThe original cDNA of SliTrk5 and other members of SliTrks were found in a study conducted to identify differentially expressed genes in mice with neural tube defects . Utill nBesides above functions in the CNS, SliTrk5 has also reported to affect the process of angiogenesis and bone formation. A study analysing markers of pork-granulosa cells cultured in vitro found that the expression of SliTrk5 was downregulated over time and proved that SliTrk5 is involved in angiogenesis and vascular development of the ovary, which can be screened as a potential marker . AdditioSliTrk5 has been identified as one of the factors modulating basic function of the physiological and pathological mechanism of CNS, such as neurite outgrowth, dendritic elaboration, synaptogenesis, and neuronal signal transmission, which was associated to the development of neuropsychiatric disorders, including obsessive-compulsive spectrum disorders (OCDs), attention deficit/hyperactivity disorder (ADHD), autism spectrum disorders (ASDs), and Parkinson's disease (PD) .Obsessive-compulsive spectrum disorder (OCD), characterized by repeated unwanted thoughts and/or repetitive behavior, is one of the most common mental disorders . ResearcAffecting the anatomy of striatumThe possible pathological mechanisms of SliTrk5 in OCD include the following:μm and greater distance from the soma in the individual medium spiny neurons of the striatum in Slitrk5 deficiency mice, either using the Sholl analysis to observe the dendritic complexity or using fractal dimension analysis to quantify how completely a neuron fills its dendritic field, all of which has reflected a decrease in synaptic connectivity. However, there was no difference in striatal cell soma area between SliTrk5 deficiency mice and their wild-type litter born larva [(2) Increasing the orbitofrontal cortex activityThe effect of SliTrk5 on the anatomy of striatum mainly contains two aspects. Firstly, the volume of the striatum was reduced in both young and aged mice whose SliTrk5 gene has been knocked out. The striatum volume relative to the whole brain estimated by Cavalieri measuring showed a significant decrease in striatum volume compared with wild-type mice, while the ratio of other brain structures to total brain volume did not change, suggesting that SliTrk5 deficiency significantly affected the anatomical structure of the striatum . These rrn larva . In conc(3) Modulating region-specific glutamatergic neurotransmissionTo investigate the role of SliTrk5 in the central nervous system, scholars evaluated the difference of baseline activity in specific brain regions between wild-type and SliTrk5 knockout mice by assessing expression of FosB, which is a transcription factor used to assess neuronal activity routinely. As a result, they found that FosB was upregulated exclusively in the orbitofrontal cortex of SliTrk5 knockout mice, which did not show in other brain regions , 74. ThiShmelkov et al. have discovered the protein amounts of glutamate receptor subunits NAMDAR2A, NAMDAR2B, Glutamate Receptor-1, and Glutamate Receptor-2 involved in excitatory neurotransmission were decreased by 20–60% in SliTrk5 deficiency mice, greatly modulating the glutamatergic neurotransmission of specific region .The sequRecent years, SliTrk5 was reported to participate in the development of brain tumors. A previous study has validated the expression of SliTrk5 is most widely expressed in different brain tumors, which was upregulated in gliomas and correlated with pathological grading , 50. SliSliTrk5 is also associated with tumorigenesis in other organs. SliTrk5 gene presented frequent genetic, epigenetic, and transcriptional alterations in colorectal neoplasia, which has been discovered by a study designed to compare and contrast the molecular profiles of laterally spreading tumors (LSTs) and colorectal cancer (CRC) . MoreoveSliTrk5 is not only involved in the tumorigenesis of the central nervous system but also related to tumors of all systems of the whole body, in which there may be some underlying common mechanisms. If we take SliTrk5 as a target for antitumor therapy in the future, there may be some unexpected surprises.The main symptoms of ADHD are inattention and/or hyperactivity/impulsivity, and SliTrk5 can regulate the hyperactivity behavior of ADHD by controlling the formation of inhibitory synapses in dopamine neurons . The posThe autism spectrum disorder (ASD) is characterized as restricted repetitive behavior, including the lower-order stereotypy and self-injury and higher-order indices of circumscribed interests and cognitive rigidity . A genomSliTrk5 can effectively reclaim activated TRKB receptors and promote BDNF-dependent signal pathway, prolonging the intensity and duration of neurotrophic factor signal in striatum neurons in the absence of BDNF supply . MoreoveIncreasing evidence has revealed that SliTrk5 plays an important role in the physiological and pathological processes of the CNS. The neurite outgrowth, dendritic branching, synaptogenesis, cell differentiation, and signal transmission of neurons are important processes for the development of CNS. SliTrk5 is an important synaptic associated protein that has a crucial regulatory effect on the above response processes of CNS. Furthermore, there have been exhibiting complex interactions between SliTrk5 and many CNS disorders, including obsessive-compulsive disorder, attention deficit/hyperactivity disorder, autism spectrum disorders, and brain gliomas. The role of SliTrk5 in the neurodegenerative diseases, such as Parkinson's disease, has been starting to come out. Taking SliTrk5 as the target and regulating its expression can alleviate neural function damage and promote the recovery of neural function, suggesting that SliTrk5 may be a promising target molecule for illustrating the mechanisms and the treatment of CNS diseases . As the


This study included randomized controlled trials (RCTs) which compared an intervention group of HIE to a control group of lower intensity exercise, or no intervention. All participants were in the acute (0–3 months) and subacute (3–6 months) stages of stroke recovery. Studies were excluded if they were not RCTs, included participants from a different stage of stroke recovery, or if the intervention did not meet the pre-defined HIE criteria. Overall, seven studies were included that used either high-intensity treadmill walking, stepping, cycling or overground walking exercises compared to either a low-intensity exercise (n = 4) or passive control condition (n = 3). Three studies reported significant improvements in 6MWT and gait speed performance, while only one showed improved BBS scores. No major adverse events were reported, although minor incidents were reported in only one study. This systematic review showed that HIE improved LL functional task performance, namely the 6MWT and gait speed. Previously, there was limited research demonstrating the efficacy of HIE early after stroke. This systematic review provides evidence that HIE may improve LL function with no significant adverse events report for stroke patients in their acute and subacute rehabilitation stages. Hence, HIE should be considered for implementation in this population, taking into account the possible benefits in terms of functional outcomes, as compared to lower intensity interventions.This systematic review investigated the effects of high-intensity exercise (HIE) on lower limb (LL) function in acute and subacute stroke patients. A systematic electronic search was performed in PubMed, CINAHL and the Web of Science from inception to 30 June 2022. Outcomes examined included LL function and measures of activities of daily living such as the Barthel index, 6 min walk test (6MWT), gait speed and Berg balance scale (BBS), adverse events and safety outcomes. The methodological quality and the quality of evidence for each study was assessed using the PEDro scale and the Risk of Bias 2 tool (RoB 2). HIE was defined as achieving at least 60% of the heart rate reserve (HRR) or VO Stroke is one of the leading causes of adult disability locally and globally, and is Singapore’s fourth leading cause of death, accounting for close to 6% of deaths . The preThe American Heart Association and American Stroke Association currently recommend aerobic exercise training for stroke survivors at least 3 days a week for 20–60 min per session, subject to the individual’s functional capacity, and resistance training 2 to 3 days a week, in the subacute and chronic stages of stroke recovery ,13. HoweExercise exerts multiple benefits on the cardiovascular, cardiorespiratory, and neurovascular systems in both healthy and clinical populations. Notably, exercise has been shown to augment neuroplasticity, resulting in functional improvements to motor  and cognThere is evidence that high-intensity exercise (HIE) can promote neuroplasticity and elicit significant improvements in aerobic fitness and peak capacity, as well as lower limb (LL) motor function  in persons post-stroke, when compared to lower intensity exercise . In addiFrom a neuroplastic standpoint, a cascade of events at the molecular, biological, and systemic level occurs almost immediately after stroke to limit any further damage to brain and initiate mechanisms for recovery . These nTherefore, to shed light on the role of high-intensity early neurorehabilitation in stroke, this systematic review aims to answer the following research questions:(1) Is HIE effective for improving LL functional outcomes in the acute and subacute stroke population (0–6 months post-stroke)?(2) Is the implementation of HIE safe in acute and subacute stroke settings?The findings from this systematic review will provide support for the role of HIE in early stroke rehabilitation and provide clinicians with safety guidelines for its implementation.This systematic review and analysis were carried out and presented according to the revised Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines , and theA systematic search for articles in the databases PubMed, Web of Science and CINAHL (EBSCOhost) was conducted up until 30 June 2022. The keywords used for the search and strategies used may be found in We included articles that fulfilled the following criteria.(1)Active or passive control group as comparator;Adults aged 18 years and above;Any form of stroke;Stage of stroke: Acute (0–3 months) and subacute (3–6 months).Randomized controlled trials (RCTs) which fulfilled the following criteria:(2)Articles had to include a detailed description of the exercise intervention;(3)The intervention group had to meet the target of HIE;(4)The control group had to have a lower intensity training, or no exercise intervention;(5)All studies had to include a LL functional outcome measure.Our exclusion criteria included unpublished and ongoing trials, and articles that were not published in English. The following sections describe the details of our inclusion criteria.This review included randomized controlled trials that included an intervention group receiving HIE and a control group receiving a physical therapy intervention of lower intensity or no specific intervention/usual care. Trials which involved more than one intervention group and crossover trials  were included in this systematic review.2 peak, 70% of maximal heart rate (HRmax), or attaining a score of 14 or more on the rate of the perceived exertion Borg scale  in accordance with the American College of Sports Medicine’s guidelines for exercise prescription [In addition, this review considered trials that evaluated the effectiveness of HIE in patients’ post-stroke. Vigorous or high-intensity was defined as achieving at least 60% of heart rate reserve (HRR) or VOcription . Exercis(1)Hyper-acute or acute phase (0–7 days);(2)Early sub-acute phase (7 days until 3 months);(3)Late sub-acute phase (3–6 months);(4)Chronic phase (>6 months).This review considered studies investigating individuals over 18 years of age with a clinical diagnosis of acute stroke   regardleAll participants included in this study suffered their stroke within 6 months of recruitment into the respective studies.Titles and abstracts of the references retrieved from the search to identify studies that satisfy the inclusion criteria were screened by two researchers (SMM and HCS) independently. Studies that did not fulfill the inclusion criteria were excluded and full text articles of potentially relevant content were retrieved. Any conflicts in study inclusion/exclusion during the title, abstract and full-text article screening were first resolved by discussion between the two researchers, and if no resolution was achieved, the recommendation of a third researcher (WPT) would be solicited. Reasons for exclusion of studies and the process of selection were recorded, and a Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) flow diagram has been presented .One researcher (SMM) was involved in extracting the data from the included studies. The primary outcome measures for this review included LL functional motor outcome measures that reflect the extent of impairments post-stroke. These outcome measures included the 6 min walk test (6MWT), gait speed, steps per day, Berg Balance scale (BBS) and the Barthel index (BI). Secondary outcomes in this study included reports of adverse events that occurred because of the intervention .www.pedro.org.au, accessed on 27 June 2022).The methodological quality of each study was independently assessed by two authors, SM and HC, by using the Physiotherapy Evidence Database (PEDro) Scale (range 0–10). Studies with a PEDro score ≥6 were considered to be of high quality; 4 or 5, of moderate quality; and <4, of low quality  to assess the risk of bias in the included studies in this systematic review . RoB 2 iA flow chart detailing the study selection process is in The seven included studies were all RCTs published from 2010 onwards ,37,38,39The details of the interventions from the included studies are presented in Four out of the seven studies ,35,38,392peak), peak HR and/or rating-of-perceived exertion (RPE) Borg scale were used to define the intensity of the exercise. Three of the seven studies measured exercise intensity using the HRR or RPE Borg scale [max, which had to be attained during the exercise session in the intervention groups.In all studies, either the heart rate reserve (HRR), peak oxygen consumption  [n = 3) [n = 2) [n = 1) [The duration of each intervention sessions ranged from 11 to 60 min. One study by Tollár, Nagy  had a sen = 4) ,37,38,39n = 4) ,35,38,39n = 4) ,36,37. I [n = 3) ,38,39, i [n = 1)  and one The primary outcome measures and results of the studies are included in Four studies investigated the effects of HIE on 6MWT ,35,36,38Three studies measured gait speed ,36,39, oOnly the study by Tollár, Nagy  found siOnly two studies ,38 lookeOnly one study  measuredOverall, five RCTs ,37,38,39All studies reported good compliance and adherence to completion of the interventions and follow-up assessments. All studies had more than 90% of subjects completed the intervention program except in the study by Pallesen and Colleagues  where thThe methodological quality of each RCT is in Based on the RoB 2 tool, the outcomes are presented in the This systematic review investigated the effects of HIE in the acute and subacute stages of stroke, as defined by acute and subacute stages (up until 6 months) of rehabilitation post-stroke, on measures of LL functional motor outcomes, safety, and adherence. In total, seven studies with 944 participants met our inclusion criteria for this systematic review. Our results indicate that five out of the seven RCTs reported improved LL functional outcomes in the HIE group, as compared to the control group, of which three had active comparator groups exercising at lower intensity ,38,39, a2 or 69% Peak HR or 13 on RPE Borg Scale) range [This systematic review, supported by similar reviews that compared HIE to control groups in other stages of stroke recovery , has shoe) range .Due to the physical impairments after stroke, stroke survivors may have difficulty reaching and sustaining high-intensity in their exercise regimes ,42, hencIndeed, our findings showed that five out of seven studies included in this systematic review reported significant improvements in LL functional outcomes following a period of HIE intervention compared to the control group, in acute to subacute stroke patients. The improvements in functional outcome included improved walking distance, gait speed, and BI (measure of function during activities of daily living), with gait speed and BI showing sustained improvements up to 6 months post-intervention. It should however be noted that the included studies focused predominantly on lower limb exercises such as high-intensity stepping, walking, cycling that maybe complimented with functional training, which is likely to explain why a larger proportion of studies reported improvements in 6MWT (three out of four studies) and gait speed (three out of three studies) as compared to BBS (only one out of three studies).n = 224, 18 months median time post-stroke) that high-intensity Upper Limb rehabilitation improved clinical measures of Upper Limb motor function , with 68.3% and 61.6% of patients reaching minimal clinical important difference in the FM-UL and ARAT, respectively even at 6 months post-intervention. Other studies utilizing various high-intensity training modalities such as resistance training [Our findings were in-line with a previous meta-analysis (of 22 included studies) conducted by Luo, Zhu  showing training , robotictraining  and stepThe use of HIE, particularly in populations with a higher risk of cardiovascular events, has often been limited, in part, due to the fear of potentially causing further adverse events from occurring. Additionally, patients may have difficulties in reaching high levels of exercise intensities, particularly when movement may be affected . LookingEven though minimal safety concerns or adverse events were reported across all studies, precautions still need to be taken to minimize risks to patients. Marzolini, Robertson  proposedThere are several limitations of this systematic review that we would like to acknowledge. Firstly, four out of the seven studies performed follow-up 2-6 months post-intervention, and this is insufficient to fully justify the improvements and effectiveness of the HIE early after stroke. Additionally, the inclusion of only seven papers that fit into our inclusion criteria is insufficient to draw any conclusions on dose-/intensity-response relationships on motor recovery following stroke. To better understand and provide evidence-based interventions of sufficient intensity to ensure optimal outcomes post-stroke, further studies need to recruit a bigger clinical population, standardize outcome measurements across studies investigating different intensities, and ensure adequate long-term follow-up. Secondly, the variations in training parameters, specifically the type of HIE rendered which varied from stepping exercises to ergometer cycling to gaming, and the types of outcome measures included in the studies were highly variable. This makes it difficult to make recommendations on the type of HIE that stroke survivors can perform and be certain if HIE will result in benefits in specific functional outcomes. Most studies to date of HIE in stroke have used either treadmill or overground stepping and walking as the mode of training, but research is emerging on other modes of HIT, including seated modalities such as the recumbent stepper and high-intensity interval training. This offers opportunities for individuals with a range of functional abilities to be included in studies investigating HIE.Our results show that HIE in acute and subacute stroke patients may be effective in improving motor function of the lower and upper limb. In addition, the included studies demonstrate that supervised HIE can be safely administered with good adherence and minor adverse events, such as muscle aches and pains. However, due to the limited studies included, the findings from these studies shown need to be interpreted with caution. Further, most of the literature available do not differentiate between the stages of stroke, and studies include all patients independent of the time since onset of stroke. As a result, very little evidence is available looking at the efficacy of higher intensity exercise early after stroke, and this then does not allow us to make proper recommendations when prescribing exercise for this population. More work in this area needs to be done, so that we can fully optimise neuro-recovery in the different stages of stroke recovery throughout their rehabilitation.


Bunyavirales and, structurally, other arboviruses suggests that these findings are broadly applicable to viruses that use a class II fusion mechanism and cause neurologic disease. La Crosse virus (LACV) is a major cause of pediatric encephalitis and aseptic meningitis in the Midwestern, Mid-Atlantic, and Southern United States, where it is an emerging pathogen. The LACV Gc glycoprotein plays a critical role in the neuropathogenesis of LACV encephalitis as the putative virus attachment protein. Previously, we identified and experimentally confirmed the location of the LACV fusion peptide within Gc and generated a panel of recombinant LACVs (rLACVs) containing mutations in the fusion peptide as well as the wild-type sequence. These rLACVs retained their ability to cause neuronal death in a primary embryonic rat neuronal culture system, despite decreased replication and fusion phenotypes. To test the role of the fusion peptide in vivo, we tested rLACVs in an age-dependent murine model of LACV encephalitis. When inoculated directly into the CNS of young adult mice (P28), the rLACV fusion peptide mutants were as neurovirulent as the rLACV engineered with a wild-type sequence, confirming the results obtained in tissue culture. In contrast, the fusion peptide mutant rLACVs were less neuroinvasive when suckling (P3) or weanling (P21) mice were inoculated peripherally, demonstrating that the LACV fusion peptide is a determinant of neuroinvasion, but not of neurovirulence. In a challenge experiment, we found that peripheral challenge of weanling (P21) mice with fusion peptide mutant rLACVs protected from a subsequent WT-LACV challenge, suggesting that mutations in the fusion peptide are an attractive target for generating live-attenuated virus vaccines. Importantly, the high degree of conservation of the fusion peptide amongst the  Bunyavirales is the largest and arguably most diverse order of viruses, with more than 477 virus species currently described and further classified in fifteen families . An. An27]. We had previously determined that mutants with reduced fusion function, such as LACV mAb escape variants V22 and V22F , replicated to lower levels than wild-type virus in the peripheral tissues  intracranially and suckling (P3) mice peripherally with the rLACVs, differentiating the properties of neuroinvasivenss and neurovirulence, as previously described . When th intracraPeripheral challenge of suckling mice (72 h old) was then used to assess the neuroinvasiveness of the rLACVs. While all of the suckling (P3) mice died regardless of which rLACV they were challenged with, the mice inoculated with the fusion peptide domain mutant rLACVs had a significantly longer survival compared to rLACV-WT challenged mice, suggesting that the fusion peptide is a determinant the neuroinvasive potential of LACV. Replication in striated muscle is one of the primary factors of neuroinvasion by enabling virus amplification and the generation of a high viremia . Indeed,Interestingly, infection with the tryptic site mutant (rLACV-R761H), which was generated as a control because it has a mutation in Gc outside the LACV fusion peptide domain, resulted in significantly shorter survival times of both young adult (P28) mice inoculated intracranially B and sucThe neuropathology of the tryptic site mutant (rLACV-R761H) was also different from rLACV-WT and the fusion peptide domain mutants. Young adult (P28) mice challenged with any of the rLACVs showed spongiform lesions as well as single cell or focal necrosis, and apoptotic bodies. However, brains of mice challenged with the tryptic site mutant rLACV-R761H did not show any perivascular cuffing A, which The susceptibility and neuropathology observed in of suckling mice could be due to an underdeveloped innate immune system, resulting in the inability to control peripheral LACV infection resulting in a high viremia. For example, suckling mice were only able to produce small amounts of interferon in only their livers when infected with Coxsackie B1 virus, while young adult (P28) mice produced interferons in all tissues that became infected . AdditioFor our rechallenge studies, weanling mice (P21) were used because they are less susceptible to peripheral challenge with LACV compared to suckling (P3) mice. Our studies showed that the fusion peptide domain mutant rLACVs provided partial protection from WT-LACV rechallenge . HoweverBunyavirales, the findings from these studies may be extrapolated to other emerging bunyaviruses, including Crimean-Congo hemorrhagic fever virus and Rift Valley fever virus with the potential to lead to the development of therapeutics and vaccines. Virus fusion peptides may provide a novel target for attenuation of viruses for vaccines or antiviral strategies. Monoclonal against the fusion peptide of influenza A virus have been shown to neutralize multiple subtypes of the virus by inhibiting the pH-dependent fusion of viral and cellular membranes ; mAb 1C9


One of the oldest mechanisms of immune defense against pathogens is through detection of foreign DNA. Since human DNA is compartmentalized into the nucleus, its presence in the cytosol heralds a potential threat. The cGAS-STING pathway is one of the most important cytosolic DNA sensing pathways and leads to interferon signaling, inflammasome activation, autophagy, and cell death. While STING signaling is protective at physiologic levels, chronic activation of this pathway can instead drive autoinflammation and autoimmunity. Here we discuss several monogenic disorders of the STING pathway that highlight its impact on both innate and adaptive immunity in the progressive loss of tolerance. The potential relevance of STING signaling in systemic lupus erythematosus is then discussed with a focus on future avenues for monitoring and targeting this pathway. One of the most ancient mechanisms of immune defense is the ability to detect and respond to foreign DNA , 2. The As we have learned more about the STING pathway, we have also begun to understand the consequences of when this pathway fails. Dysregulated STING signaling characterizes several monogenic autoinflammatory conditions due to gain-of-function in STING or loss-of-function in enzymes that clear small amounts of residual cytosolic DNA. While physiologic STING activity is fundamentally an innate immune sensor, uncontrolled STING signaling affects adaptive immunity – specifically, through promoting the development of autoreactive and dysfunctional B and T cells. As ‘DNA overload’ is also thought to be central to the pathogenesis of systemic lupus erythematosus (SLE), excess nucleic acid signaling may represent a shared origin for the development of autoimmunity. Here, we highlight biologic and pathophysiologic concepts shared amongst disorders of the STING pathway resulting in immune dysregulation and loss of tolerance.The classic cGAS-STING pathway can be broken down into four parts Figure 1(1)First, binding of double stranded DNA (dsDNA) allosterically activates cGAS to make cGAMP. cGAS contains two major DNA binding domains as well as one nucleotidyltransferase  domain. It normally exists in an autoinhibited state with inhibitory post-translational modifications, such as phosphorylation at Ser305 and glutamylation  Next, cGAMP binds to STING. Human STING is a 379 amino acid (42kDa) protein that is encoded by transmembrane protein 173 (TMEM173). STING is comprised of four transmembrane domains that anchor the protein to the endoplasmic reticulum (ER) where it interacts with the calcium sensor stromal interaction molecule 1 (STIM1). The apo form of STING exists as a homodimer that has a central binding pocket. When cGAMP is present, a single cGAMP molecule binds to this region, inducing a conformational change in the STING homodimer that permits trafficking and downstream signaling.(3) Activated STING initiates an inflammatory response. STING dimers traffic from the ER to the ER-Golgi-Intermediate Complex (ERGIC) through coatomer protein complex II (COPII) vesicles and then to the Golgi apparatus, where glycosaminoglycans enable it to polymerize along the Golgi surface  STING is ultimately degraded by autophagy (utophagy  or recyc STING isTMEM173 has recently been described leading to a plasma membrane version of STING (pm-STING) that consists of a single transmembrane domain. pm-STING appears to be internalized after extracellular CDN binding and can similarly lead to recruitment of IKK and TBK1  ectonucleotide pyrophosphatase/phosphodiesterase I (ENPP1), which can be found on the cytoplasmic membrane and the ER surface. Knock-out or blockade of ENPP1 has been shown to significantly prolong the half-life of cGAMP, and certain pathogens  have adapted to express PDEs as a means of immune escape  inhibitors could serve as targeted small molecule therapies, given their ability to block interferon signaling. In a recent literature review of SAVI, 10/20 patients who had been treated with JAK inhibitors  showed improvement in their lung disease. Benefit was more consistently seen in patients with variants in the dimerization over the cytosolic domain, although this observation requires further validation with larger patient numbers . While t-/-Irf3, -/-Irf7, and -/-Ifnar1 strains, mice proceeded to develop interstitial lung disease and vasculopathy despite a non-functional interferon pathway  crossed with  pathway , 23. Meaethality . This waethality .Given that mouse models have demonstrated that the lung pathology in SAVI can occur in the absence of interferon signaling and be mediated, in part, by T cell activity, the question remains as to how constitutive STING activity modulates the adaptive immune system. STING is highly expressed in the lung epithelium, such that gain-of-function could lead to lung-specific inflammation. This could include inflammasome activation, as STING has been shown to trigger NLRP3 inflammasome mediated cell death in myeloid cells . Gain-ofCOPA Syndrome is an autosomal dominant disorder initially described in 2015 that was named for a defect in the COPα protein that participates in the COPI heptameric  complex involved in Golgi-to-ER trafficking . While ivia COPII in SAVI, there is impaired Golgi-to-ER recycling of STING via COPI in COPA syndrome. An important distinction is that disease activity in COPA syndrome may be more sensitive to the amount of cGAMP, as this influences whether there is any activated STING at the Golgi apparatus to become trapped in the first place.In the last two years, several papers have shown evidence to support this hypothesis. It has been shown that patients with COPA syndrome have cGAS and STING dependent increases in interferon pathway activation and that STING spends a proportionally greater time in the activated state located in the Golgi apparatus , 32. ThiWhile this interferonopathy model of COPA syndrome neatly connects these two disorders and has led to case reports of using JAK inhibitors , 34, as Since COPA patients have progressive lung disease even with T cell directed therapy, it seems likely that, as with SAVI, there is an innate driven process within the lung in addition to a generalized loss of tolerance. The lung microbiota in mice has been shown to create a tonic interferon signal, which is thought to confer protection against viruses, but perhaps in COPA patients, the microbiota exacerbates disease activity as it chronically feeds cGAMP into the STING pathway . The preAicardi-Goutières Syndrome (AGS) is a genetically heterogenous disorder most commonly due to mutations in nucleases and related enzymes  that degrade DNA, RNA, or DNA : RNA hybrids , 39. PatAGS is associated with increased type I interferon production and was the first disease to bear the classification of ‘type I interferonopathy’ in 2011 , 42. Dir-/-Trex1 mice crossed with strains deficient in STING (-/-Tmem173) or interferon signaling (-/-Irf3) were rescued from death. This was also true if they were crossed with -/-Rag2 mice, supporting the critical role of T cells in disease pathogenesis. Through a series of bone marrow chimera studies using yellow fluorescent protein (YFP)-Cre reporter mice, the authors demonstrate that the type I interferon expression localized to non-hematopoietic cardiac cells of Trex1-/- mice was sufficient to signal wild-type hematopoietic (antigen presenting) cells to recruit and activate autoreactive T and B cells. These autoreactive cells were then responsible for myocardial inflammation and generation of cardiac-specific autoantibodies. In summary, their model shows that STING-triggered interferon production from non-hematopoietic tissue drives subsequent loss of T and B cell tolerance.In a study demonstrating how locally active innate immune activation drives organ specific autoimmunity, investigators developed a mouse model of AGS due to deficiency of the exonuclease Trex1 . Unlike While no AGS model fully captures the clinical phenotype in human disease, the clear interferon signature has led to two trials of JAK inhibitors under an expanded access program (NCT01724580 and NCT03047369) . Thirty-DNASE2 gene   , 52. Con-/-DNase2 mice, with no enzyme activity, are embryonic lethal due to severe anemia and liver failure. This lethality can be rescued by additional knockout of STING. While DNAseII is an endosomal nuclease, it is speculated that accumulation of dsDNA in the endo-lysosomes ultimately results in their rupture, exposing it to cGAS in the cytoplasm. This early and consistent phenotype across mice and humans is likely secondary to impaired macrophage phagocytosis of red blood cell (RBC) nuclei extruded during erythropoiesis. This leads to DNA overload, inflammation, and cell death in the liver where fetal/neonatal RBCs are produced.Mimicking the early features in human disease, -/-Ifnar1-/-DNase2 mice, deficient in interferon pathway signaling, also survive but develop an inflammatory arthritis that is based on residual STING-dependent NF-κB pathway activation  mice resulted in expanded effector memory T cell populations in the spleen and immune complex deposition in the kidney. This implied that STING-activated DCs were key contributors to loss of tolerance in lupus. To complicate the picture, a different mouse model of lupus showed the opposite effect of STING deficiency on lupus disease activity. In the MRL.Faslpr model, in which a mutation in FAS permits survival of auto-reactive lymphocytes, STING (but not IRF3) deficient mice had shorter lifespans, worse lupus nephritis, and expansion of inflammatory myeloid cell populations. Macrophages were hyper-responsive to TLR ligands due to decreases in negative regulators of TLR signaling such as A20 and suppressor of cytokine signaling 1 and 3   . The conThese opposing data reflect the variation and limitation of lupus mouse models, but they are also not entirely surprising. TLR9, which has long been studied as an important nucleic acid sensor in lupus, when entirely deleted in preclinical mouse models (similar MRL genetic background), also led to paradoxical exacerbation of lupus disease activity and decreased survival , 66. Thivia the Interferon Pathway (TULIP)-2 trial of the IFNαβ receptor antagonist, anifrolumab, there was a greater British Isles Lupus Assessment Group (BILAG)–based Composite Lupus Assessment (BICLA) response rate in the anifrolumab group vs. placebo . Response to anifrolumab did not differ by high versus low interferon gene expression at baseline  or proteins . This ca® . However, the broader relevance of the STING pathway will come from understanding its role in the context of classic autoimmune disorders. In this review, we focused on lupus as a representative scenario of systemic autoimmunity with a well-established role for aberrant nucleic acid signaling, possibly myositis , 81–83. myositis , 74, 84.The cGAS-STING pathway is recently recognized as a critical cytosolic DNA sensing pathway. However, uncontrolled signaling leads to increased autophagy, inflammasome activation, and interferon expression with subsequent loss of tolerance to self-antigens. This is best exemplified by monogenic disorders of this pathway, in which patients develop organ specific autoimmunity based on the location of the signaling burden. Future studies and new tools are required to extrapolate these themes to more common, polygenic disorders like systemic lupus erythematosus.HW and DS wrote the draft. JC and FD provided critical edits and feedback. HW and FD conceptualized the paper. 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.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.


Although samples with only one litre volume were concentrated using Al(OH)3 precipitation method and the same samples with larger volumes (5–20 L) were concentrated using the adsorption-elution technique followed by the organic flocculation method, a non-significant difference was observed between the efficiency of the two methods in all types of samples except for the drinking water samples. Based on the qualitative prevalence of studied viruses in water and wastewater samples, the number of genome copies and infectious units in the same samples, resistance to treatment processes in water and wastewater treatment plants, higher frequency of both adenoviruses and PMMoV genomes as candidate viral indices in treated sewage and drinking water was observed. The problem of having a viral genome as indices of viral pollution is that it does not express the recent viral pollution because of the longer survivability of the viral genome than the infectious units in water and wastewater. Both infectious adenovirus and infectious phiX174 bacteriophage virus showed similar efficiencies as indices for viral pollution in drinking water and treated sewage samples. On the other hand, qualitative detection of infectious PMMoV failed to express efficiently the presence/absence of infectious enteric viruses in drinking water samples. Infectious adenoviruses and infectious bacteriophage phiX174 virus may be better candidates than adenoviruses genome, polyomaviruses genome, and PMMoV genome and infectivity as viral indices for water and wastewater.The objective of this study was to compare human adenoviruses (HAdVs) genome and infectivity, polyomaviruses (JC and BK) genome (JCPyVs) and (BKPyVs), Pepper Mild Mottle Virus (PMMoV) genome and infectivity, and infectious bacteriophages as viral indices for sewage and water samples. One hundred and forty-four samples were collected from inlets and outlets of water and wastewater treatment plants (WTPs), and WWTPs within Greater Cairo from October 2015 till March 2017. Two methods of viral concentration [Aluminium hydroxide (Al(OH) Waterborne pathogens transmit diseases to around 250 million people each year resulting in 10–20 million deaths around the globe  have been identified   than those in human feces; the originating source of PMMoV in these animals is unclear  while the flow rate in Zenin was 330,000 m3/day. El-Gabal El-Asfar receives raw sewage from a large area in Cairo Governorate and Zenin receives raw sewage from a large area in El-Giza Governorate.One hundred and forty-four samples were collected from inlets and outlets of water and wastewater treatment plants (WTPs), and WWTPs within Greater Cairo. Forty-eight raw sewage samples were collected from October 2015 till September 2017 , 48 treated effluents of the same raw sewage samples, 24 raw Nile water samples of El-Giza WTP, and 24 drinking water samples of the same WTP. The samples were collected monthly. El-Gabal El-Asfar and Zenin WWTPs use an activated sludge as a treatment technology and the flow rate in El-Gabal El-Asfar was 1,700,000 cubic meters per day  containing 3% beef extract  . The pH of the solution was neutralized by adding 0.1 N HCL and the samples were kept at -70 C until used.Sewage samples (5 L of raw sewage and treated effluents) and water samples (10 L of Nile water and 20 L of drinking water) were concentrated by filtration through negatively charged nitrocellulose membranes  after addition of AlCl3) precipitation method according to Standard Methods for the Examination of Water and Wastewater  were concentrated using (Al(OH)er APHA, .Viral RNA was extracted from 140 µl of the supernatant using BIOZOL Total RNA Extraction reagent (BIOFLUX—Japan) and according to the manufacturer’s instructions and a 30 µl final volume was obtained.It was done as described previously by Kapperud et al.  and modiIt was done according to Puig et al.,  using thReal-time TaqMan PCR was performed for positive samples in the previous PCR screening. Real-time PCR was done using adenovirus@ceeramTools™ Food & Environmental kit and according to the manufacturer’s instructions using adenovirus—Q Standard (Ceeram Tools) and Mengo Extraction Control (Ceeram Tools) and using a real-time PCR thermal cycler . Raw adenovirus genome copies numbers measured by real-time RT-PCR, in duplicate, were corrected according to virus/nucleic acid extraction and RT-PCR efficiencies.It was done according to Esawy et al.  and AbdoIt was performed according to Bofill-Mas et al. . Amplifi3) precipitation method or adsorption-elution technique followed by organic flocculation method of the 144 treated and untreated sewage and water samples (the Al(OH)3 method and adsorption-elution technique followed by organic flocculation method besides the non-concentrated samples). Each sample was applied to the leaves (50 Ul/ leaf) after dusting the leaves with 600-mesh Carborandum. The leaf was supported in the palm of the left hand with the leaf apex pointing towards the wrist and the petiole downward between the second and the third fingers. The leaves were rubbed firmly but gently over the entire upper surface and then washed immediately with tap water  were used to test the infectivity of PMMoV in the water and sewage samples. Three plants were used as replicates for each concentrated sample using either (Al(OH) Walkey, . ControlIt was performed according to Zhang et al., . The paiThe PCR products of positive samples for polyomaviruses and PMMoV were sequenced. Fifty to one hundred µl of the PCR products were purified using a high pure PCR products purification kit (Qiagen) following the manufacturer’s instructions. Sequencing was performed on 1–7 µl of the purified products with an ABI prism Big dye termination cycle sequencing ready reaction kit (Applied Biosystem) using the same primers as in the PCR and following the manufacturer’s instructions. The DNA was sequenced with an ABI prism 310 automated DNA sequencer. Sequence data from both strands of the PCR products were aligned and compared using the CLUSTALW and BLAST programs (European Bioinformatics Institute).It was performed according to the standard methods for the examination of water and wastewater, 23rd edition APHA, . Bacterip < 0.05 of differences of virus recovery after (Al(OH)3) precipitation method and adsorption-elution technique followed by organic flocculation method and on the other hand, the differences of virus recovery between concentrated and non-concentrated samples.A paired Student's t test was applied to ascertain the significance at 3 precipitation method and adsorption-elution/organic flocculation technique are shown in Figs. The results of prevalence of adenovirus genome and infectious units, polyomavirus JC genome, polyomavirus BK genome, PMMoV genome and infectivity, and infectious units of phiX174 bacteriophage in raw sewage, treated effluents, Nile water, and drinking water concentrated by both Al(OH)3 precipitation technique and adsorption-elution/organic flocculation technique are shown in Fig. The results of the mean numbers of adenovirus genome and infectious units and infectious units of phiX174 bacteriophage in raw sewage, treated effluents, Nile water, and drinking water concentrated by both Al (OH)Results of the number of infectious particles (range and median) of adenovirus, PMMoV, and phiX174 bacteriophage virus in the non-concentrated raw sewage, treated effluents, Nile water, and drinking water samples are shown in Table Specificity of symptoms appeared on the leaves of pepper plants after inoculation of concentrated and non-concentrated sewage and water samples were confirmed in all positive samples (100%) using RT-PCR/sequencing Fig. . All sam3 precipitation method and adsorption-elution technique followed by an organic flocculation method did not show a significant difference in the efficiency in raw sewage, treated effluent, and raw Nile water samples. However, a significant difference in the efficiency was observed in drinking water samples. PMMoV infectivity was detected in 16.67% of the drinking water samples concentrated using the adsorption-elution technique followed by an organic flocculation method, while no infectivity was detected in the same samples concentrated using the (Al(OH)3 precipitation method. The difference of samples, volume may be the reason for the superiority of the adsorption-elution technique which twenty litres of the drinking water samples were concentrated, however, only one litre was concentrated using the (Al(OH)3 precipitation method. This difference in samples, volume was not effective with the other types of samples. This may return to the higher viral concentration in raw sewage, treated effluent, and raw Nile water samples than its concentration in the drinking water samples. Another reason is that there is no big difference between the volumes of the samples concentrated using the two methods in these types of water. A lot of previous studies showed higher viral frequency in treated and untreated sewage and river water than its frequency in the drinking water samples 3 precipitation method to concentrate viruses from drinking water samples.The objective of this study was to compare human adenoviruses genome and infectivity, polyomaviruses (JC and BK) genome (JCPyVs) and (BKPyVs), Pepper Mild Mottle Virus (PMMoV) genome and infectivity, and infectious bacteriophages as viral indices for sewage and water samples. To achieve this objective, adenovirus genome and infectious units, JCPyVs genome, BKPyVs genome, PMMoV genome, PMMoV infectivity, and infectious bacteriophages were investigated in sewage and water samples collected monthly from two WWTPs and one WTP inside Greater Cairo. The concentration of samples using (Al(OH)In this study, genomes of both adenoviruses and PMMoV have a higher frequency than the genomes of JCPyVs and BKPyVs in raw sewage and raw Nile water. Also, they have a higher resistance rate in both treated sewage and drinking water. Higher frequency of both adenoviruses and PMMoV was previously reported  precipitation method and adsorption-elution technique on nitrocellulose membranes followed by organic flocculation method have lower efficiency when concentrating bacteriophage phiX174 than their efficiencies when concentrating adenoviruses. This may give an advantage for the infectious adenoviruses as a viral index in water and wastewater. Second, both (Al(OH)3) precipitation method and adsorption-elution technique on nitrocellulose membranes followed by organic flocculation method have comparable efficiencies as viral concentration methods from raw sewage, treated effluents, and raw Nile water (river water), while, adsorption-elution technique on nitrocellulose membranes followed by organic flocculation method is better to be used with drinking water samples. Third, an integrated method that contains inoculation of concentrated water and wastewater samples in the leaves of pepper plants followed by RT-PCR is necessary to confirm the specificity of the causative agent of the appeared lesions as PMMoV.Conclusions may be summarized as follows: first, infectious adenoviruses and infectious bacteriophage phiX174 virus may be better candidates than adenoviruses genome, polyomaviruses genome, and PMMoV genome and infectivity as viral indices for water and wastewater. However, both (Al(OH)