Objective: To reveal the molecular biological mechanism of a rare Dc-variant individual using PacBio third-generation sequencing technology. Methods: ABO and Rh blood type identification, DAT, unexpected antibody screening and D antigen enhancement test were conducted by serological testing. The absorption-elution test was used to detect the e antigen. RHCE gene typing was performed by PCR-SSP, and the 1-10 exons of RHCE were sequenced by Sanger sequencing. The full-length sequences of RHCE, RHD and RHAG were detected by PacBio third-generation sequencing technology. Results: Serological findings: Blood type O, Dc-phenotype, DAT negative, unexpected antibody screening negative; enhanced D antigen expression; no detection of e antigen in the absorption-elution test. PCR-SSP genotyping indicated the presence of only the RHCE c allele. Sanger sequencing results: Exons 5-9 of RHCE were deleted, exon 1 had a heterozygous mutation at c. 48G/C, and exon 2 had five heterozygous mutations at c. 150C/T, c. 178C/A, c. 201A/G, c. 203A/G and c. 307C/T. Third-generation sequencing results: RHCE genotype was RHCE 02N. 08/RHCE-D(5-9)-CE; RHD genotype was RHD 01/RHD 01; RHAG genotype was RHAG 01/RHAG 01 (c. 808G>A and c. 861G>A). Conclusion: This Dc-individual carries the allele RHCE 02N. 08 and the novel allele RHCE-D(5-9)-CE. The findings of this study provide data support and a theoretical basis for elucidating the molecular mechanisms underlying RhCE deficiency phenotypes.

Objective To investigate the distribution and antimicrobial resistance profiles of the common pathogens isolated from urine from 2015 to 2021 in the CHINET Antimicrobial Resistance Surveillance Program.Methods The bacterial strains were isolated from urine and identified routinely in 51 hospitals across China in the CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021.Antimicrobial susceptibility was determined by Kirby-Bauer method,automatic microbiological analysis system and E-test according to the unified protocol.Results A total of 261 893 nonduplicate strains were isolated from urine specimen from 2015 to 2021,of which gram-positive bacteria accounted for 23.8％(62 219/261 893),and gram-negative bacteria 76.2％(199 674/261 893).The most common species were E.coli(46.7％),E.faecium(10.4％),K.pneumoniae(9.8％),E.faecalis(8.7％),P.mirabilis(3.5％),P.aeruginosa(3.4％),SS.agalactiae(2.6％),and E.cloacae(2.1％).The strains were more frequently isolated from inpatients versus outpatients and emergency patients,from females versus males,and from adults versus children.The prevalence of ESBLs-producing strains in E.coli,K.pneumoniae and P.mirabilis was 53.2％,52.8％and 37.0％,respectively.The prevalence of carbapenem-resistant strains in E.coli,K.pneumoniae,P.aeruginosa and A.baumannii was 1.7％,18.5％,16.4％,and 40.3％,respectively.Lower than 10％of the E.faecalis isolates were resistant to ampicillin,nitrofurantoin,linezolid,vancomycin,teicoplanin and fosfomycin.More than 90％of the E.faecium isolates were ressitant to ampicillin,levofloxacin and erythromycin.The percentage of strains resistant to vancomycin,linezolid or teicoplanin was＜2％.The E.coli,K.pneumoniae,P.aeruginosa and A.baumannii strains isolated from ICU inpatients showed significantly higher resistance rates than the corresponding strains isolated from outpatients and non-ICU inpatients.Conclusions E.coli,Enterococcus and K.pneumoniae are the most common pathogens in urinary tract infection.The bacterial species and antimicrobial resistance of urinary isolates vary with different populations.More attention should be paid to antimicrobial resistance surveillance and reduce the irrational use of antimicrobial agents.

Objective To understand the distribution and changing resistance profiles of clinical isolates of Enterococcus in hospitals across China from 2015 to 2021.Methods Antimicrobial susceptibility testing was conducted for the clinical isolates of Enterococcus according to the unified protocol of CHINET program by automated systems,Kirby-Bauer method,or E-test strip.The results were interpreted according to the Clinical & Laboratory Standards Institute(CLSI)breakpoints in 2021.WHONET 5.6 software was used for statistical analysis.Results A total of 124 565 strains of Enterococcus were isolated during the 7-year period,mainly including Enterococcus faecalis(50.7％)and Enterococcus faecalis(41.5％).The strains were mainly isolated from urinary tract specimens(46.9％±2.6％),and primarily from the patients in the department of internal medicine,surgery and ICU.E.faecium and E.faecalis strains showed low level resistance rate to vancomycin,teicoplanin and linezolid(≤3.6％).The prevalence of vancomycin-resistant E.faecalis and E.faecium was 0.1％and 1.3％,respectively.The prevalence of linezolid-resistant E.faecalis increased from 0.7％in 2015 to 3.4％in 2021,while the prevalence of linezolid-resistant E.faecium was 0.3％.Conclusions The clinical isolates of Enterococcus were still highly susceptible to vancomycin,teicoplanin,and linezolid,evidenced by a low resistance rate.However,the prevalence of linezolid-resistant E.faecalis was increasing during the 7-year period.It is necessary to strengthen antimicrobial resistance surveillance to effectively identify the emergence of antibiotic-resistant bacteria and curb the spread of resistant pathogens.

Objective To examine the changing antimicrobial resistance profile of Enterobacter spp.isolates in 53 hospitals across China from 2015 t0 2021.Methods The clinical isolates of Enterobacter spp.were collected from 53 hospitals across China during 2015-2021 and tested for antimicrobial susceptibility using Kirby-Bauer method or automated testing systems according to the CHINET unified protocol.The results were interpreted according to the breakpoints issued by the Clinical & Laboratory Standards Institute(CLSI)in 2021(M100 31st edition)and analyzed with WHONET 5.6 software.Results A total of 37 966 Enterobacter strains were isolated from 2015 to 2021.The proportion of Enterobacter isolates among all clinical isolates showed a fluctuating trend over the 7-year period,overall 2.5％in all clinical isolates amd 5.7％in Enterobacterale strains.The most frequently isolated Enterobacter species was Enterobacter cloacae,accounting for 93.7％(35 571/37 966).The strains were mainly isolated from respiratory specimens(44.4±4.6)％,followed by secretions/pus(16.4±2.3)％and urine(16.0±0.9)％.The strains from respiratory samples decreased slightly,while those from sterile body fluids increased over the 7-year period.The Enterobacter strains were mainly isolated from inpatients(92.9％),and only(7.1±0.8)％of the strains were isolated from outpatients and emergency patients.The patients in surgical wards contributed the highest number of isolates(24.4±2.9)％compared to the inpatients in any other departement.Overall,≤ 7.9％of the E.cloacae strains were resistant to amikacin,tigecycline,polymyxin B,imipenem or meropenem,while ≤5.6％of the Enterobacter asburiae strains were resistant to these antimicrobial agents.E.asburiae showed higher resistance rate to polymyxin B than E.cloacae(19.7％vs 3.9％).Overall,≤8.1％of the Enterobacter gergoviae strains were resistant to tigecycline,amikacin,meropenem,or imipenem,while 10.5％of these strains were resistant to polycolistin B.The overall prevalence of carbapenem-resistant Enterobacter was 10.0％over the 7-year period,but showing an upward trend.The resistance profiles of Enterobacter isolates varied with the department from which they were isolated and whether the patient is an adult or a child.The prevalence of carbapenem-resistant E.cloacae was the highest in the E.cloacae isolates from ICU patients.Conclusions The results of the CHINET Antimicrobial Resistance Surveillance Program indicate that the proportion of Enterobacter strains in all clinical isolates fluctuates slightly over the 7-year period from 2015 to 2021.The Enterobacter strains showed increasing resistance to multiple antimicrobial drugs,especially carbapenems over the 7-year period.

Objective To understand the changing distribution and antimicrobial resistance profiles of Proteus,Morganella and Providencia in hospitals across China from January 1,2015 to December 31,2021 in the CHINET Antimicrobial Resistance Surveillance Program.Methods Antimicrobial susceptibility testing was carried out following the unified CHINET protocol.The results were interpreted in accordance with the breakpoints in the 2021 Clinical & Laboratory Standards Institute(CLSI)M100(31 st Edition).Results A total of 32 433 Enterobacterales strains were isolated during the 7-year period,including 24 160 strains of Proteus,6 704 strains of Morganella,and 1 569 strains of Providencia.The overall number of these Enterobacterales isolates increased significantly over the 7-year period.The top 3 specimen source of these strains were urine,lower respiratory tract specimens,and wound secretions.Proteus,Morganella,and Providencia isolates showed lower resistance rates to amikacin,meropenem,cefoxitin,cefepime,cefoperazone-sulbactam,and piperacillin-tazobactam.For most of the antibiotics tested,less than 10％of the Proteus and Morganella strains were resistant,while less than 20％of the Providencia strains were resistant.The prevalence of carbapenem-resistant Enterobacterales(CRE)was 1.4％in Proteus isolates,1.9％in Morganella isolates,and 15.6％in Providencia isolates.Conclusions The overall number of clinical isolates of Proteus,Morganella and Providencia increased significantly in the 7-year period from 2015 to 2021.The prevalence of CRE strains also increased.More attention should be paid to antimicrobial resistance surveillance and rational antibiotic use so as to prevent the emergence and increase of antimicrobial resistance.

Objective To understand the changing distribution and antimicrobial resistance profiles of Klebsiella strains in 52 hospitals across China in the CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021.Methods Antimicrobial susceptibility testing was carried out according to the unified CHINET protocol.The susceptibility results were interpreted according to the breakpoints in the Clinical & Laboratory Standards Institute(CLSI)M100 document.Results A total of 241,549 nonduplicate Klebsiella strains were isolated from 2015 to 2021,including Klebsiella pneumoniae(88.0％),Klebsiella aerogenes(5.8％),Klebsiella oxytoca(5.7％),and other Klebsiella species(0.6％).Klebsiella strains were mainly isolated from respiratory tract(48.49±5.32)％.Internal medicine(22.79±3.28)％,surgery(17.98±3.10)％,and ICU(14.03±1.39)％were the top 3 departments where Klebsiella strains were most frequently isolated.K.pneumoniae isolates showed higher resistance rate to most antimicrobial agents compared to other Klebsiella species.Klebsiella isolates maintained low resistance rates to tigecycline and polymyxin B.ESBLs-producing K.pneumoniae and K.oxytoca strains showed higher resistance rates to all the antimicrobial agents tested compared to the corresponding ESBLs-nonproducing strains.The K.pneumoniae and carbapenem-resistant K.pneumoniae(CRKP)strains isolated from ICU patients demonstrated higher resistance rates to majority of the antimicrobial agents tested than the strains isolated from non-ICU patients.The CRKP strains isolated from adult patients had higher resistance rates to most of the antimicrobial agents tested than the corresponding CRKP strains isolated from paediatric patients.Conclusions The prevalence of carbapenem-resistant strains in Klebsiella isolates increased greatly from 2015 to 2021.However,the Klebsiella isolates remained highly susceptible to tigecycline and polymyxin B.Antimicrobial resistance surveillance should still be strengthened for Klebsiella strains.

Objective To investigate the changing distribution and antibiotic resistance profiles of clinical isolates of Staphylococcus in hospitals across China from 2015 to 2021.Methods Antimicrobial susceptibility testing was conducted for the clinical isolates of Staphylococcus according to the unified protocol of CHINET(China Antimicrobial Surveillance Network)using disk diffusion method and commercial automated systems.The CHINET antimicrobial resistance surveillance data from 2015 to 2021 were interpreted according to the 2021 CLSI breakpoints and analyzed using WHONET 5.6.Results During the period from 2015 to 2021,a total of 204,771 nonduplicate strains of Staphylococcus were isolated,including 136,731(66.8％)strains of Staphylococcus aureus and 68,040(33.2％)strains of coagulase-negative Staphylococcus(CNS).The proportions of S.aureus isolates and CNS isolates did not show significant change.S.aureus strains were mainly isolated from respiratory specimens(38.9±5.1)％,wound,pus and secretions(33.6±4.2)％,and blood(11.9±1.5)％.The CNS strains were predominantly isolated from blood(73.6±4.2)％,cerebrospinal fluid(12.1±2.5)％,and pleural effusion and ascites(8.4±2.1)％.S.aureus strains were mainly isolated from the patients in ICU(17.0±7.3)％,outpatient and emergency(11.6±1.7)％,and department of surgery(11.2±0.9)％,whereas CNS strains were primarily isolated from the patients in ICU(32.2±9.7)％,outpatient and emergency(12.8±4.7)％,and department of internal medicine(11.2±1.9)％.The prevalence of methicillin-resistant strains was 32.9％in S.aureus(MRSA)and 74.1％in CNS(MRCNS).Over the 7-year period,the prevalence of MRSA decreased from 42.1％to 29.2％,and the prevalence of MRCNS decreased from 82.1％to 68.2％.MRSA showed higher resistance rates to all the antimicrobial agents tested except trimethoprim-sulfamethoxazole than methicillin-susceptible S.aureus(MSSA).Over the 7-year period,MRSA strains showed decreasing resistance rates to gentamicin,rifampicin,and levofloxacin,MRCNS showed decreasing resistance rates to gentamicin,erythromycin,rifampicin,and trimethoprim-sulfamethoxazole,but increasing resistance rate to levofloxacin.No vancomycin-resistant strains were detected.The prevalence of linezolid-resistant MRCNS increased from 0.2％to 2.3％over the 7-year period.Conclusions Staphylococcus remains the major pathogen among gram-positive bacteria.MRSA and MRCNS were still the principal antibiotic-resistant gram-positive bacteria.No S.aureus isolates were found resistant to vancomycin or linezolid,but linezolid-resistant strains have been detected in MRCNS isolates,which is an issue of concern.

AIM:To investigate the mechanism of curcumin inhibiting the choroidal neovascularization(CNV)of brown Norway(BN)rats.METHODS: CNV model of 36 BN rats was established through laser photocoagulation induction, and they were divided into 6 groups with 6 rats in each group. Normal group was fed normally with no intervention, while 532nm laser photocoagulation was used to establish a experimental CNV model in BN rats. Rats after modeling were respectively intervened for 14d and divided into model group, ranibizumab group, curcumin low [100mg/(kg·d)], medium [200mg/(kg·d)], and high [400mg/(kg·d)] dose group. The model group was given intragastric administration of saline for 14d, ranibizumab(10mg/mL, 0.2mL/dose)was injected at 2d after photocoagulation with 5μL once for rats in ranibizumab group, and different concentrations of curcumin were intragastrically administrated to the rats in low, medium and high groups for 14d. Fundus photography, fundus fluorescein angiography(FFA)and indocyanine green angiography(ICGA)examination were performed at 14d after photocoagulation. Ocular histopathological specimens of rats with CNV were made, and the central thickness of CNV were observed by HE staining. Ocular histopathological specimens were made, and the expressions of AKT/p-AKT/HIF-1α/VEGF signaling pathway-related proteins were observed by immunohistochemistry. The mRNA relative expressions of AKT/HIF-1α/VEGF factor in CNV tissues were detected by RT-qPCR, and the protein expressions of AKT/p-AKT/HIF-1α/VEGF factor in CNV tissues were detected by Western-blot.RESULTS: CNV generation rates in the model group, the ranibizumab group, and the low, medium and high-dose curcumin groups were 78.18%, 73.21%, 77.19%, 75.86%, 74.55%, respectively, which were higher than 70%. The average absorbance were 182.12±6.59, 119.22±8.03, 166.45±8.33, 164.34±5.69, 149.22±6.45, respectively; the ranibizumab group was significantly lower than the model group(P<0.05); the low-dose, medium-dose and high-dose groups were significantly higher than the ranibizumab group(P<0.05), and the curcumin high-dose group was significantly lower than the model group(P<0.05). HE staining showed that the retinal tissue structure of BN rats in normal group was clear and neatly arranged. The central thickness of CNV in the ranibizumab group was significantly reduced at 14d after photocoagulation compared with the model group(P<0.05); While the curcumin high-dose group was significantly reduced compared with the model group(P<0.05), but increased when compared with ranibizumab group(P<0.05). Immunohistochemistry results showed that AKT, p-AKT, HIF-1α, and VEGF factors were negatively expressed in the retinal tissue structure of BN rats in the normal group, and no brown-yellow reactants were found. The expression of AKT, p-AKT, HIF-1α, and VEGF factors in the model group were higher than that in the normal group at 14d after photocoagulation(P<0.05); the ranibizumab group was lower than the model group(P<0.05). While the expression of the curcumin high-dose group was significantly decreased compared with the model group(P<0.05), but significantly increased when compared with ranibizumab group(P<0.05). The mRNA results showed that the relative expression levels of AKT, HIF-1α and VEGF mRNA in the model group at 14d after photocoagulation were higher than those of the normal group(P<0.05); the ranibizumab group was lower than the model group(P<0.05). While curcumin high-dose group was significantly decreased compared with the model group(P<0.05), but significantly increased when compared with ranibizumab group(P<0.05). Western-blot results showed that there was no significant difference in the relative expression of AKT protein among each experimental groups at 14d after photocoagulation. The relative expression of p-AKT protein in the model group was significantly higher than that in the normal group(P<0.05); the ranibizumab group was significantly lower than the model group(P<0.05); the curcumin high-dose group was significantly lower than the model group(P<0.05). The relative expression levels of HIF-1α protein were significantly higher in the model group than in the normal group(P<0.05), and the ranibizumab group was lower than in the model group(P<0.05). The relative expression levels of HIF-1α protein was lower in the curcumin high-dose group than in the model group(P<0.05)but higher than ranibizumab group(P<0.05). The relative expression level of VEGF protein was significantly lower in the curcumin medium/high-dose group than in the model group(P<0.05).CONCLUSION: Curcumin at 400mg/(kg·d)has an inhibitory effect on CNV in BN rats. The mechanism may be closely related to inhibiting the activation of AKT/p-AKT/HIF-1α/VEGF signaling pathways.

Objective: To explore whether hsa_circ_0000670 promotes the progression of gastric cancer by regulating the miR-515-5p/SIX1 molecular axis. Methods: The gastric cancer and adjacent normal tissues of 35 gastric cancer patients admitted to Rugao Hospital Affiliated to Nantong University from 2014 to 2015 were collected. The expression levels of circ_0000670, miR-515-5p and Sine oculis homeobox 1 (SIX1) in gastric cancer tissues and cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The correlations between circ_0000670 and miR-515-5p, miR-515-5p and SIX1, circ_0000670 and SIX1 were analyzed by the Pearson method. Patients were divided into low circ_0000670 expression group (17 cases) and high circ_0000670 expression group (18 cases) based on the median of circ_0000670 expression level, and Kaplan-Meier was used to analyze the 5-year survival of patients. Cell proliferation was assessed via clone formation assay. Cell cycle and apoptosis were detected by flow cytometry. Wound healing and Transwell assays were used to detect cell migration and invasion ability. The targeting relationship between miR-515-5p and circ_0000670 or SIX1 was confirmed by the dual luciferase reporter assay. Nude mice were injected into HGC-27 cells transfected with sh-NC or sh-circ_0000670, and the volume and weight of the transplanted tumor were measured, also, the levels of circ_0000670, miR-515-5p and SIX1 in the transplanted tumor tissue were detected. Results: The expression levels of circ_0000670 and SIX1 in gastric cancer tissues and cell lines were significantly increased (P<0.05), while the expression levels of miR-515-5p were significantly decreased (P<0.05). The survival rate of patients in the low circ_0000670 expression group (82.4%) was significantly higher than that in the high circ_0000670 expression group (28.7%, P=0.034). Circ_0000670 was negatively correlated with miR-515-5p (r=-0.846, P<0.001), and miR-515-5p was negatively correlated with SIX1 (r=-0.615, P<0.001), but circ_0000670 was positively correlated with SIX1 (r=0.814, P<0.001). Transfection of si-circ_0000670 or miR-515-5p mimic could significantly reduce the number of clone-forming cells, migration distance, migration and invasion cells (P<0.05), and increase the ratio of G(0)/G(1) phase cells, apoptosis rate and the protein level of E-cadherin (P<0.05), decreased the proportion of S-phase cells and the protein level of Vimentin (P<0.05). The dual luciferase report assay confirmed that circ_0000670 could target miR-515-5p, and miR-515-5p could bind to SIX1. Co-transfection of si-circ_0000670 and miR-515-5p inhibitor could significantly attenuate the effects of si-circ_0000670 on cell proliferation, migration, invasion, cell cycle and apoptosis (P<0.05). Co-transfection of miR-515-5p mimic and pcDNA-SIX1 could significantly reduce the effects of miR-515-5p mimic on cell proliferation, migration, invasion, cell cycle and apoptosis (P<0.05). Compared with the sh-NC group [volume=(596.20±125.46) mm(3) and weight=(538.00±114.39) g], the volume and weight of transplanted tumors in the sh-circ_0000670 group [volume=(299.20±47.58) mm 3 and weight=(289.80±48.73 g)] were significantly reduced (P<0.05), the expression levels of circ_0000670 and SIX1 were significantly reduced (P<0.05), and the expression level of miR-515-5p was significantly increased (P<0.05). Conclusion: Knockdown of circ_0000670 could inhibit cell proliferation, migration, invasion of gastric cancer cells, induce cell cycle arrest in G(0)/G(1) phase and promote cell apoptosis by regulating the miR-515-5p/SIX1 axis.
Animals ; Mice ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Mice, Nude ; MicroRNAs/genetics* ; Stomach Neoplasms/genetics*

Objective: To characterize the incidence density of systematic lupus erythematosus (SLE) in Yinzhou District of Ningbo from 2016 to 2021, and compare the age and gender specific differences. Methods: A retrospective cohort study was conducted based on the related data from 2015 to 2021 collected from the Health Information Platform of Yinzhou. Suspected SLE cases in local residents were identified by fuzzy matching of International Classification of Diseases 10^th edition code "M32" or Chinese text "lupus". The classification criteria from Systemic Lupus International Collaboration Clinics-2012 and The European League Against Rheumatism/American College of Rheumatology-2019 were used for case verification. SLE cases were identified with specific algorithm based on verification results, and new cases were identified with 1 year as the washout period. The incidence density and 95%CI were estimated by Poisson distribution. Results: From 2016 to 2021, a total of 1 551 921 permanent residents were registered in Yinzhou, in whom 51.52% were women. The M(Q₁,Q₃) age at enrollment was 40.38 (27.54, 53.54) years. The M(Q₁,Q₃) of follow-up person-years was 3.83 (0.41, 5.83) years. There were 451 new SLE cases, in which 352 were women (78.05%). The 6-year incidence density was 8.14/100 000 person-years (95%CI: 7.41/100 000 person-years-8.93/100 000 person-years) for the total population, 3.68/100 000 person-years (95%CI: 2.99/100 000 person-years-4.48/100 000 person-years) for men and 12.37/100 000 person-years (95%CI: 11.11/100 000 person-years- 13.73/100 000 person-years) for women. The incidence density in men appeared a small peak at 20-29 years old, and began to increase with age from 40 years old. The incidence density in women was highest in age group 20-29 years (16.57/100 000 person-years) and remained to be high until 30-79 years old. The incidence density of SLE in Yinzhou show no significant temporal trend from 2016 to 2021 (men: P=0.848; women: P=1.000). Conclusions: The incidence density of SLE in Yinzhou from 2016 to 2021 was similar to those of other areas in China. SLE has a high incidence in women, especially in the young and elderly, suggesting that more attention should be paid to the diagnosis and treatment of SLE in women.
Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Young Adult ; Asian People ; Incidence ; Lupus Erythematosus, Systemic/diagnosis* ; Retrospective Studies ; China/epidemiology*