Microbial communities such as those residing in the human gut are highly diverse and complex, and many with important implications for health and diseases. The effects and functions of these microbial communities are determined not only by their species compositions and diversities but also by the dynamic intra- and inter-cellular states at the transcriptional level. Powerful and scalable technologies capable of acquiring single-microbe-resolution RNA sequencing information in order to achieve a comprehensive understanding of complex microbial communities together with their hosts are therefore utterly needed. Here we report the development and utilization of a droplet-based smRNA-seq (single-microbe RNA sequencing) method capable of identifying large species varieties in human samples, which we name smRandom-seq2. Together with a triple-module computational pipeline designed for the bacteria and bacteriophage sequencing data by smRandom-seq2 in four human gut samples, we established a single-cell level bacterial transcriptional landscape of human gut microbiome, which included 29,742 single microbes and 329 unique species. Distinct adaptive response states among species in Prevotella and Roseburia genera and intrinsic adaptive strategy heterogeneity in Phascolarctobacterium succinatutens were uncovered. Additionally, we identified hundreds of novel host-phage transcriptional activity associations in the human gut microbiome. Our results indicated that smRandom-seq2 is a high-throughput and high-resolution smRNA-seq technique that is highly adaptable to complex microbial communities in real-world situations and promises new perspectives in the understanding of human microbiomes.
Humans ; Gastrointestinal Microbiome/genetics* ; Bacteriophages/physiology* ; High-Throughput Nucleotide Sequencing ; Sequence Analysis, RNA/methods* ; Bacteria/virology*

BACKGROUND:The increase in multi-drug resistant bacterial infections has become a major problem in modern healthcare due to the development of bacterial resistance to antibiotics and the development of new antibacterial alternative drug materials is of great importance. OBJECTIVE:To synthesize and perform a series of characterization of a CeO2 nanoenzyme to investigate its biocompatibility and antibacterial properties against Escherichia coli. METHODS:CeO2 nanoenzymes were synthesized using a hydrothermal method.The morphology,product composition,and chemical composition were analyzed using characterization methods such as X-ray diffraction,X-ray photoelectron spectroscopy,Fourier infrared analysis,Raman spectroscopy,scanning electron microscopy,and transmission electron microscopy.The peroxide-mimetic enzyme activity of CeO2 nanoenzymes was characterized using TMB color development assay.The toxic effect of CeO2 nanoenzymes at different concentrations(10,25,and 50 μg/mL)on mouse fibroblast L929 cells was evaluated using the CCK-8 assay.The antibacterial properties of CeO2 nanoenzymes against Escherichia coli under different conditions were evaluated using the plate coating method.Changes in intra-bacterial reactive oxygen species after treatment with different conditions were detected using a reactive oxygen species detection kit. RESULTS AND CONCLUSION:(1)The morphology of the synthesized CeO2 nanoparticles was rod-shaped,with Ce3+ accounting for 29.87%of the total Ce3+/Ce4+ and an average grain size of 7.4 nm.In a slightly acidic environment containing TMB and pH=5.5,CeO2 nanoenzymes mixed with H2O2 showed excellent peroxidase activity,but did not show peroxidase simulated activity at pH=7.4.(2)There was no statistically significant difference in the toxic effects of CeO2 nanoparticles at various mass concentrations on mouse fibroblast L929 cells.(3)In a slightly acidic environment at pH 5.5,Escherichia coli was inhibited to a certain extent in the presence of CeO2 nanoenzyme alone at a concentration of 10 μg/mL,with a decrease in CFU results of about 0.5 log(P＜0.01);in a slightly acidic environment containing 50 μmol/L H2O2,CeO2 nanoenzyme showed excellent antibacterial effects against Escherichia coli,with a decrease in Escherichia coli CFU results of by about 1.5 log(P＜0.001).After CeO2 nanoenzymes interacted with Escherichia coli,the level of reactive oxygen species in Escherichia coli increased(P＜0.05);after CeO2 nanoenzymes interacted with Escherichia coli together with H2O2,the level of reactive oxygen species in Escherichia coli increased significantly(P＜0.001).(4)The results show that the CeO2 nanoenzymes have good biocompatibility,are inherently antibacterial,and can exhibit peroxidase activity in a slightly acidic environment containing low concentrations of H2O2,and generate reactive oxygen species to kill bacteria,thus showing excellent antibacterial effects.

Objective To investigate the effect of dioscin on uric acid(UA)-induced oxidative stress injury of human renal tubular epithelial cells(HK-2)and its molecular mechanism.Methods HK-2 cells were cultured and divided into four groups:blank group(normal group),model group(uric acid-stimulation modeling),condition control group(UA+DMSO)and dioscin group(UA+dioscin).Oxidative stress injury model was induced by UA in HK-2 cells.Cells viability was detected by CCK-8.ROS level was detected by flow cytometry.Real-time PCR was used to detect the expressions of glycogen synthase kinase 3β(GSK3β),nuclear factor erythroid 2-related factor 2(Nrf2)and heme oxygenase 1(HO-1)at mRNA level,and Western Blot was used to detect the expressions of phosphorylated glycogen synthesis kinase 3β(p-GSK3β),GSK3β,Nrf2 and HO-1 at protein level.Results After stimulation by UA,HK-2 cells viability was obviously decreased,and ROS level was significantly increased(all P＜0.001).When treated with dioscin,HK-2 cells viability was obviously increased,and the ROS level of HK-2 cells was significantly decreased(all P＜0.001).The expressions of Nrf2 and HO-1 decreased at the protein and mRNA levels after stimulation with UA.But the expressions of Nrf2 and HO-1 significantly increased after treated with dioscin(all P＜0.001).Compared with the blank group,the p-GSK3β/GSK3β ratio in the model group decreased significantly at the protein level,but the p-GSK3β/GSK3β ratio increased after treated with dioscin(all P＜0.001).Conclusion Dioscin can alleviate UA-induced oxidative stress injury in HK-2 cells.The mechanism might be that dioscin can promote phosphorylation of GSK3β,and activate Nrf2/HO-1 pathway.

Objective To investigate the expression levels of serum G protein-coupled receptor-associated sorting protein 1(GASP-1),lipocalin-2(LCN2)and targeting protein for Xenopus kinesin-like protein 2(TPX2)in lung cancer patients,and their correlations with tumor-node-metastasis classification(TNM)pathological stage and prognosis.Methods A total of 92 patients with lung cancer who were admitted to General Hospital of Southern Theatre Command from April 2019 to May 2022 were selected as observation group.Meanwhile,92 healthy subjects were selected as control group.Peripheral venous blood samples of the two groups were collected to detect serum GASP-1,LCN2 and TPX2 levels.Serum GASP-1,LCN2 and TPX2 levels of lung cancer patients with different TNM stages were analyzed.According to the prognosis,the patients were divided into good prognosis group and poor prognosis group.Serum GASP-1,LCN2 and TPX2 levels were compared between the two groups.The correlations of serum GASP-1,LCN2 and TPX2 levels with TNM stage and prognosis in patients with lung cancer were investigated.Results The serum GASP-1,LCN2 and TPX2 levels in the observation group were significantly higher than those in the control group(P＜0.05).The serum levels of GASP-1,LCN2 and TPX2 in patients with TNM stage Ⅲ and Ⅳ were significantly higher than those in patients with TNM stage Ⅰand Ⅱ(P＜0.05).The levels of serum GASP-1,LCN2 and TPX2 in patients with TNM stage Ⅳ were significantly higher than those in patients with TNM stage Ⅲ(P＜0.05).Serum GASP-1,LCN2 and TPX2 levels in the good prognosis group were significantly lower than those in the poor prognosis group(P＜0.05).Serum GASP-1,LCN2 and TPX2 levels were positively correlated with TNM stage before treatment(P＜0.05),but were negatively correlated with the prognosis after treatment(P＜0.05).Conclusion Serum levels of GASP-1,LCN2 and TPX2 may be related to the TNM pathological stages of lung cancer patients.The detection of the above serum indicators after treatment can reflect the prognosis of patients to a certain extent,and provide reference for clinical practice.

Objective To explore the features of frontotemporal lobes'resting-state functional connectivity(rsFC)in preschool children with autism spectrum disorders(ASD)based on functional near-infrared spectroscopy(fNIRS)and to explore the possible neurological markers for early identification of ASD.Methods Sixty-three preschool ASD children and 72 typical development(TD)children were enrolled.Selected bilateral dorsolateral prefrontal cortex(DLPFC),bilateral premotor cortex(PMC),and bilateral temporal lobe(TL)cortex as the regions of interest(ROI).Changes of Oxyhemoglobin in the 6 ROIs in resting-state were measured by using functional near-infrared spectroscopy(fNIRS).Compared the frontotemporal rsFC strength and calculate the laterality index(LI)between two groups.Results Compared with the TD group,rsFC strength was significantly lower in the ASD group(P<0.05),and the differences existed mainly within the left ROIs(0.21±0.11 vs.0.32±0.18),right ROIs(0.16±0.16 vs.0.30±0.14),bilateral DLPFCs(0.20±0.14 vs.0.39±0.17;0.15±0.13 vs.0.36±0.13),bilateral TLs(0.15±0.14 vs.0.28±0.17;0.14±0.15 vs.0.31±0.17),and between the 10 groups of ROIs-ROIs(including right DLPFC-left DLPFC,right DLPFC-right PMC,right DLPFC-left PMC,right DLPFC-right TL,right DLPFC-left TL,left DLPFC-right PMC,left DLPFC-left PMC,left DLPFC-right TL,left DLPFC-left TL,right TL-left TL).There were a significant differences in the rsFC's laterality index of DLPFC and whole-brain between the two groups(t=2.002,P=0.047;t=3.003,P=0.003),and the ASD group showed left-lateralized connectivity.Conclusion Frontotemporal lobe's resting-state functional connectivity is abnormal in preschool children with ASD,characterized by low short-range functional connectivity of bilateral DLPFCs and TLs,low long-range functional connectivity associated with DLPFCs,and left-lateralized connectivity.

Objective:To study the clinical characteristics and compliance of early-onset gout patients by case-control analysis.Methods:A total of 111 early-onset patients (onset age ≤35 years old) were included as Group A, and 111 non-early-onset patients (onset age >35 years old) with matched disease durationwere included as Group B. The differences ofclinical characteristics, causes of acute gout attack, dairy diet habits, compliance, and misunderstanding of the disease were compared.Results:Compared with the non-early-onsetgoutpatients, the early-onset patients had a higher proportion of obesity (63 cases vs 28 cases), family history (36 cases vs 20 cases) and tophus (39 cases vs 23 cases) and higher level of VAS scores (8.5±1.3 vs 7.6±1.7; χ2=22.988, P<0.01; χ2=5.749, P=0.016; χ2=5.729, P=0.017; t=4.639, P<0.01), lowerproportionof the first metatarsophalangeal joint involvement as the initial joint involvement (45.9%, 51 cases vs 59.4%, 66 cases; χ2=4.066, P=0.044), higher proportion of the ankle involvement as the initial joint involvement (34.2%, 38 cases vs 21.6%, 24 cases; χ2=4.386, P=0.036), higher proportion of alcohol drinkers and high fructose drinkers, which was more likely to relate to alcohol intake, strenuous exercise and high fructose intakeas trigger of the flare ( χ2=6.513, P=0.011; χ2=7.126, P=0.008; χ2=1.978, P=0.160), while the proportion of regular exercisers and on diet in the family was lower ( χ2=22.887, P<0.01; t=-4.917, P<0.01). The proportion of poor diet and medication compliance in Group A was higher than that in Group B(57.7%, 64 cases vs 38.7%, 43 cases; χ2=5.207, P=0.022; χ2=5.867, P=0.015). As for the reason for poor treatment compliance, early-onset gout patients were more worry about the side-effects of drugs than non-early onset patients ( χ2=4.190, P=0.041). There was no significant difference between the two groups in the main misunderstanding of gout. Conclusion:Although early onset gout patients are young, their condition is more serious, and compliance is poorer, this group of patients should be highly valued in clinical diagnosis and treatment.

Objective:To identify the pathogenic gene mutations in a family with Leber congenital amaurosis (LCA).Methods:In October 2018, 1 patient and 3 normal family members from a LCA family was enrolled in this retrospective study. Detailed medical history of proband was obtained and fixation test, cycloplegic refraction, slit-lamp, fundus color photography and full-field ERG were performed. And other family members underwent BCVA, refraction slit-lamp, fundus biomicroscopy with the slit lamp, fundus color photography and full-field ERG. The family was investigated with a specific hereditary eye disease enrichment panel which contained 441 known pathogenic genes and based on targeted exome capture technology first to indentify the potential pathogenic genes and mutations. Then the potential pathogenic mutations were conformed by Sanger sequencing. Finally, the results were analyzed via bioinformatics analysis.Results:The proband showed no trace object from childhood, but had obvious photophobia and nystagmus. No positive changes were found in the anterior segment, vitreous and retina in both eyes. Both cone and rod system function decreased significantly in full-field ERG in both eyes. Gene tests showed the proband carried both RPGRIP1 c.1635dupA and c.3565C> T, which composited a heterozygous mutation. Bioinformatics analysis showed RPGRIP1 c.1635dupA was a pathogenic mutation, and RPGRIP1 c.3565C> T which was a novel potential pathogenic mutation in LCA.Conclusion:The compound heterozygous mutation, c.1635dupA and c.3565C> T in RPGRIP1 may be responsible for the pathogenesis in this Chinese Han LCA pedigree.

Objective To analyze systematically the existing classification criteria and assessment tools for osteoarthritis (OA).Methods Comprehensively searched and screened the available classification criteria and assessment tools reported in OA guidelines,textbooks,including secondary and original researchs.We collected and summarized the extracted data with the methods of scoping review and also used Excel software for qualitative analysis.Results A total of 63 OA guidelines,1 textbook,239 secondary or original researches,160 supplementary records were retrieved.The 5 classification criteria and 15 systematic reviews of assessment tools (855 assessment tools) were finally included.Conclusion The existing classification criteria lack a rigorous and transparent development process,and they are also too complicate to guide clinical treatment.We suggest that the development and improvement of OA classification criteria should be linked with the streamlined assessment tools,and conduct trials to test in clinical practice.

AIM: To investigate the effect of sitagliptin on the autopaghy and the expression of extracellular matrix in mesangial cells induced by advanced glycation end products (AGEs).METHODS: The cells were divided into 5 groups: control group, AGE group, and sitagliptin (5, 10 and 20 μmol/L) groups.After 48 h, the cell viability was measured by MTT assay, and the content of collagen (Col) Ⅳ in the supernatant of the cell culture was detected by ELISA.The protein levels of beclin-1, adenosine monophosphate-activated protein kinase (AMPK), p-AMPK, p70S6K and p-p70S6K were determined by Western blot.RESULTS: Compared with control group, the viability and the expression of Col IV induced by AGEs in the cultured mesangial cells were significantly increased (P<0.01).Sitagliptin decreased the viability and the expression of Col IV induced by AGEs in the mesangial cells in a dose-dependent manner.AGEs significantly inhibited the protein levels of beclin-1 and p-AMPK, but significantly increased the protein level of p-p70S6K.Compared with AGE group, sitagliptin significantly reversed the above results in a dose-dependent manner.CONCLUSION: Autophagy may mediate the protective effect of sitagliptin on mesangial cells induced by AGEs.

Objective To investigate the expression of interleukin-35 (IL-35) in serum of mice with pulmonary interstitial fibrosis.Methods Thirty-six C57BL/6 mice were divided into three groups (twelve mice in each group):control group,model group of mice with bleomycin-induced pulmonary fibrosis,glucocorticoid treatment group of mice with bleomycin-induced pulmonary fibrosis.The mice were sacrificed at day 7,day 14 and day 28 respectively,and the serum concentration of IL-35 was assayed.Statistical product and service solutions (SPSS) 17.0 statistical software was used for single factor analysis of variance and LSD-t comparison and Pearson correlation analysis was used for the comparison between each two groups.Results The serum IL-35 concentrations between groups and within groups at the time of day 7,day 14 and day 28 were compared respectively.The serum IL-35 concentration of the model group was significantly lower than the control group and the glucocorticoid treatment group at the time of day 7 (F=24.56,P＜0.05).The serum IL-35 concentration of glucocorticoid treatment group was significantly lower than the control group at the time of day 14 (F=8.85,P＜0.05),and which of glucocorticoid treatment group was significantly lower than the control group and the model group at the time of day 28 (F=36.64,P＜0.05).There was no significant difference between days 28 and day 7 within control group (t=1.03,P＞0.05).The serum IL-35 concentration of the model group at the time of day 28 was significantly higher than those of day 7 [(9.36±0.95) ng/ml vs (6.51±0.58) ng/ml,t=5.14,P＜0.05],and which of glucocorticoid treatment group was significantly lower than those of day 7 [(5.27±1.01) ng/ml vs (9.42±0.84) ng/ml,t=6.32,P＜0.05].From day 7 to day 28 the serum IL-35 concentration change of the model group and glucocorticoid treatment group showed significantly negative correlation (r =-0.743,P＜0.05).Conclusion Serum IL-35 concentration has shown an trend of increase during bleomycin-induced pulmonary fibrosis with mice model,and early glucocorticoid treatment can decrease the serum IL-35 in the model mice.