Folate receptor autoantibody (FRAA) has caught increasing attention since its discovery in biological fluids of patients with autism spectrum disorder (ASD), but quantification and understanding of its function are still in their infancy. In this study, we aimed to quantify serum binding-FRAA and explore its relation with serum folate, vitamin B12 (VB12) and ferritin. We quantitated serum binding-FRAA in 132 ASD children and 132 typically-developing (TD) children, as well as serum levels of folate, VB12 and ferritin. The results showed that serum binding-FRAA in the ASD group was significantly lower than that in the TD group (p < 0.0001). Further analysis showed that the difference between these two groups was attributed to boys in each group, not girls. There was no statistically significant difference in folate levels between the ASD and TD groups (p > 0.05). However, there was significant difference in boys between these two groups, not girls. Additionally, the combination of nitrite and binding-FRAA showed potential diagnostic value in patients with ASD (AUC > 0.7). Moreover, in the ASD group, the level of folate was consistent with that of binding-FRAA, whereas in the TD group, the binding-FRAA level was high when the folate level was low. Altogether, these differences revealed that the low serum FRAA in autistic children was mediated by multiple factors, which deserves more comprehensive investigation with larger population and mechanistic studies.

Compared to pathogens Pseudomonas aeruginosa and P. putida, P. donghuensis HYS has stronger virulence towards Caenorhabditis elegans. However, the underlying mechanisms haven’t been fully understood. The heme synthesis system is essential for Pseudomonas virulence, and former studies of HemN have focused on the synthesis of heme, while the relationship between HemN and Pseudomonas virulence were barely pursued. In this study, we hypothesized that hemN2 deficiency affected 7-hydroxytropolone (7-HT) biosynthesis and redox levels, thereby reducing bacterial virulence. There are four hemN genes in P. donghuensis HYS, and we reported for the first time that deletion of hemN2 significantly reduced the virulence of HYS towards C. elegans, whereas the reduction in virulence by the other three genes was not significant. Interestingly, hemN2 deletion significantly reduced colonization of P. donghuensis HYS in the gut of C. elegans. Further studies showed that HemN2 was regulated by GacS and participated in the virulence of P. donghuensis HYS towards C. elegans by mediating the synthesis of the virulence factor 7-HT. In addition, HemN2 and GacS regulated the virulence of P. donghuensis HYS by affecting antioxidant capacity and nitrative stress. In short, the findings that HemN2 was regulated by the Gac system and that it was involved in bacterial virulence via regulating 7-HT synthesis and redox levels were reported for the first time. These insights may enlighten further understanding of HemN-based virulence in the genus Pseudomonas.

Lonicerae Japonicae Flos (LJF) is a natural plant containing abundant antioxidant ingredients. In order to extract more antioxidants from LJF, in this study, a novel strategy was proposed for optimizing the extraction factor level by response surface methodology with a tailored deep eutectic solvent (DES) as the extraction solvent and antioxidant ability as the evaluation index. After optimizing the composition of DES and the extraction condition, the extracts obtained by our proposed method yielded better antioxidant ability (229.1–249.1 μmol TE/g DW) and higher antioxidant contents (34.2–36.5 mg GAE/g DW for total phenolics and 119.6–123.0 mg RE/g DW for total flavonoids) from LJF in 5 min without organic solvent consumption that were significantly superior to the Chinese Pharmacopoeia extraction method. The K-T solvation parameter and a scanning electron microscope were adopted to explore the extraction mechanism, and the results showed that the polarity and damage effect on plant cells of DES were crucial for the extraction of antioxidants. In addition, after combining the HPLC fingerprint and partial least squares model, chlorogenic acid, rutin, and 3,5-O-Dicaffeoylquinic acid were screened as the antioxidant Q-markers of LJF. This work demonstrates that an optimization strategy based on antioxidant ability and tailored DES has the potential to extract more antioxidants from natural plants.

Biomass carbon dots (CDs) derived from natural plants possess the advantages of low cost, photostability, and excellent biocompatibility, with potential applications in chemical sensing, bioimaging, and nanomedicine. However, the development of biomass CDs with excellent antioxidant activity and good biocompatibility is still a challenge. Herein, we propose a hypothesis for enhancing the antioxidant capacity of biomass CDs based on precursor optimization, extraction solvent, and other conditions with broccoli as the biomass. Compared to broccoli water extracts, broccoli powders, and broccoli organic solvent extracts, CDs derived from broccoli water extracts (BWE-CDs) have outstanding antioxidant properties due to the abundant C═C, carbonyl, and amino groups on their surface. After optimization of the preparation condition, the obtained BWE-CDs exhibit excellent free-radical scavenging activity with an EC50 of 68.2 μg/mL for DPPH• and 22.4 μg/mL for ABTS•+. Cytotoxicity and zebrafish embryotoxicity results indicated that BWE-CDs have lower cytotoxicity and better biocompatibility than that of CDs derived from organic solvents. In addition, BWE-CDs effectively scavenged reactive oxygen species (ROS) in A549 cells, 293T cells, and zebrafish, as well as eliminating inflammation in LPS-stimulated zebrafish. Mechanistic studies showed that the anti-inflammatory effect of BWE-CDs was dependent on the direct reaction of CDs with free radicals, the regulation of NO levels, and the upregulation of the expression of SOD and GPX-4. This work indicates that the antioxidant activity of CDs could be enhanced by using solvent extracts of biomass as precursors, and the obtained BWE-CDs exhibit characteristics of greenness, low toxicity, and excellent antioxidant and anti-inflammatory activities, which suggests the potential promising application of BWE-CDs as an antioxidant nanomedicine for inflammatory therapy.

7-Hydroxytropolone (7-HT) is a unique iron scavenger synthesized by Pseudomonas donghuensis HYS that has various biological activities in addition to functioning as a siderophore. P. donghuensis HYS is more pathogenic than P. aeruginosa toward Caenorhabditis elegans, an observation that is closely linked to the biosynthesis of 7-HT. The nonfluorescent siderophore (nfs) gene cluster is responsible for the orderly biosynthesis of 7-HT and represents a competitive advantage that contributes to the increased survival of P. donghuensis HYS; however, the regulatory mechanisms of 7-HT biosynthesis remain unclear. This study is the first to propose that the ECF σ factor has a regulatory effect on 7-HT biosynthesis. In total, 20 ECF σ factors were identified through genome-wide scanning, and their responses to extracellular ferrous ions were characterized. We found that SigW was both significantly upregulated under high-iron conditions and repressed by an adjacent anti-σ factor. RNA-Seq results suggest that the SigW/RsiW system is involved in iron metabolism and 7-HT biosynthesis. Combined with the siderophore phenotype, we also found that SigW could inhibit siderophore synthesis, and this inhibition can be relieved by RsiW. EMSA assays proved that SigW, when highly expressed, can directly bind to the promoter region of five operons of the nfs cluster to inhibit the transcription of the corresponding genes and consequently suppress 7-HT biosynthesis. In addition, SigW not only directly negatively regulates structural genes related to 7-HT synthesis but also inhibits the transcription of regulatory proteins, including of the Gac/Rsm cascade system. Taken together, our results highlight that the biosynthesis of 7-HT is negatively regulated by SigW and that the SigW/RsiW system is involved in mechanisms for the regulation of iron homeostasis in P. donghuensis HYS. As a result of this work, we identified a novel mechanism for the global negative regulation of 7-HT biosynthesis, complementing our understanding of the function of ECF σ factors in Pseudomonas.

The newly discovered iron scavenger 7-hydroxytropolone (7-HT) is secreted by Pseudomonas donghuensis HYS. In addition to possessing an iron-chelating ability, 7-HT has various other biological activities. However, 7-HT's biosynthetic pathway remains unclear. This study was the first to report that the phenylacetic acid (PAA) catabolon genes in cluster 2 are involved in the biosynthesis of 7-HT and that two genes, paaZ (orf13) and ech, are synergistically involved in the biosynthesis of 7-HT in P. donghuensis HYS. Firstly, gene knockout and a sole carbon experiment indicated that the genes orf17-21 (paaEDCBA) and orf26 (paaG) were involved in the biosynthesis of 7-HT and participated in the PAA catabolon pathway in P. donghuensis HYS; these genes were arranged in gene cluster 2 in P. donghuensis HYS. Interestingly, ORF13 was a homologous protein of PaaZ, but orf13 (paaZ) was not essential for the biosynthesis of 7-HT in P. donghuensis HYS. A genome-wide BLASTP search, including gene knockout, complemented assays, and site mutation, showed that the gene ech homologous to the ECH domain of orf13 (paaZ) is essential for the biosynthesis of 7-HT. Three key conserved residues of ech (Asp39, His44, and Gly62) were identified in P. donghuensis HYS. Furthermore, orf13 (paaZ) could not complement the role of ech in the production of 7-HT, and the single carbon experiment indicated that paaZ mainly participates in PAA catabolism. Overall, this study reveals a natural association between PAA catabolon and the biosynthesis of 7-HT in P. donghuensis HYS. These two genes have a synergistic effect and different functions: paaZ is mainly involved in the degradation of PAA, while ech is mainly related to the biosynthesis of 7-HT in P. donghuensis HYS. These findings complement our understanding of the mechanism of the biosynthesis of 7-HT in the genus Pseudomonas.

Hydrocolloids have been widely used to adjust properties of natural starches, but related research on kudzu starch is still rare. In this study, we investigated the effects of gum arabic (AG), sodium alginate (SA), locust bean gum (LG), and guar gum (GG) on kudzu starch from the perspective of its particle size, pasting, texture, rheology, dehydration rate, thermal properties, microstructure, and sensitivity to amyloglucosidase. Results showed that GG significantly increased the particle size of starch. Addition of AG led to lower peak-, final- and holding-viscosity. SA increased the retention viscosity of kudzu starch, while LG and GG increased its peak viscosity. Addition of hydrocolloids increased the hardness, chewiness, and cohesiveness of starch-hydrocolloid complexes, and reduced the dehydration rate of complex gels. Dynamic rheological data showed that the energy storage modulus (G′) was significantly higher than the loss modulus (G″). The magnitude of modulus increased with frequency, and elastic properties were better than viscous properties. Thermal analysis showed that hydrocolloids increased the starting temperature (To), and the final temperature (Tc). With addition of each of these four hydrocolloids, a more regular and porous thick-wall dense structure was formed, which effectively lowered kudzu starch's sensitivity to amyloglucosidase. It indicated that the binding of hydrocolloid to starch may slow down glucose release into blood during digestion. These results will help understand effects of natural hydrocolloid on kudzu starch, as well as expanding its application in food industry.

Polycyclic aromatic hydrocarbons (PAHs) are common carcinogens. Benzo(a)pyrene is one of the most difficult high-molecular-weight (HMW) PAHs to remove. Biodegradation has become an ideal method to eliminate PAH pollutants from the environment. The existing research is mostly limited to low-molecular-weight PAHs; there is little understanding of HMW PAHs, particularly benzo(a)pyrene. Research into the biodegradation of HMW PAHs contributes to the development of microbial metabolic mechanisms and also provides new systems for environmental treatments. Pseudomonas benzopyrenica BaP3 is a highly efficient benzo(a)pyrene-degrading strain that is isolated from soil samples, but its mechanism of degradation remains unknown. In this study, we aimed to clarify the high degradation efficiency mechanism of BaP3. The genes encoding Rhd1 and Rhd2 in strain BaP3 were characterized, and the results revealed that rhd1 was the critical factor for high degradation efficiency. Molecular docking and enzyme activity determinations confirmed this conclusion. A recombinant strain that could completely mineralize benzo(a)pyrene was also proposed for the first time. We explained the mechanism of the high-efficiency benzo(a)pyrene degradation ability of BaP3 to improve understanding of the degradation mechanism of highly toxic PAHs and to provide new solutions to practical applications via synthetic biology.

以课题组筛选出的多环芳烃高效降解菌--蜡样芽孢杆菌(Bacillus cereus)DQ01菌株为研究对象,以荧蒽为目标污染物,以牛肉膏蛋白胨培养基(LB培养基)为共代谢基质,基于同位素相对和绝对定量技术(iTRAQ)比较细菌在无机盐培养基(MSM培养基)和MSM-LB培养环境中降解荧蒽的全蛋白表达水平,旨在揭示蜡样芽孢杆菌在共代谢过程中分子代谢水平上的变化.结果共检测到64个差异表达蛋白质,其中上调表达的蛋白质有28个,下调表达的蛋白质36个.鉴定到差异蛋白的功能集中在代谢、应激反应、信息传递和调控等方面,同时对比KEGG通路富集分析的结果,也显示细菌有关能量代谢和信息传递、调控的通路产生的变化.比较细菌在是否处于共代谢下的差异蛋白表达,推测细菌在降解多环芳烃过程中的关键环节是小分子代谢和适应外界环境的通路.

Polycyclic aromatic hydrocarbons (PAHs) are degraded by the highly efficient degrading bacterium Bacillus cereus. Transmembrane transport is highly important in PAH degradation by bacteria. Surfactants are the key substances that promote PAH adsorption, uptake and transmembrane transport by Bacillus cereus. In this study, the isobaric tags for relative and absolute quantitation (iTRAQ) approach was used for high-throughput screening of key functional proteins during transmembrane fluoranthene transport by Bacillus cereus treated with Tween 20. In addition, SWISS-MODEL was used to simulate the tertiary structures of key transmembrane proteins and analyze how Tween 20 promotes transmembrane transport. Transmembrane fluoranthene transport into Bacillus cereus requires transmembrane proteins and energy. Tween 20 was observed to improve bacterial motility and transmembrane protein expression. The interior of representative transmembrane proteins is mostly composed of hydrophobic β-sheets while amphipathic α-helices are primarily distributed at their periphery. The primary reason for this configuration may be α-helices promote the aggregation of surfactants and the phospholipid bilayer and the β-sheets promote surfactant insertion into the phospholipid bilayer to enhance PAH transport into Bacillus cereus. Investigating the effect of Tween 20 on Bacillus cereus transmembrane proteins during transmembrane fluoranthene transport is important for understanding the mechanism of PAH degradation by microorganisms.

Pseudomonas donghuensis HYS is more virulent than P. aeruginosa toward Caenorhabditis elegans but the mechanism underlying virulence is unclear. This study is the first to report that the specific gene cluster gtrA/B/II in P. donghuensis HYS is involved in the virulence of this strain toward C. elegans, and there are no reports of GtrA, GtrB and GtrII in any Pseudomonas species. The pathogenicity of P. donghuensis HYS was evaluated using C. elegans as a host. Based on the prediction of virulence factors and comparative genomic analysis of P. donghuensis HYS, we identified 42 specific virulence genes in P. donghuensis HYS. Slow-killing assays of these genes showed that the gtrAB mutation had the greatest effect on the virulence of P. donghuensis HYS, and GtrA, GtrB and GtrII all positively affected P. donghuensis HYS virulence. Two critical GtrII residues (Glu47 and Lys480) were identified in P. donghuensis HYS. Transmission electron microscopy (TEM) showed that GtrA, GtrB and GtrII were involved in the glucosylation of lipopolysaccharide (LPS) O-antigen in P. donghuensis HYS. Furthermore, colony-forming unit (CFU) assays showed that GtrA, GtrB and GtrII significantly enhanced P. donghuensis HYS colonization in the gut of C. elegans, and glucosylation of LPS O-antigen and colonization in the host intestine contributed to the pathogenicity of P. donghuensis HYS. In addition, experiments using the worm mutants ZD101, KU4 and KU25 revealed a correlation between P. donghuensis HYS virulence and the TIR-1/SEK-1/PMK-1 pathways of the innate immune p38 MAPK pathway in C. elegans. In conclusion, these results reveal that the specific virulence gene cluster gtrA/B/II contributes to the unique pathogenicity of HYS compared with other pathogenic Pseudomonas, and that this process also involves C. elegans innate immunity. These findings significantly increase the available information about GtrA/GtrB/GtrII-based virulence mechanisms in the genus Pseudomonas.

Polycyclic aromatic hydrocarbons (PAHs) are hazardous pollutants that are ubiquitous in the environment. Numerous bacteria have evolved to have degrading genes or pathways to degrade PAHs. Stenotrophomonas maltophilia strain W18 was found to be able to degrade PAHs. Including 43 other complete genome sequences of S. maltophilia strains, we performed a comparative genomic analysis of 44 S. maltophilia strains by running OrthoFinder. A KEGG pathway enrichment analysis of environmental and clinical isolates of S. maltophilia revealed that environmental isolates tended to enhance gene functions such as "energy metabolism," "amino acid metabolism," "xenobiotic biodegradation and metabolism," and "folding, sorting, and degradation." The pangenome of the 44 S. maltophilia strains was open, while the core genome was estimated to reach a steady plateau. Based on gene annotations, we inferred that most of the degradation potential came from the core genome of S. maltophilia, while character genes and accessory genes also contributed to the degradation ability of S. maltophilia W18. The genes expression level of core genes, character genes and accessory genes were proved by RT-qPCR experiment, and accessory genes encoding alcohol dehydrogenase were upregulated most compared with genes with similar functions. We performed a credible comparative genomic analysis of S. maltophilia strains. S. maltophilia W18 was set as a model PAH-degrading bacterium of this species in this study, which would provide guidance for understanding and predicting the degradation mechanisms of other PAH-degrading S. maltophilia strains lacking complete genome data or waiting to be determined. IMPORTANCE This study provided the latest comparative genomic analysis on Stenotrophomonas maltophilia strains and focused on analyzing their genomic features that allow them to adapt to natural environments. In this study, we set S. maltophilia W18 as a typical PAH-degrading strain of this species. By discussing the genomic adaptative features of degrading PAH, we can predict genomic adaptative features of other S. maltophilia PAH-degrading strains since the core function of this species is stable. The gene functions of how S. maltophilia environmental isolates are enhanced for adaptation to various natural environments compared with clinical isolates have been revealed. Combined with a pangenome analysis and RT-qPCR results, we have proved that core genes, character genes, and accessory genes are all involved in PAH degradation. Accessory genes encoding alcohol dehydrogenase were upregulated most compared with core and character genes with similar functions, which suggests that PAH metabolization potential might be enhanced by horizontal gene transfer.

Symbiotic microorganisms colonize external or internal surfaces of a host depending on environmental factors, and may supply the host with special functions.More and more researchers have proven that symbiotic gut microorganisms are related to a diverse range of physiological functions of a host including immunity, nutrition, metabolism and even mental health.Thus, gut microorganisms comprise an important "microbial organ" in humans.Since the early days of microbiota research, animal models have been used frequently for their microbiota, contributing greatly to new research in this field.This review provides an overview of animals used as models in symbiotic microorganism studies, including zebrafish (Danio rerio), mice (Mus musculus), pigs (Sus scrofa domesticus), and monkeys (Macaca mulatta).We provide insight into the development and characteristics of these model animals, highlighting the advantages and disadvantages of each model, as well as any outstanding scientific achievements based on their use.We also note that honey bee (Apis), fruit fly (Drosophila) and nematode (Caenorhabditis elegans) models are emerging as more prevalent in recent gut microbiota studies.This paper will contribute to better understanding the similarities and differences between the microbiota of model animals and humans, while providing useful information for effectively implementing these animal models in future research.

17 beta-estradiol (E2) is an important sex steroid produced by ovary and brain. In mammals, E2 plays an important role in hypothalamus-pituitary-gonad axis to regulate puberty onset, however, little is known about the functional role of E2 in teleost pituitary. Using prepubertal grass carp as model, three nuclear estrogen receptors (nERs: estrogen receptor alpha, estrogen receptor beta 1, and estrogen receptor beta 2) and two G protein-coupled estrogen receptors (GPER1: GPER1a and GPER1b) were isolated from grass carp pituitary. Tissue distribution analysis indicated that both nERs and GPERs were highly detected in grass carp pituitary, which suggested that E2 should play an important role in grass carp pituitary. Using primary cultured grass carp pituitary cells as model, high-throughput RNA-seq was used to examine the E2-induced differentially expressed genes (DEGs). Transcriptomic analysis showed that E2 could significantly upregulate the expression of 28 genes in grass carp pituitary cells, which were characterized into different functions including reproduction, gonad development, and central nervous system development. Further studies confirmed that E2 could induce luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion and mRNA expression in prepubertal grass carp pituitary in vivo and in vitro. In the pituitary, LH and FSH regulation by E2 were mediated by both ER beta and GPER1. Apparently, E2-induced LH beta and FSH beta mRNA expression were mediated by adenylyl cyclase/cAMP/protein kinase A, phospholipase C/inositol 1,4,5-triphosphate/protein kinase C, and Ca2+/calmodulin/ CaM-dependent protein kinase II pathways. In addition to LH and FSH, E2 could also induce growth regulation by estrogen in breast cancer 1 (a novel regulator for pituitary development) mRNA expression in grass carp pituitary cells. These results, as a whole, suggested that E2 could play an important role in gonadotropin hormone release and pituitary development in prepubertal grass carp.