Mammalia: Chiroptera) serve as natural reservoirs for many zoonotic pathogens worldwide, including vector-borne pathogens. However, bat-associated parasitic arthropods and their microbiota are thus far not thoroughly described in many regions across the globe, nor is their role in the spillover of pathogens to other vertebrate species well understood. Basic epidemiological research is needed to disentangle the complex ecological interactions among bats, their specific ectoparasites and microorganisms they harbor. Some countries, such as Ukraine, are particularly data-deficient in this respect as the ectoparasitic fauna is poorly documented there and has never been screened for the presence of medically important microorganisms. Therefore, the aims of this study were to provide first data on this topic.Bats . Sequencing and phylogenetic analysis revealed high similarity to sequences from Rickettsia parkeri and several other Rickettsia species. Bacteria from the family Anaplasmataceae were detected in all groups of the ectoparasites , belonging to the genera Anaplasma, Ehrlichia, and Wolbachia. The detection of Bartonella spp. was successful only in fleas (Nycteridopsylla eusarca) and bat flies , representing 12.1% (29/239) of the collected ectoparasites. No DNA of Babesia or Borrelia species was identified in the samples.We report for the first time in Ukraine the molecular detection of several bacterial agents in bat ectoparasites collected from six species of bats. The data presented extend the knowledge on the distribution of ectoparasite species in bats and their involvement in potentially circulating agents pathogenic for humans and vertebrate animals.The online version contains supplementary material available at 10.1186/s13071-022-05582-3. Mammalia: Chiroptera) represent the second-most diverse order of mammals after rodents [Bats , fleas (genus Ischnopsyllus: 2 spp.), and a bat fly (Nycteribia koleantii) occurring in five bat taxa. Moreover, they examined only a small number of opportunistically collected samples , and the ectoparasites were not screened for microorganisms [Many areas, especially in Eastern Europe, are thus far lacking this type of research. For example, very little is known about bat ectoparasites in Ukraine, and the vector-borne bacteria they might carry have never been surveyed in the country –15. The The field survey was conducted in five localities divided among the three habitat types: urban November 2019), natural (August–September 2019), and rural (July 2020), representing main types of land use in the region  and the King Fisher® Flex Purification system , according to the manufacturer’s instructions. Total DNA was eluted in 100 μl elution buffer and then stored at −80 °C until further analysis.The samples were processed individually, each ectoparasite being homogenized in sterile phosphate-buffered saline (PBS) with steel beads using the Tissue Lyser II . The DNA extraction was performed from 100 μl aliquots using NucleoMagBabesia spp., Rickettsia spp., Bartonella spp., and Anaplasma/Ehrlichia spp., while only Carios vespertilionis ticks were additionally screened for DNA of Borrelia spp. The screening was done using specific primers listed in Additional file ® kit  following manufacturer’s instructions and Sanger sequenced in the Laboratory for Applied Bioinformatics and Sequencing of Viral Genomes and Transcriptomes, Institute of Diagnostic Virology, Friedrich-Loeffler-Institut.Four species of ectoparasites collected from different bat species Table  were PCRRickettsia spp. was identified in 2.9% (7/239) of ectoparasites , R. africae (MH938655), R. sibirica (KU310587), or uncultured Rickettsia sp. (MG228263). PCR based on the ompA gene followed by sequencing of positive C. vespertilionis ticks for Rickettsia indicated 100% similarity of all samples with R. parkeri (MK962698) and Rickettsia sp. (KX137902).Among the tested samples, DNA of Rickettsia ompB sequences also showed 100% similarity of the samples from this study with R. parkeri (CP040325), uncultured Rickettsia sp. (MK405417), and Rickettsia sp. (AF123720).The BLASTn analysis for the gltA, ompA, and ompB concatenated Rickettsia sequences and sequences from representative Rickettsia species available in GenBank. The analysis demonstrates that all Rickettsia sequences detected in C. vespertilionis in this study are phylogenetically closely related to R. parkeri and R. africae  was doncae Fig. a.Fig. 1PR. parkeri-like sequences in bat-associated soft ticks from Europe represents a noteworthy finding as this alphaproteobacterium is known to be associated primarily with hard ticks in the genus Amblyomma occurring in the Americas [R. parkeri sequences in tissues of Pipistrellus pipistrellus bats from China [R. parkeri sensu lato complex are well-established human pathogens, little is known about their natural reservoir hosts [The detection of Americas . Howeverom China , which wir hosts .R. parkeri-like species presently detected in Eurasia. Further research efforts, therefore, should focus on isolating and establishing the identity of the bacterium as well as elucidating its enzootic cycle. This would also be of public health relevance, keeping in mind that C. vespertilionis may occasionally bite humans in situations when their bat hosts are no longer available [Moreover, neither pathogenicity in vertebrates nor transmissibility by argasid ticks is known for the vailable .Anaplasma/Ehrlichia spp., 51% (122/239) of the samples tested positive: 4.7% (2/43) of C. vespertilionis, 56% (56/100) of Nycteridopsylla eusarca, 69.2% (54/78) of Nycteribia kolenatii, and 55.6% (10/18) of N. pedicularia  while the other sample had 99.7% similarity to uncultured Anaplasma sp. clone Erz1600 (MT601947). The phylogenetic analysis based on the 16S rRNA partial sequence indicates that the Ehrlichia sequence detected in C. vespertilionis clusters in a clade that includes Candidatus Ehrlichia shimanensis, uncultured Ehrlichia: E. minasensis or E. canis , and six sequences were 99.6–100% similar to Wolbachia endosymbiont (MH618381). Four N. pedicularia and six N. kolenatii flies positive for Anaplasma/Ehlichia spp. were also sequenced, having 99.3–100% similarity to Wolbachia endosymbiont (MH618380).Regarding DNA of p., 51% 12/239 of Anaplasmataceae such as Anaplasma phagocytophilum have been previously reported from insectivorous bats and their ticks in Europe [Ehrlichia and Anaplasma from the present study are clustering with poorly characterized species (e.g. E. minasensis) or with microorganisms not commonly associated with bats, such as E. canis or A. ovis. This implies that either a much broader range of vertebrate reservoirs and arthropod vectors support circulation of these pathogens in nature or, more parsimoniously, that the molecular markers selected for species identification have poor discriminatory capacity at this level. On the other hand, the finding of such endosymbionts as Wolbachia sp. is not surprising as these bacteria are almost universally present in many groups of arthropods and in filarial nematodes and have been reported previously in multiple bat fly species [While zoonotic n Europe , it rema species .Bartonella spp. was successful in 12.1% (29/239) ectoparasites collected from bats. All bat ticks tested negative, while N. eusarca, N. kolenatii, and N. pedicularia showed prevalence rates that varied from 7 to 21.8% and 27.8%, respectively  detected in C. vespertilionis from The Netherlands (n = 3 isolates) and uncultured Bartonella sp. isolate M451 (AJ871615) found in the blood of Nyctalus noctula from UK (n = 4 isolates). Eight N. kolenatii- and two N. pedicularia-positive samples for Bartonella 16S-23S rRNA were also sequenced. Sequence analysis indicated that three sequences from N. kolenatii had 94.5% similarity to Bartonella sp. strain 44601 (MF288119) obtained from Myotis blythii, and two sequences matched 98.7% and 99%, respectively, to Bartonella sp. strain 44718 (MF288128) from Pipistrellus pygmaeus. The other five sequences (three from N. kolenatii and two from N. pedicularia) had 96.7–97.4% similarity to uncultured Bartonella sp. clone 137 (KX420735) found in Rhinolophus ferrumequinum bat species.The detection of Bartonella gltA and 16S-23S rRNA sequences were further used to construct phylogenetic trees  and Bartonella washoensis (AF050108). The additional four obtained sequences cluster in a separate clade. The analysis based on the partial 16S-23S rRNA sequence shows that Bartonella sequences detected in N. kolenatii and N. pedicularia cluster close to uncultured Bartonella (KX420735) and Bartonella sp. (MF288119 and MF288128). All sequence data and accession nos. are shown in Additional file Bartonella sequences have been detected in many bat species and in their ectoparasites  across the world [Bartonella spp. are believed to be highly host/vector specific, the remarkable diversity of sequences belonging to this genus found in bats and their ectoparasites suggests the ancient nature and evolutionary importance of this association [Various he world , 26. Whihe world , others he world , 29. It he world . While cociation .Babesia spp., while only C. vespertilionis were screened for Borrelia spp. and were positive in 4.7% (2/43) using the 16S-23S IGS specific primers [All ectoparasite samples were negative for  primers . The folThis study offers first glimpses on the microbial diversity found in ectoparasites collected from several species of insectivorous bats in Northeast Ukraine. Our research effort creates the impetus for disentangling the vector-host-pathogen interactions among bats and their ectoparasites in an understudied part of Europe. Further studies employing larger sample sizes, greater diversity of the host and parasite species, and variable methods, including next generation sequencing, should reveal a complete and more complex picture. Given the globally changing patterns of bat distribution, their increasing proximity to humans, and the high rates of the infectious disease emergence in wildlife, domestic animals, and human populations, this basic research is important from a public health perspective as well as for conservation biology.Additional file 1: Table S1. Locations and roosting sites of bats collected for the purpose of this study. Table S2. Primers used for pathogen screening of ectoparasites. Table S3. GenBank accession numbers for sequences obtained in this study.


Mucuna pruriens has been identified to have fertility-enhancing and anti-lipid peroxidation properties. Based on these properties of M. pruriens, this study aimed to investigate the efficacy of M. pruriens seed extract in reducing sodium arsenite-induced testicular impairment and hepato-renal histopathology in rats.A significant cause of arsenic poisoning is polluted groundwater. Arsenic poisoning results in the suppression of spermatogenesis and the liver and kidneys are vulnerable to the toxic effects as well. M. pruriens  orally once daily. Subgroups 8 and 9 received a combination of NAC and aqueous seed extract  orally once daily. Following the treatment, animals were sacrificed and sperm parameters and DNA damage were evaluated. Testis, liver, and kidneys were analyzed for histopathology.The study was divided into two groups; short-term (45 days) and long-term (90 days) treatment groups and each group was divided into nine subgroups. Subgroups 1 and 2 served as normal and N-acetyl cysteine (NAC) controls, respectively. Subgroups 3–9 received sodium arsenite in the drinking water (50 mg/L). Subgroup-4 received NAC (210 mg/kg body weight [BW]) orally once daily. Subgroups 5–7 received aqueous seed extract of M. pruriens seed extract ameliorated the damaging effects of sodium arsenite with respect to tissue architecture and sperm parameters when coadministered.Sodium arsenite-induced a significant reduction in sperm parameters and increase in the abnormal architecture of spermatozoa. Histology revealed tissue necrosis. The Mucuna pruriens has beneficial effects against the deleterious effects of sodium arsenite on various tissues. Thus, M. pruriens (530 and 700 mg/kg BW) supplementation would reduce the adverse changes observed with sodium arsenite exposure. Arsenic is an all-pervading metalloid found in the earth’s crust and exists in elemental, organic, and inorganic forms. Contaminated groundwater is the major cause of arsenic poisoning . ArsenicMucuna pruriens is a leguminous plant that possesses antidiabetic and aphrodisiac effects  and 530 mg/kg BW [p < 0.01]) exhibited an increase in sperm count when compared to the sodium arsenite-only treatment group (Tables-There was a significant reduction (p < 0.001) in the sperm count after treatment with sodium arsenite compared to the normal control group. Co-administration of p Tables- and 3.M. pruriens  along with sodium arsenite significantly increased (p < 0.01) the number of motile sperm in comparison to the sodium arsenite-only treated group. Sperm motility significantly increased in the animals treated with a combination of sodium arsenite + NAC along with M. pruriens (350 and 530 mg/kg BW [both p < 0.01]) in comparison to the sodium arsenite-only treated group (Tables-The number of motile sperm significantly decreased (p < 0.001) in the sodium arsenite-only treated group compared to the normal control. Co-administration of p Tables- and 3.M. pruriens  along with sodium arsenite significantly decreased the number of abnormal sperms in comparison to the sodium arsenite only treatment group. Animals treated with sodium arsenite + NAC + M. pruriens (350 [p < 0.05] and 530 [p < 0.001] mg/kg BW) showed a significant decrease in the number of abnormal sperms when compared to sodium arsenite + NAC treatment group (Tables-Exposure to sodium arsenite significantly increased (p < 0.001) the number of abnormal sperm in comparison to the normal control. Co-administration of p Tables- and 3.M. pruriens  and co-administration of sodium arsenite + NAC+ M. pruriens (350 and 530 mg/kg BW [p < 0.001]) exhibited a significant decrease in the seminiferous tubular diameter when compared to sodium arsenite only treated group  in the seminiferous tubular diameter, indicating a distortion of the tubule when compared to the normal controls. In the long-term treatment group, co-administration of sodium arsenite + p Tables- and 3.M. pruriens 700 mg/kg BW had a significantly increased (p < 0.001) seminiferous epithelial height when compared to the sodium arsenite-only treatment group. Animals treated with sodium arsenite + NAC + M. pruriens (350 mg/kg BW) showed a significant increase (p < 0.001) in seminiferous epithelial height when compared to the sodium arsenite-only treated group. In the long-term treatment group, co-administration of sodium arsenite + M. pruriens  and co-administration of sodium arsenite + NAC + M. pruriens (350 and 530 mg/kg BW [p < 0.001]) exhibited a significant increase in the seminiferous epithelial height when compared to sodium arsenite only treated group (Tables-Sodium arsenite administration caused a significant reduction (p < 0.001) in the seminiferous epithelial height when compared to the normal control group. In the short-term treatment group, animals treated with sodium arsenite + p Tables- and 3.M. pruriens (530 and 700 mg/kg BW [p < 0.001]) along with sodium arsenite showed a reduction in the spermatozoa containing damaged DNA in comparison to the sodium arsenite-only group. The spermatozoa from the groups treated with a combination of NAC and M. pruriens (350 and 530 mg/kg BW [p < 0.001]), along with sodium arsenite, also exhibited a significant reduction in the percentage of DNA damage  in the percentage of spermatozoa carrying DNA damage when compared to normal control was observed. Animals treated with a combination of A damage .M. pruriens (350 and 530 mg/kg BW) along with sodium arsenite minimized the toxic effect of arsenic on the testis. The combination of NAC + M. pruriens (350 and 530 mg/kg BW) along with sodium arsenite showed improved structural recovery with minimal damage and intact cellular structures with an improved cell count  showed a significant reduction in seminiferous tubule diameter, seminiferous tubule epithelial height, and increased tubular lumen. Sloughed germ cells with no maturation stages were observed and the tubular basement membrane was irregular or interrupted. The cell lining of Sertoli cells was damaged and in general, there were fewer spermatozoa observed in the central core of the tubule. Co-administration of  Figures- and -2.M. pruriens (530 and 700 mg/kg BW) showed improved cellular structure. Clear sinusoidal regions were evident. Inflammatory changes were reduced. The combination of M. pruriens and + NAC showed reduced inflammatory and degenerative changes due to sodium arsenite  with sodium arsenite reduced the toxic effects of arsenic. There was damage to the cellular lining of the duct system. Higher doses of M. pruriens (530 and 700 mg/kg BW) showed minimal damage and an improved histological structure in comparison to the lower dose. The kidney tubules exhibited acidophilic cytoplasm. The combination of M. pruriens (350 and 530 mg/kg BW) + NAC showed results that were similar to sodium arsenite + M. pruriens (700 mg/kg BW)  with sodium arsenite was found to attenuate the adverse changes observed with arsenic exposure. The study suggests that a diet supplemented with M. pruriens can ameliorate the undesirable changes in rats exposed to sodium arsenite.Excessive arsenic ingestion leads to significant damage to various body tissues , making the individual prone to further complications. Sodium arsenite-exposed animals showed impaired hepatic and renal function and reproductive toxicity. The supplement of PLC, KLB, and APR: Contributed to the conception and design of the study. PLC: Performed the experiment and wrote the manuscript. PLC, KLB, and APR: Statistical analysis. KLB and APR: Reviewed the manuscript. All authors have read and approved the final manuscript.


Immunotherapy is an effective treatment option that revolutionizes the management of various cancers. Nevertheless, only a subset of patients receiving immunotherapy exhibit durable responses. Recently, numerous studies have shown that oxidative stress induced by reactive oxygen species (ROS) plays essential regulatory roles in the tumor immune response, thus regulating immunotherapeutic effects. Specifically, studies have revealed key roles of ROS in promoting the release of tumor-associated antigens, manipulating antigen presentation and recognition, regulating immune cell phenotypic differentiation, increasing immune cell tumor infiltration, preventing immune escape and diminishing immune suppression. In the present study, we briefly summarize the main classes of cancer immunotherapeutic strategies and discuss the interplay between oxidative stress and anticancer immunity, with an emphasis on the molecular mechanisms underlying the oxidative stress-regulated treatment response to cancer immunotherapy. Moreover, we highlight the therapeutic opportunities of manipulating oxidative stress to improve the antitumor immune response, which may improve the clinical outcome. Generally, the traditional treatments of cancer involve surgery, chemotherapy, radiotherapy, or molecular targeted therapy. However, even the best regimens within these strategies rarely cure late-stage tumors . At the To enhance the antitumor effect, cancer immunotherapies have been developed based on the above theories. However, only a small number of patients benefit from these immunotherapeutic strategies, largely due to the immunosuppressive mechanisms developed by tumor cells. For example, once antigens on the surface of tumor cells are recognized, the tumor cells start to overexpress suppressive cytokines, such as IL6, IL10 or TGF-β, which can block the immune system . TherefoIn this review, we present a summary of several types of cancer immunotherapy strategies. We further discuss the molecular mechanistic basis of the interplay between oxidative stress and anticancer immunity, with an emphasis on oxidative stress-regulated treatment response to cancer immunotherapy. On this basis, we highlight the therapeutic opportunities of manipulating oxidative stress to improve the antitumor immune response, which may be helpful to the development of more efficient clinical management strategies based on immunotherapy.Cancer immunotherapies have been developed based on studies of tumor escape mechanisms, which could be categorized as either active or passive immunotherapies ,9. ActivImmune checkpoint inhibitors (ICIs). Immune checkpoint inhibitors (ICIs) are one of the most broadly successful cancer immunotherapies to date [ to date . Physiol to date . The mos to date . The rec to date ,15. To a to date . Thus, d to date . Another to date . Ipilimu to date .Cancer vaccines (CVs). Cancer vaccines are subclassified as either prophylactic or therapeutic interventions, which boost the immune system to clear cancer cells. Prophylactic vaccines aim to reduce cancer incidence, morbidity and mortality. Currently, several vaccines available in clinical practice can efficiently prevent tumor progression. For instance, prophylactic cancer vaccines have been successfully used for hepatocellular carcinoma, secondary to hepatitis B virus (HBV), and squamous cell carcinoma, secondary to human papillomavirus (HPV) [us (HPV) . It is wus (HPV) . Some reus (HPV) . Currentus (HPV) . In 2010us (HPV) . In addius (HPV) ,23.Cytokine treatment. Cytokines are small proteins secreted by cells to regulate the innate and adaptive immune systems. Cytokine treatment is the first active immune therapy to be applied in the clinic, in which cytokines are directly injected to boost the immune system [+ and CD8+ T cells. Currently, IL-12 is used for the treatment of various types of cancers [e system . IL-2 is cancers ,26. Addi cancers .Monoclonal antibodies. The monoclonal antibody was the first validated treatment, aside from classical therapies. Naked antibodies are one type of therapeutic strategy that can activate apoptosis and can be used to directly trigger ADCC via NK cells. Another type is the conjugated antibody, which carries an active factor, such as a radioisotope, a toxin or a specific drug. In this case, the antibody is used as a targeting tool to deliver the effector molecule directly to the tumor [he tumor  . OVs are viruses that exist in nature or are genetically engineered to selectively replicate in tumor cells to activate immune responses and induce tumor cell lysis. Currently, many OVs are in clinical trials, most of which belong to DNA viruses, such as Herpeviridae and Adenoviridae [oviridae , and a soviridae . OVs exeoviridae . Reoviruoviridae . Researcoviridae . Moreoveoviridae ,35. A stoviridae . Talimogoviridae  . Engineered cells for ACT include T-cell receptor T cells (TCR-Ts), chimeric antigen receptor T cells (CAR-Ts) and CAR-NK cells. These cells are engineered to express fusion proteins on the cell surface, whose extracellular fragment is a monoclonal antibody-like receptor that exhibits high specificity to recognize the full antigen on a tumor cell [mor cell . The earmor cell . CAR-T cmor cell . Thus, Cmor cell . In anotmor cell . This ki2•−), and nonradical species, such as hydrogen peroxide (H2O2) [ROS are toxic intermediates or byproducts of aerobic metabolism produced by almost all organisms and cells. This type of oxygen-derived molecule includes radicals, as exemplified by hydroxyl radicals (•OH) and superoxide anions (Oe (H2O2) . Endogene (H2O2) . The twoe (H2O2) ,44,45 ane (H2O2) . In addie (H2O2) , cancer 2O2 into H2O and O2 [2•− to H2O2, which is decomposed into H2O and O2 through CAT as well as glutathione peroxidases (GPXs) [Typically, ROS act as signaling molecules in multiple cellular physiological processes at moderate levels. However, excessive ROS induce oxidative damage to biomolecules, including protein oxidation, lipid peroxidation and DNA damage . To counO and O2 . Specifis (GPXs) . Moreoves (GPXs) . Under ns (GPXs) . In addis (GPXs) ,55. Furts (GPXs) .The immune system contains innate and adaptive immunity, both playing essential roles in the maintenance of organism integrity by protecting against invading pathogens and for the elimination of transformed cells . Innate Currently, many immunotherapies  targeting or harnessing T cells have demonstrated therapeutic efficacy for a broad range of human malignancies. T cells, especially CTLs, are lymphocytes possessing specific cell-killing activity ,62. StudT cell activation. Moderate levels of ROS participate in the activation, proliferation and differentiation of T cells, whereas a high level of ROS is the main factor for T-cell inhibition. Interestingly, it was previously reported that, after a reduction in mitochondrial ROS, T cells could not be consistently activated even under CD28 or CD3 stimulation [2O2 in mitochondria can amplify the MAPK signaling pathway when stimulated by antigens, leading to T-cell activation and proliferation [+ T cells [2+ release [2+ levels results in NFAT activation that induces IL-2 gene expression and subsequent signaling events, resulting in T-cell proliferation [mulation . In addiferation . Moreove T cells . The ROS T cells ,89. Spec release . The incferation . Howeverferation ,92. Thusferation . Anotherferation ,94. TherT cell differentiation. It is well known that ROS levels increase after TCR signaling-induced T-cell activation [2O2-mediated TCR activation promotes mitochondrial ROS production, which enhances FasL expression in T cells and contributes to T-cell activation-induced cell death (AICD) [tivation . As expetivation . In conttivation   .Dendritic cells. DCs are professional antigen-presenting cells essential in the antigen-specific immune response. Immature DCs have a strong migratory ability, while mature DCs can capture various antigens and present them to T cells to initiate an antigen-specific immune response [response . ROS canresponse . Howeverresponse . Studiesresponse . These aresponse ,103. ROSresponse . Low levresponse . In addiresponse .B cells. B cells are derived from bone marrow pluripotent stem cells and are specialized antigen-presenting cells. There are various membrane surface molecules on the surface of B cells that mediate humoral immune responses by producing antibodies, such as leukocyte differentiation antigen, MHC and various membrane surface receptors [eceptors . In geneeceptors . The eleeceptors . ROS haveceptors . In addieceptors . For exaeceptors .Some immune cells can maintain peripheral tolerance and suppress antigen-specific immune responses in anticancer immunity. Tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs) and Tregs can produce excessive ROS, leading to a resistance to tumor immunotherapy through the apoptosis-related factor ligand pathway.Tumor-associated macrophages. TAMs are the most abundant immune-related cells in the TME, which is closely related to tumor development and metastasis [tastasis . TAMs cotastasis . Throughtastasis . Additiotastasis . In the tastasis . TAMs istastasis . The remtastasis . Coincidtastasis . Moreovetastasis . TherefoRegulatory T (Treg) cells. Tregs are a subtype of CD4+ T lymphocytes expressing the transcription factor forkhead box protein P3 (FOXP3). Emerging evidence has shown that the TME recruits Treg cells to exert its powerful immunosuppressive effects [+ effector T cells was blocked or reduced when ROS was eliminated by antioxidants or NADPH oxidase inhibitors [+ T cells into Treg cells. Then, the expression of FOXP3 was downregulated and ultimately lead to reduced numbers of tumor-infiltrating Treg cells [+ T cells, Tregs are less susceptible to cell death induced by oxidative stress. This finding may be ascribed to higher expression levels of antioxidative enzymes in Tregs [ effects . Generalhibitors . Additiohibitors . Metformeg cells . Converseg cells . Treatmein Tregs . In concMyeloid-derived suppressor cells. MDSCs are heterogeneous cell groups that consist of myeloid progenitor cells and immature bone marrow cells (IMCs), which are precursors of dendritic cells (DCs), macrophages and/or granulocytes with the ability to significantly suppress immune cell responses [2O2 levels in MDCS can promote their differentiation into antigen-presenting cells [2O2 scavenging with catalase induces myeloid differentiation into macrophages [esponses . Under png cells . Signifirophages . Similarrophages . TherefoROS can function as intercellular messengers for MHCII-restricted antigen presentation. T-cell activation may also be mediated by ROS derived from surrounding cells. For instance, DCs have been shown to generate ROS during antigen presentation , while tCollectively, these findings suggest that ROS could not only directly affect immune cells, but also regulate the interactions between different types of immune cells, thereby affecting the tumor immune microenvironment. In line with this, a large number of studies have shown that regulating ROS levels in a tumor-immune microenvironment can improve immunotherapeutic efficacy ,136. TheDespite the promising potential of targeting the immune system for cancer therapy, the clinical application of immunotherapy faces many challenges in terms of efficacy and safety. For instance, CAR-T against CD19 has been proven to be effective in the treatment of acute lymphoblastic leukemia, but some melanoma and non-small-cell lung cancer patients still do not benefit from these treatments ,138. Fur+ effector T cells, thus contributing to the selection and activation of antigen-specific T cells [ICD is a modality of cell death when a variety of chemical drugs and radiation are used to treat tumors. Tumor cells transform from nonimmunogenic cells to immunogenic cells to mediate the immune response ,145. The T cells . Some pr T cells . Accordi T cells . Studies T cells ,151,152. T cells . A previ T cells . CollectRecently, it has been demonstrated that both chemotherapeutics and photodynamic therapy (PDT) trigger ROS generation, which induces ER stress and promotes DAMP release and ICD cascades ,155,156.+ T cells [+ T-cell infiltration in the primary and distal tumors, and eventually the regression of both tumors could be observed [Photodynamic therapy (PDT) is a noninvasive treatment that takes advantage of ROS created by photosensitizers (PSs) to repress tumors. PSs are a class of compounds that can absorb ultraviolet rays of a certain wavelength to generate free radicals or ions and initiate photopolymerization. The ideal photosensitizer should at least satisfy the following conditions: a high chemical purity, minimal cytotoxicity and high selectivity of tumor tissues . After t T cells . It has  T cells . Moreove T cells . In addiobserved . Howeverobserved ,163. Oneobserved .+, Cu2+ and Mn4+) can generate large amounts of ROS through Fenton and Fenton-like reactions [2O2 via the Fenton reaction. The increased levels of ROS induce the secretion of proinflammatory cytokines at both the tumor site and in the systemic circulation, thereby improving the immune response. Another study has shown that magnetite IONPs polarize M2 macrophages to M1 macrophages by downregulating the expression of M2-associated arginase 1 (Arg-1) [Metal nanoparticles (MNPs) with a smaller size and higher tensile strength can penetrate deep tumors together with target materials . Many meeactions ,168,169. (Arg-1) . Similar (Arg-1) . Another (Arg-1) . In addi (Arg-1) .Cancer cells frequently face metabolic stress in the tumor microenvironment , which m2O2 [As signaling molecules or cellular toxicants, ROS are continuously generated, transformed and consumed in a variety of biochemical reactions. The traditional view links ROS to inflammation, aging and cancer. Emerging data show that ROS play important roles in immune responses. Practically, most immune cells are affected by the concentrations of ROS and thus cannot normally exert their antitumor effect in the tumor microenvironment, resulting in the resistance to some immunotherapy. Nanotechnology facilitates the development of multimodal immunotherapy strategies for cancer treatment. For example, immunotherapy based on the ROS-producing nanomaterial PDT simultaneously enhances immunity against primary and metastatic tumors . In the 2O2 ,181. Nev


Brassica species. Here, we determined the extent of genetic variation in pod orientation, described as the angles of pedicel on raceme (APR) and angles of the pod on pedicel (APP) among 136 rapeseed accessions grown across three environments of the upper, middle and lower Yangtze River in China. The APR ranged from 59° to 109°, while the APP varied from 142° to 178°. Statistical analysis showed that phenotypic variation was due to genotypic (G) and environmental (E) effects. Using the genome-wide association analysis (GWAS) approach, two QTLs for APR (qBnAPR.A02 and qBnAPR.C02) and two for APP (qBnAPP.A05 and qBnAPP.C05), having minor to moderate allelic effects (4.30% to 19.47%) were identified. RNA-seq analysis revealed 606 differentially expressed genes (DEGs) in two rapeseed accessions representing the extreme phenotypes for pod orientation and different alleles at the QTLs of APR. Three DEGs  were identified as the most likely candidates responsible for variation in pod orientation (APR). This study elucidates the genomic regions and putative candidate genes underlying pod orientation in B. napus.Spatial distribution and orientation of pods on the main raceme (stem) and branches could affect rapeseed yield. However, genomic regions underlying the pod orientation were not described in  Besides, it is used for biofuel and stock-feed production . Due to the unprecedented growth of the world’s population, increased rapeseed yield is required to meet energy demands. The rapeseed generally bears pods (siliques) on the top of the raceme (primary stem) and side branches , map-based cloning and mutant studies. For example, in rice, eritance . In addiribution . Genes sZY1 gene , and theof auxin . In Arab to rice . In maizng angle . The genng angle . Howeverng angle , and fewng angle . In addicucumber . The delcucumber . The angcucumber . Generalor above . In contor above . The podor above . The latove , Yangluo City, Hubei Province  and Lu’an City, Anhui Province  (2.2https://www.autodesk.com.cn/).Two quantitative indices related to rapeseed pod orientation were measured: the angle of the pedicel on raceme (APR) and the angle of the pod on pedicel (APP) across 136 rapeseed accessions grown in three environments. We digitally measured the pod-related angles of a GWAS panel in three independent environments. We. At the maturity , five plants of each rapeseed accession were selected from each environment investigation and measured for APR and APP using the image processing method described by 2.3@ SNP array (Illumina) as described previously  was constructed using genotype, environment, and phenotype data. The ANOVA was carried out to determine whether the genotypic effect, environmental effect and genotype×environment interaction effect exist. The multi-environment mixed linear model (MLM) in Tassel5.0 software  was used package  in R3.3. package .2.5https://ftp.ncbi.nlm.nih.gov/blast/db/FASTA/) and Swiss-port databases (https://www.expasy.org/resources/uniprotkb-swiss-prot) using blast software (https://www.arabidopsis.org/index.jsp).All genes in the candidate region were annotated in the NR  following the manufacturer’s recommendations. Libraries were sequenced on Illumina HiSeq 4000 platform, resulting in 150 bp paired-end reads. The quality control of RNA-seq reads was carried out by fastqc  and line ZS11  were collected for RNA sequencing Figure 1y fastqc  softwarey fastqc . For RNAy fastqc . StringTy fastqc  was usedy fastqc  were simy fastqc . Conside2.8https://www.ncbi.nlm.nih.gov/) and established a local protein database. Seven genes of IGT families in Arabidopsis thaliana were compared with the local protein database by BLAST (http://pfam.xfam.org/), so we used Hmmer (https://www.ncbi.nlm.nih.gov/).We downloaded candidate protein sequences involved in the pod orientation of different species from the NCBI website . The sequences were compared to the reference genome of ZS11  was calculated, and it ranged from 49.65% (APR) to 76.65% (APP). The frequency distribution of the APP and APR showed continuous variation , which indicated that different genetic factors might control variation for both measures of pod orientation.Pearson correlation analysis between angle-related traits (APR and APP) across different environments showed significant positive correlations  plots showed significant genetic associations for APP and APR across environments Figure 4We observed that phenotypes corresponding to different genotypes did not show normal distribution. Therefore, the Mann-Whitney U test Figure 53.3We selected two accessions that had extreme phenotypes: line 9319  and line ZS11   encoding SAUR-like auxin-responsive protein in Arabidopsis. BnSAUR32.C02 and AT5G53590 had poor homology and are SAUR32-like genes.We focused on three interesting DEGs which underlie QTL for pod orientation: the first two are homologous genes related to auxin synthesis. BnaA02G0138300ZS and BnaC02G0175200ZS, designated as thaliana . BnaA02GPR trait Table 3.3.4BnLAZY4.A02 gene belongs to the IGT gene family, with 167 IGT family genes in 13 species. To investigate the relationship of BnLAZY4.A02 that was expressed in the pedicel of line 9319 but not in the pedicel of line ZS11, with other species, we carried out cluster analysis using neighbour-joining approach  , in the upstream region of BnSAUR32.A02, which could affect the gene expression.To gain insights into nucleotide diversity in candidate genes involved in pod orientation, we utilised public genomic sequence data of 289 rapeseed core collections . Results4via GWAS expands the research in two ways. First, this study mined the natural variation in pod orientation traits in a diverse GWAS panel of accessions rather than two parents, used in most biparental QTL mapping studies. Secondly, we identified four QTL and candidate genes associated with pod orientation.Crop architecture is an important physiological and agronomic trait that could drive rapeseed yield. The branch angle determines the overall plant shape, while the pod angle contributes to the photosynthesis rate. The ideal plant architecture can make the best use of resources such as light, water and nutrients, achieving the maximum yield and possibly the best quality. In recent years, crop architecture has become a vital breeding aim of genetic improvement programs. Investigating pod orientation 4.1Earlier studies investigated genetic variation for pod orientation using qualitative scores: straight, inclined, flat and droop . However4.2There were statistically significant G×E interactions for both APP. APR across environments. Therefore, we employed the multi-environment-based MLM model, thus excluding the impact of the environment on the genotype-phenotype association, as opposed to the approach followed by . We idenTo identify favourable alleles for pod architecture, we sought an association between SNP marker alleles Figure 54.3Arabidopsis thaliana priori candidate genes whose role in branch and inflorescence angle has been confirmed.We followed two approaches to identify the candidate gene underlying QTL for pod orientation. Firstly, we examined genomic regions in LD (upstream and downstream 424 Kb for the A subgenome and 2.5Mb for the C subgenome). Secondly, we identified DEG underlying QTL regions. We also searched BnLAZY4.A02, BnSAUR32.A02 and BnSAUR32.C02, underlying pod orientation QTL  homologues: BnSAUR32.A02 and BnSAUR32.C02. SAURs are implicated in regulating dynamic and adaptive growth in Arabidopsis and sunflower  can be found below: JL designed and supervised the study. WW and JL collected the samples. QH provided the germplasm. YY performed genetic, and transcriptome data analyses. YY, HR and JL discussed results and prepared the manuscript. All authors contributed to the article and approved the submitted version.


Increasing evidence has shown that astrocytes are implicated in regulating oligodendrocyte myelination, but the underlying mechanisms remain largely unknown. To understand whether microRNAs in astrocytes function in regulating oligodendroglial differentiation and myelination in the developing and adult CNS, we generated inducible astrocyte-specific Dicer conditional knockout mice . By using a reporter mouse line (mT/mG), we confirmed that hGFAP-CreERT drives an efficient and astrocyte-specific recombination in the developing CNS, upon tamoxifen treatment from postnatal day 3 (P3) to P7. The Dicer deletion in astrocytes resulted in inhibited oligodendroglial differentiation and myelination in the developing CNS of Dicer cKO mice at P10 and P14, and did not alter the densities of neurons or axons, indicating that Dicer in astrocytes is required for oligodendrocyte myelination. Consequently, the Dicer deletion in astrocytes at P3 resulted in impaired spatial memory and motor coordination at the age of 9 weeks. To understand whether Dicer in astrocytes is also required for remyelination, we induced Dicer deletion in 3-month-old mice and then injected lysolecithin into the corpus callosum to induce demyelination. The Dicer deletion in astrocytes blocked remyelination in the corpus callosum 14 days after induced demyelination. Together, our results indicate that Dicer in astrocytes is required for oligodendroglia myelination in both the developing and adult CNS. In vitro evidence has shown that astrocytes can influence the differentiation, proliferation and migration of OL precursor cells (OPCs) ). All values are expressed as the mean ± SEM. Data from the acquisition phase in the Morris water maze were analyzed by separate repeated measures two-way ANOVA. For other data, statistical analysis was performed using the two-tailed unpaired t-test. Each experiment included at least three samples. P-values less than 0.05 were considered statistically significant. Significance was reported as *P < 0.05, **P < 0.01, or ***P < 0.001.The statistical significance of differences between groups was determined using GraphPad Prism 7 software (La Jolla, CA [To understand the role of astrocytes during CNS development, we used the hGFAP-CreERT mouse line to induce recombination in the developing CNS Fig. . To examg-ratios (Fig. Then we decided to assess whether Dicer deletion in astrocytes can change myelination in the developing CNS, since the mGFP-positive astrocytes seems to be closely associated with MBP-positive myelin sheaths in the 3D-reconstructed Z-stack images Fig. A. We exaios Fig. B, C, sugios Fig. D. Consisios Fig. A–C. We nios Fig. D, suggesios Fig. E, F, whiios Fig. G. These To investigate whether the Dicer deletion in astrocytes could affect neurofunction development, we examined the behavioral changes in adulthood at P60 with the induction of recombination at P3 Fig. A. The DiTo confirm the recombination efficiency in astrocytes driven by hGFAP-CreERT in adulthood, we examined the mGFP expression rate by immunostaining for GFAP in the hGFAP-CreERT; mT/mG mouse at P90, 30 days after induction from P60 (Fig. Astrocytes play an important role in governing CNS development , 27. TheEvidence has shown that astrocytes and OLs are closely associated during CNS development. Elimination of astrocytes in developing CNS causes OL death and disrupts myelination in the developing CNS, indicating that astrocytes are functionally required for OL survival . Astrocy


Antibiotic resistance and infections caused by multidrug-resistant bacteria are global health concerns. Reducing the overuse and misuse of antibiotics is the primary step toward minimizing the antibiotic resistance crisis. Thus, it is imperative to introduce and implement novel antimicrobial strategies. Recently, several alternative antimicrobials targeting oxidative stress in bacteria have been studied and shown to be promising. Oxidative stress occurs when bacterial cells fail to detoxify the excessive reactive oxygen species (ROS) accumulated in the cells. Bacteria deploy numerous defense mechanisms against oxidative stress. The oxidative stress response is not essential for the normal growth of bacteria, but it is crucial for their survival. This toxic oxidative stress is created by the host immune response or antimicrobials generating ROS. ROS possess strong oxidation potential and cause serious damage to nucleic acids, lipids, and proteins. Since ROS-based antimicrobials target multiple sites in bacteria, these antimicrobials have attracted the attention of several researchers. In this review, we present recent ROS-based alternative antimicrobials and strategies targeting oxidative stress which might help in mitigating the problem of antibiotic resistance and dissemination. Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. [Antibiotics are used in large quantities in healthcare to treat and even to prevent bacterial infections. However, excessive use of antibiotics has led to the increased development of antibiotic resistance in bacteria ,2,3. An ter spp. . These Eter spp. . Hence, ter spp. . Alternater spp. ,10,11.2−) and hydrogen peroxide (H2O2) are formed as by-products of aerobic respiration when oxygen interacts with the reduced FAD cofactor of flavoenzymes [Oxidative stress occurs when prooxidants overpower antioxidants, i.e., ROS get accumulated in a bacterial cell and exceed the cell’s capacity to readily detoxify the ROS . This caoenzymes ,12. ROS oenzymes . SeveralAntibiotics most commonly target the cell wall , protein synthesis , and DNA replication  in bacteria as their primary mechanism of action ,15. HoweE. coli [2− in the presence of O2, thereby exerting the drug’s toxicity on bacteria [Nitrofurantoin is used in the treatment of urinary tract infections typically caused by E. coli . The mecbacteria .A. baumannii, P. aeruginosa, and carbapenemase-producing Enterobacteriaceae [• [Antimicrobial peptides such as Polymyxin B (PMB) serve as antibacterial agents. PMB is mainly used to treat infections caused by Gram-negative bacteria such as eriaceae ,17. Howeeriaceae . Sampsonaceae [• ,18.Salmonella typhimurium by Arriaga-Alba and co-workers [P. aeruginosa. The induction of antioxidant enzymes on exposure to antibiotics indicated that oxidative stress in bacteria contributed to the lethality of the antibiotic [Escherichia coli was assessed by Wang and Zhao, (2010). They found out that norfloxacin was more lethal in E. coli deficient in catalase gene katG than in its isogenic mutants [ahpC E. coli mutant. Overall, these investigations indicated that treatment of E. coli with antibiotics increased superoxide levels in the bacterium. The superoxide further dismutated to H2O2 and generated the highly toxic hydroxyl radical which amplified the lethality of antibiotics. According to this study, oxidative stress contributes to the lethal effect of all three classes of tested antibiotics—namely, fluoroquinolones, β-lactams, and aminoglycosides [Oxidative stress induction was first shown to be a part of the mechanism of action of an antibiotic in -workers . Antibiotibiotic ,20. A po mutants . Similarycosides .E. coli with an objective to investigate the role of secondary damage after the removal of the initial stressor [Some studies like that by Hong and associates (2019) suggest that bacteria exposed to lethal stressors may not die during the actual treatment but post-treatment due to post-stress ROS-mediated toxicity which they confirmed with enzymatic suppression of post-stress ROS. They studied the effect of stressors in stressor ,21. It wstressor .The reports on the connection between antibiotics and ROS are contradictory. Some reports ,23,24 diAntibiotics’ efficacy is in danger because of the increasing bacterial drug resistance . However®, natural compounds like allicin and surgical honey, antimicrobial photodynamic therapy (aPDT), and non-thermal plasma (NTP) [Advances in the development of ROS-mediated antimicrobials have widened the avenues of alternative antimicrobials beyond metals and nanoparticles. Some examples include ebselen, nanoparticles, nanozymes, AGXXma (NTP) ,27,28.S. aureus [S. aureus skin lesions in rats [Ebselen is an organoselenium-based antioxidant drug that has anti-inflammatory, antioxidant, and cryoprotective properties. It inhibits TrxR in bacteria and this inhibition results in oxidative stress. Ebselen was recently shown to efficiently inhibit the growth of multi-drug resistant . aureus . Dong an in rats . Ebselen in rats .Nanoparticles are <100 nm in size. They generate high amounts of ROS which cause damage to living cells by carbonylation of proteins, peroxidation of lipids, DNA/RNA breakage, and membrane structure destruction . NanoparE. coli and S. aureus  revealing that MSN-maleamic and MSN-maleamic-Cu both triggered oxidative stress mechanisms in both Gram-positive S. aureus and Gram-negative E. coli [E. coli as compared with S. aureus. Their data indicate that MSN-maleamic (MIC = 62.5 µg/mL) works more efficiently than MSN-maleamic-Cu (MIC = 125 µg/mL) in case of E. coli. In S. aureus, the MIC for MSN-maleamic was 250 µg/mL, and >250 µg/mL in the case of MSN-maleamic-Cu [S. aureus treated with MSN-maleamic generated 50% more ROS than the untreated S. aureus while S. aureus treated with MSN-maleamic-Cu generated 30% more ROS as compared with the untreated S. aureus. In E. coli, exposure to both MSN-maleamic and MSN-maleamic-Cu led to the generation of 40% more ROS as compared with the untreated E. coli [Metals like silver and copper have been used as antimicrobials since ancient times . Diaz-Ga E. coli . The mineamic-Cu . S. aure E. coli . ROS gen E. coli . Besides E. coli .Nanozymes produce ROS and are thus promising antimicrobials . One of ®.Several metal-based antimicrobials in addition to the silver-palladium bimetallic alloy discussed above have been studied. One such metal-based novel antimicrobial is AGXX® is an antimicrobial surface coating consisting of micro galvanic elements of silver and ruthenium. It can be coated or deposited on various carriers such as cellulose, plastics, ceramics, or metals. AGXX® is only slightly cell-toxic [® has been reported so far [® catalytically produces ROS and has been shown to efficiently inhibit the growth of pathogens like Enterococcus faecalis, and methicillin-resistant S. aureus (MRSA). AGXX® also inhibited biofilm formation in MRSA [® induced general stress, heat shock (expression of Clp proteases), copper stress, and oxidative stress response in both E. faecalis and MRSA [® also interfered with iron homeostasis and upregulated several siderophore biosynthesis (sbn) genes in MRSA [S. aureus, AGXX® also caused increased protein-thiol-oxidations, protein aggregations, and an oxidized bacillithiol (BSH) redox state [AGXXll-toxic  and no rd so far . AGXX® c in MRSA ,37. Expoand MRSA ,37. AGXX in MRSA . In S. aox state ,38. The ® affects oxidative stress defenses such as superoxide dismutase (SodA), catalase (KatA), alkyl hydroperoxide reductase (AhpCF), thioredoxin/thioredoxin reductase (Trx/TrxR), MerA, and BSH. The multiple modes of action of AGXX® reduce the likelihood of the development of resistance in bacteria and give AGXX® an advantage over conventional antimicrobials [As presented in crobials .Apart from synthetic compounds, ROS-mediated natural compounds like allicin and honey have also been reported for their efficient antimicrobial activity.Allicin (from garlic) is a thiol-reactive compound; it decreases the levels of low molecular weight thiol which functions as a defense mechanism against ROS, thereby reducing the oxidative stress caused by ROS .2O2 [P. aeruginosa and also exhibits antibiofilm activity [Surgihoney Reactive Oxygen (SHRO) is a pharmaceutical honey wound gel . It uses2O2 . When apactivity .In addition to natural compounds, alternative approaches including using photodynamic therapy and non-thermal plasmas have caught attention of researchers.Vibrio fischeri and recombinant E. coli. They used 5,10,15-tris(1-Methylpyridinium-4-yl)-20-(pentafluorophenyl)-porphyrin triiodide (Tri-Py+-Me-PF) as PS to determine the recovery of the viability of bacteria and the development of resistance in bacteria. To investigate the bacterial recovery, bacteria were exposed to 5.0 µM Tri-Py+-Me-PF and were irradiated with white light for 270 min. The bacteria were then protected from the light, aliquots were collected, and bioluminescence was measured. To check if the bacteria develop any resistance toward the treatment, bacteria were exposed to 5.0 µM Tri-Py+-Me-PF and white light for 25 min. The colonies which survived the first irradiation were re-suspended in PBS. The protocol was repeated 10 times. The results suggested that aPDT efficiently damaged V. fischeri and E. coli. Neither of the bacteria recovered their viability after treatment, nor did they develop any resistance towards the treatment [Recently, aPDT has been used to eliminate bacterial biofilms. aPDT uses photosensitizers (PS) which are non-toxic dyes that can be excited by harmless visible light to generate ROS . The aPDreatment .P. aeruginosa [NTPs are plasmas with particles that are not in thermal equilibrium . Cold atruginosa . Direct ruginosa .Oxidative stress imposed by antimicrobials can cause serious damage to bacterial cells. To overcome oxidative stress, bacteria use their defense systems .2O2 accumulation [sodA and sodB which produce superoxide dismutase to protect the cells from the lethal effect of ROS. These genes are regulated by SoxRS. Exposure to H2O2 activates the OxyR regulon, which regulates the expression of protective genes such as katG and ahpC 15 [Bacteria circumvent oxidative stress using an arsenal of defenses such as detoxifying enzymes like catalase, alkyl hydroperoxide reductase, thioredoxin, and superoxide dismutase; pigments like carotenoids; metal homeostasis, DNA repair, general stress response, and SOS response ,11. Thes ahpC 15 . Iron hod by ROS .S. aureus, two siderophores Staphyloferrin A and B were found to enhance resistance to oxidative stress [E. coli, the siderophore enterobactin alleviates oxidative stress [2O2 and paraquat on E. coli to investigate siderophore-mediated regulation of oxidative stress. Enterobactin reduced the sensitivity of E. coli to both H2O2 and paraquat and the expression of enterobactin increased in E. coli when cells were exposed to H2O2 and paraquat even when iron was present in excess. The siderophore potentially neutralizes ROS [®. Siderophore biosynthesis genes were highly upregulated when the bacterium was exposed to AGXX® [® primarily acts by the generation of ROS, the induction of sbn genes (encoding siderophores) could likely be a response generated by MRSA to neutralize ROS.Iron is crucial for the growth and survival of bacteria . It is ae stress . Similare stress ,46. Oxidto AGXX® . As AGXXOxidative stress is a significant stress encountered by bacteria. An overview of oxidative stress in bacteria, the damage caused by the ROS, and the repair mechanisms used by bacteria to combat oxidative stress are shown below .Oxidative stress response in bacteria may protect them from the host immune response as well as from the antibiotic treatment. This facilitates bacteria to develop persistent infections . PersistIn summary, although using ROS-generating antimicrobials and novel strategies can be considered promising, there are some limitations to this approach. ROS cannot distinguish between bacterial and mammalian cells, and since they directly damage DNA, they can induce mutations in specific targets of antibiotics, thereby leading to antibiotic resistance and establishment of persistence ,45. Furt


Gastric cancer (GC) represents a major malignancy and is the third deathliest cancer globally. Several lines of evidence indicate that the epithelial-mesenchymal transition (EMT) has a critical function in the development of gastric cancer. Although plentiful molecular biomarkers have been identified, a precise risk model is still necessary to help doctors determine patient prognosis in GC.Gene expression data and clinical information for GC were acquired from The Cancer Genome Atlas (TCGA) database and 200 EMT-related genes (ERGs) from the Molecular Signatures Database (MSigDB). Then, ERGs correlated with patient prognosis in GC were assessed by univariable and multivariable Cox regression analyses. Next, a risk score formula was established for evaluating patient outcome in GC and validated by survival and ROC curves. In addition, Kaplan-Meier curves were generated to assess the associations of the clinicopathological data with prognosis. And a cohort from the Gene Expression Omnibus (GEO) database was used for validation.CDH6, COL5A2, ITGAV, MATN3, PLOD2, and POSTN, were identified. Based on the risk model, GC patients were assigned to the high- and low-risk groups. The results revealed that the model had good performance in predicting patient prognosis in GC.Six EMT-related genes, including We constructed a prognosis risk model for GC. Then, we verified the performance of the model, which may help doctors predict patient prognosis. Gastric cancer (GC), a worldwide health problem, represents the fifth commonest malignancy and the third deadliest cancer after lung and colorectal cancers . In 2018TSP50, CD44, and FOXA1, regulate EMT in GC [Epithelial-mesenchymal transition (EMT) represents an important molecular program in diverse cellular events such as wound healing, tissue fibrosis, and cancer progression . StudiesMT in GC –11. BecaIn this study, based on TCGA and MSigDB data, differentially expressed genes associated with EMT were selected. Then, univariable and multivariable Cox regression analyses were carried out for identifying overall survival (OS)-related genes. Next, a risk model was constructed to predict and evaluate patient prognosis in GC. We finally used additional bioinformatic and statistical methods to validate this model.https://portal.gdc.cancer.gov/repository). The RNA sequencing data of GC patients were retrieved. Clinical features, including survival status, survival time, age, gender, grade, stage, T stage, N stage, and M stage, were adopted in the current study. The “HALLMARK_EPITHELIAL_MESENCHYMAL_TRANSITION” gene list containing 200 genes was obtained from MSigDB v7.2(http://www.gsea-msigdb.org/gsea/msigdb/index.jsp). To identify differentially expressed epithelial-mesenchymal transition-related genes (ERGs), we utilized the “limma” package in the R software 4.0.2 to analyze mRNA sequencing data for these 200 ERGs, with significance threshold set at p< 0.05.The original gene expression data of the TCGA gastric dataset were acquired from the TCGA database  curves (AUCs) for 1-year, 3-year, and 5-year overall survival were utilized to determine the prediction accuracy of the constructed model.https://www.ncbi.nlm.nih.gov/geo/) for validating the prognostic value of the established risk model.In order to assess whether the above risk score independently predicts GC prognosis, univariable and multivariable Cox regression analyses were carried out. The expression levels of the six detected ERGs in normal and tumor tissues were examined, and Kaplan-Meier survival analysis based on different clinicopathological parameters was conducted. We also chose a cohort (GSE62254) from the GEO database (https://cistrome.shinyapps.io/timer/) represents a comprehensive platform that can analyze immune infiltration systematically in multiple cancers [The Tumor IMmune Estimation Resource (TIMER) database . After screening, we selected 371 gastric cancer patients for analysis. The clinical characteristics of GC patients are shown in Table P < 0.05)  were identified, which may serve as important prognosis predictors (Table COL5A2) was correlated with better prognosis (0 < HR< 1), and its overexpression may be beneficial to survival.Six genes   , and risk score were OS-associated indicators Fig. a. Subseq05) Fig. b.Fig. 3P < 0.05)   Fig. . In addi Fig. . I01) Fig. . And the01) Fig. . Further01) Fig. . The resREC8 inhibits EMT in GC cells by controlling EGR1 expression and binding to it [CRYAB) also promotes GC cell migration and invasion through EMT under the control of NF-κB signaling [Gastric cancer remains a globally important pathology, with elevated mortality. GC represents a major social burden and peritoneal metastasis is common in advanced GC cases . Postopeng to it . In adding to it . Alpha Bignaling . AlthougTo date, increasing amounts of attention focus on the construction of cancer models. The prognosis of cancer can be predicted by screening out genes related to biological processes, such as the effect of glycolysis-related genes on the prognosis of hepatocellular carcinoma and the association of autophagy-related genes with the survival of lung cancer , 20. HowCOL5A2 is related to prognosis in patients with GC [COL5A2 amounts show improved clinicopathological phenotypes [COL5A2 is also involved in the carcinogenesis of colorectal cancer [ITGAV, belonging to the integrin family of extracellular matrix receptors, is implicated in multiple cancers. Downregulation of ITGAV can cause the inhibition of GC cell proliferation, migration, and invasion, indicating a critical role for ITGAV in GC progression [MATN3, a protein-coding gene, might affect GC occurrence and development, acting as an oncogene [MATN3 is differentially expressed and aberrantly methylated in GC [PLOD2 is an important regulator of peritoneal dissemination in GC patients, leading to poor prognosis [POSTN affects cancer cell proliferation via ERK and may promote EMT [POSTN overexpression is a known risk factor for GC, inducing metastasis and aggravating malignant behavior [CDH6, a class II Cadherin, induces EMT during embryonic development and shows abnormal reactivation in malignant tumors [CDH6 may preferentially participate in the late stage of EMT, with direct association with cell invasiveness [CDH6’s function in GC remains undefined. Therefore, further research for discovering the function of CDH6 is urgently required. Besides, there are other studies on gastric cancer. Follistatin-like 1 (FSTL1) is a kind of glycoprotein, its high expression is associated with poor prognosis and has a positive correlation with immune infiltration in GC patients [Previous studies have reported the roles of these genes in GC.  with GC . In bladenotypes . COL5A2 l cancer . ITGAV, gression . MATN3, oncogene . In addied in GC . PLOD2 irognosis . POSTN amote EMT . POSTN obehavior . CDH6, at tumors . Differesiveness . Howeverpatients . CTLA-4 patients . RNAs arpatients , 35. Furpatients .So far, most studies have assessed the relationship between a single gene and prognosis in GC. Here, we constructed a dependable prognostic risk model for GC based on sequencing data for ERGs and clinicopathological features from the TCGA database. This might provide a reliable method for predicting patient survival in GC. However, some limitations are worth noting. First, we established the risk model by using the TCGA database, and further validation of this model should be carried out in vitro, in vivo, and in clinical trials. Secondly, the immune statuses of these six genes deserve further investigation.This study has developed a six-gene prognostic risk model based on EMT, which could provide a reference for assessing whether gastric cancer patients are at high risk. This risk model could help better manage patients and predict prognosis. These findings may provide novel insights into the relationship between GC and EMT.


RAFS) genes superfamily is critical for the synthesis of raffinose, which accumulates in plant leaves under abiotic stress. However, it remains unclear whether RAFS contributes to resistance to abiotic stress in plants, specifically in the Gossypium species.The Raffinose synthetase (RAFS genes from G. hirsutum, G. barbadense, G. arboreum and G. raimondii by using a series of bioinformatic methods. Phylogenetic analysis showed that the RAFS gene family in the four Gossypium species could be divided into four major clades; the relatively uniform distribution of the gene number in each species ranged from 12 to 25 based on species ploidy, most likely resulting from an ancient whole-genome polyploidization. Gene motif analysis showed that the RAFS gene structure was relatively conservative. Promoter analysis for cis-regulatory elements showed that some RAFS genes might be regulated by gibberellins and abscisic acid, which might influence their expression levels. Moreover, we further examined the functions of RAFS under cold, heat, salt and drought stress conditions, based on the expression profile and co-expression network of RAFS genes in Gossypium species. Transcriptome analysis suggested that RAFS genes in clade III are highly expressed in organs such as seed, root, cotyledon, ovule and fiber, and under abiotic stress in particular, indicating the involvement of genes belonging to clade III in resistance to abiotic stress. Gene co-expressed network analysis showed that GhRFS2A-GhRFS6A, GhRFS6D, GhRFS7D and GhRFS8A-GhRFS11A were key genes, with high expression levels under salt, drought, cold and heat stress.In this study, we identified 74 RAFS genes in Gossypium species and a theoretical basis for the identification of stress resistance materials in cotton.The findings may provide insights into the evolutionary relationships and expression patterns of The online version contains supplementary material available at 10.1186/s12859-021-04276-4. RAFS gene in peas has shown that RAFS is a transglycosidase similar to galactosidase in structure and biochemical properties [RAFS catalyzes the synthesis of raffinose, with galactinol as a galactosyl donor and sucrose as an acceptor [GOLS under abiotic stress have been well clarified in many plant species. Some GOLS are expressed under the induction of abiotic stress, such as heat shock, drought, osmotic shock and salinity [GOLS has been found to increase the content of galactinol and raffinose and then improve abiotic stress tolerance [GOLS, there have been few studies on the plant RAFS gene. The earliest reported RAFS gene was cloned from peas [RAFS genes were cloned [Arabidopsis thaliana [RAFS genes (AtRFS1-6) have been identified in Arabidopsis Thaliana [AtRFS5 has demonstrated RFS activity, while AtRFS4 has shown little raffinose synthetic capacity. Except for AtRFS4 and AtRFS5, AtRFS2 has been predicted as drought induced RAFS in Arabidopsis thaliana [RAFS genes have been identified [RAFS gene has been found to play a vital role in seed viability and longevity [RAFS can increase the content of raffinose. Despite extensive research on the role of gossyps in regulating or responding to abiotic stress in plants, the role of gossyps in drought resistance in plants remains unclear.Raffinose family oligosaccharides (RFOs) accumulating in leaves during plant development are thought to play a vital role in the stress tolerance of plants . Raffinooperties . RAFS caacceptor . The funsalinity , 8. Overolerance , 9, 10. rom peas , and late cloned , 3, 12 fe cloned , 13, rice cloned , 14 and thaliana . In partThaliana , 16, butthaliana . Based oentified . A uniquongevity . HoweverRAFS plays an important role in the plant seed acquisition of drought resistance and the extension of seed life [RAFS, especially systematic research on the RAFS gene family in Gossypium species [RAFS gene superfamily in Gossypium species, we identified RAFS genes and constructed a phylogenetic tree in the present study. We also analyzed the synteny, promoters and co-expression network associated with these RAFS genes. The constructed phylogenetic tree showed that RAFS genes could be grouped into four subfamilies. Through research of the cotton RAFS gene family, this study provides a certain theoretical basis for further research on cotton’s stress resistance mechanism and the improvement of cotton’s stress resistance.Synthetase eed life , 21. Owi species . Cotton  species . To compRAFS protein sequences of Arabidopsis, rice and maize as a query for BLAST, along with a Pfam search, a total of 74 RAFS genes in G. arboreum, G. raimondii, G. hirsutum and G. barbadense were identified. Among them, there were 12 G. raimondii RAFS genes named GrRFS1-GrRFS12, 12 G. arboreum RAFS genes named GaRFS1-GaRFS12 according to their locations on the chromosome, and 25 G. hirsutum RAFS genes named GhRFS1A-GhRFS13A and GhRFS1D-GhRFS12D based on their locations to the At and Dt sub-genomes, respectively. Moreover, 25 G. barbadense RAFS genes were named GbRFS1A-GbRFS12A, GhRFS1D-GhRFS10D and GbRFS23-GbRFS25 within the scaffold , while it sharply increased in the late stages, which suggested that these genes may be involved in responding to abiotic stresses by over-expression to increase raffinose content when subjected to abiotic stresses. Noteworthy, not all of the genes from the same clade, with similar sequences, had the same expression level and gene functions. For example, most of the other GhRFS genes were not expressed or lowly expressed in any of the abiotic stress conditions. Moreover, to clarify whether cis-acting elements influenced the expression levels, we analyzed the 2000 bp sequence upstream of the start codon of GhRFS gene. The results showed that the promoters of almost all GhRFS contained hormone-related cis-acting elements, such as GA3, except for GhRFS6A, GhRFS11D and GhRFS6A. In addition, GhRFS1A, GhRFS2D, GhRFS8A and GhRFS8D, expressed highly under abiotic stress conditions, specifically possessed cis-acting elements MYB and ANX, which proved to be relevant to abiotic stresses in plants  and Arabidopsis plants expressing ZmRFS, we found that raffinose could directly and positively regulate drought stress tolerance of plants [RAFS genes can regulate raffinose content in vegetative tissue of in cotton, which is cultivated worldwide and faces severe biotic and abiotic stresses. Raffinose participates not only in the development of plant seeds but also in the response of plants to external stresses. With the completion of cotton genome sequencing, it is very important to study cotton RAFS genes from the whole genome. In this study, we performed a whole-genome identification of candidate RAFS genes in G. hirsutum, G. barbadense, G. arboreum and G. raimondii, mainly focusing on the allotetraploid cotton G. hirsutum, with the aim of understanding its roles, their evolutionary relationships and expression in response to various abiotic stresses. The results can provide basic information and serve as an important resource for further investigations into the candidate RAFS gene.Raffinose accumulates in higher levels in plants when responding to abiotic stress . Severalthe dark , 24, 25.f plants . DespiteRAFS genes in the genomes of G. hirsutum, G. barbadense, G. arboreum and G. raimondii were identified. The phylogenetic analysis revealed that all of the candidate RAFS genes were divided into four subfamilies, and the numbers of identified candidate RAFS genes in different cotton species were almost the same based on the species ploidy. That is, tetraploid cotton had about twice the number of candidate RAFS genes as diploid cotton, for example, only 12 genes in G. barbadense and G. arboreum as compared to 25 genes in upland cotton and G. raimondii, illustrating that candidate RAFS genes were conserved during evolution and underwent large-scale expansion in higher plants. Moreover, clade I was the least abundant member of the candidate RAFS clade.A total of 74 candidate cis-elements upstream of the promoter regions can regulate gene transcription, which ultimately causes gene expression. We predicted that various cis-elements in the candidate RAFS gene promoter region were associated with plant hormone response elements and abiotic stress response elements  is a key enzyme in the synthesis process of raffinose. Although RAFS genes of some species have been identified and functionally characterized, whether RAFS contributes to resistance to abiotic stress in Gossypium species had not been investigated. In this study, we conducted a complete genome wide survey of RAFS gene family in four Gossypium species which results into identification of 74 RAFS genes distributed into evolutionary conserved four groups. RNA-seq and promoter analyses revealed genes GhRFS1A, GhRFS2D, GhRFS8A and GhRFS8D, with ANX and MYB motifs, were highly expressed in plant seeds, cotyledonsroots, and under heat, cold, salt and drought stress conditions, which implied that plant hormones may enhance the expression of genes and then contribute to resist stress. These results elucidate the potential function in response to abiotic stress for RAFS genes in Gossypium species.Raffinose synthetase  used in this study were downloaded from CottonFGD database (https://cottonfgd.org/about/download.html). A local BLASTp search [RAFS genes of Arabidopsis thaliana (AtRAFS1-AtRAFS5), Oryza sativa (OsRAFS1-OsRAFS6) and Zea mays (ZmRAFS1-ZmRAFS10) as a query against all of the protein sequences in each genome was implemented to obtain RAFS homologs with an E-value cutoff of 10−5. To identify raffinose synthetase family members more accurately, the profile hidden Markov model (PF05691) of the HMMER3.0 program [Whole-genome data of all four p search  using th program  was furthttps://itol.embl.de/). The subcellular location of the RAFS genes was predicted through the CELLO v.2.5 website (http://cello.life.nctu.edu.tw/) [MAFFT, an accurate alignment software, was used to align multiple sequences with default parameters . The neiedu.tw/) .RAFS genes were identified using the program Multiple Em for Motif Elicitation  with the following parameters: the width of a motif was between 6 and 50aa, and the number of motifs was no more than 20 [http://gsds.cbi.pku.edu.cn/) was used to draw gene structure diagrams[G. arboreum, G. raimondii, G. barbadense and G. hirsutum) were used to compile MAP files of RAFS genes, and MapInspect software was used to draw the physical location map of chromosomes.Conserved functional motifs of  than 20 . The webG. hirsutum RFS genes in all four RFS clades, the promoter sequences, including at least 2 kb of the upstream regions, were downloaded from the CottonFGD databases (http://cottonfgd.org). The conserved cis-regulatory elements in the promoter regions were predicted using the PlantCARE (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/) and PLACE (https://www.dna.affrc.go.jp/PLACE/?action=newplace) databases. Transcriptome expression data of RAFS genes in G. hirsutum TM-1 were downloaded from the NCBI database, with project number PRJNA248163, which had three biological replications per time point for each stress condition and was normalized against the control comparison group. A gene expression heatmap was produced with TBtools software. Reads were mapped to the G. hirsutum genome using Hisat2 software [RAFS genes in the development of seeds, fibers, roots, cotyledons and ovules under the conditions of cold, heat, salt and drought stress could reflect their response to abiotic stress.For software . StringTsoftware . FragmenRAFS gene family expanded through tandem duplication events or segmental duplication, a collinear analysis was completed with TBtools software [To determine whether the software .G. hirsutum under cold, heat, salt, and drought stress conditions were used to build a gene co-expression network with Pearson’s correlation coefficients (r), which were calculated using the ‘cor’ function of R package. The criteria for co-expression genes identification were set at r ≥ 0.9 or r ≤ -0.9 and P-values ≤ 0.05 [http://www.cytoscape.org/) was used to visualize the gene co-expression network with graph [Transcriptome data of s ≤ 0.05 . Cytoscath graph . The hubAdditional file 1: Table S1. Basic characteristic of RAFS genes in G. hirsutum, G. barbadense, G. arboreum and G. raimondii.Additional file 2: Table S2. Chromosomal locations of RAFS genes in G. hirsutum, G. barbadense, G. arboreum and G. raimondii.Additional file 3: Table S3. Expresstion level of RAFS genes in G. hirsutum under abiotic stress