Nanotechnology remains under continuous development. The unique, fascinating, and tunable properties of nanomaterials make them interesting for diverse applications in different fields such as medicine, agriculture, and remediation. However, knowledge about the risks associated with nanomaterials is still poorly known and presents variable results. Furthermore, the interaction of nanomaterials with biological systems and the environment still needs to be clarified. Moreover, some issues such as toxicity, bioaccumulation, and physicochemical transformations are found to be dependent on several factors such as size, capping agent, and shape, making the comparisons even more complex. This review presents a comprehensive discussion about the consequences of the use and development of nanomaterials regarding their potential risks to the environment as well as human and animal health. For this purpose, we reviewed the entire production chain from manufacturing, product development, applications, and even product disposal to raise the important implications at each stage. In addition, we present the recent developments in terms of risk management and the recycling of nanomaterials. Furthermore, the advances and limitations in the legislation and characterization of nanomaterials are also discussed. Nanotechnology involves materials at a scale ranging from 1 to 100 nm in one of its dimensions. It has brought advances in several areas such as electronics, pharmaceuticals, biotechnology, agriculture, cosmetics, and food ,5,6,7,8.The characteristics of NMs such as size, shape, surface area, stability, and the coating compounds’ intrinsic properties are essential to assess the potential risks . These fThis review presents different aspects involving nanomaterials regarding their potential risks considering the entire production chain, starting from the manufacturing process, product development, applications, and disposal . KnowingTop–down and bottom–up methodologies have been adopted to synthesize NMs through different routes such as physical, chemical, and biological. Physical and chemical methods are widely used. However, they usually require toxic solvents and generate by-products that have high-risks to the environment and harm human and animal health, hence limited applications ,13.Recently, the biogenic approach has gained notoriety. In this synthesis method, biological systems or their products are used in the process. Microorganisms, plant extracts, or isolated biomolecules such as sugars, nucleic acids, and proteins are being investigated for this purpose ,24,25,26Green chemistry aims to produce nanomaterials sustainably to reduce the maximum waste generated, energy consumption, and the concentration of precursors required. It replaces toxic compounds and solvents with benign or less toxic alternatives such as plant extracts. Another example consists of the use of water instead of organic solvents. Furthermore, reducing the steps during the synthesis process will spare wasting reagents and energy. Another essential factor to be considered is that more eco-friendly conditions of pH, temperature, and pressure are adopted during green synthesis and the establishment of protocols with reactions that generate few or no by-products. Thus, the synthesis needs to be sustainable throughout the process including the disposal ,30,31,32One of the most studied routes is using plant extracts as reducing agents and surface stabilizers. Different plant parts have already been used such as fruits ,34,35,36A variety of products containing NMs are already on the market and under development. However, through their production and utilization, these NMs can reach the environment and may cause contamination and pollution. Furthermore, how NMs are used interferes with the nature of risk exposure. Below, some of the most common applications of NMs and their main contamination targets will be presented and discussed . A revie2 and ZnONPs are used for their absorbing properties of ultraviolet (UV) radiation and hypoallergenicity. To constitute a transparent emulsion, they must have nanometric dimensions [One of NM applications is cosmetics, mainly in sunscreens. TiOmensions ,12. Anotmensions . 2 NPs have been investigated for their photocatalytic properties to eliminate organic compounds. In this context, gold nanoparticle decorated TiO2 (Au/TiO2) produced using green tea exhibited an adsorption capacity greater than 8185 mg/g of methylene blue [Nanotechnology has also emerged as a potential alternative for environmental decontamination. Various nanomaterials such as NPs, carbon nanotubes (CNTs), graphene, and nanocomposites have been extensively investigated for contaminant remediation . Studiesene blue . Carbon ene blue . In anotene blue . For thiene blue . AdditioFurthermore, many studies have reported the antimicrobial activity of different nanostructures such as silver, gold, copper, and titanium NPs ,53,54. TIn addition, NMs have offered new approaches for diagnosing diseases, imaging, administration and drug formulations as well as therapy, particularly those with antimicrobial and antitumor potential. Some NPs have demonstrated that they can be used to selectively combat tumor cells without causing significant effects on normal cells ,73,74,75NMs can also be used for drug delivery, enabling them to deliver drugs with greater efficiency, reducing toxicity, and enhancing their effects. For example, oleic acid (OA)-pluronic-coated iron oxide magnetic NPs were efficiently associated with the antitumor agent doxorubicin. This association allows for sustained drug release, and aside from the cytotoxic effect on tumor cells, was found to be dose-dependent . SimilarIn addition, the possibility of using nanotechnology in agriculture has attracted great attention. NMs can be used to increase agricultural productivity, reduce costs, and improve crop management efficiency . Nanofor2O2, uric acid, urea, neurotransmitters, hormones, drugs, metals, pesticides, viruses, bacteria, and even tumors has already been widely reported in the literature [Finally, nanotechnology has allowed for the development of nanosensors capable of detecting a variety of target analytes, which can be molecules, nucleic acids, proteins, and even cells such as bacteria ,112. Nanterature ,59,60,61A multifactorial assessment is needed to understand the possible harmful effects of NMs. In this context, it is important to highlight that the properties of NMs vary considerably depending on the particle size, shape, chemical composition, capping agent, charge, solubility, and aggregation state . NMs usuWhen in contact with the medium, NMs can adsorb components onto their surface. This affinity can originate mainly from electrostatic interactions, hydrogen, and hydrophobic bonds . The adsEspecially in biological media, NMs can be rapidly coated with proteins to form a protein corona. The latter consists of two layers : the har2) NPs were found to induce conformational changes of tubulin, thus inhibiting its polymerization properties [The composition of the biological environment can affect the dynamic protein exchange processes between the environment and the protein corona, thereby influencing the properties of the protein corona. However, NMs can adsorb many different molecules at the surface including polysaccharides, proteins, lipids, nucleic acids, metabolites, and other molecules, named the Eco-corona. In this case, protein may not be the most abundant constituent, mainly if it is formed outside an organism . The prooperties . It has operties  and fibroperties .3PO4. For example, chloride ions can interact with Ag ions to form AgCl, precipitating in the solution. As a result, fewer Ag ions are available, and the toxicity is reduced [+ release from the ions increases under high temperatures and decreases in alkaline pH. The addition of organic matter such as humic or fulvic acids can also decrease the dissolution rate of AgNPs.The dissolution process is related to the transformation of NMs such as NPs into their ion form. These ions’ release of NMs is associated with higher toxicity ,113. In  reduced . Some re reduced ,126,127.Moreover, the formation of new NMs resulting from the dissolution of another has already been proposed . In envi2+ and Mg2+ can affect the NMs’ surface charge, removing the repulsive energy, and leading to aggregation [Furthermore, NMs can interact with each other and aggregate without a stabilizing coating, or when the repulsive force is weaker than the attractive force . In addiregation . The ionregation ,129. Theregation . Larger regation .To expand the understanding of these transformations, AgNPs were studied in four different aquatic microcosms: surface water; water and sediment; water and aquatic plants; or water, sediment, and aquatic plants. Polyvinylpyrrolidone (PVP) or gum Arabic (GA) coated AgNPs were introduced into these environments. In response to the presence of AgNPs, plants released dissolved organic matter (DOM) into the water to bind Ag ions and decreased their toxicity. The plant-derived DOM had a stabilizing effect on the PVP-AgNPs but led to the removal of the GA coating from the GA-AgNPs and dissolution. These released ions bind to sediments or plant surfaces associated with thiol groups and present as oxides. In microenvironments without plants, PVP-AgNPs aggregated extensively, which was not observed for GA-AgNPs . 2 NPs. At concentrations lower than 1 mg/L, colloids in the samples led to heteroaggregates, followed by sedimentation. However, at higher concentrations (10 mg/L and 100 mg/L), homoaggregation was more important than interaction with natural colloids as being mainly responsible for aggregation. When the water samples were filtered, almost no sedimentation was observed, and the CeO2 NPs were more stable. The stability of the NPs was directly related to the initial concentration and the presence of colloids, which may be due to the increased frequency of collisions, consequently causing aggregation and sedimentation [2 with a higher ionic valence showed no marked effect on the dissolution of CuONPs at low ionic strength (10 mM) but slightly inhibited the release of Cu2+ at much higher ionic strength (100 mM). The result suggests that the ionic strength and the type of electrolyte impacted the release of CuONPs. In the absence of NOM, the nominally dissolved Cu concentration in the CuONPs suspension was only 0.16 mg/L. While in the presence of 100 mg/L NOM, the concentration of Cu was increased to 2.80 ± 0.14 mg/L for humic acid, 19.48 ± 0.41 mg/L for citric acid, and 1.73 ± 0.52 mg/L for L-cysteine, suggesting a strong effect of NOM, especially citric acid, on promoting the soluble Cu released from CuONPs. Thus, pH and NOM are critical factors in the dissolution of CuONPs, with more soluble Cu occurring at lower pH and higher NOM [Natural colloids in river water were the main ones responsible for causing the aggregation of CeOentation . In anotgher NOM . The AgNgher NOM .2 and C60 NPs. Daphnia organisms are bioindicators because they interact with large portions of water in the environment. Therefore, polluting NPs’ ingestion has tremendous potential to be affected compared to other aquatic organisms. It was observed that exposure to filtered TiO2 NPs and C60 caused mortality, in which the latter caused toxicity at much lower concentrations. The LC50 for TiO2 NPs was 5.5 ppm while that of C60 reached a much lower dose of 460 ppb. This difference was related to the NPs’ size since the C60 (0.72 nm) was significantly lower than TiO2 NPs (10–20 nm). Also, the adopted synthesis method of NPs has been found to affect their toxicity. Unfiltered sonicated NPs caused little mortality; at 25–30 times higher concentrations of TiO2 NPs, there was not much difference compared to the control. Furthermore, at a concentration of 500 ppm C60, there was only 18% mortality. These effects may result from the presence of larger NPs and clusters [In aquatic environments, NMs are present in free form or interact with organic and inorganic ligands. Meanwhile, their speciation is influenced by the physical and chemical properties of the environment. The aquatic environment is the most studied since it is the primary recipient of contaminants. Once NMs contaminate the aquatic environment, ingestion through water may occur. However, the mechanisms that living organisms respond to intoxication caused by NMs are still not well understood. An early study was devoted to assessing marine invertebrates’ toxicity toward TiOclusters . 3O4 NPs were evaluated in the microalgae Chlorella vulgaris. The results showed changes in chlorophyll content, reduced CO2 absorption, and photosynthetic rate from 50 mg/L. Oxidative stress was also noticed due to changes in stress markers’ levels. The increased levels of malondialdehyde, related to lipid peroxidation; and the reduction in glutathione levels, an antioxidant molecule that eliminates reactive oxygen species (ROS), demonstrated this finding [Algae are also essential species of the aquatic environment, as they play an indispensable role as food sources and photosynthesis. The effects of Fe finding . 2 NPs were tested against zebrafish (Danio rerio), adult specimens of Daphnia pulex, neonates of Ceriodaphnia dubia, and algae (Pseudokirchneriella subcapitata) to better understand the effects at various trophic levels. Besides, soluble salts were also tested to compare the difference between the effects of NPs and salts. It was found that both Ag- and CuNPs were toxic to all tested organisms, with the calculated LC50 ranging from 0.04 mg/L (D. pulex) to 7.2 mg/L (D. rerio fry) for AgNPs and 0.06 mg/L (D. pulex) at 0.94 mg/L (adults D. rerio) for CuNPs. Daphnia pulex was also susceptible to NiNPs . All tested NPs caused toxicity in C. dubia and P. kirchneriella at 48 and 96 h of exposure, respectively. TiO2NPs showed no toxicity in any of the tests. Based on these results, invertebrates were markedly more susceptible to NMs’ toxicity than any zebrafish stage. In general, metal salt solutions were more toxic than NPs forms [In another study, the toxicity of Ag, Cu, Co, Ni, Al, and TiOPs forms .In terrestrial ecosystems, NMs can directly or indirectly affect plants, microbial communities, and soil organisms. In plants, they can affect plant seeds’ germination and the development of plant roots and sprouts. NMs’ interaction with plants induces several physiological and morphological modifications including impaired seed germination, plant growth inhibition, reduced biomass production, and imbalanced nutrients as well as an alteration in photosynthesis and transpiration rates . NMs prePhaseolus radiatus and Sorghum bicolor growth in an agar medium, with a concentration-dependent effect. The growth rate was not affected when tested on the soil. The accumulation of Ag in the root cells was also observed [Triticum aestivum L.). The plant’s growth rate decreased markedly up to 60%, and the formation of lateral roots was stimulated. Oxidative stress associated with the presence of CuNPs has been reported [70 and MWCNTs overall rice growth stages. Black aggregates were found frequently in seeds and roots and less regularly in stems and leaves, indicating that the NPs’ absorption sequence was from the plant’s seeds and roots to the stems and leaves. The appearance of black aggregates mainly within and near the stem’s vascular system suggests that the transport of C70 occurred simultaneously with the absorption of water and nutrients in the xylem. However, the capture of MWCNTs was of little relevance, with few black aggregates appearing in the vascular system and almost none in plant tissues [AgNPs could inhibit observed . The effreported . Another tissues . B. oleracea var. capitata L.) and lettuce (L. sativa L. cv. Batavia) showed a decrease in plant weight and photosynthesis level. Additionally, CuONPs affect the stomata and were translocated to roots [2) and their equivalent ions  were tested in sweet potato (Ipomoea batatas) development. The CeO2NPs caused a small positive effect at a higher concentration (1000 mg·kg of dry weight−1) on tuber biomass, while the ZnO and CuONPs caused negative effects on tuber biomass at the same concentration. Additionally, the accumulation in both the peels and flesh was observed for all metals with no difference between the NMs and ionic treatments. This suggests that the dissolution of the NPs took place before the accumulation process [Oryza sativa L.) was strongly affected by CuONPs, reducing the water content in the shoots and roots; at a higher concentration (1000 mg·kg of dry weight−1) no plant survived to all stages, and at lower concentrations (500 mg·kg of dry weight−1), the fresh weight of grains was reduced to only 6.51% of the control values. The accumulation in the tissue plants was also seen [When exposed to CuONPs for 15 days, cabbage  treated with AgNPs and Ag+. Despite the Ag content in ion-treated plants being higher than the ones treated with AgNPs, the last caused significantly higher oxidative stress and a decrease in the glutathione and ascorbate contents [In wheat growth under hydroponic conditions, Fe noticed . When co systems . Similarcontents .Allium cepa roots. These AgNPs exhibited toxicity only at higher concentrations. AgNP-CTAB led to bigger toxicity, causing root growth inhibition and oxidative stress. The larger particle AgNP-citrate demonstrated a tendency to aggregate, thus the effects were less pronounced [Aloe vera biosynthesized AgNPs and AgNO3 reduced the growth of Brassica seedlings. This occurred due to the accumulation of Ag, which caused photosynthesis inhibition. Additionally, both led to oxidative stress, associated with increased DNA degradation and cell death. However, AgNPs were less toxic than AgNO3 due to less accumulation and better activities of the antioxidant enzymes [2 were tested against the symbiotic Alfalfa (Medicago sativa L.)—Sinorhizobium meliloti association. ZnONPs reduced the dry root biomass by 80% in all tested plants and ZnCl2 was reduced by 70–87%. Conversely, bulk ZnO increased the shoot and root biomass, while the ZnONPs increased the root length at 250 mg/kg soil, but at higher concentrations (500 and 750 mg/kg), no significant changes were observed. ZnCl2 reduced the total leaf protein in a dose-dependent and catalase activity in the stems and leaves at higher concentrations. Thus, ZnCl2 was more toxic than the ZnONPs and bulk ZnO could enhance the plant’s growth [In contrast, the ionic form of Ag was more toxic than any AgNPs with different surface coatings ) tested on onounced . Moreove enzymes . In a sis growth .2 fixation potential was reduced in all treatments, mainly for CB and MWCNT. In addition, CB and GNPs accumulated inside the root nodules [Three different carbonaceous nanomaterials, MWCNTs, graphene nanoplatelets (GNPs), or carbon black (CB) NPs were tested in the soybean plant. The soybean was grown for 39 days in soil amended with these NMs. The treated plants flowered earlier and produced up to 60–372% more flowers. In general, there were no considerable effects on the stem length, but the MWCNT-treated plants and leaves were shorter. This effect may have been in response to a stressful environment, which accelerated plant flowering to maximize the chance of flowering, at the cost of the final plant height and leaf area. Additionally, N nodules .Trigonella foenum-graecum), AgNPs promoted the growth, chlorophyll, and carotenoid content. In addition, it enhanced the number of pods and seeds and increased the antioxidant activity of the yielded seeds [Positive effects were also observed. Biosynthesized AgNPs significantly increased the root length, shoot length, fresh weight, and dry weight of rice seedlings compared to the control . After fed seeds . Other red seeds . Phaseolus vulgaris) germination and pod production. While Z-COTE® NPs did not lead to phenotypic changes, the Z-COTE HP1® NPs affected the root (~44%) and leaf length (~13%). The plant’s exposure to 125 mg/kg of Z-COTE® NPs increased the Zn content by 203%, 139%, and 76% in the nodules, stems, and leaves, respectively, while at the same concentration, Z-COTE HP1® increased Zn by 89%, 97%, and 103% in the roots, stems, and leaves, respectively. Moreover, the ZnCl2 treatment reduced the chlorophyll content by 34–46% and the leaf length by 16% at 125 mg/kg, while the bulk ZnO reduced the root length by 53% at 62.5 mg/kg [Commercially available uncoated ZnONPs (Z-COTE) and coated (Z-COTE HP1) did not affect bean  NPs and the uncoated-CeO2NPs (positively charged) [Thuja occidentalis extract–mediated green AgNPs improved the chlorophyll content, leaf number and area index, and nitrate reductase activity in Phaseolus vulgaris. However, higher doses (50 and 100 mg⋅kg) increased the proline levels, indicating higher oxidative stress [PVP-coated CeOcharged) . Thuja oe stress . Amaranthus tricolor L.), and maize (Zea mays Linn), the last were found to be more resistant to NM exposure. Important changes in the fresh weight were observed for rice with the fresh weights being reduced by 43%, 47%, 28%, 46%, and 52% for the CuO, Ag, TiO2, MWCNT, and few-layer graphene (FLG) NMs, respectively. In the case of wheat, the fresh weight was not significantly affected. In the case of the C4 plants, the fresh weight of maize was reduced across all NM exposure by 22%, 28%, 20%, 27%, and 15% for the CuO, Ag, TiO2, MWCNT, and FLG NMs, respectively. In the case of amaranth, the fresh weight was reduced by 41%, 19%, 23%, and 18% for the CuO, Ag, TiO2, and FLG NMs, respectively. In the MWCNTs, the fresh plant weight increased by 77%. A similar trend was also observed for root elongation, with the overall effect on C3 plants being the highest. In the case of the C4 plants, no significant effects on the roots were observed for amaranth, whereas Ag, TiO2, and FLG reduced the root elongation of maize by 35%, 26%, and 33%, respectively. The C3 plants reduced root elongation by 29%, 21%, and 29% for Ag, TiO2, and MWCNTs in wheat and 29%, 37%, and 29% for CuO, respectively, Ag- and TiO2NPs in rice. NM uptake and metal variation were greater in C3 species than in the C4 plants. For example, Ag concentrations in wheat and rice were at least 1.3-fold higher than that manifested in maize and amaranth. Copper levels in rice were 1.4- and 1.6-fold higher than in maize and amaranth, respectively. The copper concentrations in wheat were 1.3- and 1.5-fold greater than in maize and amaranth, respectively. The titanium content in rice was 1.6- and 3.5-fold higher than in maize and amaranth, respectively. In rice, CuONPs affected the chloroplast thylakoid structure and accumulated in the vascular sheath cells. In contrast, there were no considerable changes in the chloroplasts, and accumulation was found in the maize leaves treated with CuONPs [Comparing the C3 plants (wheat and rice), C4 plants  showed anti-inflammatory potential in models of acute inflammation and lesions caused by psoriasis in humans. Morphological changes in the concentration-dependent trend of HaCaT keratinocytes were observed. The cells became rounded, losing their characteristic epithelial cell shape and intense cytoplasmic vacuolization. These NPs accumulated in the keratinocytes, and cell viability was reduced by 50% at 79.4 µg/mL [The effects of starch-coated AgNPs were evaluated in normal human lung fibroblasts (IMR-90) and human glioblastoma cells (U251). Exposure to AgNPs caused damage to the mitochondria and increased reactive oxygen species (ROS). DNA damage was also observed. However, the tumor cell appeared to be more affected. This damage were possibly responsible for stopping the cell cycle in the G2/M phase. AgNPs were found deposited inside the mitochondria and nucleus, which can be directly related to DNA damage and mitochondrial toxicity . The cytotential . Silver .4 µg/mL .It is important to highlight that the results reported in the literature vary widely depending on the used NMs. CNTs did not lead to acute toxicity in the cell viability assays in rat NR8383 lung cells and human A549. However, increased intracellular reactive oxygen species (ROS) concentration and decreased mitochondrial membrane potential were observed . Halloys+ can interact with Cl− to form AgCl, and in the gastric acid fluid, it enters the bloodstream through systemic circulation by active transport routes for Na+ and enter the bloodstream. It is expected to bind to proteins like albumin in thiol complexes and be distributed to various tissues and organs. Additionally, it was observed that Ag-thiol complexes under UV light are photo-reduced and form new AgNPs that can react with sulfur (S) and selenium (Se). Fascinating findings were Se tarnishing of Ag surfaces and the selenide’s capacity to quickly replace sulfide in Ag2SNPs and Ag2S films through exchange reaction, leading to argyria through deposits as Ag/S/Se are located in the near-skin region. This deposition occurred preferentially in the light-affected areas [AgNPs subjected to the action of simulated saliva, gastric, and intestinal fluids showed changes in size and morphology depending on the fluid’s pH and composition. The main effect observed was aggregation, thus increasing the size of the NPs. However, low toxicity was observed for the 3D models of these tissues. Furthermore, inflammatory cytokine IL-1a decreased in epithelium models of gingival and oral tissue after repeated application of AgNPs, thus manifesting anti-inflammatory properties . Syntheted areas .2 of 15 nm in the human epidermal keratinocyte cell line HaCaT. A decrease of more than 20% in global DNA methylation was observed. In addition, the levels of methyltransferases (DNMTs) and methyl-CpG binding protein 2 also decreased [3) were vulnerable at even low concentrations (0.08 μg/mL), with a significant reduction in cell viability. Although Ag ions could impair the cell viability at concentrations of 0.08 and 0.80 μg/mL, globin gene transcription was not significantly affected compared to the Ag nanomaterial at 8 μg/mL. Additionally, it was demonstrated that over 82.1% of Ag existed in the form of NPs, with less than 17.9% as Ag ions inside MEL cells. Furthermore, the AgNPs could potentially bind to RNA polymerase but not to DNA in the in vitro transcriptional system, leading to impaired Sp6 RNA polymerase activity and reduced RNA synthesis. In vivo studies with mice also showed a large reduction associated with anemia in the embryos. The above findings showed that the Ag nanomaterial can change transcription levels through RNA polymerase inhibition [Atm, Cdk, and Gsr) and hypomethylation (Gpx) in different genes. Large AuNPs (250 nm) also changed Trp53 methylation, while 5 nm AuNPs did not significantly affect CpG methylation compared to the controls [post coitum (dpc) and euthanized after 13.5–17.5 dpc. AgNP treatment enhanced the meiotic progression of female germ cells in the fetal mouse ovaries. The RNA levels of some development-related genes  of oocytes were significantly reduced after exposure. The treatment also disrupted imprinted gene expression  in the embryos and placentas [Recent studies have shown that NMs can also affect gene transcription and have epigenetic effects ,170. Oneecreased . AgNPs whibition . Similarhibition . Anotherhibition . In anotcontrols . Sphericlacentas .In contrast, two types of carbon dots  did not cause changes in the DNA methylation pattern in human embryonic lung fibroblasts (HEL 12469), even when exposed to concentrations of 50 and 100 µg/mL, respectively, for a long duration of 24 h. However, miRNA and mRNA expressions were affected by other mechanisms associated with immune response, tumorigenesis, and cell cycle regulation after exposure to positively charged CDs. For the negatively charged CDs, it was associated with pathways related to cell proliferation, apoptosis, oxidative stress, gene expression, and cycle regulation . The mecPseudokirchinella subcapitata were exposed to AuNPs of different shapes and sizes. For a concentration of 2.9 × 1011 particles·mL−1, no toxicity was observed in the algae subjected to these exposures. However, it was observed that 10 nm spherical NPs were present in 68% of the cells in the algal population. The rod-shaped 10 × 45 nm NPs were present in just 34% of the cells. In daphnids fed with AuNP-exposed algae, the Au accumulation was 0.73%  and 1.71% (for rod-shaped) of the total Au in algae [The risks also need to be evaluated across the entire food chain, considering the trophic transfer potential. A recent study evaluated the impacts and distribution of AuNPs at different trophic levels in an aquatic food chain. Initially, the algae of the species in algae .Gammarus fossarum. Biofilms exposed for 48 h to AuNPs at two concentrations, 4.6 and 46 mg/L, were used to feed the crustacean for 7 days, with daily biofilm renewal. Both approaches generated oxidative stress in cells, mainly affecting mitochondrial respiration. Alterations in digestive enzyme activity were also reported. In the cilia of apical intestinal epithelial cells, muscle fibers, and mitochondria, the latter clearly showed a disruption of crests and internal and external membranes. Moreover, a significant difference in expression in three genes occurred: cat  and sodMn (superoxide dismutase) associated with oxidative stress; and 16S associated with mitochondrial metabolism. On the other hand, no differences in the number of metallothioneins were reported. Nevertheless, the amylase and trypsin activities were 1.45-times higher when gammarids were exposed to the highest AuNPs concentration in biofilms (46.5 mg/L) [At the next trophic level, zebrafish were fed with 10 contaminated daphnids each day for 21 days. A small percentage of AuNPs accumulated in daphnids transferred to zebrafish varied from 0.03%  to 0.48% (for rod-shaped) with the brain and liver as the target organs. Furthermore, AuNP dissolution and agglomeration in the gut of the daphnids and biodistribution in fish tissues were found to be size- and shape-dependent . Similar.5 mg/L) .13-fullerenol NPs were tested in marine ecosystems. Scenedesmus obliquus (phytoplankton) was exposed to C13-fullerenol (1 mg/L) for 36 h under continuous gentle aeration. These NPs accumulated in Scenedesmus obliquus through water exposure, where it was possible to observe black dots of 18 to 78 nm in algal cells. In Daphnia magna fed with S. obliquus, no mortality was observed. When ingested by zebrafish, fullerenol accumulated more in the intestine, followed by the liver, muscle, gills, and brain. After the 28 days of uptake and a 15 day depuration period, fullerenol NPs were mainly distributed in the intestine and liver. The gastrointestinal tract is the initial target tissue in dietary exposure, and it is expected that fullerenols can pass through intestinal membranes and enter the circulatory system, where they are subsequently distributed to other tissues. The biomagnification factor (BMF) value of fullerenes from S. obliquus to D. magna was 3.20, while the BMF value from D. magna to D. rerio was 0.54. This value is dependent on the gut passage time, assimilation, elimination ratio, and environmental conditions. Thus, the biomagnification of fullerenes was higher from the first to the second trophic level, but not to the next trophic level [D. rerio by dietary exposure. However, no considerable biomagnification was reported and the BMF for both adult and juvenile zebrafish was less than one . In addition, no mortality was observed [In another study, Cic level . For theobserved .Eisenia fetida/D. rerio/Oreochromis niloticus. E. fetida individuals were exposed to CNFs and offered to D. rerio, later feeding O. niloticus. The accumulation and cytotoxic effects occurred in O. niloticus exposed to CNFs via the food chain, and it was associated with an elevated number of erythrocyte nuclear abnormalities such as constricted erythrocyte nuclei, vacuole, blebbed, kidney-shaped, and micronucleated erythrocytes. These results suggest the occurrence of defects caused by CNFs in cell division processes. In contrast, the presence of blebbed, kidney-shaped, and notched nuclei can be inferred as precursor changes to micronuclei. Oxidative stress induction by CNFs is another possibility that could explain the erythrocyte nuclear abnormalities observed in this study [Moreover, the effects of carbon nanofibers (CNFs) across trophic levels were evaluated in a food chain: is study .2NPs from marine algae Dunaliella salina to marine crustacean Artemia salina were evaluated by waterborne (exposure of TiO2 NPs to Artemia) and dietary exposure . The crustacean was more susceptible to TiO2NPs (48 h LC50 of 4.21 mg/L) than marine algae, D. salina (48 h LC50 of 11.35 mg/L). A decrease in chlorophyll content dependent on the exposure concentration was also observed, which correlated well with the cell viability loss results. The toxicity, uptake, and accumulation were higher with waterborne exposure compared to dietary exposure. No trophic transfer from algae to Artemia occurred through dietary exposure. Waterborne exposure seemed to cause higher ROS and antioxidant enzyme production, accumulating in the midgut region, and caused the deformation of cells in the epithelial lining of the hindgut region [In another approach, the toxicity and trophic transfer potential of TiOt region .Hypostomus plecostomus. For 21 days, the fish were exposed to the biofilms earlier contaminated with 0.48 or 4.8 mg/L AuNPs. No fish mortality was noted during this experiment. The Au content could be transferred from the biofilms to fish and the bioaccumulation was organ and exposure level-dependent. All organs except the heart showed a significant Au accumulation. The digestive tract had the highest accumulation, 60- and 220-fold more than the control for 0.48 and 4.8 mg/L AuNP exposure. The kidney was second with 49- and 154-fold increases in 0.48 and 4.8 mg/L compared to the control. The brain significantly increased the gold concentration at the highest exposure level, 5-fold more than the control, whereas 0.48 mg/L was unchanged. Even in lower concentrations, histological alterations were detected in the exposed liver, spleen, and muscle of the fish. The liver exhibited a severe increase in the lipid droplets in the size and frequency with vacuolization of the hepatocyte cytoplasm. In the spleen, the control fish showed a homogeneous distribution of cell types, whereas melano-macrophage centers were observed in the exposed fish. Skeletal muscle tissue appeared disordered for the exposed fish compared to the control, with disorganized myofibrils and broken fibers leading to spaces between the fiber bundles [The potential of PEG-coated AuNPs in the trophic transfer was evaluated using an experimental benthic food chain that included two trophic levels: natural river biofilm and grazer fish  bundles .Chlamydomonas reinhardtii was exposed to AgNPs (citrate-coated and fluorogen-coated) with a concentration of 1000 μg/L. The phytoplankton internalized both, but the surface coating changed the entrance pathways. Fluorogen-coated AgNPs were internalized through endocytosis and citrate-coated AgNPs through the apical zone of the cell near the flagella. In the cells, AgNPs suffer dissolution, releasing Ag ions in the cytoplasm. The trophically transferred AgNPs to D. magna were concentrated in the gut regions due to the direct ingestion of food particles. After ingestion, about 95% of the trophically transferred Ag+ was eliminated. Retention of fluorogen-coated AgNPs by daphnids was more intense than that of the citrate-AgNPs due to their lower dissolution of Ag+ [Chlamydomonas reinhardtii, Daphnia magna, and zebrafish. It was observed that longer AgNWs were more toxic than shorter ones to both the algae and water fleas, but shorter AgNWs accumulated more than longer ones in the fish’s body .In another study, the phytoplankton n of Ag+ . The ecoEisenia andrei)—Collembola (Lobella sokamensis) food chain. The survival earthworms were assessed after the 7 days of exposition to AgNP contaminated soil . After this time, they were killed and added to the receptacles with L. sokamensis. At the lowest concentration, low bioaccumulation and negligible effects were reported. Nonetheless, the highest concentration led to juvenile mortality and enhanced the trophic transfer to Collembola, which impaired the locomotion and survival rates in a dose-dependent manner [NM toxicity through food chains in terrestrial ecosystems was studied in a soil earthworm (t manner .Escherichia coli (E. coli) and Caenorhabditis elegans (C. elegans) were used to assess the PVP-coated AgNP toxicity. The AgNPs accumulated in E. coli and transferred to the C. elegans through the food chain. The AgNPs were in the gut lumen, subcutaneous tissue, and gonad, and induced extreme toxicity including effects on reproductive integrity and life span, and germ cell death. Relative to larger NPs (75 nm), small ones (25 nm) accumulated more easily and were more toxic to the higher trophic level. This finding reinforces that smaller NMs have more toxicity . The trophic transfer potential of AuNPs was evaluated using the plant Nicotiana tabacum L. Cv Xanthi and the caterpillars (Manduca sexta). The BMF of the primary producer to a direct consumer was 6.2, 11.6, and 9.6 for the 5, 10, and 15 nm AuNP treatments, respectively [Manduca sexta) was fed with tomato leaves contaminated with 12 nm tannate-coated AuNPs. A low bioaccumulation factor (BAF) of 0.16 was observed and no evidence that the caterpillars were affected by the ingestion of AuNP contaminated plant tissue, indicating that the bioaccumulation efficiency via this pathway is not considerable for this study model [In another food chain model, ectively . The samdy model . The difOreochromis niloticus), but they potentiate the toxicity of the pesticide carbofuran more than 5-fold [D. magna in the presence of the herbicide phenanthrene, depending on the condition evaluated [D. magna when incubated with MWCNTs [D. magna. However, it increased more than ten times the toxicity of phenanthrene. No difference was observed for atrazine or methyl parathion [Another point that needs to be considered when evaluating toxicity is the effect of NMs in the presence of other contaminants. NMs could serve as carriers for other contaminants, enhancing the absorption by organisms, reactivity, bioavailability, or even present synergistic effects. However, the interaction of NMs with contaminants can also lead to positive effects. NMs can interact and adsorb contaminants, decreasing their availability, leading or not to aggregation and sedimentation, promoting the degradation of contaminants, and competing with contaminants for interaction sites . MWCNTs n 5-fold . Five tyvaluated . In addivaluated . MWCNTs h MWCNTs . Fullerearathion .2NPs were observed at 2 mg/L in embryonic abalone  but at the same concentration, the TiO2 increased the toxicity of tributyltin up to 20-fold compared with tributyltin alone, causing malformation. This enhancement was associated with tributyltin adsorption onto TiO2NP aggregates and the internalization of them by abalone embryos [2NPs also increased the toxicity of phenanthrene and cadmium (Cd2+) to A. salina at a low concentration of 5 mg/L TiO2NPs. However, at higher concentrations of 400 mg/L, the toxicity decreased by 24.5 and 57.1%, respectively. This toxicity can be associated with the pollutants’ adsorption on the surface of the TiO2NPs, TiO2-facilitated bioaccumulation of pollutants, and the aggregation and sedimentation of TiO2NPs [2NPs promoted the metabolism of pentachlorophenol to tetrachlorohydroquinone in zebrafish larvae, and the co-exposure led to higher ROS generation, eventually leading to lipid peroxidation and DNA damage [2NPs could facilitate arsenic (As) sorption on microalgae Nannochloropsis maritima within an exposure period of 24 h. This effect caused a higher As trophic transfer from the algae to A. salina nauplii. However, the retention of As in A. salina fed As-TiO2NP contaminated algae was lower than one fed As-contaminated algae after 48 h of depuration. This could be due to the reduced bioavailability of As in the presence of TiO2NPs [2 was observed. The presence of TiO2NPs improved the accumulation of Cd in carp (Cyprinus carpio). After 25 days of treatment, Cd concentration in carp significantly increased by 146% with a positive correlation between the Cd and TiO2 concentrations [No developed effects of TiO embryos . TiO2NPs TiO2NPs . In addiA damage . In addi TiO2NPs . In anottrations . In summAssessing the toxicity of NMs still presents many challenges, although some advancements in research and regulations have been made. As previously mentioned, several parameters influence the characteristics of NMs, and consequently their toxicity. Furthermore, there exists a discrepancy in the limits when comparing between study results and the creation of quality standards . To bettSince the toxicity of NMs varies according to the different physicochemical characteristics alongside the existence of different results regarding their toxicity, understanding these dynamics and establishing patterns remains a great challenge. Therefore, it is vital to consider the specific properties of each material aside from the exposure routes and the corresponding conditions. Many steps can be taken into consideration to better manage the risks of nanotechnology. The first step in assessing the potential risks of a nanomaterial is to perform in-depth and breadth characterizations. Studying its physicochemical characteristics will help to estimate the possible effects on a biological system and elucidate more precisely which ones can be modified to reduce toxicity.To understand the effects on biological systems, it is important to have a well-defined nanomaterial concentration and characterization. Fluorescent, isotopic, or radio-labeled markers could be used to determine the dosage, meanwhile, the labeling process can change the biodistribution and physicochemical properties as well as contribute to the toxicity. Moreover, some techniques such as single particle inductively coupled plasma mass spectrometry (single particle-ICP-MS) have been used to detect and quantify nanoparticles but are found to be laborious and are not suitable for some samples ,203.In addition, the existence of discrepancies between the results obtained for the same NMs and the important variety between them makes each NM unique, and extrapolating the results to others is very challenging. Despite various methods being available, there is still no broadly defined standard for assessing the toxicity of NMS in vitro and in vivo. Furthermore, the wide variety of NM characteristics can explain some discrepancies in the toxicity results. Changing just one parameter such as particle size can be enough to alter the toxicity. Moreover, other variable parameters such as exposure time, dosage, and cell lineage will also affect the results. Furthermore, it is important to know the proprieties of NMs such as the size, shape, capping agent, and speciation of the dissolved metal ions in the biological environment. It is also important to consider the production of artifacts in toxicity experiments. In general, multiple assays are based on optical detection by colorimetric and fluorimetric analyses. Thus, some NMs can interfere with these methodologies by changing parameters such as the adsorption capacity, optical properties, and hydrophobicity. In addition, some NMs can interfere with some cytotoxicity in vitro assays by scattering or absorbing light in the same spectral range of the test. Indeed, they can adsorb, deplete, and inactivate the reagents, or even interact with cell products like proteins ,205,206.Moreover, understanding the effects of NMs in complex and highly variable environments presents a great challenge. For example, evaluating all possible interactions of NMs in the atmosphere with a wide variety of contaminants, airflow, precipitation, seasonal variations, temperature, humidity, and others, remains a major challenge . The chaTo assess the potential risks of NMs to the environment, it is important to identify the emission source from their fabrication to disposal. Mathematical models can be considered as an important tool to help in examining the risks and can be used in conjunction with the experimental data for regulatory assessment. However, the amount of data from the different stages of the life cycle of NMs are still scarce and more information is needed to refine these models and obtain more accurate results . AnotherFurthermore, in regions where NMs are produced or discarded, higher concentrations in the atmosphere are expected. Some strategies can be taken to reduce possible contamination including closed reactors and emission control systems during the synthesis . AlternaSome NMs can be involved during the recycling process when associated with some products such as electronics, textiles, and construction materials, but this is poorly understood. Different processes are being developed to recover NMs from waste ,219,220.3 as an 8-h time-weighted average (TWA) of EC for the respirable range fraction [3 for fine TiO2 and 0.3 mg/m3 for ultrafine TiO2 as TWA concentrations for up to 10 h/day [Furthermore, in recent years, different countries have started to establish some laws to regulate the production, marketing, use, and disposal of NMs. The regulatory challenges are also enormous because NMs present large differences and a case-by-case assessment may not be efficient with the rapid growth of nanotechnology . For exafraction . Additio10 h/day . In Aust10 h/day . In 201710 h/day .2 as a food additive (Annex II of Regulation (EC) No. 1333/2008), but in 2021, TiO2 was no longer considered safe as a food additive. France was the first European county to ban this use for TiO2 [2 as a food color if the quantity of TiO2 does not exceed 1 percent by weight of the food.Despite these restrictions, there are already a variety of products with authorized production and sale. In 2012, the FDA approved the first clinical trial using NMs. The FDA approved the first clinical trials aiming to use AuNPs for photothermal therapy against lung cancer. European Food Safety Authority authorized TiOfor TiO2 . In the For a more adequate and standardized assessment of the risks associated with NMs, many countries and institutions have created guidelines and protocols to facilitate these analyses. The ISO/TS 12901 guides occupational health and safety measures relating to NMs including personal protection and the management of accidental releases. Furthermore, collaborative projects such as NanoReg and NanoValid have been developed to support regulatory agencies and establish protocols for the characterization and evaluation of the toxicity of NMs .It is important to highlight that nanotechnology legislation is essential to ensure the quality and safety of nanoproducts. Standardization regarding metrological and nanotoxicological parameters is essential. The International Organization for Standardization has established standards for nanotechnology with ISO/TC-229 to analyze nanotechnological aspects related to health and the environment, among other ISO standards under development . SimilarNanotechnology has enabled tremendous advances in various fields of science. Further developments are necessary to improve the existing NMs while discovering novel ones that promise benefits to different technological fields. However, as these NMs are unique with significant differences, extensive studies are needed to better understand the real risks and limitations of using NMs. Nevertheless, there still exists some discrepancies in the reported results regarding the risks associated with NMs besides the lack of comparisons associated with the standardization limits. In-depth monitoring from the synthesis to the destination will certainly allow the risks to be assessed and minimized at each stage. Green synthesis can be an important alternative for the reduction in risks. It is crucial to invest in eco-friendly fabrication methods and ensure that proper and sustainable disposal is adopted. Furthermore, the implementation of 3D cell culture can be an auspicious tool in the initial assessment of the toxicity of a nanomaterial. Broader studies that consider different environmental conditions such as high and low temperatures are fundamental in evaluating the stability of these nanostructures in a natural environment. Additionally, evaluating the timeframe in which products containing NMs maintain quality is essential, ensuring that they are only marketed within a minimum quality standard.In summary, to assess the potential risks of NMs on the environment and biological systems, it is discerning to perform in-depth and breadth characterizations of NMs and examine the potential transformations and consequences on the bioavailability and toxicity. It is also essential to assess the routes and duration of exposure, stability, and interaction with other contaminants or by-products as well as carry out bioassays that provide more practical information. Nevertheless, it is important to highlight that nanotechnology can benefit society and economic development .


Intraflagellar transport (IFT) is a microtubule-based system that supports the assembly and maintenance of cilia. The dysfunction of IFT leads to ciliopathies of variable severity. Two of the IFT-B components are the paralogue proteins TTC30A and TTC30B. To investigate whether these proteins constitute redundant functions, CRISPR/Cas9 was used to generate single TTC30A or B and double-knockout hTERT-RPE1 cells. Ciliogenesis assays showed the redundancy of both proteins while the polyglutamylation of cilia was affected in single knockouts. The localization of other IFT components was not affected by the depletion of a single paralogue. A loss of both proteins led to a severe ciliogenesis defect, resulting in no cilia formation, which was rescued by TTC30A or B. The redundancy can be explained by the highly similar interaction patterns of the paralogues; both equally interact with the IFT-B machinery. Our study demonstrates that a loss of one TTC30 paralogue can mostly be compensated by the other, thus preventing severe ciliary defects. However, cells assemble shorter cilia, which are potentially limited in their function, especially because of impaired polyglutamylation. A complete loss of both proteins leads to a deficit in IFT complex B integrity followed by disrupted IFT and subsequently no cilia formation. Chlamydomonas reinhardtii IFT70/DYF-1 orthologue with IFT52 )ntegrity . Howeverntegrity ,44.flr), C. elegans (DYF1) [C. reinhardtii [flr described that can compensate for phenotypes caused by mutation or complete loss. However, in more complex organisms , paralogue TTC30 proteins can be found [Xenopus tropicalis, TTC30A was linked to ciliary chondrodysplasia with polycystic kidney disease [These findings underline, in combination with MS data, on the one hand, the need of these two paralogues with similar and non-overlapping functions, and on the other hand the importance of an intact IFT complex B itself. A loss of function in one protein can be to some degree compensated for, due to redundancy, by the other protein, but not completely. Cells still form a cilium but one that is shorter, and that is potentially limited in its functions. A complete loss of both proteins, such as the loss of other IFT proteins, leads to disrupted intraflagellar transport and, subsequently, no cilia formation at all, which is most likely a lethal condition during development ,46,47. Fs (DYF1)  and C. rnhardtii . Interesbe found ,48. Ther disease , leading disease , is a ma disease ,49. Howe disease . The mil


Backgrounds: Concomitant abdominal aortic aneurysms (AAA) and gastrointestinal malignancies are uncommon. Endovascular repair (EVAR) is widely used to treat AAA. However, no consensus exists on the optimal strategy for treating AAA when associated with pancreatic adenocarcinoma. In addition, only few reports of pancreaticoduodenectomy (PD) after EVAR exist. Presentation of case: A pancreatic tumor was detected during follow-up after EVAR for AAA in an 83-year-old female patient. The diagnosis was high-grade intraepithelial neoplasia. Modified pylorus-preserving pancreaticoduodenectomy was safely performed. The patient recovered moderately and was discharged two weeks after surgery. The pathological diagnosis was middle-grade pancreatic ductal adenocarcinoma. The patient survived for 24 months with no recurrence or cardiovascular complications. Conclusions: Conducting periodic follow-ups after AAA surgery is helpful for the early discovery of gastrointestinal tumors. EVAR surgery is safe and feasible and thus recommended for AAA patients with pancreatic cancer, although it may increase the risk of cancer. The stage of malignancy and post-EVAR medical history can be valuable in evaluating the benefits of pancreatic surgery for such cases. Pancreatic cancer is one of the most malignant gastrointestinal tumors with insidious onset and rapid progression. This cancer is characterized by high aggressiveness, metastasis, and recurrence rate. According to the latest data from the National Cancer Institute, the five-year survival rate of pancreatic cancer is only 11.5% .Case reports of abdominal aortic aneurysms (AAAs) complicated with pancreatic cancer are rare in the medical literature. Previously, surgical resection was the recommended treatment for abdominal aorta. However, a simultaneous resection of AAA and tumors of the digestive system can enormously increase the trauma and risks of artificial vessel infection. In addition, staged resection of AAA and tumor lesions can result in adhesion in the area of the first operation, leading to postoperative complications that increase the difficulty of the second operation. This problem is very significant in the case of pancreatic lesions, considering the anatomy of the pancreas and how close it is to the aorta.With the widespread use of endovascular repair (EVAR), perioperative mortality is significantly reduced, and postoperative recovery time is shortened accordingly ,4. In 202, respectively. She had undergone EVAR due to AAA at the age of 73 years old. After that, she had regular follow-up visits. The AAA was located on the infrarenal aorta . The left and right hepatic arteries had normal blood flow and originated from the proper hepatic artery without vascular variation. Modified PPPD procedure was then performed along with regional lymph node dissection (0/12), including no. 8a, 8p, 12a, 12p, 12b, 12c, 13a, and 13b. An intraoperative histological analysis revealed a negative pancreatic transection margin (R0). The patient recovered uneventfully without complications and was discharged from the hospital 12 days after surgery. Postoperative pathological analysis reported moderately differentiated ductal adenocarcinoma of the pancreas (T2N0M0), 3.5 × 3.3 × 3.3 cm in size. No regional lymph node metastases or perineural infiltrations were observed. The patient survived for 24 months with no recurrence or cardiovascular complications.It is controversial to simultaneously treat concomitant AAA and malignancies in the adjacent region, especially pancreatic tumors. We systematically searched online literature databases  for articles containing “gastrointestinal tumor”, “pancreatic cancer”, and “abdominal aortic aneurysm.” We identified a total of 20 cases of pancreatic lesions with AAA and summarized the patients’ age, gender, aneurysm size, treatment method, interval time, pathology, tumor stage, and prognosis  5,6,7,8,14,15,16It is widely recognized that risk profiles of AAA and pancreatic neoplasia are similar, which may influence the prevalence and workup of pancreatic tumor in patients with AAA . PatientAs the number of middle-aged and elderly residents increases, the incidence of AAA and pancreatic cancer is consistently on the rise. Interestingly, risk factors for AAA and pancreatic cancer overlap, such as smoking history, prevalence in males, and advanced age ,19. TherPreviously, researchers believed that when pancreatic cancer and AAA co-occurred, PD should be performed first followed by AAA resection. Deiparine et al.  reportedWith the growing research interest in pancreatic cancer, more and more surgeons are realizing the importance of the meso-pancreas ,34,35,36Previous literature has reported that the circumferential resection margin (CRM) consists of the ventral and dorsal pancreatic surfaces, as well as the medial pancreatic margin ,39. AddiThe effects of chemotherapy on aneurysms should also be considered. Russwurm et al.  reportedRegular follow-ups after AAA surgery can help in the early detection of gastrointestinal tumors. Although EVAR may increase the burden of cancer at a later stage , it is recommended for patients with complicated AAA pancreatic cancer. It is also necessary to evaluate whether patients can benefit from pancreatic surgery after EVAR by considering the stage of malignant tumor and their medical history. For patients with advanced pancreatic cancer, most patients receiving two types of treatments usually have a better prognosis, and those receiving only one kind of treatment also have a better prognosis than those without treatment. However, some treatments, such as chemotherapy, may accelerate the growth of AAA and thus should be carefully evaluated.


Catharanthus roseus). Both of these alkaloids' accumulations are known to be influenced by salicylic acid (SA). The transcriptome data to reveal the induction effect (s) of SA, however, seem restricted at this time. In this study, the de novo approach of transcriptome assembly was performed on the RNA-Sequencing (RNA-Seq) data in C. roseus. The outcome demonstrated that SA treatment boosted the expression of all the genes in the Terpenoid Indole Alkaloids (TIAs) pathway that produces the vinblastine and vincristine alkaloids. These outcomes supported the time-course measurements of vincristine alkaloid, the end product of the TIAs pathway, and demonstrated that SA spray had a positive impact on transcription and alkaloid synthesis. Additionally, the abundance of transcription factor families including bHLH, C3H, C2H2, MYB, MYB-related, AP2/ ERF, NAC, bZIP, and WRKY suggests a role for a variety of transcription families in response to the SA stimuli. Di-nucleotide and tri-nucleotide SSRs were the most prevalent SSR markers in microsatellite analyses, making up 39% and 34% of all SSR markers, respectively, out of the 77,192 total SSRs discovered.The anti-cancer vinblastine and vincristine alkaloids can only be naturally found in periwinkle ( Catharanthus roseus, a diploid (2n = 16) plant native to Madagascar, belongs to the Apocynaceae family. Despite the fact that it is grown as an ornamental plant in many tropical countries2, commercial alkaloids are rich in the aerial and underground tissues of this medicinal herb5.6. Regarding the plant kingdom, C. roseus solely synthesize vinblastine and vincristine alkaloids which are used in cancer treatments8. However, the synthesis rate and accumulation of both alkaloids in C. roseus are low and insufficient for the economic benefit of pharmaceutical industries. To this end, attempts to synthesize the aforementioned alkaloids, have not been successful yet11. The C. roseus valuable alkaloids are synthesized in the Terpenoid Indole Alkaloids (TIAs) pathway. It's interesting to note that the plant's developmental stages and the cell's reaction to biotic and abiotic stimuli alter the expression of TIAs pathway genes13. Many transcriptional activators and repressors regulate the transcription level of TIAs pathway genes17. Therefore, revealing the biosynthetic pathway of TIAs and identification of the regulators of these compounds can be useful in metabolic pathway engineering8. In recent years, several studies have been performed to identify genes and regulators of the TIAs pathway19. Despite tremendous advances, there is a need to understand and identify other genes and regulators involved in the TIAs pathwayA wide range of plant driven alkaloids such as ajmalicine, serpentine, vinblastine and vincristine have been extensively used in clinical therapy20. Recent NGS platforms have revolutionized sequencing methods to identify uncharacterized genes/proteins involved in complex pathways7. For instance, using the de novo method, Verma, et al.1 investigated the transcripts of leaf, flower and root tissues in C. roseus, resulting in the identification of 59,220 unique transcripts with an average length of 1284 bp. Almost 65% of the transcripts showed homology to the deposited sequences in the databases, while 35% of the unigenes were specific to the bisindole alkaloids synthesis activity in the plant leaves and roots organs, whereas enzymes associated with vindoline and vinblastine synthesis were solely found in the plant aerial tissues. Furthermore, Verma, et al.1 identified 11,620 simple sequence repeats (SSRs) markers and 1820 transcription factor (TFs) in C. roseus.The use of Next-Generation Sequencing (NGS) technology in analyzing genomic and transcriptomic resources in all organisms, including non-model species, enables the identification of genes involved in secondary metabolism pathways7 generated a partial genome assembly for C. roseus aiming to understand the monoterpenoid indole alkaloids (MIAs) biosynthesis pathway7. In the past 10 years,, not only the MIA pathway leading to the formation of strictosidine, but several downstream pathway branches, and previously undescribed TFs and transporters have been characterized22.Kellner, et al.24. It has already been reported that most alkaloid regulators in C. roseus are induced by JA. Nonetheless, they exert cell-specific effects, since some TFs exclusively regulate the specific part of the MIA pathway22.Jasmonic acid (JA) and salicylic acid (SA) among other elicitors have been shown to alter the expression of TIAs pathway genes26. Spraying SA on C. roseus leaves resulted in a significant increase in the expression of TIAs pathway genes, which in turn caused the accumulation of catharanthine, vinblastine and vindoline alkaloids27. Also, spraying SA on leaves in C. roseus plants facing water28 and salinity25 stresses, reduced the effects of stress and increased the total amount of alkaloids, more specifically a significant increase in vincristine, and vinblastine production rate. Nevertheless, despite the beneficial effects of the application of SA, higher concentrations (> 1 mM) of SA resulted in decreasing the amount of alkaloids, suggesting that the induction role of SA seems to depend on the concentration of SA29. Therefore, it can be anticipated that the expression pattern of associated TIAs pathway genes following SA treatments by RNA-Seq, will further facilitate the unravelling and identifying genes involved in the biosynthesis of complex molecules such as vincristine and vinblastine.It has been documented that SA induces the expression of TIAs pathway genes and the synthesis of vindoline, vinblastine and vincristine alkaloidsIn order to generate a comprehensive transcriptome of C. roseus leaves in response to 0.1 mM SA treatment 24-h post-treatment, cDNA libraries were constructed in three biological replicates and sequenced. In total, 191,022,812 bases were generated by RNA-Seq , 47,398 (83%) unitranscripts were allocated to Eudicots based on GhostKOALA analysis30. According to GhostKOALA server, genetic information processing had the highest number in the functional classification of Protein families. Sections such as secondary compounds, amino acid metabolism, and terpenoid metabolism were also identified  protein databases. Out of 74,301 transcripts identified, 44,688 (60%), 36,292 (49%), 45,111 (61%), and 22,919 (31%) from all the libraries were annotated against KAAS, Uniprot, NCBI NR protein, and GO databases, respectively  pathway. Information and names of genes with different expressions in the biosynthetic pathway of the aromatic amino acids tryptophan, phenylalanine and tyrosine can be seen in supplementary C. roseus and is not synthesized in other plants such as Arabidopsis, which has been used as a template in the analysis. For this reason, the protein sequences of TIAs pathway enzymes were obtained from the UniProtKB/Swiss-Prot. The C. roseus database was prepared for the TIAs pathway and the sequence of transcripts was compared and analyzed. As a result of this analysis, all genes in the TIAs pathway were identified 35. Overall, 77,192 SSRs were identified. Among these SSRs, the di-nucleotides and tri-nucleotide SSRs were the most abundant SSR markers, accounting for 39% and 34% of the total SSR markers, respectively 47 showed that MeJA treatments induced the expression patterns of TIAs pathway genes in Catharanthus roseus seedlings. Even though MeJA and JA seem to share similar effects on the overall expression of TIAs genes, according to Rischer et al.(2006), using C. roseus cell suspension cultures showed that JA induced the expression of some TIAs biosynthetic genes, such as G10H, SGD, and TDC, but the expression of genes encoding MAT and DAT did not change. Taken together and in comparison to our results, MeJA, JA and SA treatments seem to target similar TIAs genes expression. However, further analysis is needed to unveil the effects of all three organic compounds in a single study at the same time.It was found that many of up-regulated genes in 48. In the present study, 1895 TFs were identified that 710 transcription factors had differential expression. bHLH, C3H, C2H2, MYB, MYB-related, AP2/ERF, NAC, bZIP and WRKY families had the highest number of TF members in C. roseus6. Interestingly, Verma, et al.1 identified bHLH, MYB, AP2/ ERF and WRKY TFs in a C. roseus transcriptome study. Also, the study of the effect of methyl jasmonate and ethylene on the synthesis of TIAs led to the identification of WRKY and ABC TFs8. It is well documented that the synthesis of terpenoid indole alkaloids is modulated by TFs related to the structural genes of the TIAs pathway50. For example, two TFs, ORCA2 and ORCA3, as members of the AP2/ ERF transcription factor family, are crucial regulators TIAs Pathway51. The overexpression of ORCA2 significantly increased the expression of many genes in the TIAs pathway resulting in a higher accumulation of catharanthine, ajmalicine, serpentine, tabersonine52 and vindoline53. In addition to the role of TFs belonging to the AP2/ ERF family, it has been shown that CrMYB2 overexpression in the hairy roots of C. roseus alters the expression of ORCAs and also increases the amount of catharanthine and tabersonine54. In fact, the bHLH transcription factor regulates the expression of structural genes, whereas ORCA3 does not regulate structural genes and may play redundant roles22. WRKY1 transcription factor binds to the Wbox element in the promoter of the Tdc gene, regulating the expression level of As, Dxs, Sls, Sgp and Tdc genes, consequently, and modulating the production of serpentine and decreased catharanthine. Therefore, WRKY is a good candidate for regulating metabolites such as serpentine and ajmalicine production14.TFs play an important role by modulating the expression level of their target genes. NGS methods with the aim of bioinformatics analysis have made it possible to identify both TFs and target proteins in different species56. Out of 77,192 SSR markers identified by MISA, three and two nucleotide repeats had the highest number of SSRs with 34% and 39%, respectively. The presented results were consistent with the results of Verma, et al.1 where the highest percentage of SSRs belonged to two and three nucleotide repeats.SSR markers are co-dominant species-specific markers which are used in breeding programs to study genetic diversity, and kinship relationships germplasm analysisC. roseus as the only source of vinblastine sulfate and vincristine sulfate drugs, it is also considered as a model plant in studying secondary metabolites production. This is the first study dealing with the expression analysis of TIAs pathway genes following SA treatment. The expression of all genes in the TIAs pathway producing anti-cancer alkaloids vinblastine and vincristine increased after SA treatment. These results were consistent with the measurements of vincristine alkaloid  over time and showed that SA spray had a positive effect on both transcription and production of alkaloids. The result of this study can be used as a way to increase the anti-cancer alkaloids in C. roseus.In addition to the importance of the The pipeline of study is shown in Fig. Catharanthus roseus (L.) G. Don ‘Pacifica XP Burgundy Halo Vinca’ seeds were purchased from Pan American Seed Company . The C. roseus seeds were planted in pots (10 cm diameter × 10 cm height) and kept in a growth chamber . At the beginning of flowering stage . The collection of plant material, complies with relevant institutional, national, and international guidelines and legislation. Plant materials were then immediately frozen in the liquid nitrogen and stored at − 80 °C for RNA extraction. Based on the results of qRT-PCR analysis from our previous study27  according to the manufacturer’s instructions. The extracted RNA samples were treated with DNase I enzyme to remove possible DNA contaminations. The quantity and quality of RNA samples were analyzed by using the NanoDrop 2000c Spectrophotometer  and 1% agarose gel electrophoresis, respectively. Next, high-quality RNA samples , were sent to the sequencing company . Furthermore, quality controls and the RNA integrity number (RIN) were assessed for all RNA samples. cDNA preparation and sequencing were performed with the use of standard Illumina kits according to the manufacturer’s instructions . Finally, paired-end sequencing was performed by Illumina HiSeq 2500 platform with a read length of 150 nucleotide.59. Trimmomatic 0.3660 was employed to remove adapters, nucleotides and low quality readings as well as to check the quality of the nucleotides reads. De novo assembly was conducted using Trinity 2.11.061 by default parameters, k-mer length of 32 and minimum transcripts length of 300 bp. Trinity transcripts were clustered using CD-HIT by identity cutoff of 95%62. Transcriptome completeness was assessed using the bioinformatics tool BUSCO v3  to obtain the percentage of single-copy orthologues represented in three datasets edicots_odb1063. After integrating the readings, to identify the candidate Coding Sequences (CDS), all sequences were translated into protein sequences by TransDecoder tool (http://transdecoder.github.io). To find homology for identified transcripts and translated peptides, BLASTX and BLASTP were performed, respectively.Sequencing data quality assessment was performed by using FastQC 0.11.20http://trinotate.github.io/), functional annotation of the C. roseus transcriptome was done. Next, the assembled unigenes of C. roseus were blasted against NR proteins (http://www.ncbi.nlm.nih.gov/) and UniProtKB/Swiss-Prot (https://www.uniprot.org/) databases. Then, HMMER software was used to predict protein domains against the Pfam database. Also, signal peptide and transmembrane domains were identified using the SignalP5.0 server. Biological pathway information (Available in the Kyoto Encyclopedia of Genes and Genomes (KEEG) database) using GhostKOALA (KEGG Orthology And Links Annotation)30  and KEGG Automatic Annotation Server (KAAS) (https://www.genome.jp/kegg/kaas/) servers, gene ontology terms and functional classifications  related to identified homologous genes were extracted from the WEGO (Web Gene Ontology Annotation Plot) Web tool (http://wego.genomics.org.cn/).Using a Trinotate annotation pipeline  were obtained. Differentially expressed transcripts were blasted against the C. roseus protein database to re-evaluate the expression changes of the genes in the target pathway. Finally, the diagram of the genes involved in the TIAs pathway was plotted.Protein sequences related to the 27.For qRT- PCR validation, Total RNA extraction and cDNA synthetize were carried out. The full details of the qRT-PCR analysis and the results were published in Soltani et al.66 with slight modifications. Briefly, leaf samples were dried in the shade and pulverized leaf tissues were three times suspended in 80% methanol solution. After drying, 10 ml of water was added to the samples and the pH was adjusted to 3.5. The aqueous phase was removed by washing three times with chloroform, the pH was raised to 8.5 by adding ammonia and after three washes with chloroform, the chloroform phase was removed. Methanol was added to the dried sample and stored in the refrigerator for further analysis.The samples were prepared according to Singh, et al.67.High-performance liquid chromatography was performed using Eurospher II 100–5 C18 column with precolumn (Column 250 * 4.6 mm) and UV detector (model K-2600) with an HPLC device (KNAUER WellChrom model) at 254 ɳm. The mobile phase consisted of methanol: diammonium dihydrogen phosphate (5 mM) with a composition of 71:29 (pH = 7.3) and a flow rate of 1 mL/min68 in R software 4.1.065 to statistically compare changes of vinblastine and vincristine alkaloids content at the different sampling points. The Least Significant Difference (LSD) test was employed to compare treatment means at a confidence of 95%.One-way analysis of variance (ANOVA) was done by using Agricolae package 1.3–5C. roseus TFs families, the TF database Plant TFDB 5.0 was used69. The MISA web (https://webblast.ipk-gatersleben.de/misa/)35 was used to identify the C. roseus microsatellites.For the identification of Supplementary Information.


Background: Sodium-glucose cotransporter-2 inhibitors (SGLT2 inhibitors) are a relatively new class of medications used for the management of type II diabetes mellitus targeting the kidneys. Within the last decade, several warnings have been issued regarding the development of severe genitourinary infections, including necrotizing fasciitis, or Fournier’s gangrene, in those with pre-existing type II diabetes and concomitant use of this drug class.Objective: The purpose of this review is to highlight and discuss the factors contributing to the development of Fournier’s gangrene, its pathogenesis, and a review of existing literature describing patient outcomes, treatment, and future directions regarding early detection of this complication.Methods: Articles and studies addressing effective treatment adherence and key factors contributing to Fournier’s gangrene with SGLT2 inhibitors were identified by effective keyword searches in PubMed Central, Google Scholar, and Cochrane, as well as the references found within these articles.Results: Using the keywords provided, 55 case reports, review articles, and meta-analysis reports written within the last 20 years were utilized as the source of the data presented in this systematic review article. Fournier’s gangrene (FG) is a surgical emergency condition in which extensive necrosis of soft tissue and fascia around the genital and perineal regions ,2. EscheSGLT-2 inhibitors and mechanism of action and its side effects Sodium-glucose cotransporter-2 (SGLT-2) inhibitors are commonly orally administered glucose-lowering drugs for adult patients with type-2 diabetes mellitus (T2DM). It is a relatively new subtype of the oral antidiabetic drug, which was first introduced in 2013 . The sodSeveral clinical trials have shown that this group of drugs has a better cardiovascular outcome in terms of multiple hospitalizations and mortality associated with heart failure . Not onlThe FDA and the European Medicine Agency-approved SGLT-2 inhibitors include canagliflozin, dapagliflozin, ertugliflozin, and empagliflozin. Since these drugs increase urinary glucose excretion, expected side effects include glycosuria or polyuria . RecentlFournier’s gangrene and its association with SGLT-2 inhibitors: pathogenesisE. coli, K. pneumoniae, B. fragilis, and Staphylococcus aureus [Fournier's gangrene is a rare necrotizing fasciitis localized within the perineum and genitourinary region. FG is an aggressive infection that spreads rapidly and affects the tissues surrounding the muscles, nerves, fat, and blood vessels of the perineum, resulting in multi-organ dysfunction, septic shock, and even death . Therefos aureus . The moss aureus .The pathophysiology of T2DM places patients at a higher risk of urinary tract and genital infections, despite the fact that the exact pathophysiologic processes of FG development are unknown. Glucosuria, immunological dysfunction, and enhanced bacterial adhesion to uroepithelium all contribute to this increased risk . IncreasDiscussion of cases associated with Fournier’s gangrene and SGLT-2 inhibitorsAt the end of August 2018, the U.S. Food and Drug Administration (FDA) fitted out a black box warning with reference to a possible association between Fournier’s gangrene and SGLT-2 inhibitors . A totalStudies have shown that the incidence of FG is roughly 1.6 per 100,000 among males  ,22,23 anIn Sweden, the first case of FG was reported in 2016. A literature review was performed among eight patients who were diagnosed with FG after starting an SGLT2 inhibitor. Of these eight patients, five were being treated with dapagliflozin, two patients were receiving empagliflozin; and one patient was receiving canagliflozin. The patients' ages ranged from 39 to 72, and six out of eight of the patients were obese. Additionally, four out of eight patients were smokers. The development of FG from the initiation of SGLT2 inhibitors varied from 10 days to six years, indicating significantly varied .Another review/study showed that SGLT2 inhibitors were first launched in India in April 2015. From April 2015 to July 2018, 23 FG cases were identified. All the patients were males who were in their 50s and 60s. Of those, 70% had T2DM. All the patients required surgical intervention (the severity index was 5.08 ± 4.6) and three of them needed surgery more than once. Five patients needed to be shifted to the ICU, and three patients died from multiple organ dysfunction from sepsis .Other than literature reviews, it is important to scrutinize the voluntary reports to have an overall better comprehension of the true incidence and prevalence of FG. The FAERS database has records of the suspected adverse outcomes reported by consumers, health professionals, and manufacturers. According to one study, 319 FG cases were reported by healthcare professionals, and among them, 43.00% were submitted by physicians. Consumers reported only 193 (35.61%) cases. Studies have also shown that the number of reported FG cases has jumped from 1 in 2014 to 407 in 2019, especially after the FDA warning in 2018. An increasing number of diabetes patients and the increasing popularity of using SGLT2 inhibitors may contribute to this huge escalation . In thisBy soliciting disproportionality analysis and Bayesian analysis in SGLT2 inhibitors and FG incidents in the FAERS database from January 2004 to September 2019 , studiesWhen it comes to risk factors and comorbidities, a number of conditions are of concern; among them, pre-existing diabetes 20-70%) is the most concerning factor ,24. Stud0-70% is Diagnosis and managementUnfortunately for physicians, the diagnosis of FG is largely based on clinical findings; there are no specific imaging or laboratory studies that can rule in or rule out FG or any necrotizing soft-tissue infection (NSTI) for that matter. This indicates that detailed investigations with a high index of suspicion are crucial in patients with risk factors for FG development . HoweverTo combat the high mortality and swift progression of FG, several algorithms and calculators have been developed to enhance disease recognition and reduce disease severity . These tWhile imaging alone is not sufficient to rule out an NSTI or FG, it is certainly useful in planning surgical management. Specifically, computed tomography (CT) imaging has a diagnostic sensitivity of 88.5% and a specificity of 93.3% for NSTIs ,29,34. CA. hydrophila and can further propagate an NSTI if not mitigated [Since a majority of FG cases appear to be polymicrobial, an effective antibiotic cocktail should include coverage for streptococcal, staphylococcal, and Gram-negative bacteria; coliforms; Pseudomonas; Bacteroides; and Clostridium. Furthermore, coverage of MRSA species is recommended either with vancomycin or linezolid in conjunction with a carbapenem or beta-lactamase inhibitor Table  22,38,3,338,39. Emergent surgical debridement, broad-spectrum antibiotics, and appropriate resuscitation with intravenous fluids and vasopressor medications are the cornerstones of FG therapy ,41. ThisSeverity and outcomeGiven the extremely rapid progression and severe mortality rates of FG, there are several algorithms that have been developed to facilitate early detection and, subsequently, early treatment of FG. These tools include the Fournier's Gangrene Severity Index (FGSI), Uludag Fournier Gangrene Severity Index (UFGSI), Age-Adjusted Charlson Comorbidity Index (ACCI), the Laboratory Risk Indicator for Necrotizing Fasciitis (LRINEC) score, the Combined Urology and Plastics Index (CUPI) and the Platelet Mass Index (PMI) Score . The LRIThe simplified Fournier’s Gangrene Severity Index is the most commonly used scoring system to determine the severity of FG-specific necrotizing infections developed by Lin et al. ,31. The The goal for patient outcomes in those with FG is to save the patient from sepsis and to minimize sexual and urinary dysfunction as much as possible . While pThose with a later diagnosis of FG and prolonged delays getting to the operating room required more aggressive surgical debridement and longer hospital stays ,41. One Things to consider before starting SGLT-2 inhibitors in diabetic patientsType 2 diabetes mellitus (T2DM) is a condition resulting from a defect in the secretory function of pancreatic beta cells. Many medications are available to treat T2DM. SGLT-2 inhibitors are the newer group of oral antihyperglycemic medications which reduce blood sugar by increasing urinary glucose excretion. At present, T2DM is managed with either monotherapy with oral agents such as metformin or combination therapy with other groups of medicine. SGLT-2 inhibitors  are used with metformin to control blood glucose levels. Moreover, SGLT-2 inhibitors are part of the newest antidiabetic medication approved by the Food and Drug Administration .Preinitiation Safety ScreeningSGLT-2 inhibitors are the second-line drugs to control blood glucose levels along with other groups of antidiabetic medications ,44. The Besides the many advantages of taking SGLT-2 inhibitors, patients with type 1 DM, peripheral arterial disease, bladder malignancy, and severe renal impairment are not suitable candidates to take this group of medications. In those cases, patients may develop euglycemic diabetic ketoacidosis, limb amputation, or acute kidney injury, respectively. Furthermore, it is also not recommended for hepatic impairment patients -44,46. TSusceptibility to Fournier's Gangrene in SGLT-2 Inhibitor UsersThe most devastating adverse effect of SGLT-2 inhibitors is FG. Case reports prepared by Onder et al. and Kumar et al. have shown that the incidence of FG was more prevalent in uncontrolled T2DM patients who had poor genital hygiene due to morbid obesity ,50. OtheLimitationsSGLT2 inhibitors are a relatively new class of antihyperglycemic agents found to be efficient in improving glycemic control. They are also authorized to be used in mitigating major adverse cardiovascular occurrences among adults with type-2 diabetes and have existing cardiovascular complications. Moreover, hospitalization rates for patients with heart failure were lower and kidney disease progression was reduced when SGLT2 inhibitors were administered. Diabetes patients are at high risk of developing gangrene, and one of the side effects of SGLT2 is also gangrene. But considering the evidence we have so far, the beneficial effects outweigh the negative side effects ,53-55.FG exhibits extensive mortality and morbidity, especially in diabetic patients, which warrants the importance of prompt diagnosis of it. For SGLT2 inhibitor users, it is crucial for all clinicians to consider FG as a complication given its rapidly progressing nature, though a strong causal relationship has not been established in the literature. Being a promising class of antihyperglycemic agents, SGLT2 inhibitors have become an appealing medication modality since they have an advantageous cardiac and renal outcome. Significant microvascular complications and relative immunosuppression in DM could be blamed for the resulting FG, which can roughly be potentiated by SGLT2 use. This life-threatening necrotizing infection of the perineum in SGLT2 users demands further research to validate whether this medication is directly causing or only provoking the condition.


Cucumis sativus var. hardwickii (HD) in four kinds of soils. We found that soil type, cultivation time, and genotype affected the composition and structure of cucumber rhizosphere bacterial communities. Notably, HD showed better physiological features in sandy soil and sandy loam soil than it did in black soil and farm soil at 50 days post-sowing, which was due to its stronger recruitment ability to Nitrospira, Nocardioides, Bacillus, and Gaiella in sandy soil, and more Tumebacillus, Nitrospira, and Paenibacillus in sandy loam soil. Meanwhile, we also found that HD showed a better recruiting capacity for these bacterial genera than XT in both sandy soil and sandy loam soil. Functional predictions indicated that these bacteria might have had stronger root colonization ability and then promoted the growth of cucumbers by enhancing nitrogen metabolism and active metabolite secretion. In this study, our findings provided a better insight into the relationship between cucumber phenotype, genotype, and the rhizosphere bacterial community, which will offer valuable theoretical references for rhizosphere microbiota studies and its future application in agriculture.The composition and structure of the rhizosphere microbiome is affected by many factors, including soil type, genotype, and cultivation time of the plant. However, the interaction mechanisms among these factors are largely unclear. We use culture-independent 16S rRNA amplicon sequencing to investigate the rhizosphere bacterial composition and the structure of cultivated cucumber Xintaimici (XT) and wild-type cucumber  The rhizosphere refers to a narrow zone surrounding plant roots in which chemical and biological features of the soil are affected by the structure and exudates of the roots . The intArabidopsis thaliana were strongly affected by soil types  is one of the most important vegetables in the world, with extremely high economic value  were provided by the Functional Gene Research Group of the Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences. We collected four kinds of soil from three different cities in China. Sandy and sandy loam soils were collected from Shouguang City, Shandong Province, China . Black soil was collected from Linghai City, Liaoning Province, China , and farm soil was collected from Beijing, China .The seeds of cultivated cucumber Xintaimici (XT) and wild-type cucumber 4 1 mM, CaCl2 4 mM, KNO3 10 mM, NH4H2PO4 1 mM, KCL 10 mM, Ca(H2PO4) ⋅ 2H2O] 0.5 mM and trace element powder purchased from Coolabern 74.93 mg/L) every 4 days. No chemical fertilizers or pesticides were used in this experiment.The seeds of XT and HD were surface sterilized in 75% ethanol for 15 s and 0.3% sodium hypochlorite for 15 min, then washed six times with sterile distilled water. The seeds were germinated on a sterile paper and incubated at 28°C in the dark until the roots reached 1 cm (24 h). After germination, the cucumber seedlings were transplanted into pots with four different kinds of soils and grown in the greenhouse of the Southern District of the Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, China  under the conditions of 16/8 h light/dark and 28°C/18°C day/night. The pots were watered every 2 days with sterilized water and sterilized with modified 1/4 Hoagland’s nutrient solution [1X Hoagland’s nutrient solution: MgSOPhysiological features were measured at 30 days post-sowing (30 DPS) and 50 days post-sowing (50 DPS) of XT and HD in all four kinds of soil. Specifically, plant height was measured from the base of the shoot to the tip of the tallest leaf; the stem diameter was measured with a caliper just below the cotyledon scar; and the leaf area was calculated with this formula: leaf area = leaf length (leaf base notch to leaf tip) × leaf width (the widest part of the leaf) × 0.87. At each time point, for each soil type and cucumber genotype, triplicate samples were taken.Rhizosphere soil samples of XT and HD in four kinds of soils were collected at 30 and 50 DPS. At each time point, for each soil type and cucumber genotype, triplicate samples were collected. Roots with tightly attached rhizosphere soil were placed in a sterile tube with 25 ml of sterile water and vortexed for 30 s. Then, we transferred the supernatant to another new tube and centrifuged it for 10 min at 10,000 rpm. Following that, the supernatant was discarded, and the sample was collected from the bottom of the tube and then stored at −80°C before DNA extraction.The sterilized soil of sandy loam soil was obtained by autoclaving it three times for 30 min. After germination, both XT and HD seedlings were transplanted into pots containing sterilized and non-sterilized sandy loam soil, respectively, and grown in a greenhouse. Physiological features were measured at 40 days post-sowing (40 DPS). The triplicate samples were taken per treatment.DNA was extracted from 0.25 g of each soil sample using the DNeasy PowerSoil Pro Kit (QIAGEN HB-2485) by following the manufacturer’s instructions. The concentration and quality of the DNA were evaluated by the NanoDrop 2000 spectrophotometer . The V3–V4 region of the 16S rRNA gene was amplified using the universal primers (341F: 5′-CCTACGGGNBGCASCAG-3′ and 806R: 5′-GACTACNVGGGTATCTAATCC-3′) . Every 5via the q2−feature−classifier . One-way ANOVA and two-tailed Student’s To explore the phenotype variation of cucumbers with distinctive genotypes in different soils, we cultivated two types of cucumbers, cultivated cucumber XT and wild-type cucumber HD, on four different kinds of soil, including sandy soil, sandy loam soil, black soil, and farm soil .p < 0.05). The average height and leaf area of HD in sandy soil increased by 197.48 and 149.45% compared to that in black soil and 183.89 and 231.27% compared to that in farm soil, respectively. The average leaf area in sandy loam soil increased by 132.72% compared with black soil and 209.05% compared with farm soil, respectively  per sample on average from 72 samples .Principal coordinate analysis (PCoA) of Bray–Curtis distance was performed to reveal the overall dissimilarity of rhizosphere bacterial composition. The result showed that four kinds of soils were distributed into four clusters, which were separated along the second coordinate axis . We founAccording to the sequencing results, over 90% of ASVs can be assigned to the phylum level on average . For allNocardioides, Bacillus, and Gemmata, which may play an important role in cucumber growth; the abundance of Gaiella in sandy soil is higher than in other soils, and there are more Rubellimicrobium and Tumebacillus in sandy soil and sandy loam soil than in black and farm soil. For black soil, it contained more Pedospheraceae, Streptomyces, Gemmatimonas, and Bryobacter than sandy soil, sandy loam soil, and farm soil. Module B contained more Nannocystis, Ohtaekwangia, Arenimonas, Lureolibacter, and Cellvibrio in farm soil than in black soil at 30 DPS; conversely, there were more Massilia, Gemmatimonas, and Flavisolibacter in black soil than in farm soil. The relative abundance of Spingomonas and Lysobacter in Module B is greater than in Module A; these genera may help cucumber development at an early stage in black soil and farm soil.At the genus level, the top 50 genera of all samples were selected for further investigation of cucumber rhizosphere bacterial composition ; we geneSince we found that HD grown in sandy soil and sandy loam soil was significantly better than that in black soil, farm soil, and Bray–Curtis distance suggested that sandy soil and sandy loam soil had similar bacterial communities , we, theTo further explore which bacteria may promote the growth of HD in sandy soil and sandy loam soil, we chose the top 20 genera (Top 20)  and the Nocardioides, Bacillus, Rubellimicrobium, Gaiella, Tumebacillus, and Nitrospira in sandy soil was higher than that in black soil, and farm soil  in the rhizosphere of HD was more than four times higher than that in black soil (0.22 ± 0.12%) and farm soil (0.33 ± 0.15%)  ; the rel ± 0.15% .Nocardioides ASV1755 and Bacillus ASV341  showed the same tendency. At the same time, HD recruits more of these ASVs than XT in sandy soil (Tumebacillus ASV2711 (0.77%), Paenibacillus ASV1874 (1.05%), Pseudomonas ASV2144 (0.76%), and Luteolibacter ASV1499 (0.62%) than in black soil and farm soil; the relative abundances of these rhizosphere bacterial ASVs of HD were higher than that of XT  and late (50 DPS) stages of cucumber cultivation, the structures of the rhizosphere bacterial community in sandy soil and sandy loam soil were similar, but there was no such trend in black soil and farm soil. Therefore, we speculate that cucumbers had formed a stable rhizosphere bacterial community in sandy soil and sandy loam soil at an early stage, while in black soil and farm soil, cucumbers needed longer to form such a stable bacterial community. Therefore, this phenomenon might be caused by the joint effects of the soil type and plant cultivation time. A similar phenomenon was also found in wheat; the rhizosphere microbial community was relatively stable during the seedling, tillering, and flowering stages; however, significant microbial structure changes were observed during the mature stage .Nitrospira, Nocardioides, Gaiella, Bacillus, Tumebacillus, and Paenibacillus for sandy soil and sandy loam soil  can be found below: YQ, XY, and SH designed the study. ZL, YZ, and YQ performed the experiments and analyzed the 16S rRNA sequencing data. YL provided technical support to this study. ZL, YZ, YQ, and XY wrote the manuscript. SH provided critical comments and edited the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.