Plants recruit specific microorganisms to live inside and outside their roots that provide essential functions for plant growth and health. The study of the microbial communities living in close association with plants helps in understanding the mechanisms involved in these beneficial interactions. Currently, most of the research in this field has been focusing on the description of the taxonomic composition of the microbiome. Therefore, a focus on the plant-associated microbiome functions is pivotal for the development of novel agricultural practices which, in turn, will increase plant fitness. Recent advances in microbiome research using model plant species started to shed light on the functions of specific microorganisms and the underlying mechanisms of plant–microbial interaction. Here, we review (1) microbiome-mediated functions associated with plant growth and protection, (2) insights from native and agricultural habitats that can be used to improve soil health and crop productivity, (3) current -omics and new approaches for studying the plant microbiome, and (4) challenges and future perspectives for exploiting the plant microbiome for beneficial outcomes. We posit that integrated approaches will help in translating fundamental knowledge into agricultural practices. The plant microbiome comprises a plethora of beneficial, commensal, and pathogenic microorganisms that play important roles in plant growth and health . AdvanceMicrobacterium, Stenotrophomonas, and Xanthomonas, were found to be enriched in the rhizosphere of Arabidopsis thaliana upon foliar defense activation by the downy mildew pathogen Hyaloperonospora arabidopsidis  encoding for a polyketide synthase-non-ribosomal peptide synthetase enzyme in the genome of the isolated Flavobacterium, and site-directed mutagenesis in the BGC confirmed its involvement in disease suppression biotic stresses or plants growing in unfavorable environments can provide insights into microbial and plant traits that allow them to withstand such conditions. Wild plants display higher tolerance to abiotic  and biotic  stresses when compared with domesticated plants. These wild relatives have been exploited by plant breeders as sources of resistance genes and specific traits . The devAgricultural practices also impact the plant microbiome composition and functions , 2018 wiPlants alter the (a)biotic properties of the environment where they grow via the production of root exudates. To some extent, these properties will also impact the growth and health of the current and future plant generations . This prTo study the complexity of genetic, microbial, and metabolic factors that impact the plant microbiome, comprehensive systems biology approaches are needed . Metagenin situ are still limited and complex owing to the dynamics and heterogeneity of the soil environment. Soil metabolites are composed of both plant and microbial secreted molecules and their production is influenced by the soil properties and by the interactions of soil microorganisms with host plants (1H NMR) revealed a higher abundance of sugar-derived molecules in suppressive soils than non-suppressive soils, and the latter also contained higher amounts of lipids and terpenes  that antagonize the pathogen.The plant microbiome encompasses distinct microbial groups, such as bacteria, fungi, viruses, algae, and protozoa. Currently, the majority of microbiome studies had focused on bacterial and fungal communities. However, plant interactions with other members of the microbiome as well as interactions across these microorganisms determine the overall diversity and functioning. Recent studies have shown that protists play important roles in the soil microbiome and in plant health . For exap to 80% . The effA number of studies have validated the role of specific microbial synthetic communities (SynComs) using culturable microbial consortia on plant drought tolerance , soil diIn addition, the identification of substrate preferences might contribute to the culture of microorganisms which can be further used as SynComs. For example, a study coupling genomics and exometabolomics showed that chemical succession in the rhizosphere interacts with metabolite substrate preferences by microorganisms that are predictable from genome sequences . This apThe current challenge for exploiting and applying microbiome-mediated functions that positively influence plant growth and protection across diverse environments rely on critical aspects of context dependency. In other words, these beneficial effects are largely dependent on plant genotypes, microbial interactions, soil types, management practices as well as interactions among these factors . AlthougIn agriculture, microbial inoculation of single strains with beneficial traits has been used for disease management and promoting yield gain . HoweverOur understanding of the plant microbiome has vastly increased in the past decade. Integrated approaches, such as different multi-omics and microbiome engineering strategies, have greatly contributed to a better understanding of the organization and dynamics of plant-associated microbial communities. In addition, plant–soil feedbacks opened a new avenue for improvement of agricultural practices using knowledge obtained from natural ecosystems. Plant breeding programs have traditionally focused on exploring genetic variability of the crops for higher productivity and stress resistance, often neglecting the importance of beneficial interactions between microorganisms and plants. Therefore, future strategies for plant breeding should take plant microbial symbionts as life-long bodyguards into consideration. These proposed integrated approaches will provide solutions for exploring and exploiting plant–microbiome interactions for improving the sustainability and productivity of global agriculture.CS, FZ, VJC, and VC conceived and wrote the manuscript. All authors read and approved the final version of the article.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The COVID-19 pandemic is a time of many predictions and rapid learning. Early on, the World Health Organization (WHO) identified Ethiopia as one of thirteen countries in Africa as a top priority for COVID-19 preparedness . DespiteThe complex interplay between biology, health policy, health human resources, sociocultural factors, and political decision-making influence pandemic outcomes, although these interactions are challenging to track. Nations with seemingly similar sociocultural and political structures have taken drastically different measures to address COVID-19, suggesting that even within large geographical regions, local context matters . Sub-SahWe employed a three-step process early in the pandemic to understand the pattern of COVID-19 in Ethiopia: a focus group to explore explanatory themes, an analysis of current evidence, and a panel discussion with a larger audience of experts. The single focus group was conducted on April 25, 2020 and consisted of in-country experts including those with clinical, public health and health system leadership and policy backgrounds, who are actively engaged in the COVID-19 pandemic control program at institutional and national levels. The goal of the focus group discussion was to draw upon the multi-faceted perspectives of participants to identify explanations and rationales for Ethiopia’s low COVID-19 rate. Second, an extensive search of the academic literature was done to identify evidence for the identified explanatory themes. We searched Medline, Embase and the Chinese Biomedical Literature Database using keywords identified in the focus group without limiting our selection to specific study designs. We also searched for pre-print publications from medRxiv. Finally, we combined information from the focus group and the literature review to engage with a multidisciplinary panel of over 80 high level academic and policy experts and health professionals from Ethiopia on May 8, 2020. The aim of this two-hour virtual panel discussion was to validate findings from the focus group and literature review and to develop recommendations for the ongoing monitoring and tracking of COVID-19 in Ethiopia. During the panel discussion, first, the results of the focus group and literature synthesis were presented; second, a senior infectious disease expert, pulmonologist and epidemiologist provided feedback on findings; and finally, feedback and input from the remaining discussion participants of the panel was taken.The focus group discussion identified 30 possible contributors to the status of the epidemic in Ethiopia. These possible contributors were then categorized into four potential explanatory interpretations , number of tests performed for each confirmed case, and case fatality rate. All three variables can help us understand the true spread of the virus . The numThe second possible explanation proposed that the Ethiopian population may have more immunity and/or lower susceptibility to COVID-19. Preliminary evidence suggested that Ethiopia may have lower susceptibility for the exponential transmission of COVID-19. Potential contributors to the slow spread of COVID-19 in the country (and across the sub-Saharan region) were that Ethiopia is a tropical country with high average temperatures; the capital city is at high altitude; the high prevalence of tuberculosis and malaria may have a negative association with COVID-19 ; a univeThe third possible explanation proposed that the early implementation of non-pharmacological public health interventions (NPIs) in Ethiopia has slowed the progression of the epidemic. These early interventions included mandatory quarantining of passengers from abroad, closing borders to prevent travel in and out of the country, closing schools, banning mass gatherings, and early health education campaigns . Each ofThe fourth possible explanation proposed that Ethiopia remains in a different phase of the epidemiological curve than elsewhere. Little is known about the initial phases of the epidemiological curve in countries that experienced earlier outbreaks, and to what extent cases were present but undetected. It is also possible that Ethiopia has experienced a different shape in the epidemiological curve with a longer and flatter first wave. However, observation in other countries has suggested that while the rate of growth may be variable, increased cases is the norm. While the rate of growth has been recently increasing, and COVID-19 cases may well eventually catch up with the rest of the world, it is noteworthy that Ethiopia has shown a sustained pattern of slow growth over an extended period of time.During the panel discussion the three invited panelists  and the panel discussion participants corroborated and validated the literature synthesis findings. The overall consensus was that Ethiopia has experienced a slow and delayed epidemic trajectory because of the combined contributions and interaction of testing-related factors, contextual host-agent dynamic and NPIs. Despite this slow growth, the panel accordingly recommended that it would be extremely unwise to risk complacency, and ongoing monitoring of all epidemiological indicators is essential. Policy makers should continue to advance a coordinated strategy to reduce the impact of COVID-19 should it become more severe at later stages. This approach, which brought together infectious disease, pulmonology and epidemiology experts from public health and academic institutions, provided insight and direction for making reasoned decisions that incorporated contextual realities within Ethiopia. We hope that other countries with population and contextual diversity might learn from our strategies in ways that allow them to prepare for the unpredictable future phases of COVID-19.


A crosstalk established between tumor microenvironment and tumor cells leads to contribution or inhibition of tumor progression. Mesenchymal stem cells (MSCs) are critical cells that fundamentally participate in modulation of the tumor microenvironment, and have been reported to be able to regulate and determine the final destination of tumor cell. Conflicting functions have been attributed to the activity of MSCs in the tumor microenvironment; they can confer a tumorigenic or anti-tumor potential to the tumor cells. Nonetheless, MSCs have been associated with a potential to modulate the tumor microenvironment in favouring the suppression of cancer cells, and promising results have been reported from the preclinical as well as clinical studies. Among the favourable behaviours of MSCs, are releasing mediators (like exosomes) and their natural migrative potential to tumor sites, allowing efficient drug delivering and, thereby, efficient targeting of migrating tumor cells. Additionally, angiogenesis of tumor tissue has been characterized as a key feature of tumors for growth and metastasis. Upon introduction of first anti-angiogenic therapy by a monoclonal antibody, attentions have been drawn toward manipulation of angiogenesis as an attractive strategy for cancer therapy. After that, a wide effort has been put on improving the approaches for cancer therapy through interfering with tumor angiogenesis. In this article, we attempted to have an overview on recent findings with respect to promising potential of MSCs in cancer therapy and had emphasis on the implementing MSCs to improve them against the suppression of angiogenesis in tumor tissue, hence, impeding the tumor progression. Given the importance of cancer, research on timely diagnosis and effective treatment in cancer field is on the fast track , 2. Old This review paper endeavoured to go through the recent findings with regard to promising potential of MSCs in cancer therapy and have emphasis on the implementing MSCs to improve them against the suppression of angiogenesis in tumor tissue.In addition to bone marrow, MSCs have been extracted from multiple tissues, such as adipose tissues, skeletal muscle, dental pulp, placenta, synovial membranes and umbilical cord . Some imMobilization from the bone marrow and other organs is the very necessary role for MSC homing to tumors. Endogenous MSCs have been demonstrated to mobilize from the bone marrow and other tissues to the peripheral tissues through various injury conditions, such as normoxia, inflammatory conditions, and hypoxia . It has Tumor microenvironment influences on the plasticity of MSC phenotype and responses through diverse stimuli, leading to polarization of MSCs and obtaining specific characteristics . RegardiNumerous mediators have been identified that play critical roles in the cross-talk between MSCs, tumor microenvironment, and tumor cells Table . By induThe communication of MSCs with tumor microenvironment might either supress or promote the tumor progression. Although most of researches aiming to exert MSCs in cancer therapy have focused on the tumor suppressor properties of MSCs, these cells may also promote tumor advancement. In vivo and in vitro experiments revealed that human MSCs were able to enhance the metastasis and growth of tumor cell in a mice model of osteosarcoma . AdditioExosomes are extracellular vesicles (EVs) that are generated in the endosomal compartment of eukaryotic cells . ExosomeProdrug suicide gene therapy by different MSCs can deliver the chemotherapeutic drug and activate the prodrug directly within the tumor to generate toxic products for tumor cells , 48. AdiThe composition and biological activities as well as the therapeutic potential of MSC-derived EVs in cancer have been extensively reviewed by Xunian and Kalluri . The virGraphene oxide (GO) is a versatile platform for bioimaging, tissue engineering, gene delivery, and drug delivery, that is functionalized with carboxylic acid . The impIn recent years, different treatment strategies have been proposed for the treatment of cancers. Generation of new blood vessels (neoangiogenesis) from pre-existing vasculature during the embryogenesis is characterized as a fundamental process that leads to development of concomitant vascular network . NeoangiConsidering the participation of angiogenesis and vascularization the development and progression of cancers, it seems that inhibition of tumor angiogenesis confers a therapeutic strategy. Basically, inhibition of growth factors/signalling pathways necessary for progression and growth of ECs is regarded as a vital approach to supers the tumor angiogenesis , 73. It It was shown that treatment of mice with human tumor by an anti-VEGF neutralizing monoclonal antibody (mAb) resulted in fundamental repression of the tumor expansion in the animal . BevacizUpon successful experiences, several agents inhibiting the VEGF pathway have been developed that are in the clinical trial evaluations, which target either VEGF or the related receptor. AS a chimeric soluble receptor of VEGF, VEGF-TrapR1R2 contain functional sections of VEGFR1 and VEGFR2 , and theInhibition of VEGF signalling pathway by blocking the VEGFR has been accompanied with promising outcomes , 83. ForConflicting outcomes have been approached with respect to the efficacy of VEGF inhibitors between preclinical models and clinical trials. Through clinical trials, it has been observed that the efficacy of mAbs might be different compared with the small anti-angiogenesis blockers. Studies have revealed that bevacizumab combined with SOC had suitable results in individuals with metastatic BC , CRC 7878, and NBesides VEGF blockers, vascular disrupting agents (VDAs) have also been developed to inhibit angiogenesis. VDAs action in inhibition of the tumor development is through stimulation vascular collapse, leading to hypoxia and, therefore, cancer cells necrosis . As a VDLigation of growth factors to their corresponding receptors leads to acceleration of the process of angiogenesis, resulting in activation and signal transduction through receptor tyrosine kinases (RTKs) . A numbeAngiogenesis is a key event involved in the perpetuation of the growth and progression of tumor cells . EmployiExosomes derived from MSCs are able to deliver miRs into the target cells with the aim of modulation of angiogenesis in tumors. Exposure of MSCs with the exosome secretion blocker GW4869 led to reduced levels of pro-angiomiRs in the MSC-derived medium . These oThe inflammatory setting of tumor microenvironment induces the local recruitment of MSCs. Pro-inflammatory mediators produced in the tumor microenvironment might influence on the phenotype and functional characteristics of MSCs that finally results in MSC-mediated angiogenesis of tumor. In vitro investigations demonstrated that tumor necrosis factor (TNF)-α, as an inflammatory cytokine, was able to stimulate the expression of VEGF in the intra-pancreatic tissue-derived (IPTD) MSCs. Moreover, the secreted VEGF was able to enhance angiogenesis in the human endothelial cells . Yang etOver the course of past decade, there has been a quick progression on the application of cells in treatment of cancers, and MSCs-based therapies has been on the fast track even further. Despite the general anti-tumor characteristics of MSCs, these cells conferred a potential for the personalized cell-based treatments due to the ease of availability with little invasive implementations on the individuals along with a great potential of expansion in vitro . At the The 2013 phase I/II clinical trial conducted in the Hospital Universitario Niño Jesús, Madrid, Spain, evaluated the application of bone marrow-derived autologous MSCs that were transduced with the oncolytic adenovirus Celyvir for treating refractory and metastatic solid tumors both in adults and children . In thisThe Treatment of Advanced Gastrointestinal Tumors with Genetically Modified Autologous Mesenchymal Stromal Cells (TREAT-ME1) phase I/II clinical trial delivered herpes simplex virus thymidine kinase (HSV-TK) by genetically modified autologous MSCs as a delivery vehicle and monitored the safety and efficacy of the treatment . A phaseAmong the clinical trials, nine studies assessed the potential of MSCs in the amelioration of adverse outcomes during the conventional anti-cancer therapies, for example radiation-induced hemorrhagic cystitis (NCT02814864), radiotherapy-induced xerostomia NCT03874572), anthracyclines-induced cardiomyopathy (NCT02509156), cisplatin-induced acute renal failure (NCT01275612). As a result, MSC-associated cell-based cancer therapy confer a therapeutic approach in the treatment of side effects by other anti-cancer therapies, along with direct application of MSCs in cancer therapy. Although a promising progression have been made with respect to exploring the potential of MSCs in cancer therapy, this therapeutic system is in its infancy and it will take a great deal of work to transform the preclinical and clinical findings into real clinical practice (Table 874572, aAlthough application of modified MSCs has conferred a novel and promising treatment option in cancer therapy, a number of challenging issues have been raised that might limit their use. As an instance, proper homing of MSCs in the target sites as well as their survival after engraftment are potential limitations. Biomaterial scaffolds may promote the suitable engraftment of the delivered MSCs in the target regions –126. ForMSCs can preferentially migrate into the tumor tissues and have transactions with various cells in the tumor microenvironment. These cells are easily accessible, do not induce immunological responses, can be simply manipulated in vitro without requirement for immortalization; therefore, they are the most suitable choices for cell-based treatments in cancers. In spite of progress in obtaining sufficient amount of autologous and allogeneic MSCs from bone marrow, adipose tissue, umbilical cord blood, and local tissues, nonetheless, their regulation and kinetics requires further elucidation. Moreover, engineering the exosomes released by MSCs may offer another promising technique in MSC-based cancer therapy.In spite of providing novel and attracting therapeutic system in cancer therapy, MSCs has been accompanied by a number of challenging issues and limitations. Efficient homing of MSCs in the target regions and their survival after engraftment seem to be challenges in the way toward MSC-based cancer therapy. Nonetheless, a number of recent studies have tried to utilize the biomaterials to come up with solutions for such seatbacks. It should be noted that we just started to understand the behaviours of MSCs and exert them in the clinical trials. We need to be armed with identification of new hypotheses in exertion of anti-angiogenic agents that have little adverse effects. Reaching to an optimal anti-angiogenic treatment strategy requires elaborated solutions. Manipulation of MSCs for precise delivering of the angiogenesis inhibitors could be still the first option. That notwithstanding, searching for novel molecular targets as well as agents with less side effects may help to improve the anti-angiogenesis therapy of tumors in the future.


MYC, SRC, FOS), tumor suppressor genes , apoptosis regulators , transcription regulators , genes involved in DNA repair mechanisms , in cell proliferation , in the immune response , and in other significant biological processes . Further research on the genomic profile of pregnancy-associated breast cancer is urgently required in order to identify potential biomarkers facilitating early-stage diagnosis and individualized therapy.Breast cancer is the most common malignancy diagnosed during pregnancy. Strong data on the genomic profile of pregnancy-associated breast cancer are lacking. This systematic review aims to integrate and analyze all existing data from the literature regarding the genomic background and the gene mutational patterns of pregnancy-associated breast cancer. Using various genomic analysis methods, multiple differentially expressed genes and numerous non-silent mutations have been detected. More particularly, our review demonstrates the aberrant expression of several oncogenes (e.g.,  Breast cancer is the most common type of malignancy diagnosed in women. Its incidence is notably rising with increasing age , 2. BreaPABC management consists a real challenge for physicians as both the mother and the fetus may be critically damaged , 6. Due HER-2 overexpression  searched the literature and another pair of investigators (AMK and EZ) independently extracted data from each eligible study. In case of disagreement between the members of each pair, team consensus was obtained after consulting the principal designers of the study (FZ and MAD). The articles included in this systematic review had to meet certain inclusion criteria: (1) studies highlighting the genomic profile of PABC, including mutational patterns, (2) studies based on the analysis of biological samples and/or bioinformatic approaches or computational algorithms with data originating from databases, (3) articles written in the English language. Publications were excluded if they met one or more of the following criteria: (1) animal studies without subsequent validation in human specimens, (2) reviews of literature, comments, letters or duplicate publications.The search strategy retrieved 23 articles. Of these, 16 were omitted based on the exclusion criteria and 7 were eligible –17. WhilAs far as the genomic analysis approach is concerned, four studies were based on formalin-fixed paraffin embedded (FFPE) tissue analysis –13, 18, MYC  and the BRCA1 gene , tumor suppressor genes , apoptosis regulators , transcription regulators , genes involved in DNA repair mechanisms , in cell proliferation , in the immune response  and in other significant biological processes . The most significant studies on the genomic profile of PABC are shortly presented.This review aims to systematically summarize all existing data from the literature regarding the molecular nature of pregnancy-associated breast cancer. Multiple differentially expressed genes and numerous non-silent mutations have been detected. More particularly, our review demonstrates the aberrant expression of several oncogenes ; of note, alterations in the biological functions of glycosylation are correlated with breast carcinogenesis and metastasis  and the serotonin receptor signaling pathway  was statistically associated with DFS in the multivariate model.Azim et al. also evaluated the biological pathways of PABC aiming to define the prognostic value of the different molecular aberrations . Even th pathway , 23. UsiPTEN and on the levels of the apoptosis regulators PDCD4 and BCL2 in PABC, and on their role as potential markers of poor prognosis  at the BRCA2 gene when compared to non-PABC cases, suggesting an initial genetic event in the pathogenesis of PABC  rate in PABC, particularly in triple negative molecular subtype and pre-menopausal status. The researchers indicated that ASB6 may have an essential role as a prognostic biomarker and a therapeutic target in PABC management  may be recognized as promising therapeutic targets ensuring no harm to pregnancy progress and fetal development.Last but not least, Thanmalagan et al. attempted to explain the biological profile of the disease and to improve the diagnostic and therapeutic tools by analyzing the microarray profile dataset by Harvell et al. , 19. In Among the limitations of this review, it should be stressed that our conclusions are based on studies that utilized heterogeneous genomic approaches , different sample preparation methods and sample types . Additionally, no correlation among the genomic profile and the clinicopathological characteristics of PABC was examined in the majority of the studies included in our review. Furthermore, the number of eligible articles was limited due to the rarity of the disease. Thus, we are not allowed to draw definite conclusions and formulate recommendations; further studies should be conducted to confirm the abovementioned observations.In conclusion, several studies on PABC indicate an adverse prognostic outcome for the disease that is correlated with its unexplained molecular nature. Highlighting the genomic background of PABC and analyzing all the biological pathways will further facilitate the identification of novel biomarkers defining women among the general population who are at high-risk of developing PABC. Our review systematically summarizes all available data on the distinct genomic profile of PABC offering valuable insight into PABC biological characteristics; this approach may eventually serve as a significant resource for further research in the field and elucidate PABC underlying mechanisms for the disclosure of new diagnostic, prognostic and therapeutic targets. Further research in the field of pregnancy-associated breast cancer is highly recommended as its rate is expected to increase substantially in the upcoming years.All datasets generated for this study are included in the article/supplementary material.AMK and EZ were the writers of the article. MM, EZ, and AMK performed the literature search and data extraction from all studies examined. FZ and MAD contributed to the conception and design of the study and to the revision of the manuscript. All authors have read and approved the final manuscript.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.


In the present Research Topic, the red blood cell (RBC) flow-affecting properties are examined from different angles of view, revealing the complexity of these characteristics and their role in ensuring proper blood circulation.2, which is carried out mainly in the blood capillaries. The ability of blood to circulate is largely determined by the flow-affecting properties of RBCs . Under the flow, RBCs export vasoactive mediators in response to deformation and other physiological or pathological stimuli (McMahon). This mediated response to the flow promotes efficient circulation and, in turn, optimizes gas exchanges. These flow-affecting RBC properties relate mainly to cell deformability , their adherence to wall endothelial cells (ECs) (McMahon) and self-aggregation .The primary role of RBCs is the transport of respiratory gases, oxygen, and COJani et al.). RBC deformability has been postulated to be a significant determinant of RBCs survival  or under different pathological conditions, RBC deformability can be impaired. This phenomenon is induced by modification of cell membrane or cytosol composition and membrane structure, for instance, by the alteration of the scaffolding interactions between membrane and cytoskeleton, and the ion or water contents of RBCs  and the increased concentrations of acute-phase proteins, particularly fibrinogen, results in enhanced erythrocyte aggregation  role in the blood circulation, these features also affect each other. Thus, Balushi et al., treatment with antioxidants can repair these features.Over their lifespan, erythrocytes are regularly exposed to the oxidative stress (OS) and undergo various oxidative damages due to the high oxygen tension in arterial blood and their abundant heme iron content, etc. .Impairment of the flow-affecting properties of human erythrocyte was associated with the pathophysiology of various diseases, such as sepsis, thalassemia, cerebral malaria, stroke, sickle cell anemia, and diabetes. This relationship has been substantiated by Lee et al.).Under normal conditions, RBC deformability allows individual cells to traverse nutritive capillaries. Decreased deformability will result in impaired perfusion and gas exchange in peripheral tissues. Hence, RBC deformability can be regarded as a potential index to diagnose specific diseases  combined the capabilities of intravascular microscopy with numerical simulation. Spectrum of in-vitro techniques that can be used for characterization of RBCs flow-affecting properties is very broad. This list includes laser ektacytometry , laser tweezers (Lee et al., Depond et al. discuss the usefulness of these methods for the characterization of the RBC deformability for malaria patients.Most of the relevant studies have been provided by under Thus, we can conclude that the features discussed above can act independently and synergistically, affecting blood circulation, and thus, their deterioration can be considered as a powerful catalyst for circulatory disorders.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Bridge structures nationwide across the United States are aging and in need of repair or, in some cases, full replacement. Repair decisions are common among bridge owners because of the economic feasibility when compared to the higher cost of full replacement of damaged or deteriorated bridge components such as girders. Using a proper repair approach, as a long-term or just a short-term solution, can lead to benefits that could not be achieved otherwise such as considerable savings in both time and cost. Additionally, an appropriate repair approach can help avoid adverse environmental impacts, interruptions to service, overburdening of nearby infrastructure, and local opposition to construction. The main objective of this paper is to provide a synthesis of the repair methods and materials for reinforced concrete bridge girders proposed in research studies, i.e., state-of-the-art as well as state-of-the-practice established methods. Different steps in the general repair procedure are explained first. Next, a detailed description of three common bridge girder deficiencies, i.e., shear, flexural, and fire damage, is provided. For each damage type, the main causes and common solutions found in the literature are presented. The authors then provide specific recommendations to each repair procedure. This is intended to enable researchers, engineers, and decision makers to compare the available repair methods more conveniently to find the optimal repair approach for specific projects based on economic and environmental requirements as well as structural and construction conditions. A majority of the United States’ transportation infrastructure is over 50 years old . Among tThe available options applicable to a bridge with damaged girders are “leave and monitor”, “repair”, or “replacement” of the girders. Harries et al.  classifiThe main objective of this study is to gather the information about different materials and methods of bridge girder repair implementation that have been used so far in practice or merely proposed through research projects. More than 200 studies have been reviewed and summarized here with a focus on the repair of reinforced concrete (RC) bridge girders which contribute to more than 60% of the bridge inventory in the US . For thiThe main steps in a typical repair procedure can be summarized in eight steps: (1) inspection and monitoring; (2) making the first decision on choosing a repair material; (3) making the second decision on choosing the repair method; (4) surface preparation; (5) repair design (6) application of repair material; (7) prestressing of repair material, if applicable; (8) anchorage system; and (9) strand splicing, if needed. A summary of the state-of-the-art and practice on each of the eight steps is presented next.This may be performed on a periodic or usage basis, or motivated by reports of damage or extreme loading to determine the severity of the damage, cause, and prognosis ,23. The If repair is needed, then the next step is to choose an appropriate repair material. Availability and durability of the material, ease of handling on site, cost-effectiveness, type and condition of the structural element, and the targeted enhancement in the structure are factors that should be considered in making this decision . Common Fiber-reinforced composites, as shown in Depending on the type of the matrix used, fiber-reinforced composites are categorized into three groups: polymeric composites, cement-based composites, and hybrid composites.Polymeric composites: Polymeric composites  are fibers embedded in a polymeric resin such as unsaturated polyester, epoxy, vinylester, phenolic, and polyurethane resins. Epoxy resins are the most common matrix in structural repair applications due to their characteristics such as good adhesive properties, low shrinkage during curing, and resistance to environmental degradation ,33. PopuCement-based composites: Use of fibers with cementitious matrix instead of epoxy resin was proposed in the early 1980s, but did not gain that much attention until the late 1990s. Cementitious matrix has several advantages over conventional epoxy resins in terms of fire resistance, performance under ultraviolet (UV) radiation, permeability, thermal reversibility, and cost. Additionally, having similar mechanical, chemical, and physical properties as the concrete substrate is another advantage. Unlike the FRP installation safety requirement because of the characteristics of the used resins, installation of cementitious matrix can be done merely by simple troweling techniques and protective equipment for typical concrete applications. Further, unlike FRP, cementitious matrix can be applied to wet surfaces, and thus the project will not be affected by weather conditions as much. The different types of cement-based composites include sprayed concrete, textile-reinforced mortar (TRM), textile-reinforced concrete (TRC), fiber-reinforced concrete (FRC), fiber-reinforced cementitious mortar (FRCM), and mineral-based composites (MBC) ,36,37,38Hybrid composites: Different fiber-based composites can be combined together as a hybrid to enhance the strengthening efficiency. For example, they can provide a progressive failure pattern to compensate for the loss of ductility that is observed in traditional fiber-reinforced materials . AdditioUsing steel plates for the strengthening of concrete elements started in the 1960s as a fast and economical solution compared to the demolition of the structure. Ever since, it has been traditionally used for the strengthening of structural concrete members in different forms such as rods, bars, tendons, plates, strand splice systems, and steel jackets. The main benefit of steel material is its well-known properties. Nonetheless, using steel comes with the common major concerns on poor corrosion resistance and difficulty in handling on site ,29.Other less common or emerging materials used for the repair of RC bridge girders are ultra-high-performance fiber-reinforced concrete (UHPFRC or just UHPC), aluminum alloy, ferrocement, shotcrete, innovative concrete mixes made with waste material, and coatings and sealers. Each of these materials has been used in a number of repair projects due to their specific benefits and the availability of the material ,43,44,45After the repair material is chosen, the next decision is to choose a proper way for the application of the material to the damaged girder. There are several factors affecting this decision ,46,47:  or circular cross-section (bars), manufactured using the pultrusion process. Bars can be sandblasted or deformed, but studies have indicated that deformed bars have a better bond performance. Further, it has been demonstrated by some researchers that strips can lead to a more effective repair since they provide an increased surface area between the FRP and adhesive interface, with strips failing in tension rupture and achieving full composite action with the concrete ,55. FiguThe NSM method improves the fire resistance and impact damage resistance of the strengthened member compared to the EB method since the repair material is placed inside of the concrete instead of being exposed on the concrete surface, and also as a result of the increased contact area. Additionally, there is no change of dimensions in this technique, keeping the cross-section of the girder the same ,50,54. HIn the case of overheight vehicle collision and damage to the prestressed girder strands, NSM might not be the best repair choice. This is because the thickness of the strand splicing equipment used for repairing the prestressing strands can conflict with cutting grooves into the concrete surface in particular locations . AdditioEven when using the NSM method, premature debonding can still occur, resulting in incomplete use of the tensile capacity of the common FRP repair material. This is more likely to happen in beams with T-shaped or I-shaped cross-sections. Additionally, studies have indicated that in the NSM method, detachment of the cover concrete in which the NSM reinforcement is used might occur which prevents the repair approach to work in full capacity. Therefore, it might not be possible to fully utilize the tensile strength of the repair material using the NSM or EB methods, unless proper anchorage is provided. This motivated another method which is embedding reinforcing materials well inside the girder, i.e., into the concrete core beyond the cover or surface grooves, to increase the bonding. This is because the concrete core handles the stress transfer to the strengthening material, and compared to the concrete cover, it can provide better confinement, and thus improved bond behavior. Additionally, in this deep embedment technique, protection against fire and vandalism is even more effective than the NSM method ,57. FiguSurface preparation, i.e., cleaning and roughening the surfaces of composites is a critical step in the repair process which can improve bond strength. An improperly prepared surface can result in debonding or delamination. Sandblasting, water jetting, grinding, brushing, air pressure, rounding of corners, pressure washing the concrete surface, surface patching, and nylon peel-ply techniques are commonly used for this purpose. Failure in proper surface preparation can result in damage to the repair material due the delamination of the concrete substrate ,59,60,61It is recommended to remove slightly more concrete rather than too little unless it affects the bond of prestressed strands. If patching is going to be done after unsound concrete removal, the chipped area should at least be one inch deep and should have edges as straight as possible, at right angles to the surface. Air-driven chipping guns or a portable power saw can be used for cutting the concrete. However, care should be taken not to damage the strands or the reinforcement .In case there are cracks on the girder, they should be filled with proper materials, i.e., patching. There are several patching methods and the most common five ones are discussed. (1) The dry pack method: which is suitable for holes having a depth nearly equal to the smallest dimension of the section, such as the core or bolt holes. The method should not be used on shallow surfaces or for filling a hole that extends entirely through the section or member. (2) The mortar patch method: which is appropriate for concrete members with shallow defects that require a thin layer of patching material such as in honeycombs, surface voids, or areas where concrete has been pulled away with the formwork. (3) The concrete replacement method: where the defective concrete is replaced with machine-mixed concrete that will become integral with the base concrete. This is preferred when there is a void extending entirely through the section, or if the defect goes beyond the reinforcement layer, or in general if the volume is large. (4) Synthetic patching: this method is beneficial where Portland cement patches are difficult or impractical to apply. Examples are patching at freezing temperatures or patching very shallow surface defects. In these situations, epoxy- and latex-based products can be used. Epoxies can be used for a variety of purposes such as bonding agent, binder for patching mortar, adhesive for replacing large broken pieces, or as a crack repair material. Small deep holes can be patched with low-viscosity epoxy and sand, whereas shallower patches require higher-viscosity epoxy and are more expensive. Although epoxies offer excellent bond and rapid strength development, they are hard to finish and usually result in a color difference between the patch and the base concrete. Therefore, it is suggested that epoxy mortars be used only in situations where exceptional durability and strength are required. Latex materials are used in mortar to increase its tensile strength, decrease its shrinkage, and improve its bond to the base concrete, thus helping to avoid patch failure due to differential shrinkage of the patch. Latex is especially useful in situations where feathered edges cannot be avoided. (5) Epoxy injection: this method is used to repair cracks or fill honeycombed areas of moderate size and depth. It becomes an important part of the repair process, specifically for corrosion-damaged girders in which cracking and spalling of the concrete is commonplace. Epoxy injection should be done only by appropriately trained personnel ,62. FiguAs part of the surface preparation, the surface of the concrete is usually polished until fine aggregates are exposed . This imThe concrete surface should be cleaned before the application of the repair material. This can be done using a variety of methods including pressurized air and acetone or water jetting and pressure washing . It is uIn order to increase the performance of the repair that will be applied on the concrete substrate, a primer might be applied to the concrete surface. One example procedure is presented in , where aAASHTO Guide Specifications for Design of Bonded FRP Systems for Repair and Strengthening of Concrete Bridge Elements (latest version is 2013).AC125 Acceptance Criteria for Concrete and Reinforced and Unreinforced Masonry Strengthening Using Externally Bonded Fiber-Reinforced Polymer (FRP) Composite Systems (latest version is 2012). AC125 is issued by ICC Evaluation Service to establish minimum requirements for the issuance of evaluation reports on FRP composite systems under the 2012, 2009, and 2006 International Building Code (IBC) and the 1997 Uniform Building Code (UBC).ACI 440.3R Guide Test Methods for Fiber-Reinforced Polymers (FRPs) for Reinforcing or Strengthening Concrete Structures (latest version is 2004).ACI 440R Report on Fiber-Reinforced Polymer (FRP) Reinforcement for Concrete Structures (latest version is 2007).ACI SP-215 Field Applications of FRP Reinforcement: Case Studies (latest version is 2003).ISIS Design Manual No. 4, FRP Rehabilitation of Reinforced Concrete Structures, issued by the Canadian Network of Centers of Excellence on Intelligent Sensing for Innovative Structures (latest version is 2008).Design of the repair is an important step to make sure that the chosen repair material is applied in a configuration that can provide sufficient strength and durability. Different design approaches might be required depending on the type of the damage, extent of the damage, the expected durability of the repair, availability of the resources for the application of the repair, etc. Additionally, design optimization techniques might be used to achieve the most efficient outcome . In the The next step after surface preparation is the application of the repair material. Depending on the repair approach being used, i.e., EB technique, NSM method, or embedded reinforcement, the repair material should be applied in different ways and configurations. The process for the application of the repair material for each method is briefly described in this section, while the repair configuration, which mostly depends on the type of girder deficiency, is described in The EB repair techniques using FRP materials are usually implemented in three ways: 1) wet layup, (2) pre-preg, or (3) pre-cured. In the wet layup approach see , the res wet layuFirst, grooves are made into the concrete surface, and the concrete in between the cuts is chiseled away. Then, the groove is cleaned, and dust is removed using compressed air. In order to have a clean final appearance, tape can be applied to the sides of the grooves. The strengthening material  is fastened into the groove using a filler material . Finally, the adhesive surface is leveled using a trowel and the tape is removed prior to the curing of the adhesive . The proTo increase the efficiency of the repair, the material for both EB and NSM methods can be prestressed. Prestressing was first utilized for strengthening bridges in the 1950s . It enabFor cases of high peeling or shear stress, an anchorage system might be used in order to delay the debonding of the strengthening system such as FRP materials. A proper anchorage system might allow the use of a strengthening plan that otherwise would not meet the design code provisions, allowing the repair material to continue carrying load even after debonding occurs and thereby increasing its contribution. It can enable greater strengthening or the use of a wider range of possible configurations and material properties. Different anchorage systems have been introduced so far depending on the strengthening approach that they are used with. Some examples include: additional horizontal strips of the repair material, embedment of the repair material into the beam flange through precut grooves with adhesive bonding, various mechanical anchorage systems involving bolts and plates, and fan-shaped textile-based anchors ,34,71,73The fiber-based anchors have the advantage of being light-weight and non-corrosive. Additionally, since the use of FRP-based or textile-based materials is commonplace for girder repair, using a compatible anchor material is also advantageous . A drawbfpu). In order to increase their efficiency, the splices should be staggered as illustrated in When one or more prestressing strands in a prestressed girder are damaged, strand splicing can be used to do the repair. It is a fast, efficient, and cheap repair method for reconnecting damage or broken prestressing strands in order to restore the prestressing force. Strand splices alone cannot be relied on for fully restoring the ultimate strength of the strands or the element that is being repaired. That is because they are limited to developing 85% of the nominal strength of the strands they are joining  concrete crushing, (2) prestressed strand and/or steel reinforcement being exposed which makes them more susceptible to further damage and corrosion, and (3) prestressed strand loss. Harries, Kasan  indicateFor flexural repair using the EB technique, it is often recommended that the repair material is applied to the soffit of the girder since the material near the neutral axis is less efficient in strengthening the element . This isIn this method, similar to the flexural repair using the EB technique, the repair material should be applied to the soffit of the girder through grooves to have the best efficiency. This section provides recommendations for the repair of girders with flexural deficiency, based on the repair case studies found in 151 studies, including both research and real-life applications of the repairs which are both critical for achieving a sufficient understanding of a repair technique. Eight different methods were found in the literature: (1) EB FRP sheets on the soffit + EB FRP U-wraps or complete wraps ,151,152;The most common anchorage system found in the literature is transverse U-wraps evenly spaced along the entire length of the girder or at parts where it is necessary. The U-wraps enhance the bond behavior of the FRP sheet attached to the tension side of the girder as a means of flexural strengthening. They also help in reducing crack propagation in the concrete section. While continuous CFRP U-wraps are also common for the repair of impact-damaged girders, Graeff  showed tThe most commonly used material for the repair of impact-damaged girders seen in the literature is CFRP in the form of sheets and strips. However, as mentioned before, the choice of the repair approach, including the repair material and repair adhesive, highly depends on the specific project requirements and available resources. Based on the previous discussion, the recommended repair process for flexural or impact damage is presented in Fire hazards for bridges are caused by crashing vehicles, burning of fuels in the vicinity of the bridge, arson, and wildfire . Fire daHigh temperatures resulting from a fire hazard can adversely affect the performance of epoxy resins. Therefore, in the case of considering FRP-based repair and epoxy resin materials for a bridge, ACI 440 recommends ignoring the capacity contribution of the FRP. This is a major issue in the utilization of the most common materials, i.e., FRP materials, in the repair of bridge structures. Two proposed solutions to this problem are: (1) the use of cement-based adhesives instead of epoxy, which has been exhibited in part of the shotcrete repair of a bridge in Texas after intensive fire damage ; or (2) Based on the brief discussion above, if an appropriate repair option is needed for a bridge prone to fire hazard or a fire-critical bridge, one of the following three recommended approaches can be considered: (1) use of FRP-based repair with a cement-based adhesive instead of epoxy; (2) use of FRP-based repair with a cement-based fireproof coating; or (3) use of cement-based repair such as shotcrete with or without complementary FRP repairs such as confining FRP U-wraps.Bridge girder repair is an important topic due to the aging of bridge structures in the US and because full girder replacement is not favorable due to the associated higher costs and longer downtime causing traffic shutdowns and inconvenience. The goal of this study is to provide a comprehensive reference for researchers, engineers, and decision makers to compare different repair approaches for RC bridge girders, and find the best method that suites their specific repair problem to provide the highest economy, efficiency, and safety. Three main girder deficiencies in bridge structures are identified and discussed herein, which are shear deficiencies due to corrosion and exposure of the shear reinforcement, flexural deficiencies due to vehicle impact damage, and fire damage.The choice of a repair approach is highly dependent on the specific conditions of a project . However, the following methods were found to be the most common and efficient means of repair, and hence recommended in case a quick decision is needed. For shear repair, either EB CFRP U-wrap  or NSM CFRP laminates on the web with no need for anchorage or above-head installation can be used. For flexural repair, EB CFRP sheets on the soffit can be used in addition to EB discontinuous CFRP U-wraps. For fire repair, the use of cement-based adhesives instead of epoxy and/or fireproofing of FRP layers is recommended.


The functioning of a cell at various organizational levels is determined by the interactions between macromolecules that promote cellular organelle formation and orchestrate metabolic pathways via the control of enzymatic activities. Although highly specific and relatively stable protein-protein, protein-DNA, and protein-RNA interactions are traditionally suggested as the drivers for cellular function realization, recent advances in the discovery of weak multivalent interactions have uncovered the role of so-called macromolecule condensates. These structures, which are highly divergent in size, composition, function, and cellular localization are predominantly formed by liquid-liquid phase separation (LLPS): a physical-chemical process where an initially homogenous solution turns into two distinct phases, one of which contains the major portion of the dissolved macromolecules and the other one containing the solvent. In a living cell, LLPS drives the formation of membrane-less organelles such as the nucleolus, nuclear bodies, and viral replication factories and facilitates the assembly of complex macromolecule aggregates possessing regulatory, structural, and enzymatic functions. Here, we discuss the role of LLPS in the spatial organization of eukaryotic chromatin and regulation of gene expression in normal and pathological conditions. The eukaryotic cell nucleus contains several functional compartments, such as the nucleolus, speckles, Cajal bodies, and PML bodies . These cLLPS occurs when the concentration of interacting molecules reaches a threshold value, which can be achieved within a local neighborhood due to a gathering of interacting molecules on a platform possessing specific affinity sites for macromolecules capable of forming phase condensates . This prLLPS first drew attention in connection with the assembly of the so-called nuclear bodies, such as the nucleolus, nuclear speckles, Cajal bodies, and ND10 bodies ,38. NuclThe role of LLPS in global nuclear space organization is likely not restricted to the assembly of nuclear bodies. The recent model of eukaryotic cell nucleus organization discriminates the chromatin domain represented by chromosomal territories and the interchromatin compartment (IC) ,43. The 3D genome organization has become a hot topic in molecular biology because it has been demonstrated that for transcription activation remote enhancers should establish spatial contact with target promoters, a process that is only possible at the 3D genome level ,49,50. IChromatin fiber can be regarded as a polymer comprising alternating blocks of different nature . UndSeveral observations show that LLPS is essential for the assembly of both constitutive and facultative heterochromatin. It has long been assumed that the dense packaging of DNA in heterochromatin physically prevents DNA accessibility to transcription factors and components of transcription machinery ,69. ReceFurthermore, some essential genes are located in heterochromatin and are transcribed . It thusAnother nuclear protein that is likely to contribute to LLPS in the pericentromeric chromatin is Scaffold Associated Factor B (SAFB), previously known as nuclear matrix protein . SAFB inAlthough all of the above-discussed observations suggest the role of LLPS in heterochromatin formation, the typical heterochromatin domains possess some features that distinguish them from liquid condensates. These domains do not necessarily have a round shape, and their crucial components, such as HP1, do not rapidly exchange with the nucleoplasmic pull. It is, therefore, possible that after being initially assembled, the LLPS heterochromatic domains eventually undergo gelation ,75. GelaWhatever the mechanism of heterochromatin domain formation , it is likely to be controlled by epigenetic modifications of histone tails. Of note, nucleosome arrays lacking these tails do not produce liquid droplets. Acetylation of histone tails in pre-formed droplets causes dissolving of droplets . BridginAnalysis of Hi-C maps has demonstrated that spatial interactions between remote genomic elements exist preferentially within A and B chromatin compartments but to a much lesser extent between these compartments .Active chromatin is less densely packed as compared to heterochromatin. Within the A chromatin compartment, there are no general organizers, such as HP1 or Polycomb proteins. However, long-distance interactions within the A compartment are common and, furthermore, in some cells, they are more pronounced than long-distance interactions within the B compartment . Most ofBesides transcriptional factories, nuclear speckles also represent LLPS-derived compartments that attract active genes ,94,95. AThe list of liquid nuclear compartments to which various genes may be recruited is not limited by transcription factories and nuclear speckles. A subset of genes is attracted to Cajal bodies ,100, andAn important feature of the transcription control system in higher eukaryotes is the presence of remote enhancers that can be located hundreds of kilobases away from target promoters ,105. TheBeing brought to spatial proximity, liquid compartments associated with an enhancer and a promoter fuse, giving rise to a common compartment rich in proteins necessary for effective transcription initiation  107,116116. Of nA characteristic feature of many eukaryotic promoters is the presence of paused Pol II elongation complexes . The regRecent observations suggest that distinct liquid condensates are formed at gene bodies in the course of transcription ,131. TheThe LLPS process being determined by the structure of cellular proteins and RNAs is affected by genetic mutations in a number of severe diseases, such as neurodegenerative disorders  and cancThe same is true for Rett syndrome (RTT), which is a postnatal neurodevelopmental disorder characterized by mental disability and autism-like symptoms. In RTT, different mutations in the C-terminal IDR and structured methyl-binding domain of the methyl CpG binding protein 2 (MeCP2) suppress the ability of this protein to form condensates in vivo . In partThe expansion of short nucleotide repeats is another example of genetic deficits affecting LLPS. It has been shown that CAG multiplication, which is characteristic of Huntington’s disease and spinocerebellar ataxias, promotes RNA aggregation in vivo and in vitro . CAG-conThe importance of simple physicochemical processes in the organization of living systems has long been known. A good example is the formation of membranes, which is guided by simple physicochemical processes. The role of such processes in the formation of non-membranous structures became apparent only relatively recently after the demonstration of the importance of forces arising under conditions of macromolecular crowding for maintaining the integrity of various structures in the cell nucleus. Separation of liquid phases is another physicochemical phenomenon that plays an extremely important role in the functioning of living systems due to the fact that it is this phenomenon that underlies the compartmentalization of the intracellular space. Compartmentalization is one of the fundamental characteristics of a living cell. Without intention to discuss various theories of the origin of life, we only note that all of these theories postulate the separation of a certain protocell from the surrounding inanimate matter. It is also easy to see that cell compartmentalization became more complicated with evolution. Functional compartmentalization of the eukaryotic cell nucleus is interconnected with the spatial genome organization and underlies the realization of genome activities such as replication and transcription. Various regulatory events, such as establishing enhancer-promoter communication, also rely on dynamic compartmentalization manifested by the assembly of activating compartments via LLPS. To this end, it is not surprising that compromising LLPS results in the development of various diseases, including cancer. The ambitious task is, therefore, to learn how one can modify and control LLPS within living cells. Of course, this control must be targeted. As discussed above, in many cases, the macromolecules undergoing LLPS are gathered on a certain platform, which may be protein, RNA, or even DNA. The clearest method to disrupt liquid condensates of a particular type is, thus, to target this platform. The approach would depend on the nature and stability of a platform, the number of interacting molecules, and the properties of interacting sites. One solution is to suppress the synthesis of a platform (if the platform is RNA or protein). However, the possibility of blocking interacting sites with some drugs may also be considered. Whatever solution is found, we may expect that drugs affecting the integrity of macromolecular condensates will eventually be developed. The cell nucleus is an equilibrium system stabilized by various interactions of different nature.


Flavivirus genus of the Flaviviridae family, are a major public health threat in tropical and subtropical regions, and have recently become a medical concern in temperate zones. Most flaviviruses are classified as zoonotic viruses. Human flavivirus infections can be asymptomatic, responsible for unspecific symptoms in the first few days following infection, or responsible for severe complications potentially resulting in death. During the first days following symptom onset, laboratory diagnosis of acute human flavivirus infection is mainly based on molecular detection of the viral genome by RT-PCR methods, followed by the capture of specific antibodies using serological tests after the first week of infection. The detection of antibodies that have virus neutralizing activity can be used to confirm flavivirus infection. However, human flavivirus infections induce the production of cross-reactive antibodies, often making serology inconclusive. Indeed, serological diagnosis of flavivirus infection can be hampered by a patient’s history of flavivirus exposure, particularly in regions where multiple antigenically related flaviviruses co-circulate. We focus our mini review on conventional immunoassays that allow the diagnosis of major flavivirus-associated human infections in basic, routine and high-profile central health centers; and the interpretation of diagnostic serology tests for patients living within different epidemiological situations.Arthropod-borne viruses (arboviruses) belonging to the  Most ars (DENV) ,4, West s (DENV) , or Zikas (DENV) ,8. Reemes (DENV) , or ticks (DENV) , have nos (DENV) . Medicals (DENV) . This was (DENV)  and Euros (DENV)  from thes (DENV)  in Medits (DENV)  and reces (DENV) . To prevs (DENV) ,18. Flavs (DENV) , WNV [20s (DENV)  as well s (DENV) , can comFlavivirus genus [Flavivirus-associated human disease diagnosis requires laboratory testing, either through direct detection of the infecting agent, or the detection of antibodies directed against the infecting virus. The detection of viral RNA using PCR-based techniques offers excellent detection performance of flaviviruses in biological fluids. Most of the time, viral RNA amplification techniques still require clinical laboratories with advanced technology. Antibody tests detect antibodies directed against flavivirus antigens, mainly the envelope E protein, and, depending on the serological assay, can detect different immunoglobulin classes including IgM, IgG, and IgA. Interpretation of serological results can be challenging, principally due to an extensive cross-antigenic reactivity between the members of the us genus ,27,28,29Routine diagnosis of flavivirus-associated human diseases relies on the detection of the pathogen, its nucleic acids or specific viral antigens during the acute phase of the disease followed by the capture of specific antibodies at least one week after the infection. Advantages and limitations of each method and the window of detection are detailed in Flavivirus-associated human diseases can be reliably confirmed by the detection of viral nucleic acids using conventional reverse transcription (RT)-PCR or real-time RT-qPCR. The molecular diagnosis of flavivirus infection is dependent on the presence of viral nucleic acids in the sampled biological material. The RT-PCR assay, which is used routinely on blood specimens, can also be used on urine samples for ZIKV ,31 and WAlthough RT-PCR is a sensitive and specific method that allows rapid and reliable diagnosis of acute phase flavivirus in human diseases, its efficacy is mainly limited to the acute phase of infection (a few days post infection), and can be impacted by briefness of the viremic period or a low viral load in the blood in patients infected with flaviviruses such as WNV  or ZIKV The early diagnosis of dengue can also be performed using immunocapture of viral protein NS1 in the bloodstream using commercially available kits . UnfortuAedes mosquito C6/36 cells are well-recognized methods for the confirmation of flavivirus infections, but their use is usually restricted to reference laboratories [In addition, virus isolation methods in cell lines such as African green monkey Vero cells and ratories .Specific immunoglobulin class M antibodies are usually detectable from the first week to 3 months post infection, and immunoglobulin class G antibodies are usually detectable from 2 weeks to several months or years. However, important variations can take place in the kinetics of antibody response depending on the biological assay used, viral antigens targeted for the antibody capture, immunological status in relation to a previous flavivirus exposure either by natural infection or vaccination, primary and secondary flavivirus-related human diseases, and individual immunological backgrounds.Specific antibodies can be detected in CSF for flaviviruses responsible for severe neurological complications, especially tick-borne encephalitis virus (TBEV) , WNV 6,,6,25 or The main limitation of flavivirus serological assays is antibody cross-reactivity ,27,28,29External quality assessment for WNV , DENV 5 or TBEV For ZIKV serology, cross reactions using first line serological assays are so promiscuous that official guidelines recommend the confirmation of all positive and inconclusive results by confirmatory serological assays . Indeed,DENV-1 to DENV-4 infections display similar symptoms, and antibody cross-reactivity makes simple antigen-based discrimination difficult between these four serotypes . DiffereSerological assay alone cannot discriminate primary from secondary DENV infections known to cause more severe disease that could relate to antibody-dependent enhancement phenomena ,53.The detection of neutralizing anti-flavivirus antibodies is correlated with the presence of the specific IgGs in blood specimens. Conventional plaque-reduction neutralization tests (PRNT) and virus neutralization assays (VNA) in microplates are considered the “gold standard” in discriminant flavivirus serology . HoweverAlthough virus neutralization assays are the reference tests for serological diagnosis of flavivirus-associated human diseases, serological cross reactions between the flaviviruses are also reported with this assay  and variOne major advance in the diagnosis of flavivirus human-associated infections is the development of multiplex assays that can simultaneously detect antibodies directed against several flaviviruses in the same run. High-density microarrays containing peptides derived from various flaviviruses have been developed to improve the sensitive and specificity of flavivirus serodiagnosis . MicrospA wide range of methodological designs have been applied to serosurvey studies and the lack of standardization represents a major limitation in the interpretation of results, as demonstrated for DENV or ZIKV .Diagnosis should take into account the access to laboratory tools, the epidemiology of arbovirus diseases in the region where the patient is supposed to have been infected, and the past exposure to flaviviruses.Most countries that experience high levels of endemic flavivirus circulation have limited laboratory capabilities. It is not unusual that only rapid serological diagnosis test kits are available at the point of care testing ,77. ConsIn countries with limited laboratory capacities, ELISA assays are usually not available and there is no access to confirmatory virus neutralization assays. This is one of the main limitations of the implementation of international recommendations and guidelines that focus on gold standard assays but do not take test and facility availability into consideration.In high income countries, even if tests are available and access to reference laboratory possible, the issue is that commercial serological assays have not been developed for each flavivirus of medical concern. Multiple flaviviruses co-circulate in most areas of endemic diseases , and co-Another consequence of co-circulation is the high risk of co-infection and a positive result for a flavivirus does not preclude infection with another one .In endemic areas of several flaviviruses, the rate of serologic positivity against such pathogens is so elevated that most serological assays are unreliable for the diagnosis of a flavivirus infection. This becomes even more intricate in regions where confirmatory serologic diagnosis tests are still lacking or when most people have been vaccinated for flavivirus-associated diseases such as dengue, yellow fever and Japanese encephalitis. Implementation of multiplex microsphere immunoassays (MIA) using native or recombinant viral antigens could be appropriate for resolving the complexity of flavivirus diagnostic serology in endemic areas ,84,85,86Most serological assays against flaviviruses are evaluated in terms of specificity and sensitivity, but not in predictive value, which is the probability that the test gives the correct diagnosis ,88,89. PAs mentioned above, because the prevalence of flavivirus-associated human diseases is highly variable in time and space in endemic areas, interpretation of results requires detailed knowledge of the local epidemiology at the moment the patient has been exposed.As the window of detection of flavivirus RNA, of specific IgM and IgG antibodies, is determined relative to the onset of symptoms, determination of the exact timing of the symptoms is key to using the correct diagnostic test. Usually the onset of symptoms can be clearly identified in infections such as dengue because there is a rapid onset of high fever. However, in some flavivirus infections such as Zika, there is often no fever or a low grade fever without acute onset of symptoms . If therAs most flavivirus-associated human diseases are asymptomatic or responsible for mild symptoms, patients may have past unrecognized disease. Indeed, in endemic areas it should be considered that all residents may have cross-reactive antibodies.In flavivirus endemic regions where the populations are engaged in a vaccination program such as yellow fever in Africa and South America and Japanese encephalitis in South East Asia, or enrolled in a clinical trial of a candidate vaccine against a flavivirus of medical interest, interpretation of serological testing necessitates available information on the patient’s vaccination history.To illustrate the complexity in interpreting flavivirus serological results, we will consider the example of French Polynesia (South Pacific region), which suffers from high levels of flavivirus circulation, is a major tourist area, and has been identified as a hub for the spread of medically-important flaviviruses including Zika in 2013–2014 .DENV was the only flavivirus circulating in French Polynesia until the emergence of ZIKV in 2013. DENV and ZIKV co-circulated from 2013 to 2014, and since mid-2014 DENV is the only circulating flavivirus .Until the emergence of ZIKV in French Polynesia, the available serology tests for DENV provided reliable diagnosis for the disease among a large majority of residents. During the period of co-circulation of DENV and ZIKV, there was great difficulty in the serological diagnosis of both diseases due to a strong cross-antigenic reactivity between the two flaviviruses.At the end of the ZIKV epidemic in French Polynesia, where at least one-half of the residents developed ZIKV antibodies , the serFor a traveler returning from French Polynesia, the situation is quite different. As has been observed with residents, a serology positive for DENV was specific prior to the emergence of ZIKV, whereas after the emergence, interpretation was confounded for the whole period of virus co-circulation due to the risk of cross-reactive antibodies. However, after the period of co-circulation, a positive serology could once again be considered specific for DENV (excepted if the traveler has a history of travel to another endemic area for flaviviruses or has been vaccinated against another flavivirus).Indeed, the same serological test can have high or low predictive values, depending on the circulating rate of flaviviruses in the concerned area at the time of infection.Flavivirus-associated human diseases are a growing cause of morbidity and mortality across the world and their area of distribution is expanding. Flaviviruses show a large amount of cross-reactivity in serological diagnostic tests, triggered by natural infection or vaccination. Cross-antigenic reactivity combined with a large overlap in clinical syndromes, co-circulation of different flaviviruses, and poor access to advanced laboratory diagnosis tools for serological confirmation make serological diagnosis of flavivirus-associated human diseases a great challenge. Most international guidelines and recommendations rely on gold standard assays, such as the virus neutralization assay, that unfortunately are not available in most areas with high endemic flavivirus circulation. We have identified an urgent need for more specific and sensitive serological methods allowing reliable standardized diagnosis for the timely management of flavivirus-associated human diseases.Due to the frequent unreliability of current flavivirus serological tests, it is strongly encouraged to use molecular diagnosis in the first days following the onset of symptoms. However, we are convinced that implementation of multiplex serological assays based on the use of recombinant viral antigens will open avenues for the development of new specific and sensitive diagnostic tools for flavivirus detection in the near future ,72,73,74


The use of nanoparticles (NP) in diagnosis and treatment of many human diseases, including cancer, is of increasing interest. However, cytotoxic effects of NPs on cells and the uptake efficiency significantly limit their use in clinical practice. The physico-chemical properties of NPs including surface composition, superficial charge, size and shape are considered the key factors that affect the biocompatibility and uptake efficiency of these nanoplatforms. Thanks to the possibility of modifying physico-chemical properties of NPs, it is possible to improve their biocompatibility and uptake efficiency through the functionalization of the NP surface. In this review, we summarize some of the most recent studies in which NP surface modification enhances biocompatibility and uptake. Furthermore, the most used techniques used to assess biocompatibility and uptake are also reported. Nanoparticles (NPs) are ultrafine particles with a size between 10 and 500 nm composed of different organic and/or inorganic materials . These pWhen NPs are used in biomedical applications two very important characteristics must be considered: toxicity and cellular uptake . In factThe use of NPs has grown exponentially in the last 15 years especially for cancer treatment, because of their capability to perform high precision tasks, such as the delivery of drugs and imaging contrast agents (CAs) directly to tumor cells , by usinThe use of NPs for active targeting of drugs and/or contrast agents is based on the interaction of properly modified NPs surface with a molecular target overexpressed by cells and tissues. The molecules used to modify the NPs surface include small proteins, peptides, antibodies, aptamers, and oligosaccharides . FurtherThe focus of this review is to summarize the most used techniques to evaluate NPs toxicity and uptake and to describe the most useful strategies to avoid these issues through the modification of the NPs surface.NPs can be applied to cancer for two main purposes: diagnosis and therapy . The usaMedical imaging techniques have evolved very quickly over the past 20 years, overall in cancer disease, to identify very little tumor masses for an early diagnosis . This evFor cancer therapy, there are currently numerous agents available depending on the disease stage, location, type of tissue, age, and conditions of the patient . The useThe surface modification of nanoparticles is a powerful methodology to fix or attenuate issues related to NPs toxicity and uptake, since both phenomena are closely related to the NPs surface composition.2, R-COOH, etc.), useful to bind biological molecules. For silica NPs, the most used linkers are aminosilanes that introduce an amino group on the NPs surface for the next bio-conjugation  and it is specially used with metallic and silica NPs . Non-covIn vivo experiments confirmed the results obtained in vitro, showing that the DOX-loaded NPs inhibited the growth of tumor more efficiently compared to free drug and also reduced side-effects , scansion electron microscopy (SEM), and Raman microscopy. These procedures, unlike label-based ones, have the advantage of not requiring the use of fluorophores that could affect the NPs size or chemical properties. Furthermore, using labeled-NPs complicates the discrimination of internalized NPs or those attached to the cell membrane. Conversely, TEM and SEM imaging, despite offering high-resolution down to the cellular organelle scale, are relatively expensive and time-consuming. Furthermore, TEM and SEM, unlike Raman microscopy, are destructive imaging approaches.2 nanoparticles for the cell membrane, compared to bare NPs, in A549 cells and human monocytes. Furthermore, TEM allowed to identify the nanoparticles in endosomes. TEM analysis is widely used to study nanoparticle uptake and cellular localization. in vitro and in vivo cellular imaging. This methodology was used by Among label-free techniques, Raman microscopy presents several advantages useful to study nanoparticles uptake. Compared to SEM and TEM analysis, Raman microscopy requires minimum sample preparation and allows Label-based techniques include methods that exploit a fluorescent signal emitted by the nanoparticles. This signal can be attributed to an intrinsic property of the NPs  or to thThe use of this approach to study NPs cellular uptake has several advantages like the ease of use, no need or simple specimen preparation, possibility to analyze live-cells and to perform time-lapse acquisitions. However, fluorescence methods do not allow a quantitative analysis, but only comparison between different experimental conditions . This is due to the fact that the signal is not absolute because its intensity depends on the excitation source, the number of fluorophores per NP, the quantum yield of fluorophores or the NP itself, and the sensitivity of the detector .The most used label-based techniques to study NPs cellular uptake are confocal fluorescent microscopy (CFM) and flow cytometry (FC).Owing to very high resolution, contrast and penetration depth, CFM allows the detection of very low amounts of NPs and to localize them to cellular compartments. 2 NPs uptake in NIH/3T3 and A549 cell lines. The authors observed that NIH/3T3 cells internalized the TiO2 NPs more efficiently than A549 cells, confirming the importance of the cell type in the uptake behavior. Flow cytometry can also be used to investigate the internalization pathway of the nanoparticles. The use of flow cytometry to study NPs uptake is widely used, despite the impossibility of discriminating NPs attached on cellular surface from the internalized ones. To study NPs biocompatibility, there are several aspects, summarized in The physiochemical properties of NPs, like shape, size, charge, material and surface chemical groups, influence their toxicity and uptake efficiency. Some of these, such as surface charge and chemical groups, can be easily modified by surface modification.in vitro and in vivo applications. 3O4 and SiO2 NPs increased the biocompatibility when PEGylated NPs were used at high concentrations (200 μg/mL) with murine neural stem cells, unlike bare Fe3O4&SiO2 that caused a viability reduction of about 50% already at a dose of 20 μg/mL. This study is a clear example of the high convenient properties of PEG to increase NPs biocompatibility. Furthermore, the presence of PEG on the NPs surface also improves the hemocompatibility, as observed with chromium-doped zinc gallate and diatomite based NPs  and pinocytosis . The latter can be distinguished in clathrin-dependent endocytosis, caveolae-dependent endocytosis, macropinocytosis, and clathrin, and caveolae-independent endocytosis. The modification of the physicochemical properties of NPs surface can be exploited to enhance cellular uptake.PEG, usually tested to enhance biocompatibility, can also be explored to increase NPs uptake. The PEG monomer is about 0.35 nm in length with a molecular weight of about 2 kDa, and can be synthesized at different lengths. This molecule reduces aggregation and increases NPs stability in biological fluids . Further2). In particular, when longer PEG molecules and a high density were used to functionalize gold NPs, a reduction in the non-specific protein adsorption was observed. However, this kind of modification resulted in a decrease in the uptake of NPs in all cell lines tested.As reported by 2) that at physiological pH (about 7.4) are positively charged. 2) affects the cellular uptake of NPs in the HaCaT cell line and in skins explants. Negatively charged SiO2 NPs showed lower uptake levels compared to positive 3-aminopropyl-trimethoxysiliane (APS) modified NPs. Conversely, 2 groups on the surface enhanced toxicity, related to the high reactivity of the amino groups. The higher uptake and toxicity of positively charged NPs, compared to negatively charged NPs, has also been demonstrated by Other molecules used to modify NPs surface involve changes in the superficial charge. Generally, these consist of amino ending molecules  . An examIn Another strategy widely explored to increase NPs uptake is based on the use of cell penetrating peptides (CPPs). These molecules are composed of a specific aminoacidic sequence, usually polycationic or amphipathic structures, that enhance NPs uptake. Due to the importance of enhancing the uptake of NPs, research into alternative molecules that bind on the NPs surface is very active. In recent years, surface modification of NPs was mainly aimed at active targeting and cellular uptake by exploiting the specific interactions of NPs surface ligands with a wide range of receptors overexpressed in cancer cells . The molin vitro and in vivo. Results showed an elevated selective uptake in N87 human gastric cancer cells of mAb-gold NPs when compared to bare NPs. In vivo experiments indicated accumulation of mAb-gold NPs in tumor sites and a reduction of non-specific uptake at others anatomical districts. Another receptor widely used to perform active uptake in cancer cells is the epidermal growth factor receptor (EGFR). As reported by in vivo model. A further example of mAb modified NPs to improve cellular internalization was reported by Monoclonal antibodies (mAbs) used to perform active uptake of NPs are extensively studied due to their high stability, specificity and binding ability . FathianAntibodies are very useful molecules to enhance specific NPs uptake, but they have high molecular weight (about 150 kDa), and this could be an issue in bioconjugation process, above all with smaller NPs (<10 nm). The use of the antibody antigen-binding fragments (Fabs) to perform NPs active uptake is a viable alternative, as reported by in vivo showed that GE11-Evo-NPs allow higher evodiamine concentration in tumor after intravenous administration, compared to the drug alone. Furthermore, the accumulation of GE11-Evo-NPs in tumor masses in vivo was higher than Evo-NPs. Another study where a peptide was used to enhance active NPs accumulation is reported by A viable alternative to the use of antibodies and Fabs to modify NPs surface is represented by peptides that bind with high affinity to specific receptors . Indeed,The use of natural proteins that interact with cancer cell receptors has been widely investigated, as reported in several recent studies . ScheereFurthermore, other molecules are currently being explored to modify NPs surface and to perform active uptake. Among these, aptamers represent a useful molecule since are cheap and easy to synthetize . Aptamerin vivo experiments showed that the enhanced accumulation into tumor tissue was confirmed. The use of carbohydrates was also explored for targeted gene-delivery in macrophages by Exploitation of simple molecules to enhance active uptake also includes the use of carbohydrates . Among tIn addition to the molecules already mentioned, there are other small molecules useful to perform NPs active uptake . Khan etIn the last years, the development of nanoparticles functionalization strategies is considerably grown, due to potential applications of NPs in nanomedicine. Surface modification is evolved from the use of simple molecules (PEG), to the aim to decrease toxicity, clearance and immune response, to more specific and complex ligands, in order to increase specificity and efficacy. The evolution of the NPs surface bioconjugation went hand in hand with the identification of specific cellular targets. The possibility to tune the NPs physico-chemical properties in combination with the knowledge of cancer biology makes possible the use of these nano-platforms in biomedical applications, both for therapy and diagnostic, providing a great contribution to the advance of nanomedicine.Uptake and biocompatibility are two of the most important features of a usable nanoplatform for medical applications, and overcoming issues related to these two aspects is the first goal in the development of NPs. For this reason, the study of toxicity and cellular uptake is the first step during the biological assessment of NPs.in vivo and in vitro, due to the modification of surface charge and to the inactivation of reactive chemical groups that can affect cellular membrane stability. Moreover, the addition of specific molecules can also enhance NPs passive and active uptake, reducing systemic toxicity in vivo and allowing high precision therapy and/or diagnosis. The binding of molecules on the NP surface can be obtained by covalent and non-covalent approaches. The former is widely used to bind proteins, antibodies, aptamers and peptides exploited to enhance uptake and to perform active targeting, while non-covalent interactions are generally used for loading of drugs and for all molecules that must be released in the cells.The NPs surface modification is a powerful instrument to enhance uptake and biocompatibility, as confirmed by the vast amount of scientific papers focused on this topic. These studies demonstrate that the conjugation of molecules on the NPs surface can effectively enhance biocompatibility both (n)) could be necessary to stave off the ligand from NP surface. The use of spacers also has an effect on conjugation density. In fact, in order to bind a sufficient amount of molecules to the NPs it is necessary to reduce steric hindrance. As reported in numerous studies, the conjugation density is a very important parameter that can affect the behavior of NPs. Moreover, it is important to note that the conjugation of some molecules, usually used to enhance uptake, can also increase the NPs biocompatibility.The protocols for the conjugation of molecules on NPs surface depend on the nanomaterial used and on the available functional groups, for this reason the use of linker molecules or the modification of NPs and/or ligands in order to obtain a stable conjugation could be necessary. Furthermore, some molecules used to functionalize NPs have high molecular weight due to the overall size of the proteins and the use of a spacer  the addition of big size molecules could change the size of the NPs, thus influencing uptake; (II) some molecules used for active targeting could change their 3D structure during the functionalization protocol and to lose the ability to bind target molecules; (III) the conjugation with non-covalent bonds could result in an unstable surface modification due to the influence of environmental conditions like pH and ionic strength; (IV) the conjugation density and the orientation of ligands on NPs surface is a key parameter to enhance uptake; (V) the reduction of toxicity could be due to a reduction in NPs uptake; and (VI) positively charged NPs are usually better internalized by cells.In conclusion, through the handling of surface characteristics, the nanoparticles can be transformed in smart platforms, containing therapeutic and imaging agents as well as stealth property, delivering drugs to specific tissues and providing controlled release therapy. This targeted and sustained drug delivery decreases the drug related toxicity and the frequency of treatments. Nanoparticles have proven useful in the treatment of cancer, and many other diseases, also providing advancement in diagnostic and theranostic applications.GS and AL designed the review. All authors wrote, read and approved the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The human intestinal tract is colonized by a resilient integrated ecosystem represented by a complex consortium of trillions of microbes. Overall, five phyla dominate the bacterial composition of the gut microbiota with the 90% of phylotypes identified in Firmicutes and Bacteroidetes, whereas Actinobacteria, Proteobacteria, and Verrucomicrobia are minor components. Each microorganism within the gut encodes unique metabolic functions which taken together comprise the gut microbiome. Indeed, the metabolic potential of the gut metagenome, or the sum of microbial encoded genes is about 100-fold that of the human genome, and even with functional redundancy between different microbial species, represents a metabolic potential rivaling that of the liver both in chemical diversity and impact on host health [The gut microbiota is a highly dynamic system with its density and composition affected by many exogenous  and endogenous  influencing factors. All of the environmental factors may uniquely influence the microbiota composition, but it has been shown that diet, in particular, plays a major role in determining the composition and changes of the gut microbiota .Alistipes, Bilophila, and Bacteroides) and decreased the levels of Firmicutes that metabolize dietary plant polysaccharides producing butyrate . Western diets, which are mainly omnivore-type diets apparently lead to a composition of the microbiota that is more associated with different types of disease. On contrary, the Mediterranean diet (MD) that has a very large fiber content and bioavailability, particularly in terms of insoluble fiber, being more than twofold higher than in a typical Western diet, has been associated with a beneficial microbiome related metabolomics profiles [Specifically, compositional pattern of the gut microbiota has been associated with habitual diets. In 2014, David et al.  demonstrOther specific changes have been observed in therapeutical dietary interventions such as low-fermentable, oligo-, di-, mono-saccharides and polyols (FODMAPs), ketogenic (KD), and gluten-free (GFD) diets prescribed in irritable bowel syndrome (IBS), celiac disease (CD), or neurological disorders (ND). These kinds of diets are characterized by a reduction or exclusion of a specific nutrient from the entire dietary pattern. Despite these alimentary regimens showing beneficial effects on disease symptoms, they can affect microbiota composition, especially if they are protracted for a long time .However, due to the complexity of the food composition  and their interactions, it is very difficult to identify the effects of single dietary nutrients in the context of the different dietary patterns in habitual or therapeutical diets. In vitro models are used ad preclinical tools to assess the impact of specific food components on human gut mucosa by mimicking digestion, absorption and intestinal fermentation. Several in silico models could be interconnected and used in the future to predict the effect of food systems on various metabolic responses. The huge number of scientific reports about in vitro gastrointestinal digestion systems in the latest 40 years indicated the interest in understanding the behavior of food and food components during human digestion . HoweverBifidobacterium, Lactobacillus, Roseburia, Bacteroides, Akkermansia, butyrate-producing Fecalibacterium, and Ruminococcus at the genus level, which are associated with various health benefits, whereas diets containing more than 44% energy from fat increase the ratio of F/B, which can be attenuated by red wine/tea-derived polyphenols. Major groups of polyphenols assayed in both in vitro and preclinical studies have shown their ability to modulate the gut microbiota to a beneficial pool characterized by the abundance of Bifidobacterium, Lactobacillus, Akkermansia, and Fecalibacterium sp. The beneficial mechanisms observed in those studies were mainly attributed to the production of SCFAs and other bacterial metabolites that contributed towards positive changes in gut health and reducing the inflammatory process, thereby improving systemic disease status. A very interesting review article published by Yang in Nutrients  summarizThe authors synthesized the role of some micronutrients  in modulating gut microbiota toward a healthy phenotype but underline a scarcity in the literature that explores the role of minerals and trace elements in modulating gut microbiota. The use of over-the counter dietary supplements of vitamins and minerals is increasing among the public; it is therefore extremely useful to understand their potential interaction with the gut microbiota which is not always positive. Following the work of Yang et al.  a few inThe final question regarding the impact of the total diet on gut microbiota remains partly unanswered but we have many elements that will be useful to depict the picture. Diet, especially high intake of fermentable fibers and plant polyphenols, appears to regulate microbial activities within the gut, supporting regulatory guidelines encouraging increased consumption of whole-plant foods , and providing the scientific rationale for the design of efficacious prebiotics. Due to complexity of multifactorial diet–microbiota studies, novel tools for extracting useful information from large data sets (machine learning) are required to predict specific food consumption pattern by measuring bacterial biomarkers, combined with fecal food metabolites and indicators of the immune and mucosal status of the gut . This hoHealth care professionals should be educated on the important role of dietary choices in modulating gut microbiota for both preventive and therapeutically purposes. The maintenance of a healthy gut microbiota, via nutrition and use of food supplements, could lead to the reduction of the metabolic risk associated to dangerous lifestyles.