RTX toxin enhances the survival of Vibrio vulnificus during infection by protecting the organism from phagocytosis

Vibrio vulnificus is a marine bacterium causing serious septicemia and wound infection in humans. It produces an RTX toxin that can lyse a variety of cells and is important for virulence in mice. In this study, we explored the role of RTX in pathogenesis by characterizing an RTX-deficient mutant. This mutant showed an ～2-log reduction in virulence for mice infected by various routes. Survival of the mutant at the infection site and subsequent spread into the bloodstream were impaired. In mice pretreated with cyclophosphamide to deplete the neutrophils, both the virulence and survival at the infection site of this mutant were enhanced. This mutant was further shown to be more readily cleared from the macrophage-rich mouse peritoneal cavity and phagocytosed by murine macrophages. These findings suggest that the RTX of V. vulnificus is required for bacterial survival during infection by protecting the organism from phagocytosis.     

联合应用抗菌药物治疗创伤弧菌感染的实验研究

目的　探讨联合应用抗菌药物对创伤弧菌感染小鼠的治疗效果。方法　创伤弧菌 ( 6.0× 10 8cfu/ml,0 .6ml/10g小鼠 )经腹腔注射感染小鼠后 1h、2h、3h ,分别以人体常用治疗量的抗菌药物亚胺培南、氯霉素、多西环素 (灌胃 )、奈替米星、头孢哌酮、哌拉西林、左旋氧氟沙星等单独或联合用药 ,观察存活小鼠数及小鼠脏器的超微结构变化。结果　创伤弧菌感染早期上述 7种抗菌药物均有显著治疗作用 ,氯霉素、奈替米星、头孢哌酮、左旋氧氟沙星疗效最好 ,亚胺培南治疗效果较差。头孢哌酮 +左旋氧氟沙星、头孢哌酮 +奈替米星、奈替米星 +多西环素联用能显著增加治疗效果 ,小鼠主要脏器超微结构损伤恢复正常。结论　及早选用氯霉素、奈替米星、头孢哌酮、左旋氧氟沙星对创伤弧菌感染小鼠有较好的治疗效果 ,头孢哌酮与左旋氧氟沙星、头孢哌酮与奈替米星、奈替米星与多西环素联用治疗效果更好     

聚合酶链反应检测O1群霍乱弧菌肠毒素A亚单位基因

为了加强对霍乱的防治，探索对霍乱弧菌的快速和敏感的检测方法，建立了一种快速标本处理和聚合酶链反应扩增Ｏ１群霍乱弧菌肠毒素Ａ亚单位基因区的诊断方法。     

湘西地区女性泌尿生殖道沙眼衣原体感染状况及基因分型初步研究

目的对湘西地区女性泌尿生殖道沙眼衣原体的感染状况及基因分型进行初步研究。方法利用单克隆抗体直接免疫荧光法（DFA）对100例宫颈病变患者及77位健康女性进行筛查,对沙眼衣原体感染者的宫颈管内拭子标本直接进行套式PCR,扩增主要外膜蛋白基因（Omp1）第4可变区（VS4）,对将扩增的Omp1VS4基因片段进行测序、分型,并进行同源性分析。结果DFA检测177名妇女的宫颈拭子标本,36份沙眼衣原体阳性,29份阳性标本扩增出277bp目的DNA。经基因测序,共检出8个基因型,其中E型9株（31.0%）、F型7株（24.1%）、G型5株（17.2%）、D型3株（10.3%）、H型2株（6.9%）、J型1株（3.5%）、Ba型1株（3.5%）、K型1株（3.5%）,4例存在Omp1基因变异。结论湘西地区女性存在生殖道沙眼衣原体感染,利用对基因的多态性分析可以对沙眼衣原体感染流行情况作出分析。     

大劣按蚊含硫脂蛋白基因在约氏疟原虫感染中的作用

目的分析大劣按蚊含硫脂蛋白(TEP1)基因在约氏疟原虫感染过程中的作用。方法根据GenBank中大劣按蚊TEP1基因序列设计带有T7启动子的引物,以大劣按蚊cDNA为模板,进行PCR扩增,纯化产物,体外转录试剂盒合成AdTEP1双链RNA。羽化1～2日的雌性大劣按蚊分3组(200只/组):TEP1干扰组、绿色荧光蛋白(EGFP)干扰组和对照组。TEP1、EGFP干扰组按蚊分别进行胸部微量注射AdTEP1双链RNA、EGFP双链RNA各147 ng,对照组未处理。干扰后3d,每组取15只按蚊,去头,以大劣按蚊核糖蛋白S7(AdS7)为内参进行半定量PCR,检测干扰效果。干扰后4d,用约氏疟原虫BY256荧光株感染3组大劣按蚊。感染后9d,每组各解剖25只蚊胃,记录感染率和感染度。结果对照组和EGFP干扰组AdTEP1的表达条带亮度基本一致,而TEP1干扰组AdTEP1的表达条带亮度非常微弱。感染后9 d,蚊胃卵囊数统计学分析表明,对照组、EGFP干扰组和TEP1干扰组感染率分别为(24±2.83)%、(24±0.71)%和(80±3.54)%;感染度分别为0.32±0.7、0.44±0.85和5.52±4.84...     

Resiniferatoxin modulates the Th1 immune response and protects the host during intestinal nematode infection

In the early stage of the intestinal phase of Trichinella spiralis infection, the host triggers a Th1‐type immune response with the aim of eliminating the parasite. However, this response damages the host which favours the survival of the parasite. In the search for novel pharmacological strategies that inhibit the Th1 immune response and assist the host against T. spiralis infection, a recent study showed that resiniferatoxin had anti‐inflammatory activity contributed to the host in T. spiralis infection. In this study, we evaluated whether RTX modulates the host immune response through the inhibition of Th1 cytokines in the intestinal phase. In addition, it was determined whether the treatment with RTX affects the infectivity of T. spiralis‐L1 and the development of the T. spiralis life cycle. Our results show that RTX decreased serum levels of IL‐12, INF‐γ, IL‐1β, TNF‐α and parasite burden on muscle tissue. It was observed that T. spiralis‐L1 treated with RTX decreased their infectivity affecting the development of the T. spiralis life cycle in mouse. These results demonstrate that RTX is able to inhibit the production of Th1 cytokines, contributing to the defence against T. spiralis, which places it as a potential drug modulator of the immune response.     

Identification of genetic variants of the IL‐22 gene in association with an altered risk of COPD susceptibility

The incidence of chronic obstructive pulmonary disease (COPD) is related to the interaction between environmental exposure and genetic factors. Far more than 15% of smokers eventually develop COPD. In addition to smoking, genetic susceptibility may be another factor in the development of COPD. IL‐22 and its receptors are increased in human and experimental COPD and contribute to pathogenesis. Here, we conducted a case–control study to evaluate the association between IL‐22 tag‐single nucleotide polymorphisms (SNPs) and COPD risk. Four tag‐SNPs (rs2227478, rs2227481, rs2227484 and rs2227485) were identified according to linkage disequilibrium (LD) analysis in 30 healthy controls. A total of 513 COPD cases and 504 controls were recruited to perform an association study between these four tag‐SNPs and COPD risk. We found that the “C” allele of rs2227478T>C and the “T” allele of rs2227481C>T were obviously related to decreased COPD susceptibility. Genetic model analysis showed that rs2227478T>C and rs2227481C>T were significantly associated with a decreased risk of COPD under dominant models after adjusting for the above factors. In the recessive model, rs2227485T>C was obviously associated with decreased COPD risk. Our data showed that only rs2227485T>C was associated with a decreased COPD risk after Bonferroni correction. The eQTL analysis showed that rs2227485T>C was significantly associated with IL‐22 expression. The pGL4‐rs2227485‐C gene reporter had a higher promoter activity than pGL4‐rs2227485‐T. In our study, rs2227485T>C, located in the promoter region of IL‐22, was associated with a decreased risk of COPD and increased IL‐22 promoter activity, suggesting that this variant might modulate COPD susceptibility.     

1例早期HIV感染者体内病毒准种的各基因片段分析

目的比较1例早期艾滋病病毒（HIV）感染者体内病毒不同基因片段准种复杂度。方法使用套式聚合酶链反应（Nested-PCR）,扩增HIV感染者外周血淋巴细胞中病毒的gag、pol、vif-vpr和env区的部分基因,通过克隆、测序方法观察该感染者体内病毒准种分布情况,比较不同基因片段检测病毒准种的可靠性。结果不同基因片段检测病毒准种结果显示,该早期感染者体内的病毒不同基因片段显示的准种基因离散率不同,其中gag区显示出的病毒准种复杂度最高,反映为该区段病毒准种的基因离散率最高,为0．008;其他各区基因（pol、vif-vpr、env）的离散率依次为0．006、0．001、0．000。结论该HIV感染者体内病毒准种的基因中,gag区基因的突变率较高,提示Gag在机体感染HIV早期承受的免疫压力较高,因此在病毒准种研究中,gag区应该作为重要的基因区域。     

Identified OAS3 gene variants associated with coexistence of HBsAg and anti‐HBs in chronic HBV infection

The underlying mechanism of coexistence of hepatitis B surface antigen (HBsAg) and hepatitis B surface antigen antibody (anti‐HBs) is still controversial. To identify the host genetic factors related to this unusual clinical phenomenon, a two‐stage study was conducted in the Chinese Han population. In the first stage, we performed a case‐control (1:1) age‐ and gender‐matched study of 101 cases with concurrent HBsAg and anti‐HBs and 102 controls with negative HBsAg and positive anti‐HBs using whole exome sequencing. In the second validation stage, we directly sequence the 16 exons on the OAS3 gene in two dependent cohorts of 48 cases and 200 controls. Although, in the first stage, a genome‐wide association study of 58,563 polymorphism variants in 101 cases and 102 controls found no significant loci (P‐value ≤ .05/58563), and neither locus achieved a conservative genome‐wide significance threshold (P‐value ≤ 5e‐08), gene‐based burden analysis showed that OAS3 gene rare variants were associated with the coexistence of HBsAg and anti‐HBs. (P‐value = 4.127e‐06 ≤ 0.05/6994). A total of 16 rare variants were screened out from 21 cases and 3 controls. In the second validation stage, one case with a stop‐gained rare variant was identified. Fisher’s exact test of all 149 cases and 302 controls showed that the rare coding sequence mutations were more frequent in cases vs controls (P‐value = 7.299e‐09, OR = 17.27, 95% CI [5.01‐58.72]). Protein‐coding rare variations on the OAS3 gene are associated with the coexistence of HBsAg and anti‐HBs in patients with chronic HBV infection in Chinese Han population.     

HMGB1 gene polymorphisms in patients with chronic hepatitis B virus infection

AIM: To characterize high mobility group box chromosomal protein 1(HMGB1) polymorphisms in patients infected with hepatitis B virus(HBV) and determine the different patterns in patient subgroups.METHODS: A total of 1495 unrelated Han Chinese HBV carriers were recruited in this hospital-based casecontrol study.The HMGB1 1176 G/C polymorphism was genotyped by polymerase chain reaction-restriction fragment length polymorphism assay.RESULTS: A significant association was observed between HMGB1 1176 G/C polymorphism and outcome of HBV infection.The subjects bearing 1176G/G genotype had an increased risk of susceptibility to chronic hepatitis B,liver cirrhosis and severe hepatitis B when compared with those bearing at least one 1176C allele.CONCLUSION: Patients with 1176G/G genotype of HMGB1 gene are more likely to have a progressive status in HBV infection.     

Association testing of common variants in the insulin receptor substrate-1 gene (IRS1) with type 2 diabetes

Aims/hypothesis Activation of the insulin receptor substrate-1 (IRS1) is a key initial step in the insulin signalling pathway. Despite several reports of association of the G972R polymorphism in its gene IRS1 with type 2 diabetes, we and others have not observed this association in well-powered samples. However, other nearby variants might account for the putative association signal. Subjects and methods We characterised the haplotype map of IRS1 and selected 20 markers designed to capture common variations in the region. We genotyped this comprehensive set of markers in several family-based and case-control samples of European descent totalling 12,129 subjects. Results In an initial sample of 2,235 North American and Polish case-control pairs, the minor allele of the rs934167 polymorphism showed nominal evidence of association with type 2 diabetes (odds ratio [OR] 1.25, 95% CI 1.03–1.51, p = 0.03). This association showed a trend in the same direction in 7,659 Scandinavian samples (OR 1.16, 95% CI 0.96–1.39, p = 0.059). The combined OR was 1.20 (p = 0.008), but statistical correction for the number of variants examined yielded a p value of 0.086. We detected no differences across rs934167 genotypes in insulin-related quantitative traits. Conclusions/interpretation Our data do not support an association of common variants in IRS1 with type 2 diabetes in populations of European descent. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorised users. D. Altshuler and L. Groop jointly supervised this project.     

Associations of SRD5A1 gene variants and testosterone with dysglycemia: Henan Rural Cohort study

Background and aimMultiple studies support a complex relationship between testosterone and type 2 diabetes mellitus (T2DM) and the transformation of testosterone is affected by several reductases. Thus, we aimed to explore the associations of steroid-5α-reductase type 1 (SRD5A1) gene polymorphism with impaired fasting glucose (IFG) and T2DM and the interactive effects of testosterone and genotypes on glycometabolism.Methods and resultsA case–control study including 2365 participants was performed. Genomic DNA was extracted from the whole blood and genotyped for the SRD5A1 single nucleotide polymorphisms (SNP) rs1691053. Multivariable logistic regression and linear regression were performed to estimate the associations of SRD5A1 rs1691053 alleles and genotypes with glycometabolism. Generalized linear models were used to investigate the modulatory effects of serum testosterone on glycometabolism indexes in males.After multivariable adjustment, the odds ratio (OR) of homozygous CC genotypes in male carriers was 2.62 (95%CI: 1.11–6.18) for IFG. Furthermore, significant associations of SRD5A1 rs1691053 polymorphisms with adverse indices of glycometabolism were observed in males. Interestingly, the opposite associations in females were observed. The interactive associations of SNP and testosterone were found and mutations were more likely to lead unfavorable metabolic phenotypes.ConclusionThese results showed that SRD5A1 rs1691053 gene polymorphism was independently associated with glycometabolism. The interaction between a genetic polymorphism from SRD5A1 and testosterone involved glycometabolism was identified in males. Although this preliminary data should be replicated with other rigorous researches, it highlighted the importance of the SNP-testosterone interaction over the present of glycometabolism.     

Two variants located in the upstream enhancer region of human UCP1 gene affect gene expression and are correlated with human longevity

The brown fat specific UnCoupling Protein 1 (UCP1) is involved in thermogenesis, a process by which energy is dissipated as heat in response to cold stress and excess of caloric intake. Thermogenesis has potential implications for body mass control and cellular fat metabolism. In fact, in humans, the variability of the UCP1 gene is associated with obesity, fat gain and metabolism. Since regulation of metabolism is one of the key-pathways in lifespan extension, we tested the possible effects of UCP1 variability on survival.Two polymorphisms (A-3826G and C-3740A), falling in the upstream promoter region of UCP1, were analyzed in a sample of 910 subjects from southern Italy (475 women and 435 men; age range 40–109). By analyzing haplotype specific survival functions we found that the A-C haplotype favors survival in the elderly. Consistently, transfection experiments showed that the luciferase activity of the construct containing the A-C haplotype was significantly higher than that containing the G-A haplotype. Interestingly, the different UCP1 haplotypes responded differently to hormonal stimuli. The results we present suggest a correlation between the activity of UCP1 and human survival, indicating once again the intricacy of mechanisms involved in energy production, storage and consumption as the key to understanding human aging and longevity.     

ABCA1基因多态性与冠心病易感性的关联研究

目的探讨三磷酸腺苷结合盒转运子A1（ATP binding cassette transporter1,ABCA1）基因R219K及M883I单核苷酸多态性（SNP）位点与脂代谢和冠心病（coronary heart disease,CHD）易感性的关系。方法以医院为基础的病例-对照研究。经冠状动脉造影确诊的冠心病病例224例,同一地区正常对照248例。分别以PCR-RFLP和PIRA-PCR对ABCA1第219密码子G→A（Arg219Lys）和第883G→A（Met883Ile）密码子多态进行检测,比较不同基因型与个体血脂水平和冠心病患病风险的关系。结果吸烟、高血压和高血糖是冠心病的独立危险因素。与携带219RR基因型者比较,携带至少1个219K等位基因者（即RK和KK基因型）冠心病患病风险显著降低59％（OR = 0．41,95％ CI = 0．27～0．61）。而在883位点,II基因型携带者患冠心病率较低（OR = 0．54,95％ CI = 0．26～1．11）。而两位点联合作用分析发现与携带219RR,883MM或883MI基因型者相比较,携带其他组合基因型的个体冠心病患病风险降低61％（OR = 0．39,95％ CI = 0．26～0．60）。另外,对照组中携带219K等位基因者血清HDL-C水平显著高于219K非携带者（P = 0．037）,提示Arg219Lys位点的多态改变主要通过改变HDL-C水平影响个体冠心病的患病风险。结论ABCA1 R219K可能与中国汉族人群冠心病遗传易感性有关,血清高密度脂蛋白可能是其作用靶点。