几种级别动物的膜菌群分析和比较

在我们的研究中,分析和比较了普通大鼠、SPF大鼠及悉生大鼠的盲肠膜菌群和内容物菌群中的5类菌,其中悉生大鼠的盲肠内容物中肠杆菌数为8.09±0.26,肠球菌数为7.59±0.26,类杆菌数为10.1±0.34,乳杆菌数为9.62±0.58,双歧杆菌数为10.32±0.18;而普通大鼠盲肠内容物的5类菌分别为5.79±0.52、6.13±0.84、8.76±0.5、9.74±0.68和10.10±0.78,其中肠杆菌和类杆菌数差异显著(P<0.01)。SPF大鼠盲肠内容物中5类菌分别为5.29±、3.47±0.93、8.48±0.71、8.40±0.48和10.06±0.44,与悉生大鼠相比肠杆菌及肠球菌差异显著。     

鲟鳇鱼网箱养殖环境微生物菌群结构及潜在病原菌分析

为探究鲟鳇鱼网箱养殖环境中的细菌群落结构组成和潜在致病菌的种类与数量,文章用高通量测序对中华鲟Acipenser sinensis、施氏鲟Acipenser schrenckii、达氏鳇Huso dauricus、大杂交鲟A. schrenckii♂×H.dauricus♀等网箱养殖水体的细菌群落结构特征和潜在病原菌属进行分析,同时应用实时荧光定量PCR方法对鲟鳇鱼养殖水体中的典型病原菌种进行定量检测。研究结果表明,在同一水域同一管理条件下,不同品种鲟鳇鱼网箱养殖水体中细菌群落结构组成相似度高,优势菌门由Proteobacteria、Actinobacteria、Bacteroidetes、Verrucomicrobia和Planctomycetes组成,占到所有样品序列总数的88.81%—93.94%;在属水平上,各样品未分类的细菌(unclassified)占比较大,共发现鲟鳇鱼潜在致病细菌菌属7个:Aeromonas、Acinetobacter、Chryseobacterium、Edwardsiella、Flavobacterium、Pseudomonas和Vibrio。实时荧光...     

光唇鱼仔稚幼鱼肠道菌群与养殖水体细菌群落的相关性

采用16S rRNA高通量测序,系统研究了光唇鱼(Acrossocheilus fasciatus)仔鱼、稚鱼和幼鱼肠道菌群组成及与同时期养殖水体细菌群落的相关性。研究结果表明,光唇鱼仔鱼、稚鱼和幼鱼的肠道菌群中Chao1指数和Shannon指数均没有显著变化(P>0.05);而随着光唇鱼幼体的发育,养殖水体中Chao1指数和Shannon指数呈现显著下降趋势(P<0.05)。光唇鱼仔鱼和稚鱼肠道菌群中的优势菌门由变形菌门(Proteobacteria)、拟杆菌门(Bacteroidetes)和厚壁菌门(Firmicutes)组成,而同时期养殖水体中优势菌门为变形菌门;光唇鱼幼鱼肠道菌群中的优势菌门为梭杆菌门(Fusobacteria)和变形菌门,同时期养殖水体中优势菌门由变形菌门、拟杆菌门和梭杆菌门组成。线性回归分析结果显示,随光唇鱼幼体发育,在光唇鱼肠道菌群中变形菌门、厚壁菌门和梭杆菌门相对丰度的时序变化趋势与其在养殖水体中相同。在属水平上,光唇鱼仔鱼、稚鱼肠道菌群中优势菌属均为醋酸杆菌属(Acetobacter),而幼鱼肠道菌群中优势菌属为鲸杆菌属(Cetobacte...     

双相情感障碍患者健康状况与肠道菌群的相关性分析

探讨双相情感障碍（BD）患者健康状况与肠道菌群的相关性,为该类患者的治疗提供参考。

收集我院BD患者（BD组）和健康人群（健康对照组）的粪便样本,使用QIAamp^® DNA粪便抽提试剂盒进行粪便样本的总DNA提取,随后进行16S rRNA基因测序,使用主成分分析（PCoA）探究健康对照组和BD组患者肠道菌群多样性情况。采用BD患者自我报告健康问卷评估患者的健康状况。

BD患者肠道菌群Shannon指数（P = 0.020 5）、Chao指数（P = 0.025 1）和Simpson指数（P = 0.020 8）显著低于健康对照组（均P＜0.05）。加权的Unifrac PCoA分析表明,BD组患者肠道菌群相似性低于健康对照组。两组对象肠道菌群门水平上有差异的菌群为放线菌门和厚壁菌门;属、种水平上的差异菌群有普氏粪杆菌、梭状芽胞杆菌和瘤胃球菌属。两组对象在功能性身体健康（PCS）、心理健康（MCS）、睡眠质量（PSQI）、抑郁（PHQ-9）、焦虑（GAD-7）和狂躁（ASRM）的平均总成绩上差异均有统计学意义（均P＜0.05）。BD患者的功能性身体健康得分与肠道放线菌门、厚壁菌门、拟杆菌属、梭状芽胞杆菌呈负相关,与普氏粪杆菌以及瘤胃球菌属呈正相关。

健康对照组和BD组患者的肠道菌群结构大致相似,BD患者的肠道菌群多样性低于健康对照组,BD患者健康状况与肠道菌群存在一定的相关性。

     

聚乳酸材料在不同土壤环境中生物降解的菌群结构分析

【目的】评价聚乳酸(Polylactic acid,PLA)材料在不同土壤环境中自然降解的效果,通过对3种不同土壤菌群结构的分析,找到能够对聚乳酸材料有降解作用的优势菌群。【方法】通过扫描电镜、断裂拉伸强度和CO2释放量测定来评价3种土壤对PLA材料的降解效果,并运用高通量测序技术,对3种土壤细菌群落进行基因组测序分析,检测3个样本细菌群落的差异性。【结果】PLA材料在沼泽地、芒果林地和稻田中的生物降解率分别为13.7%、10.6%和4.5%。3种土壤的样品分别获得11 110、11 236和8 848个OTU,共涉及细菌域的9个主要门和16个主要科。其中沼泽地土壤的微生物群落丰富度和多样性最高,稻田土壤最低。【结论】结合土壤的降解效果,土壤中生物群落丰富度和多样性越高,对PLA材料的降解作用越好。同时变形菌门(Proteobacteria)和拟杆菌门(Bacteroidetes)是降解聚乳酸材料的优势菌群。在科水平上,黄杆菌科(Flavobacteriaceae)、丛毛单胞菌科(Comamonadaceae)和噬纤维菌科(Cytophagaceae)的微生物对聚乳酸材料的降解最有潜力。这一研究成果为能有效降解聚乳酸材料的微生物资源的开发提供了理论依据。     

烧伤患者创面与环境中病原微生物耐药性及相关性的研究

了解烧伤科病人创面感染、环境监测的主要致病菌及其耐药性,探讨二者的关系。运用Sceptor鉴定仪对检出菌进行鉴定,K-B法对主要致病菌进行药敏试验。结果表明,引起烧伤创面感染的主要致病菌为不动杆菌(24.6%)、MRSA(19.8%)、绿脓杆菌(16.7%),烧伤科病房环境分离出的主要细菌为不动杆菌(17.7%)、绿脓杆菌(17.7%)、MRSA(13.3%),创面分离的致病菌与环境检出的细菌具有一定的相关性。特别是MRSA、MRSE,从创面与环境二者分离出的菌株具有相似的抗菌谱,这说明环境因素特别是医陪护人员的手表、物表成为该菌在烧伤病房传播的重要环节。     

人工湿地中指示与病原微生物动态分布及相关性分析

选择总大肠菌群(total coliforms)、粪大肠菌群(fecal coliforms)、粪链球菌(fecal streptococci)、大肠杆菌(Escherichia coli)、产气荚膜梭菌(Clostridium perfringens)和沙门氏菌(Salmonella spp.)6种指示和病原微生物,采用多管发酵法和倾注平板法研究其在自由表面流人工湿地中的去除效果、相关性及动态分布特征.结果表明:人工湿地进水中沙门氏菌数量最多,其次为总大肠菌群、粪大肠菌群、产气荚膜梭菌和大肠杆菌,粪链球菌数量最低;人工湿地对不同指示和病原微生物去除效果存在差异,对总大肠菌群、粪大肠菌群和大肠杆菌去除效果最好,年平均去除率分别为94.1％、93.4％和84.0％;对沙门氏菌和产气荚膜梭菌去除效果较差,年平均去除率分别为55.8％和63.6％,并且存在较大波动;出水中粪链球菌数量高于进水;人工湿地对指示和病原微生物的去除效果在不同季节间无显著差异,但迸水中总大肠菌群数量、进出水中沙门氏菌数量及进水中产气英膜梭菌数量在不同季节间存在差异.相关性分析表明,总大肠菌群、粪大肠菌群以及大肠杆菌之间存在较强相关性,沙门氏菌和粪大肠菌群以及产气荚膜梭菌和总大肠菌群、粪大肠菌群间存在弱相关性.     

典型对虾养殖水体中参与硝化与反硝化过程的微生物群落结构

【目的】本研究旨在分析典型虾塘养殖水体中参与氮循环关键过程的菌群多样性,为指导实际对虾养殖水体中NH 4+和NO 2-的微生物降解、水体氮素污染控制以及虾塘养殖氮素循环的有效管理提供科学依据。【方法】使用聚合酶链式反应及变性梯度凝胶电泳技术(Polymerase Chain Reaction-Denaturing Gradient GelElectrophoresis,PCR-DGGE)从8个不同地点的虾塘水样中确定代表性水样,以此为典型水样进行研究,构建了氨单加氧酶基因(amoA)、亚硝酸盐氧化还原酶基因(nxrA)、亚硝酸盐还原酶基因(nirS)的克隆文库。利用限制性片段长度多态性(Restriction Fragment Length Polymorphism,RFLP)技术将克隆文库进行酶切分析。【结果】通过序列多态性分析,表明amoA基因克隆文库中所有序列都属于变形杆菌门β亚纲(β-Proteobacteria),分别为亚硝化单细胞菌属(Nitrosomonas)(81%)和亚硝化螺旋菌属(Nitrosospira)(19%)2个属。nxrA基因克隆文库检测到α-Proteobacteria和δ-Proteobacteria两个亚纲,其中硝化杆菌属(Nitrobacter)是优势菌群,占整个文库的92%,仅有一个类群属于δ亚纲的脱硫杆菌科(Desulfobacteraceae)(8%)。nirS基因文库群落结构相对于amoA和nxrA基因文库较复杂,分别为α-Proteobacteria、β-Proteobacteria亚纲和Actinobacteria,序列分析表明,25%的类群为固氮弧菌属(Azoarcus),25%的类群为(Polymorphum),20%的类群为需氧去氮菌属(Thauera),10%的类群为(Sophophora),10%的的类群为链霉菌属(Streptomyces),5%的类群为(Brachymonas),5%的类群为(Ruegeria)。【结论】典型虾塘养殖水环境中氮素循环关键过程的菌群多样性丰富,其中亚硝化单胞菌属(Nitrosomonas)和硝化杆菌属(Nitrobacter)分别是此环境中主要的氨氧化作用推动者和亚硝酸盐氧化作用推动者,而在反硝化重要环节中,固氮弧菌属等多种菌群都起着推动作用。     

肠道菌群与过敏性紫癜性肾炎患儿肾功能及免疫功能的相关性研究

探讨肠道菌群与过敏性紫癜性肾炎患儿肾功能及免疫功能的相关性研究。

选取2021年9月—2023年9月于本院治疗的85例过敏性紫癜性肾炎患儿,标记为研究组。并纳入同期进行健康体检的68例同年龄段健康儿童作为对照组。采集患者空腹静脉血和粪便,采用qRT-PCR法定量分析肠道中大肠杆菌和双歧杆菌的数量,计算双歧杆菌/大肠杆菌（B/E）比值。使用全自动特定蛋白分析系统ARRAY360和流式细胞仪检测IgM、IgG、IgA、外周血CD4⁺ T淋巴细胞和CD8⁺ T淋巴细胞,计算CD4⁺/CD8⁺比值。过敏性紫癜性肾炎患儿肠道菌群与肾功能及免疫功能之间的相关性分析采用Pearson分析。

两组的性别、年龄、体质量指数差异均无统计学意义（P＜0.05）。与对照组相比,研究组大肠杆菌数量增加（P＜0.05）,血肌酐、尿素氮、血清尿酸、IgM、IgG、IgA、CD8⁺ T细胞水平升高（P＜0.05）,但双歧杆菌数量降低（P＜0.05）,B/E比值、CD4⁺ T细胞、CD4⁺/CD8⁺比值显著降低（P＜0.05）。过敏性紫癜性肾炎患儿肠道中的大肠杆菌数量分别与血肌酐、尿素氮、血清尿酸、IgM、IgG、IgA、CD8⁺ T细胞水平呈正相关（P＜0.001）,与CD4⁺ T细胞水平、CD4⁺/CD8⁺比值呈负相关（P＜0.001）;双歧杆菌、B/E比值分别与血肌酐、尿素氮、血清尿酸、IgM、IgG、IgA、CD8⁺ T细胞水平呈负相关（P＜0.001）,与CD4⁺ T细胞、CD4⁺/CD8⁺比值呈正相关（P＜0.001）。

过敏性紫癜性肾炎患儿肠道B/E比值显著降低,并与其肾功能及免疫功能指标存在相关性。

     

贝类中不同种类病原生物的快速检测方法

为适应进口鲜活贝类的快速检验需求,有必要建立快速检测多种病原生物的方法.针对进口鲜活水产品中常见的细菌、病毒和原生动物等不同种类的病原生物,设计了退火温度相近的特异PCR引物,建立了能够同时定性检测不同种类病原生物的方法.在此基础上,针对不同病原生物设计了特异的荧光引物,建立了检测贝类中不同病原生物的荧光定量PCR体系,并进行了特异性与重复性试验.与传统的检测方法相比较,该方法检测贝类中的病原生物更为快速准确,结果直观,可以满足口岸进口水产品快速检测的需要.     

Simultaneous Detection of Enteric Bacteria by QPCR in Surface Waters Located in an Urban Area

A rapid quantitative polymerase chain reaction (QPCR) method was developed for simultaneous detection of enteric bacteria from surface waters by utilizing a pair of universal primers that targeted four bacteria strains, namely Shigella dysenteriae, Vibrio cholerae, Salmonella typhimurium, and Escherichia coli. It was estimated that the QPCR method had a 94% confidence, and a detection limit as 2.7 Escherichia coli cells per sample in undiluted DNA extracts. The QPCR method was applied for the bacteriological examination of several surface waters in the urban area of Xi'an, China, and comparison was made with the conventional bacteria indicators determined by conventional membrane filter (MF) method. As a result, the calibrator cell equivalents (CCE) determined by QPCR was 2.2 to 5 times of the total coliform CFU, and the characteristics of the bacterial quality of different waters could be well presented by the QPCR results with a higher sensitivity. The coefficient of variation (CV) of data obtained by QPCR was smaller than that by traditional MF method, indicating a more stable analysis result. The QPCR method established by this study has manifest advantages over conventional methods in its rapidness and sensibility for the detection of pathogenic bacteria from surface water. It would provide a more reliable approach for the assessment of bacteriological risk of water environment.     

宁波市城区内河肠道病原细菌定量 PCR 分析

了解城市内河中病原菌含量对城市生态安全具有重要意义。应用定量PCR（ qPCR）方法,对宁波市城区6条重要内河水体中的病原菌含量及主要水质指标进行分析。结果表明,祖关山河、苏家河、南北河、后西河、史魏家河和直落河每升水体中含有病原菌基因的拷贝数分别为：3.34×105、1.47×105、4.68×105、1.75×106、9.65×106和1.33×106;且病原菌基因的含量与水体溶解氧负相关,与总氮、氨氮、浊度、总磷、高锰酸钾指数、叶绿素a和电导率呈正相关关系,但与水体透明度、温度和酸碱度无明显关系。定量PCR分析结果可望为评价城市内河水体质量提供重要参考。     

Survival of enteric bacteria and coliphage MS2 in pure human urine

Aims:  The survival of Escherichia coli , Salmonella enterica serovar Typhimurium, Enterococcus faecalis and coliphage MS2 was studied in stored, fresh and diluted (1 : 1) human urine at 15 and 30°C.
Methods and Results:  Survival rate was studied by the plate count method. All the organisms showed rapid inactivation in stored urine, but they survived better in diluted and fresh urine. The high pH level and temperature were the major factors found to influence the survival of the micro-organisms with the survival rate being higher at 15°C than at 30°C.
Conclusions:  The destruction of all micro-organisms in stored urine required <1 week at 30°C. Thus, the storage of urine is a useful way to reduce the risk of contamination while using urine as a fertilizer.
Significance and Impact of the Study:  The urine fertilization is aimed for the developing countries and the high temperatures in these countries may hasten the destruction of micro-organisms in urine. On the contrary, a higher survival rate of these organisms in fresh and diluted urine is a public health concern because the dilution of urine with water is likely to happen during flushing.     

致病菌体内诱导基因的功能分析—体内表达技术的应用

病原菌体内诱导的基因在致病过程中起重要作用,体内表达技术是一类很有前景的研究体内诱导基因的技术,本介绍了体内表达技术的基本原理,类型以及在致病菌体内诱导基因方面的研究进展和应用前景。     

Induction of cytokine formation by human intestinal bacteria in gut epithelial cell lines

Aims: To investigate the effects of human gut micro‐organisms on cytokine production by human intestinal cell lines. Methods and Results: Quantitative real‐time PCR assays were developed to measure the production of pro‐inflammatory (IL‐1α, IL‐6, IL‐18 and TNFα) and anti‐inflammatory (TGF‐β1, TGF‐β2, TGF‐β3, IL‐4 and IL‐10) cytokines in HT‐29 and Caco‐2 cell lines. They were co‐cultured with a range of mucosal bacteria isolated from ulcerative colitis patients, together with lactobacilli and bifidobacteria obtained from healthy people. HT‐29 cells were also co‐cultured with Campylobacter jejuni, enterotoxigenic Escherichia coli (ETEC), enteropathogenic E. coli and Salmonella typhimurium. The majority of commensal bacteria tested suppressed the expression of anti‐inflammatory cytokine mRNA, increased IL‐18, reduced IL‐1α, and with the exception of nonpathogenic E. coli, reduced TNF‐α. All overtly pathogenic species increased both pro‐inflammatory and anti‐inflammatory cytokine mRNA. Conclusion: Commensal and pathogenic species induced fundamentally different cytokine responses in human intestinal epithelial cell lines. Significance and Impact of the Study: Interactions between commensal bacteria tested in this study and the innate immune system were shown to be anti‐inflammatory in nature, in contrast to the pathogenic organisms investigated. These data contribute towards our understanding of how potential probiotic species can be used to suppress the pro‐inflammatory response in inflammatory bowel disease.     

Sanger和Pyrosequencing测序法分析健康成人口腔菌群

目的 对比Sanger和Pyrosequencing测序法分析健康人口腔菌群组成。方法 收集6例健康成人唾液、舌背、黏膜、龈上及龈下菌斑并构建16S rRNA基因文库,分别用Sanger和Pyrosequencing测序法分析。结果 Sanger测序所得已知的序列有5,794条（占6,535总序列数88.7%）、75个属,396个序列划分操作分类单元（operational taxonomic units,OTUs,占总OTUs的61.4%）。Pyrosequencing测序所得已知的序列有10,771条（占11,103总序列数97.0%）、66个属,322个OTUs（占总OTUs的68.0%）。Sanger和Pyrosequencing测序法所得口腔菌群在门、属的水平分布趋势基本一致,但在种的水平分布差异显著。Sanger和Pyrosequencing测序法构建的口腔菌群文库均匀度值分别为0.016和0.007,说明Pyrosequencing分析口腔菌群物种数量分布比Sanger测序方法的文库均匀性稍差,但优势种更显著。结论 Pyrosequencing测序时所构建基因文库能代表口腔菌群的多样性且经济、省时,可以应用于口腔细菌物种的分析。     

厌氧氨氧化启动过程Anammox菌富集规律和差异分析

为明确厌氧氨氧化反应器启动过程中Anammox菌的富集规律,采用荧光原位杂交(FISH)和实时荧光定量PCR(q-PCR)分析技术,对未添加填料、添加多面空心球以及添加竹炭的3个UASB反应器厌氧氨氧化启动过程中Anammox菌的增长规律进行分析。研究表明,Anammox菌的相对数量和绝对数量均随着启动时间呈逐渐递增趋势,在稳定运行阶段的第123天,无填料、多面空心球和竹炭反应器中Anammox菌分别占总细菌的23.3%、32.6%和43.7%,单位VSS污泥中Anammox菌16S rRNA基因拷贝数分别为(25.64±2.76)×107、(47.12±2.76)×107和(577.99±27.25)×107拷贝数g–1 VSS。竹炭反应器中Anammox菌最大生长率和最短倍增时间分别为0.064 d?1和10.8 d,最大生长率分别是无填料和多面空心球反应器的1.78倍和1.88倍。因此,填料添加特别是竹炭的添加可极大地促进Anammox菌的选择性生长和繁殖。FISH和q-PCR分析技术均适用于Anammox菌的富集规律研究,但因其检测目标存在差异,造成两者表征结果有所不同。     

Hepatitis E virus‐based evaluation of a virion concentration method and detection of enteric viruses in environmental samples by multiplex nested RT‐PCR

Aims: The prevalence of enteric viruses in drinking and river water samples collected from Pune, India was assessed. During an outbreak of HEV in a small town near pune, water samples were screened for enteric viruses. Methods and Results: The water samples were subjected to adsorption–elution‐based virus concentration protocol followed by multiplex nested PCR. Among 64 Mutha river samples, 49 (76·56%) were positive for Hepatitis A Virus, 36 (56·25%) were positive for Rotavirus, 33 (51·56%) were positive for Enterovirus and 16 (25%) were positive for Hepatitis E Virus RNA. Only enterovirus RNA was detected in 2/662 (0·3%) drinking water samples, and the samples from the city’s water reservoir tested negative for all four viruses. HEV RNA was detected in three out of four river water samples during HEV outbreak and partial sequences from patients and water sample were identical. Conclusions: The study suggests absence of enteric viruses both in the source and in the purified water samples from Pune city, not allowing evaluation of the purification system and documents high prevalence of enteric viruses in river water, posing threat to the community. Significance and Impact of the Study: The rapid, sensitive and relatively inexpensive protocol developed for virological evaluation of water seems extremely useful and should be adapted for evaluating viral contamination of water for human consumption. This will lead to development of adequate control measures thereby reducing disease burden because of enteric viruses.