Evaluation of circulating insulin-like growth factor-1, heart-type fatty acid-binding protein, and endotrophin levels as prognostic markers of COVID-19 infection severity.

BACKGROUND: Coronavirus Disease 2019 (COVID-19) is a worldwide pandemic challenge spreading enormously within a few months. COVID-19 is characterized by the over-activation of the immune system causing cytokine storm. Insulin-like growth factor-1 (IGF-1) pathway can regulate the immune response via interaction with various implicated cytokines. Heart-type fatty acid-binding protein (H-FABP) has been shown to promote inflammation. Given the fact that coronavirus infections induce cytokines secretion leading to inflammatory lung injury, it has been suggested that H-FABP levels are affected by COVID-19 severity. Moreover, endotrophin (ETP), the cleavage product of collagen VI, may be an indicator of an overactive repair process and fibrosis, considering that viral infection may predispose or exacerbate existing respiratory conditions, including pulmonary fibrosis. This study aims to assess the prognostic capacity of circulating IGF-1, HFABP, and ETP, levels for COVID-19 severity progression in Egyptian patients. METHODS: The study cohort included 107 viral RNA-positive patients and an equivalent number of control individuals with no clinical signs of infection. Clinical assessments included profiling of CBC; serum iron; liver and kidney functions; inflammatory markers. Circulating levels of IGF-1; H-FABP, and ETP were estimated using the corresponding ELISA kits. RESULTS: No statistical difference in the body mass index was detected between the healthy and control groups, while the mean age of infected patients was significantly higher (P = 0.0162) than the control. Patients generally showed elevated levels of inflammatory markers including CRP and ESR concomitant with elevated serum ferritin; D dimer and procalcitonin levels, besides the COVID-19 characteristic lymphopenia and hypoxemia were also frequent. Logistic regression analysis revealed that oxygen saturation; serum IGF-1, and H-FABP can significantly predict the infection progression (P < 0.001 each). Both serum IGF-1 and H-FABP as well as O2 saturation showed remarkable prognostic potentials in terms of large AUC values, high sensitivity/specificity values, and wide confidence interval. The calculated threshold for severity prognosis was 25.5 ng/mL; 19.5 ng/mL, 94.5, % and for IGF-1, H-FABP, and O2 saturation; respectively. The calculated thresholds of serum IGF-1; H-FABP, and O2 saturation showed positive and negative value ranges of 79-91% and 72-97%; respectively, with 66-95%, 83-94% sensitivity, and specificity; respectively. CONCLUSION: The calculated cut-off values of serum IGF-1 and H-FABP represent a promising non-invasive prognostic tool that would facilitate the risk stratification in COVID-19 patients, and control the morbidity/mortality associated with progressive infection.

Confidence in COVID-19 vaccine effectiveness and safety and its effect on vaccine uptake in Tanzania: A community-based cross-sectional study.

COVID-19 is a major public health threat associated with increased disease burden, mortality, and economic loss to countries and communities. Safe and efficacious COVID-19 vaccines are key in halting and reversing the pandemic. Low confidence in vaccines has been one of the factors leading to hesitancy. We aimed to assess the COVID-19 vaccine confidence (safety and effectiveness), associated factors, and its effects on vaccine uptake among general community members in Tanzania. This was a community-based cross-sectional survey conducted from December 2021 to April 2022 in six regions of Tanzania mainland and two regions in Zanzibar. Participants were interviewed using an electronic questionnaire. Multiple logistic regression models estimated odds ratios (ORs) and 95% confidence interval (CI) for factors associated with vaccine confidence. All analyses were performed using SPSS version 25.0. The study enrolled 3470 general Tanzanian community members; their mean age was 40.3 (standard deviation ±14.9) years, and 34% were males. The proportion of COVID-19 vaccine confidence was 54.6%. Geographical region, residence area, COVID-19 disease risk perception, and good knowledge of COVID-19 vaccines were significantly associated with COVID-19 vaccine confidence. Confidence in COVID-19 vaccines was associated with over three times higher odds of vaccine uptake. Confidence in COVID-19 vaccines was low in Tanzania. Innovative community engagement strategies and region-specific interventions are needed to improve comprehensive knowledge and address community perceptions and attitudes toward COVID-19 vaccines.

Atypical Presentation of Burkitt Lymphoma With Isolated Peritoneal Involvement and Association With Refractory Type B Lactic Acidosis and Hypoglycemia Secondary to the Warburg Effect.

Lactic acidosis is considered to be one of the most common causes of high anion gap metabolic acidosis in hospitalized patients. Warburg effect can present with type B lactic acidosis and is considered to be a rare but well-known complication of hematological malignancies. Here, we present the case of a 39-year-old male who had type B lactic acidosis and recurrent hypoglycemia secondary to newly diagnosed Burkitt lymphoma. This case highlights the importance of considering malignancy workup in any case of unexplained type B lactic acidosis with vague clinical presentation, which can aid in early diagnosis and management.

The impact of COVID 19 on the outcomes of thrombectomy in stroke patients: A systematic review and meta-analysis.

We aimed to conduct the current meta-analysis to provide better insight into the efficacy of mechanical thrombectomy (MT) in managing COVID-19 patients suffering from a stroke. An electronic search was conducted through eight databases for collecting the current evidence about the efficacy of MT in stroke patients with COVID-19 until 18 December 2021. The results were reported as the pooled prevalence rates and the odds ratios (ORs), with their corresponding 95% confidence intervals (CI). Out of 648 records, we included nine studies. The prevalence of stroke patients with COVID-19 who received MT treatment was with TICI ≥2b 79% (95%CI: 73-85), symptomatic intracranial haemorrhage 6% (95%CI: 3-11), parenchymal haematoma type 1, 11.1% (95%CI: 5-23), and mortality 29% (95%CI: 24-35). On further comparison of MT procedure between stroke patients with COVID 19 to those without COVID-19, we found no significant difference in terms of TICI ≥2b score (OR: 0.85; 95%CI: 0.03-23; p = 0.9). However, we found that stroke patients with COVID-19 had a significantly higher mortality rate than stroke patients without COVID-19 after MT procedure (OR: 2.99; 95%CI: 2.01-4.45; p < 0.001). Stroke patients with COVID-19 can be safely and effectively treated with MT, with comparable reperfusion and complication rates to those without the disease.

Development and clinical evaluation of commercial glucose meter coupled with nanofiber based immuno-platform for self-diagnosis of SARS-CoV-2 in saliva

Rapid diagnostics for the diagnosis of COVID-19 are urgently needed in offices, residences, and other public places due to the new Covid epidemic stages. A portable and easy-to-use immunosensing platform was developed and evaluated for a point-of-care and self-detection of SARS-CoV-2 spike protein without the need for extraction, separation, or amplification steps using clinically isolated samples (n = 40 samples). The sensing platform was fabricated based on functionalized nylon nanofibrous membranes and a commercial glucose meter to enable easy deployment of the sensing technology. The fabrication of the immunoreaction vial using nylon nanofibrous membranes as a support matrix for the tethering of antibodies significantly improved the sensitivity of the detection platform in contrast to the use of conventional nylon casted membranes. The sensitivity of the nanofibrous membrane attached antibody was at least an order of magnitude higher (∼12 times) compared to the sensitivity of detection with regular casted membrane-based immunoreaction vial. The feasibility of the designed sensing platform was investigated using saliva as a non-invasive and self-administered sample for the diagnosis of SARS-CoV-2. With a detection limit of 9 ng mL−1 and no pretreatment processes required, the sensing platform demonstrated its suitability for the direct detection of SARS-CoV-2 spike protein in the spiked saliva samples. In addition, the developed platform depicted high agreement with RT-qPCR data in the analysis of the clinical samples with good stability over the storage time and reusability for three cycles with maintaining more than 95% of its original activity. [Display omitted] • Diagnosis of COVID-19 in human saliva using commercial glucose meter. • High sensitivity with broad dynamic range was achieved using nanofibrous membrane. • Successfully analyzing of clinical samples in agreement with RT-qPCR. • Potential application for self- and on-site diagnosis without sample pretreatment. • The sensing platform depicted reusability for 3 cycles and good storage stability. [ FROM AUTHOR]

Consequences of SARS-CoV-2 Infection in Pregnant Women and Their Infants: A Systematic Review.

Coronavirus disease 2019 (COVID-19) is a worldwide health problem, particularly for pregnant women. This review assesses the effects of COVID-19 on pregnant women and their infants. A systematic search was performed of studies published on PubMed, Web of Science, Google Scholar, and Embase from January 2020 to January 2021, without restriction by language. This review included 27 studies (22 from China, one from the United States, one from Honduras, one from Italy, one from Iran, and one from Spain), which cumulatively evaluated 386 pregnant women with clinically confirmed COVID-19 and their 334 newborns. Of the 386 pregnant women, 356 had already delivered their infants, four had medical abortions at the time of research, 28 were still pregnant, and two died from COVID-19 before they were able to give birth. Cesarean sections were performed on 71% of pregnant women with COVID-19 to give birth. Fever and cough were common symptoms among women. Premature rupture of membranes, distress, and preterm birth were pregnancy complications. Low birth weight and a short gestational age were common outcomes for newborns. The common laboratory findings among pregnant women were lymphopenia, leukocytosis, and elevated levels of C-reactive protein. Chest computed tomography revealed abnormal viral lung changes in 73.3% of women. Eleven infants tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. There was no evidence of vertical transmission. Most infants were observed to have lymphopenia and thrombocytopenia. The clinical features of pregnant women were found to be similar to those of generally infected patients. There is evidence of adverse pregnancy and neonatal outcomes caused by COVID-19.

Development and clinical evaluation of commercial glucose meter coupled with nanofiber based immuno-platform for self-diagnosis of SARS-CoV-2 in saliva

Rapid diagnostics for the diagnosis of COVID-19 are urgently needed in offices, residences, and other public places due to the new Covid epidemic stages. A portable and easy-to-use immunosensing platform was developed and evaluated for a point-of-care and self-detection of SARS-CoV-2 spike protein without the need for extraction, separation, or amplification steps using clinically isolated samples (n = 40 samples). The sensing platform was fabricated based on functionalized nylon nanofibrous membranes and a commercial glucose meter to enable easy deployment of the sensing technology. The fabrication of the immunoreaction vial using nylon nanofibrous membranes as a support matrix for the tethering of antibodies significantly improved the sensitivity of the detection platform in contrast to the use of conventional naylon casted membranes. The sensitivity of the nanofibrous membrane attached antibody was at least an order of magnitude higher (∼12 times) compared to the sensitivity of detection with regular casted membrane-based immunoreaction vial. The feasibility of the designed sensing platform was investigated using saliva as a non-invasive and self-administered sample for the diagnosis of SARS-CoV-2. With a detection limit of 9 ng mL−1 and no pretreatment processes required, the sensing platform demonstrated its suitability for the direct detection of SARS-CoV-2 spike protein in the spiked saliva samples. In addition, the developed platform depicted high agreement with RT-qPCR data in the analysis of the clinical samples with good stability over the storage time and reusability for three cycles with maintaining more than 95% of its original activity.

Influence of Manual Diaphragm Release Technique Combined with Inspiratory Muscle Training on Selected Persistent Symptoms in Men with Post-Covid-19 Syndrome: A Randomized Controlled Trial.

OBJECTIVE: To determine whether the addition of manual diaphragm release to an inspiratory muscle training programme is more  effective than inspiratory muscle training alone in reducing blood pressure, dyspnoea, fatigue, and aerobic performance capacity in men with post-COVID-19 syndrome. DESIGN: A prospective, randomized-controlled trial. SETTING: Chest Disease Department, Outpatient Clinic, Cairo University, Egypt. PARTICIPANTS: Fifty-two men with post-COVID-19 syndrome were allocated randomly to the study and control groups. INTERVENTION: The study group underwent diaphragm release plus inspiratory muscle training, whereas the control group received inspiratory muscle training only. OUTCOME MEASURES: All patients were assessed with the following measures at baseline and 6 weeks postintervention: maximum static inspiratory pressure for inspiratory muscle strength, peripheral arterial blood pressure, Modified Medical Research Council scale for dyspnoea, Fatigue Severity Scale, serum lactate level, and 6-min walk test distance for aerobic performance. RESULTS: All outcome measures showed a significant improvement in favour of the study group (p < 0.001) over the control group. However, maximum static inspiratory pressure increased significantly, by 48.17% (p < 0.001) in the study group with no significant change in the control group. CONCLUSION: Addition of manual diaphragm release to an inspiratory muscle training programme potentiates the role of inspiratory muscle training in the management of men with symptomatic post-COVID-19 syndrome.

Changes in physical activity during COVID-19 pandemic among Saudi Arabians: Results from a cross-sectional study.

Background and Aims: The COVID-19 pandemic and the resultant change in sedentary behaviors have had immense health, economic, and social implications globally. As governments worldwide imposed lockdowns and curfews, the amount of time spent indoors greatly increased. This lead to a dramatic change in physical activity (PA) levels and profound consequences on daily routines. Our study aimed to investigate patterns of PA during the COVID-19 pandemic among adults residing in Saudi Arabia. Methods: This cross-sectional survey-based study aimed to investigate patterns of PA during the COVID-19 pandemic among adults residing in Saudi Arabia. The International Physical Activity Questionnaire was utilized to measure participants' PA levels between April 2021 and May 2021. Participants were then classified into three groups according to their PA level, and their PA levels and sedentary behaviors were analyzed. Results: We surveyed 463 participants, 315 (68%) of which were female and 134 (32%) of which were male with a median age of 23 (interquartile range, 21-35) years. Moderate-to-high PA was reported by 257 (55.7%) of the participants. There was a significant decrease in PA during the COVID-19 pandemic and resultant lockdowns among the participants (p = 0.04), with higher rates of sedentary behavior among males than females (p = 0.14). Conclusions: The decline in PA is a profound challenge of the COVID-19 pandemic that needs to be addressed by health practitioners and policymakers. Our study highlights the decline in PA levels seen during the COVID-19 pandemic and the importance of promotional programs and interventions to increase PA among the Saudi Arabian population without compromising the essential health restrictions and social distancing.

A validated LC-MS/MS method for determination of antiviral prodrug molnupiravir in human plasma and its application for a pharmacokinetic modeling study in healthy Egyptian volunteers.

A fully validated, simple, rapid and reproducible liquid chromatography-tandem mass spectrometry method was developed to determine NHC (N-hydroxycytidine), the active metabolite of Molnupiravir (MOL) in human plasma; one of the limited treatment options for SARS-CoV-2 in plasma of healthy volunteers. The internal standard (IS) used was ribavirin. The extraction of analyte and IS from plasma was performed using acetonitrile as a solvent for protein precipitation. Agilent Zorbax Eclipse plus C18, 4.6 × 150 mm, (5 µm) was used for chromatographic separation using a mixture of methanol0.2 % acetic acid (5:95, v/v) as a mobile phase that was pumped at a flow rate of 0.9 mL/min. Detection was performed on a triple quadrupole mass spectrometer operating in multiple reaction monitoring (MRM) employing positive ESI interface using API4500 triple quadrupole tandem mass spectrometer system, with the transitions set at m/z 260.10 â†’ 128.10 and 245.10 â†’ 113.20 for NHC and IS respectively. Method validation was performed in accordance with United States FDA bioanalytical guidance. The concentration range of 20.0-10000.0 ng/mL was used to establish linearity via weighted linear regression approach (1/x2). Moreover, the analyzed pharmacokinetic data from twelve Egyptian healthy volunteers were used to develop a population pharmacokinetic model for NHC. The developed model was used to perform simulations and evaluate the current MOL dosing recommendations through calculating the maximum concentration (Cmax) "the safety metric" and area under the curve (AUC0-12 h) "the efficacy metric" for 1000 virtual subjects. Geometric mean ratios (GMR) with their associated 90% confidence intervals (CI) compared to literature values were computed. Geometric means of simulation-based Cmax and AUC0-12 were 3827 ng/mL (GMR = 1.05; 90% CI = 0.96-1.15) and 9320 ng.h/mL (GMR = 1.04; 90% CI = 0.97-1.11), respectively indicating that current MOL dosage can achieve the therapeutic targets and dose adjustment may not be required for the Egyptian population. The developed model could be used in the future to refine MOL dosage once further therapeutic targets are identified.

Worry from contracting COVID-19 infection and its stigma among Egyptian health care providers

BackgroundsHealthcare providers (HCPs) in COVID-19 epidemic face stressful workload of disease management, shortage of protective equipment and high risk of infection and mortality. These stressors affect greatly their mental health. The aim is to identify working conditions among Egyptian HCPs during COVID-19 epidemic as well as stigma and worry perceptions from contracting COVID-19 infection and their predictors.MethodsA cross-sectional study was conducted among 565 HCPs. Data was collected through Google online self-administered questionnaire comprised seven parts: demographics characteristics, knowledge and attitude of COVID-19, working condition, worry of contracting COVID-19 at work, discrimination intention at work for COVID-19 patients, stigma assessment using impact stigma, and internalized shame scales.ResultsThe vast majority of HCPs (94.7%) were worried from contracting COVID-19 at work. Risk factors for perceiving severe worry from contracting COVID-19 were expecting infection as a severe illness, believing that infection will not be successfully controlled, improbability to continue working during the pandemic even if in a well/fit health, high discrimination intention and impact stigma scales. Significantly high impact stigma scores were detected among those aged < 30 years, females, workers primarily in sites susceptible for contracting COVID-19 infection, those had severe worry from contracting infection at work, and high internalized shame scale. The risk factors for perceiving higher internalized shame scores were not having a previous experience in working during a pandemic, high discrimination intention towards COVID-19 patients and high impact stigma scale.ConclusionsConsiderable levels of worry and stigma were detected among Egyptian HCPs during COVID-19 outbreak. The psychological aspect of health care providers should not be overlooked during epidemic;appropriate institutional mental health support should be provided especially for young HCPs, those without previous work experience in epidemic and those who work in high-risk units. Raising the community awareness about contribution of HCPs in fighting the epidemic might decrease stigmatization action toward HCPs.

A Definitive Prognostication System for Patients With Thoracic Malignancies Diagnosed With Coronavirus Disease 2019: An Update From the TERAVOLT Registry.

INTRODUCTION: Patients with thoracic malignancies are at increased risk for mortality from coronavirus disease 2019 (COVID-19), and a large number of intertwined prognostic variables have been identified so far. METHODS: Capitalizing data from the Thoracic Cancers International COVID-19 Collaboration (TERAVOLT) registry, a global study created with the aim of describing the impact of COVID-19 in patients with thoracic malignancies, we used a clustering approach, a fast-backward step-down selection procedure, and a tree-based model to screen and optimize a broad panel of demographics and clinical COVID-19 and cancer characteristics. RESULTS: As of April 15, 2021, a total of 1491 consecutive eligible patients from 18 countries were included in the analysis. With a mean observation period of 42 days, 361 events were reported with an all-cause case fatality rate of 24.2%. The clustering procedure screened 73 covariates in 13 clusters. A further multivariable logistic regression for the association between clusters and death was performed, resulting in five clusters significantly associated with the outcome. The fast-backward step-down selection procedure then identified the following seven major determinants of death: Eastern Cooperative Oncology Group-performance status (ECOG-PS) (OR = 2.47, 1.87-3.26), neutrophil count (OR = 2.46, 1.76-3.44), serum procalcitonin (OR = 2.37, 1.64-3.43), development of pneumonia (OR = 1.95, 1.48-2.58), C-reactive protein (OR = 1.90, 1.43-2.51), tumor stage at COVID-19 diagnosis (OR = 1.97, 1.46-2.66), and age (OR = 1.71, 1.29-2.26). The receiver operating characteristic analysis for death of the selected model confirmed its diagnostic ability (area under the receiver operating curve = 0.78, 95% confidence interval: 0.75-0.81). The nomogram was able to classify the COVID-19 mortality in an interval ranging from 8% to 90%, and the tree-based model recognized ECOG-PS, neutrophil count, and c-reactive protein as the major determinants of prognosis. CONCLUSIONS: From 73 variables analyzed, seven major determinants of death have been identified. Poor ECOG-PS was found to have the strongest association with poor outcome from COVID-19. With our analysis, we provide clinicians with a definitive prognostication system to help determine the risk of mortality for patients with thoracic malignancies and COVID-19.

Pfizer-BioNTech COVID-19 Vaccine (BNT162b2) Side Effects: A Systematic Review.

Vaccinations prevented severe clinical complications of COVID-19. It was considered a vital component of living endemically with COVID-19. The Pfizer-BioNTech vaccine is the first mRNA-based vaccination that enhances immunity. Resulting in various adverse effects that may emerge after vaccination. This systematic review was undertaken to assess the Pfizer-BioNTech vaccine side effects by reviewing the previous studies. A total of 107 PubMed and Google Scholar publications were screened for Pfizer-BioNTech COVID-19 vaccine side effects. Fourteen articles met the study inclusion criteria. The included searching terms were a combination of "Pfizer vaccine and Side effects," "BioNTech vaccine and side effects," and "BNT162b2 vaccine and side effects," as well as all synonyms. The total number of participants in the 14 studies was 10,632 participants. Average of the most frequent side effects of 14 studies were injection site pain 77.34%, fatigue 43%, muscle pain 39.67%, local swelling 33.57%, headache 33.27%, joint pain 25.75%, chills 18.34%, fever 18%, itching 9.38%, lymph nodes swelling 7.86%, nausea 7.58%, dyspnea 7.86%,and diarrhea 6.36%. The average side effects after the first dose were 79% compared with 84% after the second dose. The average occurs side effects in females at 69.8% compared with males 30.2%. Our study reveals that side effects after the Pfizer-BioNTech vaccine are common, but they are usually mild and self-limited. Local reactions like pain at the injection site are the most common. Anaphylactic shock or severe reactions are rare. We hope that our results will reassure the public that the benefits of vaccination far exceed the dangers. Also, help reduce vaccine hesitancy among individuals worried about vaccine safety and possible adverse effects.

Inflammatory biomarkers and severity of COVID-19: Cross sectional study among Egyptian patients.

The newly emerging coronavirus disease 2019 (COVID-19) is characterized by multisystem inflammatory syndrome. The development of SARS-CoV-2 complications usually starts within few days following infection, and the severity of the disease determines its outcome. Vitamin D insufficiency is associated with risk of lung infections, also cell-based studies reported the ability of vitamin D to control enveloped virus growth. We aimed to investigate the relationship between the most eminent inflammatory biomarkers and the level of vitamin D aiming to provide a tool for early diagnosis and prediction of disease progression. The current study was approved by Research Ethics Committee (REC), Kasr Al-Ainy. After confirmation of being COVID-19 by PCR, the admitted patients were categorized as mild-moderate, and severe-critically ill based on clinical and radiologic data. The total levels of serum 25(OH)D, as well as other pro-inflammatory biomarkers were measured and were analyzed by receiver operating characteristic curve (ROC) analysis for detection of their association with COVID-19 disease severity and to determine their sensitivity and specificity at optimum cutoff points. The area under the curve (AUC) ROC for predicting COVID-19 disease severity was the highest (of 0.97) for vitamin D, inflammatory cytokines, liver enzymes, ferritin, and D-Dimer. In addition, high serum levels of creatinine, and elevated liver enzymes associated with severe-critical COVID-19. The low 25(OH)D was associated with the disease severity.

Emphasizing the link between blood types in multi-ethnic disparities and COVID-19 infection in Makkah, Saudi Arabia.

OBJECTIVES: To analyze the impact and distribution of blood groups in different ethnicities and the extent of susceptibility to infection with COVID-19 in Makkah, Saudi Arabia. METHODS: A retrospective study was performed on 4,609 COVID-19 patients from five ethnic groups to assess the impact and distribution of different blood types and susceptibility to COVID-19 infection. The study was carried out between November 2020 and June 2021 in the College of Medicine, Umm Al-Qura University in collaboration with the General Directorate of Health Affairs, Makkah, Saudi Arabia. RESULTS: Blood group (A, B, and O) distributions in 2,617 COVID-19 patients with local control populations was done. Our study found that in both Saudi and non-Saudi populations, blood groups O and A were associated with higher infection rates, whereas blood group AB was associated with lower infection rates (p=0.0001). COVID-19 seems to be associated with blood groups A, B, and AB (RR=3.23, 95% CI=2.702-3.821, p=0.0001). COVID-19 risk was lower in people with O blood group (RR=0.783, 95% CI=0.733-0.836, p=0.0001). South Asians had higher odds of COVID-19 infection when compared to Saudi cases and other ethnic groups (OR=1.12, 95 % CI: 1.074-1.24, p=0.04). CONCLUSION: We emphasize that COVID-19 infection is not proportional among ethnically related blood groups. Notably, RhD-negative protect against COVID-19, whereas A and O blood types are more susceptible. Thus, when assessing COVID-19 prognosis and vaccination priority, blood groups A and O are critical.