Half of the specimens of each group (n = 12) were exposed to fatigue with cyclic mechanical loading  and simultaneous thermocycling (5–55 °C). Single-load-to-failure testing  was performed for all specimens without/with fatigue application. Data analysis was performed using ANOVA, Tukey’s post-hoc test, two-sample t-test and Bonferroni correction (p < 0.05). Results: All specimens survived fatigue exposure. Significant differences in failure loads were detected among groups (p ≤ 0.004). Materials Z-HT and Z-ST showed the highest failure loads followed by Z-XT, RL and E. The influence of fatigue was only significant for material RL. Conclusions: All types of tested materials exceeded clinically acceptable failure load values higher than 900N and can be recommended for clinical use. Z-HT and Z-ST appear to be highly reliable towards fatigue. Rapid-layer design of PICN and YZ-HT might be an interesting treatment concept for posterior implant SCs.This laboratory study aimed to evaluate the thermo-mechanical fatigue behavior and failure modes of monolithic and rapid-layer posterior single-crowns (SCs) supported by zirconia implants. Methods: 120 all-ceramic crowns supported by one-piece zirconia implants  were divided into five groups ( Today, ceramic implants can be considered a valid treatment addendum in the portfolio of dental implantology with osseointegration capacities and survival rates similar to titanium implants ,2,3. EncAs most of the studies solely focus on soft- and hard-tissue integration of zirconia implants, little evidence is published on the long-term behavior of their prosthetic counterparts ,11. HoweTo overcome the risk of chipping for veneered restorations, an increased application of monolithic materials can be observed in fixed tooth- and implant prosthodontics ,15. ReduBesides reinforced glass ceramics, resin-matrix ceramics (RMC) and zirconia ceramics are considered to be suitable materials for monolithic application. So far, zirconia was mainly used as a framework material due to its high flexural strength but opaque appearance . RecentlCAD/CAM RMCs and especially polymer-infiltrated ceramic networks (PICN) were introduced as more flexible and elastic materials, combining positive material properties of ceramics and polymers ,22. Due Yet, neither in vitro nor clinical studies analyzing the performance of monolithic translucent zirconia restorations or a restorative combination of PICN with zirconia on ceramic implants could be identified. Hence, the performance of these materials on ceramic implants remains to be elucidated. Therefore, the purpose of this in vitro study was to evaluate the thermo-mechanical fatigue behavior and failure modes of monolithic and rapid-layer posterior translucent zirconia and PICN single crowns supported by one-piece zirconia implants. The tested null hypotheses assumed: (i) type of restoration material and (ii) fatigue exposure do not significantly influence the failure load of posterior all-ceramic implant crowns. Monolithic 3Y-TZP implant restorations served as control. n = 24 specimens each) according to the type of restoration material ;Test group Z-ST: 4Y-TZP monolithic “super translucent” zirconia crown ;Test group Z-XT: 5Y-TZP monolithic “extra translucent” zirconia crown ;Test group E: monolithic polymer-infiltrated ceramic network crown ;Test group RL (rapid layer): polymer-infiltrated ceramic network  “table top” adhesively bonded to a 3Y-TZP framework .A total of 120 one-piece zirconia implants  were divided into four test groups and one control group . The master model was scanned  and a master design of a mandibular molar crown  was used for all monolithic crowns to ensure identical and comparable test specimens A–D. The All zirconia implants  were perpendicularly embedded in polyester resin  with a simulation of 0.5–1 mm bone resorption between the implant neck and resin surface. Hence, clinical conditions with reported marginal bone losses of 0.70–0.79 mm after 1 year ,25 and 0PICN implant restoration of Group E and PICN “table top” of Group RL were first cleaned with 70% ethanol and pretreated with 5% hydrofluoric acid , rinsed with air–water spray (30 s) and afterwards subjected to ultrasonic bath  and air-dried. The intaglio crown surface of Group Z-HT, Z-ST and Z-XT were first air-particle abraded with 50 μm aluminum oxide at a pressure of 2 bar prior to cementation. The zirconia framework of Group RL was treated accordingly. All zirconia restorations  were afterwards ultrasonically cleaned with 70% ethanol for 3 min and chemically modified with an MDP-containing primer . PICN table tops of Group RL were adhesively luted with a self-curing resin composite cement  to the pretreated zirconia substructure. The primer (Clearfil Ceramic Primer Plus) was also applied to PICN restorations and to the implant abutment of the investigated one-piece ceramic implant. n = 12 each) were exposed to cyclic mechanical loading and simultaneous thermocycling  in a chewing simulator . An occlusal load of 198 n at 1.2 million chewing cycles with a frequency of 1.6 Hz was applied to the mesio-lingual cusp of the restoration, simulating a clinical 5-year exposure under clinically relevant conditions , but the main effect of restoration material  and its interaction with fatigue  were highly significant.Fatigue did not have a significant main effect across restorative material types [F = 2.05, p < 0.001) failure loads with fatigue (p = 0.607), Z-ST (p = 0.414), Z-XT (p = 0.047) and E (p = 0.413). To further elucidate the interaction, t-tests for the pairwise comparison between specimens with and without fatigue exposure for each restorative material type were performed. The only significant (Bonferroni corrected) difference was found for RL, which showed significantly higher ( fatigue . No signThe highest mean failure load was detected for group Z-HT0 (8450 N) without fatigue (Z-HT0>Z-ST0>Z-XT0>RL0>E0) and for group Z-ST1 (8557 N) with fatigue (Z-ST1>Z-HT1>Z-XT1>RL1>E1). Material E showed the lowest mean failure load of all tested restorative materials both without and with fatigue application.p ≤ 0.004), irrespective of fatigue, but there was no significant difference between Z-HT and Z-ST (p = 1).Tukey’s post-hoc tests revealed that restorative materials Z-HT and Z-ST showed significantly higher failure loads relative to all other three restorative materials tested  (p = 1) where no significance was found for fatigue  . The onl fatigue .All monolithic crowns of Z-XT and E revealed catastrophic bulk fractures after single load to failure. Monolithic zirconia crowns of Z-HT and Z-ST predominately suffered from catastrophic bulk fractures. Cracks occurred on four specimens of material Z-HT and on one of Z-ST. Both subgroups of material RL (RL0 and RL1) solely showed fractures of the PICN table top, whereas the underlying zirconia framework revealed no fractures A,B.For all monolithic materials C,D, fracIn contrast, for material RL C,D, fracThis in vitro study investigated the failure load and fatigue behavior of different designs of monolithic translucent zirconia, PICN and rapid-layer posterior implant crowns on one-piece zirconia implants. The tested null hypothesis was rejected as (i) type of restoration material and (ii) fatigue application in part (only for RL) affected the failure load.Dynamic loading and accelerated aging tests are a reliable method to provide essential information on limitations and lifetime predictions of ceramic restorations . The appp = 1) of failure loads between 3Y-TZP and 4Y-TZP monolithic zirconia ceramics was found. Monolithic 4Y-TZP zirconia (Z-ST) could therefore be a preferable choice of material for monolithic posterior implant-supported single crowns, since it seems as durable as 3Y-TZP but with better translucency and esthetics [The highest failure loads were recorded for monolithic 3Y-TZP (Z-HT) and 4Y-TZP (Z-ST) zirconia crowns due to their high flexural strength and fracture toughness . The measthetics . In vitro studies reported loading capacities for 3Y-TZP monolithic zirconia crowns (Vita YZ T) above 6000 N  on one-pComparable failure loads of 1162 N , 1297 N A significant influence of fatigue was observed only for PICN/3Y-TZP rapid-layer implant crowns, where fatigue significantly increased after thermomechanical loading. A possible reason of higher failure loads for fatigued rapid-layer PICN/3Y-TZP implant crowns of group RL1 compared to RL0 might be a complete polymerization of the adhesive resin interface during thermomechanical loading resulting in a strength increase and durable bond between the PICN and the underlying 3Y-TZP (YZ-HT) framework. In general, a 1-day water storage at 37 °C prior to thermal cycling is recommended to allow a complete polymerization of the adhesive interface . Failure and fractographic analysis revealed that under load, all monolithic crowns were stretched apart with a fracture onset probably starting in the fissure. In the fissure, the material is subjected to tensile stresses, which is the most unfavorable loading case and leads finally to a bulk fracture. In contrast, material RL shows a different loading situation and fracture pattern. The PICN table tops are predominately loaded under pressure and the adhesive interface is more or less loaded in a right angle, resulting in slight shearing between the base of the PICN table top and the underlying 3Y-TZP framework. This loading scenario already leads to a high initial failure load, since the PICN table top is well supported by the 3Y-TZP zirconia framework and is primarily loaded under pressure.If now the polymerization of the resin cement continues during the chewing simulation, this situation is improved by two effects: (1) the compressive strength of the luting material increases and better supports the PICN table tops ; (2) sheTime-dependent fatigue fractures can occur due to repetitive cyclic loading, which leads to subcritical crack propagation and ultimately to restoration failure . OcclusaTranslucent 4Y- and 5Y-TZP ceramics showed a similar wear behavior to 3Y-TZP zirconia in a recent in vitro study , which iOne limitation of the current study is the laboratory design itself, which cannot fully represent the clinical environment. Moreover, load-to-fracture tests have been criticized in the past ,51, and So far, no comparable data for the proposed rapid-layer design on ceramic implants are available. Comparing materials E and RL, it seems that the underlying zirconia framework reinforced and doubled the failure loads. This restorative combination could merge the advantages of the PICN material with a high-strength 3Y-TZP or 4Y-TZP substructure. The PICN could offer the benefits of a tooth-like wear behavior , an abilFatigue did only influence the failure load of the restorative concept RL (higher failure loads with fatigue).All tested materials showed higher failure loads (>1750 N) than normal physiological occlusal loads in the posterior region (200–900N) and can be recommended for clinical use.Irrespective of the material used in monolithic application, zirconia  and PICN restorations failed mostly from bulk fractures. When PICN was used as a table top, fracture was limited to PICN, whereas the 3Y-TZP framework remained intact.PICN materials, especially in the proposed rapid-layer design, might be an interesting restorative treatment concept for zirconia implants due to their facilitated replaceability.Based on the present results and within the limits of this in vitro study, the following conclusions can be drawn:


In people with calf muscle weakness, the stiffness of dorsal leaf spring ankle–foot orthoses (DLS-AFO) needs to be individualized to maximize its effect on walking. Orthotic suppliers may recommend a certain stiffness based on body weight and activity level. However, it is unknown whether these recommendations are sufficient to yield the optimal stiffness for the individual. Therefore, we assessed whether the stiffness following the supplier’s recommendation of the Carbon Ankle7 (CA7) dorsal leaf matched the experimentally optimized AFO stiffness.Thirty-four persons with calf muscle weakness were included and provided a new DLS-AFO of which the stiffness could be varied by changing the CA7®  dorsal leaf. For five different stiffness levels, including the supplier recommended stiffness, gait biomechanics, walking energy cost and speed were assessed. Based on these measures, the individual experimentally optimal AFO stiffness was selected.p < 0.001) and a higher speed . Additionally, a larger ankle range of motion  and higher ankle power  were found with the experimentally optimized stiffness compared to the supplier recommended stiffness.In only 8 of 34 (23%) participants, the supplier recommended stiffness matched the experimentally optimized AFO stiffness, the latter being on average 1.2 ± 1.3 Nm/degree more flexible. The DLS-AFO with an experimentally optimized stiffness resulted in a significantly lower walking energy cost (− 0.21 ± 0.26 J/kg/m, In people with calf muscle weakness, current supplier’s recommendations for the CA7 stiffness level result in the provision of DLS-AFOs that are too stiff and only achieve 80% of the reduction in energy cost achieved with an individual optimized stiffness. It is recommended to experimentally optimize the CA7 stiffness in people with calf muscle weakness in order to maximize treatment outcomes.Trial registration Nederlands Trial Register 5170. Registration date: May 7th 2015. http://www.trialregister.nl/trialreg/admin/rctview.asp?TC=5170.The online version contains supplementary material available at 10.1186/s12984-021-00890-8. Persons with neuromuscular disorders like Charcot–Marie–Tooth disease and poliomyelitis often exhibit weakness of their calf muscles. Calf muscle weakness changes the gait pattern, and typically leads to excessive ankle dorsiflexion, persistent knee flexion and reduced ankle push-off power during stance , 2. ThesTo improve walking in persons with calf muscle weakness, dorsal leaf spring ankle–foot orthoses (DLS-AFOs) can be provided with the aim to restrict the ankle dorsiflexion angle by providing an external plantar flexion moment. This external moment is proportional to the bending and stiffness of the leaf spring –5. If thThe effects of DLS-AFOs on restricting the ankle dorsiflexion angle and supporting ankle power depends largely on the AFO’s ankle stiffness , 6, 10. Previously, we demonstrated that individualization of the DLS-AFO stiffness in persons with calf muscle weakness resulted not only in a lower walking energy cost, but also in better treatment outcomes in terms of perceived fatigue and walking satisfaction, compared to AFOs provided in usual care . In thisFor this study we used data from the PROOF-AFO trial, which was an observational study on the effect of optimizing the AFO ankle stiffness in people with calf muscle weakness . The proThe main inclusion criterion for the PROOF-AFO trial was the presence of non-spastic calf muscle weakness  due to a neuromuscular disease or nerve damage. Other inclusion criteria were: aged 18 years or older, using an AFO or orthopedic shoes in daily life; able to walk for at least 6 min; and weight below 120 kg as this was the maximum for the intervention AFO according to the suppliers recommendation. Exclusion criteria were knee extensor weakness, which required use of a knee–ankle–foot orthosis and not being able to reach more than 0 degrees of ankle dorsiflexion (pes equinus) during weight bearing.Participants were provided a new DLS-AFO Fig. , for whi2) and carbon dioxide production (VCO2) were simultaneously measured . If needed, participants were allowed to use an assistive device, e.g. crutch or cane. To avoid interference with the test, participants were instructed to withhold the intake of food and sugar holding beverages in the two hours before the test.Walking energy cost and speed while walking without AFO and with the five AFO stiffness levels were measured during a 6-min walk test on a 35-m oval track. During the test, oxygen consumption , while ground reaction forces were measured using four force plates . Measurements for each stiffness condition were repeated until three valid trials were recorded .Manual muscle strength of the plantar flexors, dorsiflexors and knee extensors was assessed by a trained physician, and scored according to the Medical Research Council (MRC) scale .2, VCO2 and speed were constant was visually determined for the last three minutes of the test. Walking energy cost was calculated by dividing the mean energy consumption  by the The timing of foot-strike and toe-off were determined using force-plate data. Data were processed within VICON Nexus . Ankle angle, moment and power were time normalized (0–100% of the gait cycles) and averaged across trials using Matlab . Additionally, the maximum ankle angle, moment and power, and maximal positive hip power during early stance and pre-swing for each gait cycle were calculated. We selected the ankle parameters as they are clinically meaningful affected by AFO stiffness , while tThe optimal CA7 AFO stiffness was individually selected based on a pre-defined optimization procedure. The selection was primarily based on walking energy cost as optimization metric and, secondarily, on walking speed and a clinical appraisal of the gait pattern by three independent assessors who were unaware of the supplier recommendations. In case the assessors selected different optimal AFO stiffness levels, the optimal stiffness was selected by consensus. An extensive description of the optimization procedure has been published elsewhere .In clinical practice, the CA7 stiffness is determined on the basis of user’s body weight and activity level, according to a classification matrix as provided by the supplier, see Fig. We analysed the agreement between the clinically selected AFO stiffness following the supplier classification matrix and the experimentally selected optimal AFO stiffness s with Cohen’s Kappa. Differences in energy cost and comfortable speed between walking with no AFO, supplier recommended AFO stiffness and the experimentally selected optimal AFO stiffness were tested with paired t-tests. Differences in effect-size between unilateral and bilateral affected patients were tested with independent t-tests.Additionally, we divided participants in a group of high, medium and low responders to the experimental optimization based on their energy cost, in order to control whether a difference in stiffness explains the effects. In high responders, energy cost reduced by more than 10%, in medium responders energy cost reduced between 5 and 10%, while in low responders energy cost reduced less than 5%. Individual effects are presented in the Additional file Differences between the supplier recommended AFO stiffness and experimental selected optimal AFO stiffness on ankle angle, moment and power were tested with a multilevel linear mixed model to account for the presence of unilateral and bilateral affected patients. The mixed model consisted of three levels; participant (third level), leg (second level), and condition (first level). To model differences between participants while walking with the supplier recommended AFO, a random intercept was incorporated, and to model differences in effect of the optimization a random slope was added. The analysis was performed in MLwiN 2.34 . For the ankle moment and power, participants walking with an assistive device during the gait analysis were excluded because no valid ground reaction forces could be measured.To explain the difference in walking energy cost and speed between the supplier recommended and experimental optimized AFO stiffness, the relation between the difference in ankle power and difference in walking energy cost and speed between conditions was assessed with Pearson’s correlation. This was done for unilateral and bilateral affected subjects combined and separately. For bilateral affected subjects, the average difference in ankle power for the two legs was used. Participants for whom the orthotic-supplier recommended AFO stiffness and the experimental selected AFO stiffness were the same, were excluded from this particular analysis.Of the 37 participants included in the PROOF-AFO trial, three participants were left out from the analysis as the stiffness for their right and left leg were optimized separately. Sociodemographic and clinical characteristics of the remaining 34 participants are presented in Table p = 0.187).The stiffness of the supplier recommended and experimental optimized AFO stiffness corresponded in 8 23.5%) of 34 participants and differed in the remaining 26 participants (Table 3.5% of 3p < 0.001). This difference in stiffness was not significantly different between unilateral (1.1 ± 1.7) and bilateral (1.5 ± 1.2) affected participants (p = 0.457).In 20 (59%) of 34 participants the experimental optimized AFO stiffness was between 2 and 4 Nm/degree, in 9 (26%) participants it was 4.3 Nm/degree, while in only five participants it was above 5 Nm/degree (Table p < 0.001). This means that compared to walking without AFO the supplier recommended stiffness only achieves 82% of its potential effect on energy cost , experimental: 4.08 ± 0.78 J/kg/m, p < 0.001) . The improvement was not significantly different between unilateral (5.9%) and bilateral (4.2%) affected patients (p = 0.357).The experimental optimized AFO stiffness significantly reduced walking energy cost by an additional 4.9% compared to the supplier recommended stiffness . The experimental optimized AFO stiffness increased walking speed further to 1.10 ± 0.18 (+ 2.5%), which was significantly higher compared to the supplier recommended stiffness (p = 0.003). No difference in effect was found when excluding participants with the same recommended as experimentally optimized AFO or between unilateral (+ 3.6%) and bilateral (+ 1.8%) affected patients (p = 0.107).Regarding walking speed, the supplier recommended AFO stiffness significantly increased speed by 0.21 ± 0.18 m/s (24.1%) compared to no AFO  participants were high-responders, in whom energy cost reduced by 0.64 ± 0.2 J/kg/m while speed increased by 0.06 ± 0.05 m/s. In these cases, the experimentally optimized stiffness was 2.5 ± 0.5 Nm/degree lower compared to the supplier recommendation. Eight (23%) participants were medium-responders. In this group, energy cost reduced by 0.32 ± 0.05 J/kg/m and speed increased by 0.04 ± 0.05 m/s, while the experimentally optimized stiffness was 1.6 ± 1.0 N/m degree more flexible. Twelve (35%) participants were low-responders. Their energy cost reduced by 0.06 ± 0.12 J/kg/m and speed increased by + 0.00 ± 0.04 m/s and stiffness was 1.4 ± 1.7 Nm/degree lower compared to the supplier recommended stiffness.p = 0.146) or maximal ankle moment (p = 0.716) during terminal stance. Ankle range of motion (p < 0.001) and ankle power (p < 0.001) were both significantly higher for the experimental optimized AFO stiffness compared to the supplier recommended AFO stiffness (see Table p > 0.248).No significant differences between the supplier recommended and experimental optimized AFO stiffness were found for maximal ankle dorsiflexion angle  and for bilateral affected subjects separately , no significant relationships between difference in ankle power and difference in walking energy cost or speed were found. When only evaluating unilateral affected subjects, increase in ankle power with the experimental optimized AFO compared to the supplier recommended AFO stiffness related significantly with reduction in walking energy cost  .For the unilateral and bilateral affected subjects combined ), the experimentally optimized AFO stiffness was more flexible, resulting in an additional absolute 5% reduction, in walking energy cost compared to the supplier recommended stiffness level. This effect is considered large and meaningful given that DLS-AFOs as provided in usual care reduce energy cost by 7 to 10% compared to no AFO, although the stiffness of dorsal leaf AFOs provided in usual care is often low . Moreove7%), the The experimentally optimized AFO was more flexible and consequently resulted in a larger ankle range of motion and ankle power, which corresponds with previous work , 6. AppaThe aforementioned effects of experimental stiffness optimization clearly demonstrate that the selection of the individual optimal DLS-AFO stiffness solely on the basis of body weight is not sufficient to select the stiffness resulting in the best walking performance on level ground in people with calf muscle weakness. Other orthotic supplier’s recommend certain AFOs solely by type of weakness or impairment, but lack details on for example severity of weakness. Hence, such metrics are also unspecific and are unlikely to perform better in recommending the individual optimal stiffness. In order to improve the AFO stiffness provision, research should focus on creating comprehensive recommendations including all relevant factors to select the individual optimal AFO stiffness. Besides body weight and type of impairment, previous work indicated that higher walking speeds  and seveMeanwhile, to maximize walking performance on level ground, experimental stiffness optimization of CA7 dorsal leaf spring AFOs should be performed in clinical practice instead of following the suppliers’ recommendations, as long as they are not proven to be sufficient to predict the optimal stiffness. To reduce optimization time, costs and patient burden of extensive repetitive measurement procedures, we propose to limit the testing in usual care to the three stiffness levels between 2.8 and 4.3 Nm/degree (R5 to R3 of the CA7 series). These stiffness levels were optimal in almost 90% of our subjects, and higher stiffness levels were only optimal for subjects with a body weight above 80 kg and a slow walking speed without AFO of less than 1 m/s. Therefore, in these heavier and slower walkers, we advise the testing of stiffness levels 3.5 and 5.3 Nm/degree (R4 to R2 of the CA7 series). In the future, fast experimental optimizations might be achieved by human-in-the-loop optimizations , althougAlthough the current study demonstrates a large additional effect of optimizing the AFO stiffness on energy cost, part of this might be explained by the fact that we, among other variables, optimized towards energy cost. Additionally, we did not test whether the experimentally optimized AFO performed better in other walking conditions encountered during daily life. However, the experimentally optimized AFO was more flexible compared to the supplier recommended AFO and, therefore, imposes fewer restrictions in ankle motion. This likely leads to a better walking performance during locomotion conditions such as walking uphill and walking stairs, but meanwhile for a worse performance during conditions where the demands of the AFO are higher than during level walking, such as loaded walking. In short, the current study demonstrates that in people walking on level ground better outcomes can be achieved with AFOs more flexible than recommended by the orthotic supplier, but when prescribing an AFO stiffness in clinical practice individual circumstances and walking conditions should be taken into account.In people with calf muscle weakness, experimentally optimizing the CA7 stiffness outperforms current body weight-based supplier recommendations with regard to improvement in walking energy cost and speed. Current supplier recommendations result in the provision of AFOs that are stiffer than necessary and only achieve 80% of the potential reduction in energy cost, partly explained by the reduced ankle power associated with the higher stiffness. To better match the CA7 stiffness to the individual patient, thereby improving treatment outcomes, we recommend to experimentally optimize the AFO stiffness in people with calf muscle weakness. Additionally, our results suggest that precision orthotics, i.e. matching the AFO mechanics to the patient's pathomechanics, could potentially serve a wider range of patients.Additional file 1: Table. Individual effects of the experimentally optimized stiffness versus the supplier recommended stiffness.


Our in silico reverse vaccinology (RV) approach consisted of two parts: (1) analysis of the selected viral proteins based on annotated cellular location, antigenicity, allele coverage, epitope density, and mutation density and (2) analysis of the various aspects of the epitopes, including antigenicity, allele coverage, IFN-γ induction, toxicity, host homology, and site mutational density. After performing a mutation analysis based on the contemporary mutational amino acid substitutions observed in the viral variants, 13 potential epitopes were selected as subunit vaccine candidates. Despite mutational amino acid substitutions, most epitope sequences were predicted to retain immunogenicity without toxicity and host homology. Our RV approach using an in silico pipeline may potentially reduce the time required for effective vaccine development and can be applicable for vaccine development for other pathogenic diseases as well. One of the most effective strategies for eliminating new and emerging infectious diseases is effective immunization. The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) warrants the need for a maximum coverage vaccine. Moreover, mutations that arise within the virus have a significant impact on the vaccination strategy. Here, we built a comprehensive  SARS-CoV-2 first emerged in China, specifically in Wuhan, in December 2019 .Betacoronavirus of the Coronaviridae family, which also includes the severe acute respiratory syndrome coronavirus and the Middle East respiratory syndrome coronavirus. SARS-CoV-2 is an enveloped, positive-stranded RNA virus with a genome size of approximately 29.8 kb that encodes 29 different proteins [SARS-CoV-2 belongs to the genus of proteins ,4,5. Theproteins ,7. The vWith the lack of effective antiviral treatments, there is an urgent need for a protective vaccine that is effective against different strains of SARS-CoV-2 and with wide immunization coverage across the world’s nations and ethnic groups. Several projects for effective vaccine development have been undertaken, and some of these vaccines are now commercially available for human administration ,13,14,15in silico approach to discover novel B- and T-cell epitopes has become the preferred strategy. This approach is considered one of the most effective for discovering antigens by screening the entire microbial proteome using various prediction tools [in silico vaccine candidate identification approach, designated as reverse vaccinology (RV), starts with the genetic material of the selected pathogen with the subsequent performance of rational computational predictions to come up with a manageable list of targets to be validated experimentally. This approach significantly reduces the time needed to develop a vaccine and provides reasonable targets that are identified based on the selection criteria. RV has been frequently applied with success to discover vaccine candidates for various types of pathogenic microbes (mostly bacteria) [Since discovering antigens and epitopes by an experimental approach can be very tedious, expensive, and time-consuming, using an on tools . The in acteria) ,19,20,21in silico studies have been carried out with the aim of identifying SARS-CoV-2 epitopes, most of these studies focused on individual proteins—mainly the spike protein—or a small subset of SARS-CoV-2 proteins [Although several proteins ,28,29,30proteins ,27,30. Iproteins ,32,33.In the present study, we propose a comprehensive reverse-vaccinology workflow that can be used to identify and shortlist potential T- and B-cell epitopes. Starting with the analysis of three aspects of viral proteins, namely cellular location, antigenicity, and epitope density, antigenic proteins were selected, among which candidate epitopes were subsequently identified. The identified candidate epitopes were further analyzed based on their predicted antigenicity, allele coverage, and induction of interferon-gamma (IFN-γ). Furthermore, mutation analysis of all variant isolates was performed to determine the conservation of the selected epitopes. Our comprehensive approach includes both horizontal (across the whole coding regions of the genome) and vertical (across many sequenced genomes) screening for B- and T-cell epitopes. Based on our results, we provide a shortlist of the candidate epitopes for designing a potential vaccine against SARS-CoV-2, suggesting the usefulness of a comprehensive RV workflow that can be applied to other pathogens as well.https://www.ncbi.nlm.nih.gov/genome).The full multi-FASTA format protein sequences of SARS-CoV-2 were downloaded from the Microbial Genomes Resources at NCBI (https://www.uniprot.org/). Proteins that were annotated as viral membrane proteins were selected as prospective B-cell immunogenic targets. In contrast, proteins that were annotated as host membrane proteins were selected as prospective T-cell activators.Proteins were selected based on their annotated location using the Uniport database (http://scratch.proteomics.ics.uci.edu/index.html). The online tool Vaxijen predicts antigenicity based on the physicochemical properties of proteins, which is independent of sequence alignment (http://www.ddg-pharmfac.net/vaxijen/VaxiJen/VaxiJen.html). The option for the selection of the target organism was set for virus with the default cutoff score of 0.4. Next, two values obtained from the two tools were averaged and normalized, in which unity was the highest possible value and zero was the lowest possible value.Prediction of the antigenicity of the selected proteins was carried out using two web tools: AntigenPro and Vaxijen v2.0 ,35. Antihttps://www.iedb.org) epitope-prediction tools were applied to both the B- and T-cell candidates, as previously described [The Immune Epitope Database  was used to predict IFN-γ induction [The VaxiJen v2.0 was used to prioritize epitopes based on their antigenicity using virus as the target organism with a default cutoff score of 0.4, and epitopes with 0.4 or higher scores were selected for further analyses. In addition, for the T-cell epitopes, the stringency was increased for CD4(+) by reducing the consensus percentile cutoff to 10% , and for the CD8(+) T-cell epitopes, allele coverage was assessed among the 12 supertypes. Furthermore, the IFNepitope online tool (nduction .http://crdd.osdd.net/raghava/toxinpred/) [https://blast.ncbi.nlm.nih.gov/Blast.cgi). These steps were performed to exclude selected epitopes that were predicted to be toxic or had host (human) homology.Next, the ToxinPred online tool was used to predict the potential toxicity of the selected epitopes (inpred/) . In addihttp://huanglab.phys.hust.edu.cn/hpepdock/) [Toll-like receptors (TLRs) detect conserved pathogen-associated molecular patterns (PAMPs) of pathogens including viruses . Among Tepdock/) . TLR4 waepdock/) .http://cov-glue.cvr.gla.uk) and were then categorized for further analysis [https://www.gisaid.org/) [Mutations in SARS-CoV-2 viral variants at the time of the analysis were collected from the CoV-GLUE resource  ,45. MutaThe site mutation density within the selected epitopes  was used to heighten the selection criterion in order to maximize the effectivity range of the selected epitopes. The site mutation density was calculated at each amino acid replacement within the epitope region by dividing the number of sequences at the amino acid site with 100,000 sequences ; site mutation density = /100,000. After performing the mutation analysis using the site mutation density, the selected epitopes were divided into three categories. The first category consisted of epitope sequences with sites with a mutation density of less than 0.01. The epitopes of this category were accepted. The second category consisted of epitopes that had more than 3 sites with a mutation density greater than 0.5; the epitopes of this category were excluded. The third category comprised epitopes that had 1–3 sites with a mutation density between 0.01–0.5. The epitopes of this category were considered for reanalysis. The third category was subjected to reanalysis with online tools, such as Vaxijen, ToxinPred, and BLAST, to check the effect of the mutation in the epitope sequence on the epitope’s immunogenicity, toxicity, and host homology, respectively.https://predictprotein.org/) and PEP-FOLD3 (https://bioserv.rpbs.univ-paris-diderot.fr/services/PEP-FOLD3/) online tools [de novo prediction and analysis tools and provides a 3D prediction of the selected proteins and short peptides. In addition, PEP-FOLD3 was used to predict the structure of epitopes from the third category, which resulted from the mutation analysis of the epitopes, to compare epitopes with or without mutations.Among the selected immunogenic proteins, ORF10, which encodes 38 amino acids with no known homolog protein, was subjected to structural and functional prediction using PredictProtein  without the need for using whole-pathogen entities. Thus, the selection of antigenic part(s) is one of the important steps in vaccine development, as it affects the effectiveness of the vaccine. Given that protein-based subunit vaccines are to be developed for immunization against the current outbreak of COVID-19, our selected proteins can be useful for the development of protein subunit-based vaccines.In the first step, the viral proteome was examined, and candidate proteins were selected according to their annotated cellular locations based on the Uniprot database annotation. Proteins that were found on the surface of the viral membrane were considered B-cell antigens . Three pProtein antigenicity of the selected B-cell and T-cell antigens was predicted using both AntigenPro and VaxiJen online tools  T-cell epitopes and 105 CD8(+) T-cell epitopes. Next, epitope density was calculated for each selected protein, and the values are summarized in B-cell epitope densities for three virion membrane proteins  were found to have similar scores at around 0.03, while ORF10 had the highest epitope density with a value of 0.131, which is 4-fold higher than that of the other three proteins. The CD4(+) epitope densities of T-cell antigens ranged from 0.015 to 0.062 (nsp6 (0.062), nsp4 (0.044), ORF3a (0.036), ORF10 (0.026), and low values for both spike protein and nsp3 . Thus, the spike protein and nsp3 are less likely to activate CD4(+) cells (helper T cells) because both proteins were predicted to contain fewer CD4(+) epitopes compared to the other antigenic proteins . In contrast, CD8(+) epitope densities ranged from 0.058 to 0.131 for the T-cell antigens; the highest value was obtained for ORF10 (0.131), followed by the nsp4 protein (0.080). Thus, ORF10 and nsp4 are very likely to activate CD8(+) cells . As for the allele coverage score, the lowest score (0.25) was assigned to ORF10, followed by nsp6 with a score of 0.75. The remaining three proteins  exhibited the highest score of 1.0 with maximum allele coverage, suggesting that these three proteins have a wider coverage within the population.Another type of subunit vaccine, the epitope-based vaccine, relies on the fundamental immunogenic components of the antigens, which are mainly responsible for protective and specific immune responses. Thus, our next step was to analyze the epitopes in our selected antigens, which were SARS-CoV-2 proteins selected in the preceding screening steps. Seven epitopes identified from the previous step using the IEDB online tool were first subjected to the VaxiJen online tool to predict their antigenicity scores with a cutoff > 0.4. Among the B-cell epitopes , the spiIt is well known that IFN-γ plays a crucial role in defense activation and the regulation of pathways to elicit the antiviral activity of CD8(+) and CD4(+) T cells ,49. In aMoreover, T-cell epitopes were analyzed for their TLR4 binding using the HPEPDOCK server; as a control, a synthetic lipopolysaccharide peptide was used . On the other hand, five epitopes were subjected to reanalysis for their antigenicity, toxicity, and host homology using new sequences with the highest frequent replacement : nucleocapsid phosphoprotein epitopes  ,72. Alth105RTRSMWSFNPETN117 and 168ITVATSRTLSYYKLGASQRVAGDSGFAA195, and the ORF3a-derived epitope, 66KKRWQLALSKGVHFV80 (the matched sequence is underlined). In addition, an epitope derived from the spike protein—258WTAGAAAYY266—has been identified in several in silico studies [Recently, a peptide microarray study of the SARS-CoV-2 proteome was performed to analyze antibody interactions at amino acid resolution . Among t studies ,75 and w studies . Moreove studies ,78,79. Ain silico approach to screen vaccine candidates, we obtained effective candidates that can be used for SARS-CoV-2 vaccine development. Our in silico RV approach was composed of two parts: (1) analysis of the various functional aspects of viral proteins, including antigenicity, allele coverage, epitope density, and mutation density, was carried out for the selected proteins based on annotated cellular location; and (2) epitopes were analyzed for antigenicity, allele coverage, IFN-γ induction, toxicity, host homology, and site mutational density. By narrowing down the list of candidate epitopes with mutational replacements, we offer a list of 13 epitopes as subunit vaccine candidates. The candidates identified using our approach can be further tested as recombinant proteins in in vitro or in vivo studies to validate their antigenicity and avoid unwanted side effects such as antibody-dependent enhancement.In conclusion, using an


This cross-sectional study investigates web tracking prevalence and characteristics on medical journal websites. This information can be used to serve clinicians advertising targeted to medical specialties and areas of professional interest inferred from their browsing histories, potentially contributing to undue pharmaceutical industry influence on clinical practice.2 Thus, we investigated web tracking prevalence and characteristics on medical journal websites.Web tracking tools allow third-party advertisers to compile detailed information about individuals based on their browsing behavior.3 We visited each journal’s home page using webXray, a tool that detects third-party tracking on websites.1 For each journal’s home page, we recorded third-party data requests, which are important because they initiate data transfers from a user’s computer to third parties. We also recorded the presence of third-party cookies, data stored on a user’s computer that frequently serve as persistent identifiers and enable third parties to track individuals across multiple websites.In this cross-sectional study, we identified all journals with an impact factor of 2.0 or higher in clinically relevant subcategories of the Web of Science’s life sciences and biomedical category.STROBE) reporting guidelines for cross-sectional studies. We performed χ2 and Kruskal-Wallis tests to examine associations between journal third-party data requests or cookies and impact factor. All statistical tests were 2-tailed and used a P value of .05 as a threshold for significance. Analyses were conducted in Stata, version 16 (StataCorp LLC).We calculated the percentage of journals with a third-party data request or cookie and the median number of data requests and cookies per journal home page overall and by journal impact factor. We calculated the most prevalent tracking entities across all web pages. We performed Google searches for the top 5 most prevalent tracking entities’ advertising policies and marketing segment disclosures to determine whether they allowed pharmaceutical advertising and medical profession–specific ad targeting. Data were analyzed in October 2021. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology  medical journal home pages included a third-party data request, and 1239  included a third-party cookie, without significant differences by impact factor . Journal4 and Adobe Inc,5 both of which allow medical profession–specific ad targeting. However, all 5 top tracking entities allow pharmaceutical advertising.Nearly all (1593 of 1605 [99%]) journal home pages included a data request from a third-party entity owned by Alphabet, Google’s parent company. Data requests from entities owned by Twitter, Facebook, Oracle, and Adobe occurred on at least 40% of journal home pages . Marketi6This cross-sectional study demonstrated that 99% of medical journals with an impact factor of 2.0 or higher expose visitors to third-party tracking by entities that work with pharmaceutical advertisers. Although similar levels of tracking have been found in health-related websites, tracking on journal websites raises distinctive policy concerns because it may facilitate targeted advertising to clinicians. While targeted advertising can increase knowledge of new therapeutics, it can also sway clinicians’ prescribing patterns toward therapies with limited evidence of efficacy and cost-effectiveness.This study had limitations. Results may not be generalizable to medical journals with an impact factor of 2.0 or lower. Additionally, marketing segment disclosures for 3 of 5 top tracking entities could not be located, and those that were located may be out of date. Finally, this study did not assess how accessing articles through library proxies may alter tracking.Given growing concerns over digital health privacy risks and pharmaceutical advertising to clinicians, medical journal editors and publishers should monitor and assess the potential impact of third-party tracking on journal websites. Further research is needed to determine how tracking information influences targeted advertising to clinicians.


Gelatinization, retrogradation and gel properties of wheat starch–wheat bran arabinoxylan (WS–WBAX) complexes have been evaluated. The results of rapid viscosity analyzer (RVA), differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR) confirmed that WBAX samples with larger Mw and branching degree (HWBAX) significantly impeded gelatinization process of starch by effectively reducing the amount of water available for starch gelatinization. DSC analysis showed that both molecular characteristics and additive amount of WBAX samples have an effect on the long-term retrogradation behavior of starch. For the rheological studies of WS–WBAX mixed gels, the elastic moduli (G’) and shear viscosity of WS–WBAX mixed gels increased with the increase in additive amount of WBAX. WS–HWBAX mixed gels exhibited the lower G’ compared with starch gels containing WBAX with lower Mw and branching degree (LWBAX) at the same amount. The scanning electron micrographs (SEM) revealed that the microstructures of WS–WBAX mixed gels were mainly affected by the amount of WBAX, but hardly by the molecular characteristics of WBAX. Texture profile analysis (TPA) showed that the cohesiveness of fresh WS–WBAX mixed gels became larger with an increase in the WBAX addition amount. The hardness of WS–WBAX mixed gels tended to increase over the 14-day storage. Starch is the main component in wheat, contributing to the characteristics of wheat-based foods that include moisture retention, viscosity, texture, taste and shelf-life . The quaRetrogradation is a recrystallization process in which disaggregated amylose and amylopectin molecules in gelatinized starches reassociate to form ordered structures . Starch Arabinoxylan (AX) is a non-starch polysaccharide isolated from cereal bran. It consists of a linear backbone of β-1,4-D-xylopyranose to which α-L-arabino furanose substituents are attached through O-2 and/or O-3 . AdditioThe objective of this study was to investigate the gelatinization, retrogradation behaviors and gel properties of wheat starch–wheat bran arabinoxylan (WBAX) mixed systems. In our study, the arabinoxylan with different molecular characteristics ) was obtained by alkaline extraction with graded ethanol precipitation. The gelatinization and retrogradation behavior of starch systems in the presence of the WBAX with different molecular characteristics and additive amounts were investigated using rapid viscosity analysis and thermal analysis. The gel properties of starch–arabinoxylan complexes were studied using scanning electron microscopic (SEM), rheological analysis and texture analysis. We hope to provide a theoretical basis for the functions of wheat bran arabinoxylan to control the physical properties of starchy foods.The RVA characteristics of wheat starch–wheat bran arabinoxylan with higher Mw and branching degree (WS–HWBAX) and wheat starch–wheat bran arabinoxylan with lower Mw and branching degree (WS–LWBAX) complexes are presented in The pasting temperature of WS–WBAX complexes with 1% WBAX was significantly lower than those of WS systems in the presence of 0.25% of WBAX. This could be attributed to the increase in the effective concentration of starch in the continuous phase . It was Setback (SB) value indicates the short-term retrogradation degree of starch paste during the cooling process, especially the recrystallization and rearrangement of amylose molecules ,30. The o), peak temperature (Tp), conclusion temperature (Tc) and endothermic enthalpy (ΔHgel), calculated from the DSC thermograms are listed in o, Tp, Tc. The ΔHgel values of starch pastes containing the HWBAX were significantly lower than those of pastes containing LWBAX at the same amount, indicating WS–HWBAX complexes exhibited lower gelatinization degree. The lower gelatinization degree upon addition of HWBAX may be resulting from the limited water availability for starch gelatinization owing to the greater hydration capacity of HWBAX [o, Tp and Tc) of WS–LWBAX complexes were independent of the additive amount of LWBAX, whereas there was a significant increase in To, Tp and Tc when the HWBAX addition increased from 0.25% to 1%. In sum, the pasting properties of WS–WBAX complexes were affected by both the addition level and molecular characteristics of WBAX molecules.The onset temperature , loss modulus (G″) and loss tangent (tanδ = G″/G′) of the gels as functions of frequency for WS–HWBAX and WS–LWBAX samples are presented in The values of G′ for the starch–WBAX mixed gels increased with the increase in WBAX concentration, indicating that apparent gel strength of starch-containing pastes was enhanced by increasing the concentration of WBAXs. The starch gels with HWBAX exhibited the lower G′ as compared with starch paste containing the same amount of LWBAX at the same angular frequency. Liu et al. (2012) reported that before the recrystallization of starch gel occurred, the size of the crystalline lamellae in amylopectin cluster was observed to rise with increasing amylose content and the rigidity of starch granule structure was in proportion to its amylose content but in inverse proportion to the degree of granule swelling . HWBAX iThe scanning electron micrographs (SEM) of cross-sections of different starch gels are shown in The retrogradation degree of gels decreased as the concentration of LWBAX increased, while the opposite trend was found for HWBAX . The degThe textural properties of starch gel samples in terms of gel hardness and cohesiveness were measured on 0 day and 14 days during refrigerated storage . The harCohesiveness indicates the ability of a sample to withstand deformations. When the WBAX addition amount increased, the cohesiveness of the fresh starch-containing gels tended to be larger. The leaching of amylose out of the starch granules and their reassociation play the major role in formation of a cohesive starch gel. Starch gels with 1% of WBAX had lower gelatinization temperature and higher breakdown as compared with starch gels containing 0.25% of WBAX. Large quantities of amylose chains could leach out of the granules to form an interconnected network; therefore, the cohesiveness values of starch gel with 1% of WBAX were larger. Interestingly, there was no significant difference in cohesiveness of all WS–WBAX mixed gels retrograded for 14 days.Both molecular characteristics and additive amounts of WBAX samples have an effect on the gelatinization, long-term retrogradation and gel properties of starch–WBAX complexes. HWBAX had a stronger inhibitory effect on the gelatinization process of starch as compared with LWBAX. LWBAX–WS complexes exhibited a lower long-term retrogradation degree than HWBAX-WS complexes. For fresh WS–WBAX mixed gels, the gel strength and shear viscosity of gels increased with increasing the amount of WBAX. WS–HWBAX mixed gels exhibited lower gel strength (G’) than starch gels containing LWBAX at the same amount. The microstructures of WS–WBAX mixed gels were mainly affected by the amount of WBAX, but hardly by the molecular characteristics of WBAX. The cohesiveness of WS–WBAX mixed gels became larger with an increase in the WBAX addition amounts. For WS–WBAX mixed gels retrograded at 4 °C for 14 days, both hardness and cohesiveness of WS–WBAX mixed gels tended to increase. Overall, results of the current study suggest that WBAX has great potential to modify the properties of starchy foods in the industry.w/w) moisture; 0.20% (w/w) ash content) was purchased from Shanghai Xiangben Industrial Co., LTD., China. The WBAX with lower Mw and branching degree , and WBAX with higher Mw and branching degree  were obtained through alkaline extraction with graded ethanol precipitation. Water was purified with a Milli-Q-Plus system . Potassium bromide (KBr) was purchased from Aladdin Industrial Corporation .Commercial wheat starch (11.0% (w/w)) to make a total weight of 28.0 g. The starch-WBAX suspension was run in the RVA instrument using the Standard Method 2 programmed in the Thermocline Software. Briefly, the suspensions were equilibrated at 50 °C for 1 min, and then raised to 95 °C within 9 min. They were maintained at 95 °C for 5 min, cooled to 50 °C at the same rate, held at 50 °C for 2 min. The rotation speed of the plastic paddle was at 960 rpm for the first 10 s, and then a lower constant speed (160 rpm) was used for the rest of the testing period. Each sample was run at least in duplicate. The Thermocline for Windows software was used to obtain pasting viscosity parameters .Pasting viscosity characteristics of starch–WBAX complexes were determined using a Rapid Visco Analyzer . Wheat starches  were added to an aluminum canister containing LWBAX or HWBAX solutions . The samples were prepared in KBr tablet before measurement. The spectra were recorded from 600 to 4000 cm2 gas, with an empty aluminum pan as a reference. The onset, peak and conclusion temperatures  of gelatinization and gelatinization enthalpy (Hgel), were analyzed using Shimadzu TA-60 analysis software. For long-term retrogradation studies, these pans with the gelatinized samples were stored at 4 °C for 14 days before being re-scanned using the same temperature profile, and the retrogradation enthalpy (ΔHret) was automatically evaluated. The retrogradation ratios (%) were calculated as the ratio of retrogradation enthalpy (∆Hret) to the gelatinization enthalpy (∆Hgel).The gelatinization and long-term retrogradation properties of starch/WBAX mixtures were measured using DSC . Starch powder (2.5 ± 0.1 mg) was prepared with 7.5 μL of WBAX solutions in hermetically sealed aluminum pans and then equilibrated for 24 h at 4 °C. In the DSC measurements, the prepared pans were heated from 20 to 130 °C at a heating rate of 10 °C /min under a continuous flow of dry N−1. Dynamic shear rheological properties were obtained from frequency sweeps over the range of 0.1–10 rad·s−1 at 1% strain. The 1% strain was in the linear viscoelastic region for each sample. TA rheometer data analysis software was used to obtain the experimental data and to calculate the apparent viscosity, the storage modulus (G′), loss modulus (G″) and loss tangent (tanδ = G″/G′). All rheological measurements were performed in triplicate.The resulting arabinoxylan–starch mixed pastes were cooled to room temperature (25 °C), which took 30 min to obtain the fresh gels. The rheological properties of starch–WBAX fresh gels were determined under both steady and dynamic shear. The steady shear rheological properties were obtained with a rheometer , using a parallel plate system (4 cm diameter) at a gap of 1000 μm. For steady shear measurements, the sample was loaded onto the plate of the rheometer and then sheared continuously from 1 to 100 sFor scanning electron microscopy (SEM) observation, the starch–arabinoxylan fresh gels sliced into thin pieces were freeze-dried after immersing in liquid nitrogen for 5 min. Lyophilized slices were attached to the SEM aluminum plate through the double-sided conductive carbon tape. After coating with gold, the microstructure of the cross-section of gels was observed under a scanning electron microscope  at an accelerating voltage of 20 kV.Textural properties of the starch–arabinoxylan mixed gels after 0 day and 14 days storage were determined with a TA-XT plus texture analyzer . Each starch gel tested was cut from the center into cubes of 20 mm. Samples were compressed twice to 25% of original sample height using TA4/1000 probe at a pre-test, test, and post-test speed of 0.8 mm/s. The trigger type is ‘auto’ with trigger force of 5.0 g. Four texture profile analysis (TPA) parameters were calculated from the force/time curve: hardness and cohesiveness. Three replicate samples were tested and their average values with SD were calculated.The data obtained were expressed as mean ± standard deviation. Significant differences were between values at a α level of 0.05.


A person’s financial well-being (FWB) is the complete contentment gained from one’s present financial condition. This has a powerful impact on the entire achievement of an employee’s “well-being.” Researchers, financial analysts, financial planners, educationists, and economists have explored the “enablers” to improve employees’ living standards by investigating the possible “FWB” resources for decades. There is no literature available to show the connection between social capital theory, social exchange theory (SET), social cognitive theory (SCT), financial literacy and FWB, and employees’ financial knowledge sharing a moderator to expand the complete FWB. Financial well-being (FWB) has recently become a hot topic . The firThe present research helps understand the important aspects that can aid employees’ FWB in an organization or workplace. There is a need for improving ways of “financial management,” particularly for the working employees because of the augmented levels of the present individual “indebtedness,” and focus more on an individual’s obligation for “financial planning” between them . This reThe present study carries out the bibliometric analysis of literature studies relating to FWB. The present research talks about the following questions regarding the research conducted on the printed material on FWB: (1) What is the pattern of annual publication trends? (2) What is the pattern of collaboration and co-citation trends? (3) Which are the most productive countries? (4) Which are the most trending scientific journals? (5) Which are the most frequently used author keywords and themes in the literature? The responses to these questions aid in directing the present study. The author has gone a step ahead and has combined a conceptual model to help future researchers better. This will further facilitate a better understanding of the FWB.The present study has created a conceptual model that covers significant predictors that allow employees’ FWB with the aid of significant theories like social capital. These theories were widely talked about in sectors like knowledge sharing (KS), health, well-being, etc. . Thus, tThe following is the paper’s arrangement: section “Research Methodology” talks about the research methodology, and section “Bibliometric Analysis” introduces the bibliometric analysis of the active literature by recognizing the main aspects that impact the employees’ FWB in the organization by suggesting a theoretical model derived from social theories for forthcoming studies. Significant theories are examined in the theoretical underpinning in section “Theoretical Underpinning and Literature Review.” Section “Predictors and Moderators of Financial Well-being” familiarizes the vital determinants and a moderator of FWB. Section “Conceptual Model” suggests a conceptual model framework and, finally, the conclusion is presented in section “Conclusion.”To carry out a bibliometric review of the published FWB articles, data were collected from the web of science (WoS). The present research used the preferred reporting items for systematic review (PRISMA) standard . KeywordThis research incorporated bibliometric analysis on 192 scientific research papers that were culled out from the online WoS database relating to the theme “FWB;” bibliometric R-package and biblioshiny were applied to attain and document the quantitative data relating to the numerous articles chosen; and based on a detailed review, this study proposes a conceptual model and testable propositions.Bibliometric methods apply “bibliographic data” from databases available online. The bibliometric analysis is based on data that permits scientific research and presents a complete vision of scientific research. The data relating to the bibliography is readily available and has augmented bibliometric reviews in distinct studies. This method has been applied to numerous researches. Many software tools are available and are used by scientometrics to examine the bibliometric data. The bibliometrics select a tool depending on the kind of analysis needed. Bibliometrix can be applied for both examining and planning bibliographic data. Knowledge workers can evaluate, alter, and enhance Bibliometrix as it is an “open-source software” written in R-packages, a prominent society of developers and users, and presently has over 16,000 software packages. Bibliometrix can, thus, be applied as part of a superior and broad data analysis workflow. The necessary details of extracted documents by WoS for bibliometric analysis are shown in Information on 192 articles that were published between 2001 and 2021 was culled out from the WoS database. The “analysis period” spreads over 20 years of scientific production, but the actual enhancement in published articles happened in the last 4 years . The annThe authors examined the occurrence of the keywords “time-dependent occurrence.” Emerging Adulthood and Journal of Family and Economic Issues have a strong association with the literature relating to FWB’s primary research themes, particularly “FWB,” “well-being,” “financial socialization,” and “financial satisfaction.”The organization of the co-citation network depicts that Social Indicators Research and Journal of Family and Economic Issues are presented.The typological themes are presented on a two-dimensional plot in the thematic map . KeywordThe historiographic analysis was conducted based on the downloaded data, and a chronological map was generated from the most pertinent citations . The firThe bibliometrix R-package uses the conceptual structure function to conduct multiple correspondence analysis (MCA) for drawing a conceptual structure of the field and K-means clustering to recognize a bunch of documents that convey general ideas. MCA is an exploratory multivariate technique for the graphical and numerical analysis of multivariate categorical data. Homogeneity analysis of an indicator matrix is conducted by MCA to get a low-dimensional Euclidean representation of the earlier data. The words are marked on a two-dimensional map. The author has gone a step ahead and introduced a conceptual model. The present study has created a conceptual model covering significant predictors that allow employees’ FWB to aid significant theories like social capital, social exchange theory (SET), and social cognitive theory (SCT). Thus, understanding theories and their constructs can aid as critical factors in fostering the FWB of employees in the organization.According to the social capital theory, the social association between individuals is “productive resources.” Social capital depicts the association between individuals residing in a specific society. Social capital is described as “the sum of the actual and potential resources embedded within, available through, and derived from the network of relationships possessed by an individual or a social unit” . FurtherFrom Social exchange theory is one of the most significant theoretical concepts to understand people’s reactions. There are sequences of communications that create responsibilities in social exchange. These communications are inter-reliant and dependent on another person’s action . Such coPositive social exchange can provide joint advantages to the organization and employees . To compThe source of SCT is the health sciences. It presents a “human agency model” where people practically “self-reflect, self-regulate, and self-organize” . Martin Financial well-being is studied extensively in economics, psychology, and services marketing, yet the description or dimensions of this notion are not accepted globally in the literature . FWB hasThese processes can be calculated impartially as presented with the help of the profits, assets worth, etc. . Yet, itSignificant facilitators and a moderator relating to social capital theory, SET, SCT, financial literacy, and KS derived from the literature review are presented in this section. It also gains testable proposals for upcoming studies.Reciprocity is an essential characteristic of social exchange and social life. Reciprocity includes the giving back of advantages to another person for-profits attained . GeneralP1: Reciprocity is connected with FWB in a positive and significant manner.Social capital is a defensive source for the addition to social networks . This caP2: Social network is connected with FWB in a positive and significant manner.Sometimes, social trust is seen as an outcome and a catalyst of networks that are used recurrently . Vital sP3: Social trust is connected with FWB in a positive and significant manner.As per the SCT, in a social-learning context, self-efficacy connects persons and the surroundings, and the communication between them . It explP4: Financial self-efficacy is connected with FWB in a positive and significant manner.Generally, financial literacy involves providing people with the information and cognitive talents required to understand the economic division and manage their monetary matters . PsycholP5: Financial literacy is connected with FWB in a positive and significant manner.Knowledge sharing means when a persons’ attitude relates to imparting their information to other people . AccordiP6: KS moderates the relationship between financial literacy and FWB.A conceptual model is suggested in this literature review-based research to clarify aspects of employees’ FWB. This model is based on three critical social theories. There are two social capital dimensions based on social capital theory, namely, social network  and social trust  with reciprocity that is based on SET. Financial self-efficacy based on “SCT and financial literacy” is the independent variable that promotes employee’s FWB. Many authors have shown the significance of “financial literacy” for promoting “FWB” . In FiguFinancial well-being is being discussed everywhere. FWB is considered significant for academicians, public policy officials, financial managers, and employers. There are low levels of FWB that many employees around the globe experience. FWB contributes significantly to an individual’s happiness. Because of this, there are many workplace FWB programs to address this problem. Thus, the present research is significant in considering the significant factors that can help to improve employees’ FWB in the organization.Based on SET supposition, organizations’ optimistic and helpful actions toward their employees and/or their representatives help set up the superior quality of associations that confers a duty on an employee to respond positively in a helpful manner. Thus, a manager should appreciate the best probable conversation so that employees are optimistic and the returns are good FWB from their employer. Promoting the employee’s FWB is a type of social accountability, where acts are done to help a larger society. Thus, supporting FWB aids the “corporate social responsibility” (CSR) objectives of an organization to present an excellent organizational picture and trust . Many stThe first half of 2020 has impacted both domestic and business due to COVID-19. It is a rigorous investigation of a person’s economic flexibility. Legislators put economic flexibility and FWB at the forefront of the schedule and offer a chance to re-concentrate on the critical aspects of financial education. Also, people/small businesses and their monetary circumstances have faced economic and monetary issues resulting from the health crisis during the COVID-19 pandemic. The extended criticism of financial activity and the long impacts of disrupted monetary connections may affect work, profits, and persons’ economic flexibility. Legislators and organizations have provided momentary advantages to the unemployed and/or easy admission to a person’s money at funded/sponsored price, and provide “credit holidays” where banks are required to postpone the full loan installment payment dates . Also, tThere is a new “employability-based psychological contract,” which mentions no lifetime service in one company or firm . EmployeThe association between FWB “determinants” and knowledge sharing as a “moderator” will help both industries and employees plan methods to promote FWB between employees. Thus, it is suggested to determine and authenticate the planned hypothetical-based model in the forthcoming study using proposals as a “testable” proposition. Chances to conduct such researches will be there in the developed and developing countries employing diverse methods.As far as rules and propositions are concerned, financial professionals and policymakers might suggest methods of growing “financial self-efficacy,” “financial literacy,” and significant “social capital dimensions” to aid the population financially, i.e., both vulnerable and invulnerable, and control pressure relating to finances in employment. The present research has two drawbacks. The first is that only the WoS database was employed for bibliometric analysis, and secondly, there is a need for an empirical and systematic review for securing a detailed insight on the subject. Thus, according to the authors, it will be best to conduct a systematic review followed by significant empirical re-examination.AT: conceptualization, methodology, validation, and original draft preparation. AT and VG: visualization and reviewing and editing. VG: supervision. Both 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.


Surgical reconstruction in anterior cruciate ligament (ACL) ruptures has proven to be a highly effective technique that usually provides satisfactory results. However, despite the majority of patients recovering their function after this procedure, ACL reconstruction (ACLR) is still imperfect. To improve these results, various biological augmentation (BA) techniques have been employed mostly in animal models. They include: (1) growth factors , (2) mesenchymal stem cells, (3) autologous tissue, (4) various pharmaceuticals , (5) biophysical/environmental methods , (6) biomaterials , and (7) gene therapy. All of them have shown good results in experimental studies; however, the clinical studies on BA published so far are highly heterogeneous and have a low degree of evidence. The most widely used technique to date is platelet-rich plasma. My position is that orthopedic surgeons must be very cautious when considering using PRP or other BA methods in ACLR. Anterior cruciate ligament (ACL) ruptures are frequent injuries. In the United States, an estimated 200,000 such ruptures occur annually . ACL recThe outcome of an ACLR depends on the graft healing response, i.e., osseointegration at the graft-tunnel interface and remodeling of the intra-articular graft, known as “ligamentization” . The natIn 2021, Sherman et al. presented the results of a survey on global trends in ACLR conducted with the members of the Anterior Cruciate Ligament Study Group, showing that 21% of them employed some type of biological augmentation (BA) in primary ACLR .The aim of this article is to answer the following question: is BA necessary in ACLR? To this end, we conducted a narrative review of the most recent literature on the subject.There are several types of BA techniques that have been used in ACLR . PlateleThe most investigated BA techniques to date in animal studies are biomaterials, growth factors, stem cells, autologous tissue, gene therapy , pharmaceuticals, biophysical interventions, and combinations of these techniques. According to Hexter et al., the methodological quality scores of the animal studies ranged from 4/8 to 8/8, with 97% of the publications scoring at least 5 points  .In the study by Hexter et al., 80% of the publications evaluated growth factors, all of them in the form of platelet concentrates, such as PRP. The rest studied platelet-rich fibrin matrices, autologous serum, platelet leukocyte gels, and autologous platelet concentrates. In most publications, the BA technique was employed during the surgical procedure, although a number of studies used the technique postoperatively. Postoperative treatment with shock waves, joint injections of growth factors in the form of PRP, and autologous serum have also been investigated .According to Hexter, most studies related to BA of ACLR have focused on grafting, tunneling, and the intraoperative application of biological material. However, a number of studies have focused specifically on bone tunnels. Approximately 50% of the publications reported positive results; these included studies on platelet concentrates, calcium phosphate hybridized tendons, and ESWT. A number of clinical studies using remnant repair and bone substitutes have shown mixed results, with positive radiological results but no difference in clinical outcomes. Studies on platelet concentrates and bone marrow-derived mesenchymal stem cells (BMSCs) have shown no beneficial effects for the BA technique . In termMost publications refer to growth factors, used individually or as platelet concentrates  Table 1Table 1.The growth factors investigated in animals include bone morphogenetic protein (BMP), epidermal growth factor, granulocyte colony-stimulating factor, basic fibroblast growth factor, transforming growth factor-β (TGF-β), hepatocyte growth factor, and vascular endothelial growth factor . PlateleIn 2020, Momaya et al. evaluated the cost of PRP and stem cell injections and determined that the mean cost of a PRP injection in the United States was USD 707 , and the mean cost of a stem cell injection was USD 2728  .Growth factors are thought to enhance the healing process and can be used to target graft healing, both on the graft-tunnel interface and to promote intra-articular ligamentization .Numerous types of mesenchymal stem cells (MSCs) have been reported to enhance healing at the graft-tunnel interface in experimental studies . Such cells can be isolated during surgery through bone marrow aspiration or cultured prior to surgery . Table 2Current evidence indicates that when the ACL ruptures, locally released CD34+ vascular MSCs contribute to the rupture’s healing . An impoIn a series of 128 patients, Ouanezar et al. observed that ACLR using the single anteromedial bundle biological augmentation technique significantly improved clinical and functional outcomes. The mean follow-up was 31.7 months . Twenty-four (18.7%) revision surgeries were required: 10 meniscal procedures, 7 ACL revisions, 5 arthroscopies for cyclops lesions, 1 microfracture, and 1 manipulation under anesthesia. The comparative laxity between the healthy and operated sides and the rates of reoperation, graft failure, and cyclops lesions were not significantly different between the ≥50% or <50% ACL remnant groups .A published study indicated that the periosteum is a source of regenerative cells that can promote osteogenesis and chondrogenesis; that is, the use of periosteal cell graft healing could theoretically recreate a direct type of fibrocartilaginous enthesis . Table 3A number of drugs that could modulate the inflammatory response after ACL rupture and enhance graft healing have been investigated in several animal models .Using rabbits undergoing bilateral ACLR injected with a matrix metalloproteinase-inhibitor alpha-2-macroglobulin in the knees, Demirag et al. studied the role of this inhibitor in graft healing. At 5 weeks’ follow-up, the knees in which the inhibitor was injected showed a more mature graft-tunnel interface, a decrease in matrix metalloproteinases in the synovial fluid, and improved biomechanical strength .The effect of bisphosphonates on osseointegration after ACL rupture has also been studied in a rat model of ACLR. At 6 weeks of follow-up, the local or systemic administration of alendronate resulted in improved bone tunnel mineralization, decreased bone loss in the peri-tunnel, and increased graft integration in the tunnel .In a study comparing rats treated with daily subcutaneous parathyroid hormone against control rats, computed tomography scans showed greater thickness and better trabecular bone microarchitecture in the parathyroid hormone-treated animals than in the control animals treated with saline injections .In a study of rabbits, Yeh et al. used hyperbaric oxygen to improve graft neovascularization. Animals treated with hyperbaric oxygen showed improved neovascularization, osseointegration, and biomechanical properties .A number of published studies on animal models have reported positive results with the use of low-intensity pulsed ultrasound . One aniIn a rabbit model, Wang et al. observed that ESWT applied to the tibial tunnel immediately after ACLR improved healing at the graft-tunnel interface, both histologically and biomechanically, at 8 and 24 weeks of follow-up . Table 5This section will discuss the four key technologies related to biomaterials: biological fixation methods, biological coatings, biosynthetic bone substitutes, and osteoconductive materials .A number of experimental studies on ACLR performed in rabbits have shown that magnesium-based interference screws can accelerate graft mineralization, promote bone formation in the periscrew region, and improve extensor tendon osseointegration ,38. In aStudies have shown that the graft-tunnel interface healing of artificial grafts can be improved by using various types of coatings, including chitin , bioglasAnimal studies have shown positive results using biosynthetic bone substitutes, such as demineralized bone matrix (DBM) and recombinant bone xenograft. These biomaterials appear to improve graft tunnel interface healing due to their osteoinductive and osteoconductive properties . DBM hasCalcium phosphate (CaP) is a resorbable and osteoconductive biomaterial that has shown positive results in several animal models for assessing graft healing . FurtherMost of the recently published studies on gene therapy to improve graft healing in ACLR have been based on MSCs ,54. In tAccording to Hexter et al., goals for augmentation of ACLR comprise easing graft to-bone healing, optimizing the ligamentization process, and rendering further stability while the graft is temporarily frail during remodeling. Several growth factors and bioactive molecules are encountered in a number of platelet preparations such as PRP, fibrin clot, and autologous conditioned serum . PRP is Some of these, comprising PDGF, VEGF, and TGFβ, have been involved in both graft-to-bone healing and graft maturation and remodeling. Platelet preparations have been the topic of many clinical studies trying to augment these processes, but the outcomes remain variable and unverified. In a few prospective randomized controlled studies, intraoperative local administration of PRP gel to the graft and tunnels was associated with better healing characteristics on postoperative MRIs when compared with controls ,57,58.p < 0.001), suggesting that PRP might have hastened the graft maturation process [p = 0.019) and 2.5 months (p = 0.008) postoperatively, suggesting a positive influence on graft-to-bone healing and incorporation [Radice et al. found that ACLR augmented with PRP accomplished intra-articular segment signal homogeneity on T1- and T2-weighted MRI sequences in 48% of the time needed by the control group . Most patients  returned to their preinjury level of competition, while 7 (55) patients sustained a reinjury requiring revision surgery. These authors thought that these outcomes compared favorably with the 25% rate of reinjury and revision among pediatric and adolescent athletes published by other authors ,67. One Augmentation with amnion-based matrices might render an option to collagen scaffolds that already have helpful growth factors and bioactive substances . WoodallA recently published study on a sheep model of ACLR has once again demonstrated that BMSCs and PRP significantly improve graft maturation . AlthougIn 2020 Yu et al. have introduced and validated a metal-free, reproducible, and reliable mouse model of ACLR surgery as an efficacious method for a better understanding of molecular mechanisms of graft-tunnel healing after ACLR . One hunp < 0.05); however, this was not sustained at 120 min. Vancomycin-treated tendon tissue demonstrated no significant increase in apoptotic gene expression, or apoptotic protein levels in tissue or supernatant, ex vivo. Vancomycin was associated with a decrease in inflammatory proteins from treated tendon supernatants . Vancomycin ACL wrap did not alter the molecular structure of the ACL hamstring graft and may ameliorate graft integrity [According to Atherton et al. the utilization of the vancomycin wrap to pretreat the hamstring graft in ACLR has significantly diminished rates of postoperative infection. However, it was not known whether this antibiotic treatment (5 mg/mL concentration) affects the molecular composition of the graft. In a controlled laboratory study, surplus hamstring tendon collected after routine ACLR surgery was utilized for in vitro cell culture and ex vivo tissue experiments . Vancomyntegrity .All BA methods analyzed in this article have shown good outcomes in animal studies; however, the clinical studies on BA published so far are highly heterogeneous and have a low degree of evidence.Iorio et al. reported a randomized controlled trial with 40 patients examining the clinical and radiographic impact of hamstring autograft augmentation with nanohydroxyapatite to ease graft-to-bone healing. Lysholm, Tegner, IKDC scores, and KT-1000 arthrometer readings, did not differ significantly between the study group and the control group. However, radiographic parameters related to graft strength, interface incorporation, and bony remodeling showed a tendency toward better outcomes with nanohydroxyapatite augmentation .In two different randomized controlled trials with minimum 2-year follow-up, Mutsuzaki et al. found better outcomes after ACLR with calcium phosphate hybridized hamstring autograft. Significantly better Lysholm scores at 2-year follow-up were found with calcium phosphate- hybridized hamstring autograft compared with controls, as well as significantly less laxity on KT-1000 arthrometer testing at 1 and 2 years postoperatively, and significantly less bone tunnel enlargement in both the femur and tibia. Therefore, calcium phosphate hybridization was shown to avert bone tunnel enlargement in anatomic hamstring autograft ACLR. However, the clinical significance of this finding remains uncertain ,79.ClinicalTrials.gov identifier: NCT03704376, accessed on 7 November 2021), Bailey et al. observed that adding intraoperative PRP did not improve the self-reported knee function, functional performance, or time to return to the preinjury activity level for patients undergoing ACLR with meniscal repair. The use of PRP also had negative consequences in terms of the postoperative recovery of knee range of motion. Based on these data, the authors stated that orthopedic surgeons should cautiously consider the application of PRP during surgery for intra-articular knee injuries [In 2021, Bailey et al. analyzed the effect of intraoperative PRP on postoperative knee function and complications after ACLR with meniscal repair at 2 years of follow-up. In a level 3 evidence study  used a single-bundle technique for ACLR and that more than 50% chose hamstring autograft as the primary graft of choice . More thIt is important to highlight the discrepancy between 2 recent articles: one on an experimental study stating that orthobiologics can accelerate biological events related to tendon allograft incorporation  and anotn = 5). Group 1 received 1 × 107 allogeneic BMSCs in 2 mL of fibrin seal; group 2 received 12 mL of PRP in a plasma clot injected into the graft and bone tunnels, and group 3 (control) received no adjunctive treatment. At autopsy performed at 3 months, a graft maturation score was calculated by summing the graft integrity, synovial coverage and vascularity, graft thickness and apparent tension, and synovial sealing at the tunnel apertures. The signal-noise quotient (SNQ) and fibrous interzone were analyzed by magnetic resonance imaging  to assess intra-articular graft maturation and tendon-bone healing, respectively. Spearman’s rank correlation coefficient (r) of the SNQ, the graft maturation score at autopsy, and the bone tunnel diameter were analyzed. The BMSC group (p = 0.01) and the PRP group (p = 0.03) obtained a significantly higher graft maturation score than the control group. The BMSC group scored significantly higher for synovial sealing at the tunnel apertures (p = 0.03) than the control group. The graft maturation score at autopsy correlated significantly with the SNQ . Femoral tunnel diameter at the opening  and mid-portion  were positively correlated with the SNQ [In the sheep model described by Hexter et al. in 2020, the authors observed that BMSCs and PRP significantly enhanced graft maturation, indicating that orthobiologics might accelerate biological events related to tendon allograft incorporation . They al the SNQ .p < 0.001). No other differences in postoperative complications were observed [In contrast, the retrospective matched case-control study published Bailey et al. analyzed 324 patients (162 PRP patients and 162 control patients) and concluded that the added use of intraoperative PRP did not improve the analyzed parameters . Patientobserved .Regarding the use of BA in orthopedic sports surgery, Looney et al. stated that although the diversity and availability of new biologic technologies continue to increase, the literature remains inconclusive as to the optimal indications for its use . Long-teIn a retrospective review, McMillan et al. analyzed 14 patients operated on for ACLR using an augmented quadruple-stranded hamstring allograft with a type I resorbable collagen matrix impregnated with PRP. The authors observed that their technique produced good early clinical results (at the 2-year follow-up) and had no ACLR failures. This novel ACLR approach could improve the incorporation of biologic grafts into host bone tunnels . HoweverFuture studies are needed to assess the efficacy of BA techniques in ACLR and to determine whether a combination of various therapies could be beneficial in clinical practice. Despite the promising results obtained in animal models with all BA methods, they are still experimental. More clinical studies that meet the minimum quality standards are needed to determine whether current and emerging BA techniques in ACLR are truly beneficial to patients. Orthopedic surgeons should be very cautious when considering using BA in ACLR and should rely on proven scientific data and not get swept up by the BA fad, the clinical efficacy of which has yet to be proven.