被污染的工业产品中常见微生物种类及耐药性分析

为了工业产品中微生物污染治理及其防腐体系的构建提供科学依据,收集了被微生物污染的工业产品,并对其进行分离、鉴定和分类,再通过测定杀菌剂的最低抑菌浓度(MIC)来评估微生物的抗药水平。污染微生物中革兰氏阴性菌约占47.59%,革兰氏阳性菌约占32.62%,主要为芽孢杆菌属、假单胞菌属和伯克霍尔德氏菌;污染真菌约占19.8%,主要为链格孢霉属、曲霉属、青霉属。MIC结果表明,分离的洋葱伯克霍尔德氏菌对卡松、布罗波尔、DMDMH抗性均高于标准菌株,分离的铜绿假单胞菌、洋葱伯克霍尔德氏菌和黑曲霉对DMDMH的抗性均高于标准菌株。导致工业产品污染的微生物种类较多,应根据其种类、理化性状、防腐剂杀菌机理不同进行治理。     

常见变质水性工业产品中的微生物种类和耐药性分析

为了科学治理常见变质水性工业产品中的腐败微生物,对其进行分离、鉴定和分类,同时以卡松、布罗波尔、甲基异噻唑啉酮和苯并异噻唑啉酮四种杀菌防腐剂对6种标准细菌菌株的最小抑菌浓度(MIC)作为耐药性标准,对腐败细菌进行耐药性评估分析。结果显示,日化用品变质样中革兰氏阴性细菌约占80.00%(克雷伯氏菌属、假单胞菌属、伯克霍尔德氏菌属等),革兰氏阳性细菌约占16.82%(芽孢杆菌属最多);而涂料及胶粘剂变质样中革兰氏阴性细菌约占53.33%(假单胞菌属、固氮菌属等),革兰氏阳性细菌约占38.27%(主要含芽孢杆菌属)。两者的耐药菌均主要为假单胞菌属且易对甲基异噻唑啉酮和苯并异噻唑啉酮产生耐药性。结论认为,导致产品微生物污染的因素很多,一方面由细菌产生耐药性引起,另一方面也与空气、水样和原材料污染等外部因素密切相关,但也不排除细菌自身一些生理、遗传、代谢特征等因素所致,即工业变质产品中的腐败微生物与耐药性微生物有相关性但是概念及含义并不相同。     

北京城市空气细菌群落结构与动态变化特征

在北京市选择3个典型的功能区,通过定点系统取样,运用BIOLOG鉴定技术,着重研究了北京城市空气细菌的群落结构与动态变化特征。结果表明:北京市空气中革兰氏阳性菌明显多于革兰氏阴性菌,约占80%~85%,其中阳性球菌占总数的50%~55%。不同功能区共发现47属空气细菌,其中革兰氏阳性菌31属,革兰氏阴性菌16属。优势菌属依次为微球菌属(Micrococcus)、葡萄球菌属(Staphylococcus)、芽孢杆菌属(Bacillus)、棒杆菌属(Corynebacterium)和假单胞菌属(Pseudomonas),它们总和约占50%。在优势菌属中,微球菌属约占总数的20%~30%,是北京市空气中比例最大的菌属,假单胞菌属约占2.5%~5.0%。文教区和交通干线细菌浓度明显高于公园绿地(P<0.01);并且文教区和交通干线空气细菌浓度四季变化特征显著,夏季和秋季较高,春季和冬季较低,公园绿地空气细菌浓度四季没有显著差异;3个功能区13:00时的细菌浓度明显低于9:00时和17:00。     

北京市夏季空气微生物群落结构和生态分布

着重研究北京市夏季空气微生物的群落结构和生态分布特征。结果表明 :北京市夏季空气中革兰氏阳性菌明显多于革兰氏阴性菌 ,约占 70 %～ 85 % ,其中阳性球菌占总数的占 35 %～ 4 5 %。 3个功能区 (文教区、交通干线和公园绿地 )共发现 30属空气细菌 ,其中革兰氏阳性菌 2 0属 ,革兰氏阴性菌 10属。优势细菌属为微球菌属 (Micrococcus)、芽孢杆菌属 (Bacillus)、葡萄球菌属 (Staphylococcus)和假单胞菌属 (Pseudomonas)。 3个功能区共出现 10属空气真菌 ,优势菌属枝孢属 (Cladosporium)、链格孢属 (Alternaria)、无孢菌 (nonsporing)、青霉属 (Penicillium)和曲霉属 (Aspergillus) ,其中枝孢属是绝对优势菌属 ,占总数的4 8.2 %。空气细菌浓度交通干线和文教区明显高于公园绿地 ,而空气真菌浓度公园绿地和文教区明显高于交通干线。空气细菌浓度一日中 13:0 0时较低 ,0 9:0 0时和 17:0 0较高     

塔河天然胡杨林地可培养细菌的生态分布研究

目的：研究塔里木河天然胡杨林部分地区可培养细菌的生态分布。方法：通过塔里木河胡杨林采样,可培养菌分离及16S rDNA序列鉴定。结果：从3种不同样品（水样、土样和胡杨树杆分泌物）中分离筛选了22株细菌,其中17株菌为革兰氏阳性菌,5株为革兰氏阴性菌。根据生理生化特征与16S rDNA序列分析结果表明,其中15株菌属于芽孢杆菌属,4株属于不动杆菌属、假单胞菌属、动性球菌属和Agrococcus属各含有1个分离株。结论：塔里木河胡杨林可培养微生物中芽孢杆菌比较丰富,其中有3个可能的新种。     

革兰氏阴性不发酵杆菌的鉴定及其对抗生素的敏感性

本文报道从临床病人和医院内环境分离的165株革兰氏阴性不发酵杆菌的鉴定,以及这些菌对常用抗生素敏感性试验的结果。165株菌归属为五个菌属：假单胞菌属、不动杆菌属、莫拉氏菌属、无色杆菌属和产碱菌属。绝大部分菌株对于常用抗生素耐药。讨论了细菌鉴定方法和程序。提出这些菌与临床感染具有密切关系,耐药菌株对于感染症治疗和医院内感染带来的问题。     

不同生境下五唇兰根部可培养内生细菌多样性研究

兰科植物内生细菌与菌根真菌的协作对宿主植物的生长、抗病、抗逆及植物修复环境能力等具有重要意义,揭示其内生细菌多样性及与生境之间的关系有助于阐明兰科植物的适应与进化机制。本研究基于16SrDNA序列分析探讨了不同生境下东南亚特有种五唇兰根部可培养内生细菌多样性及其空间异质性。结果表明:从不同生境下五唇兰根部共分离出内生细菌59株,其中从土生型五唇兰根部分离出内生细菌45株(76.27%),从石生型五唇兰根部分离出内生细菌14株(23.73%);基于内生细菌16SrDNA序列同源性分析及构建的系统发育树显示,五唇兰根部内生细菌分属于7属,即芽孢杆菌属(Bacillus)、伯克氏菌属(Burkholderia)、草酸菌属(Pandoraea)、土壤杆菌属(Agrobacterium)、类芽孢杆菌属(Paenibacillus)、泛菌属(Pantoea)、欧文氏菌属(Erwinia),其中优势属为芽孢杆菌属,次优势属为泛菌属和伯克氏菌属;多样性分析显示,土生型五唇兰根部内生细菌群落的Shannon多样性指数大于石生型五唇兰,不同生境下五唇兰根部内生细菌群落结构差异极显著(P0.01)。土生型五唇兰根部内生细菌群落优势属为芽孢杆菌属和泛菌属,石生型五唇兰根部内生细菌群落优势属为芽孢杆菌属和伯克氏菌属。     

蕙兰根内可培养细菌的物种多样性

以MS基本培养基添加蕙兰菌根浸出液制成的培养基进行分离培养的方法,从野生蕙兰（Cymbidium faberi）根部首次分离到内生细菌。经过分离纯化培养获得纯菌株27株。经过16S rDNA基因序列测序,并与GenBank数据比对,其相似性均在98%以上,分析鉴定结果表明,存活的22株菌可分为8属14种。分别隶属于伯克氏菌属（Burkholderia）、假单胞菌属（Pseudomonas）、芽胞杆菌属（Bacillus）、Leifsonia属、贪食菌属（Variovorax）、欧文氏菌属（Erwinia）、Duganella属和不动杆菌属（Acinetobacter）。对这些菌株进行分离培养及鉴定有助于理解兰花与微生物之间的相互作用关系,为开发利用这些微生物开辟新的思路。     

大庆油田油藏采出水的细菌群落结构

采用ARDRA (扩增性rDNA限制性酶切片段多态性分析)技术对大庆油田聚驱、水驱和过渡带3种油藏采出水中的细菌群落的基因组总DNA的16S rDNA克隆文库进行分析,研究了细菌群落结构.结果表明：随机挑取的596个阳性克隆可分为85个操作分类单元 (OTUs),其中聚驱、水驱和过渡带文库分别含有28、41和33个.通过对优势OTUs测序,并与GenBank进行序列比对,发现油藏采出水中的优势菌群为不动杆菌属、弓形杆菌属、厚壁菌门、假单胞菌属和硫磺单胞菌属.聚驱样品中细菌群落组成最简单,优势菌群为不动杆菌属,占库容的85%,假单胞菌属占7%;水驱样品中的优势菌群也是不动杆菌属,占库容的62%,假单胞菌属和硫磺单胞菌属各占20%和6%;过渡带文库的细菌群落的优势菌群为弓形杆菌属,占库容的50%,不动杆菌属和厚壁菌门各占19%和18%.     

腐败变质鸡蛋内容物细菌群落多样性分析

为研究腐败变质鸡蛋内容物细菌群落,本研究以蛋白液化(J)、发臭(JC)的鸡蛋为样品,鸡蛋内容物细菌基因组DNA为模板进行16S rDNA(V4+V5)区域Miseq测序、多样性指数分析及物种组成分析。结果表明,J样品和JC样品分别得到了42 550条和38 477条高质量序列,可归为19个操作分类单元(OTU),分属于2门、14科、19属、23种;多样性指数分析表明J样品的细菌丰度小于JC样品,而群落多样性高于JC样品。两样品的优势菌群均为变形菌门(Proteobacteria)肠杆菌科(Enterobacteriaceae)的柠檬酸杆菌属(Citrobacter)、沙雷氏菌属(Serratia)和莫拉菌科(Moraxellaceae)不动杆菌属(Acinetobacter)、假单胞菌科(Pseudomonadaceae)假单胞菌属(Pseudomonas)细菌以及厚壁菌门(Firmicutes)芽孢杆菌科(Bacillaceae)芽孢杆菌属(Bacillus)和肉杆菌科(Carnobacteriaceae)肉杆菌属(Carnobacterium)的细菌;其中,J样品中芽孢杆菌属(Bacillus)相对丰度最大(40.10%),但在种水平上未分类;而JC样品中主要是柠檬酸杆菌属(Citrobacter)相对丰度最大(81.23%),多为吉伦氏柠檬酸杆菌(Citrobacter gillenii)、埃希氏-志贺氏菌属(Escherichia-Shigella)和肠球菌属(Enterococcus)等序列极少,丰度很低;在种水平上23个种中,有14个种为未分类或未培养,同一个种分属于不同OTU。本研究得出,腐败变质鸡蛋内容物细菌种群复杂,其菌群组成及丰度能对鸡蛋腐败变质造成影响,为进一步研究鸡蛋主要腐败菌及其控制提供了参考。     

Diversity of bacterial endophytes in roots of Mexican husk tomato plants (Physalis ixocarpa) and their detection in the rhizosphere

Endophytic bacterial diversity was estimated in Mexican husk tomato plant roots by amplified rDNA restriction analysis and sequence homology comparison of the 16S rDNA genes. Sixteen operational taxonomic units from the 16S rDNA root library were identified based on sequence analysis, including the classes Gammaproteobacteria, Betaproteobacteria, Actinobacteria, and Bacilli. The predominant genera were Stenotrophomonas (21.9%), Microbacterium (17.1%), Burkholderia (14.3%), Bacillus (14.3%), and Pseudomonas (10.5%). In a 16S rDNA gene library of the same plant species' rhizosphere, only common soil bacteria, including Stenotrophomonas, Burkholderia, Bacillus, and Pseudomonas, were detected. We suggest that the endophytic bacterial diversity within the roots of Mexican husk tomato plants is a subset of the rhizosphere bacterial population, dominated by a few genera.     

黑龙江大豆胞囊线虫胞囊可培养细菌多样性

【目的】了解黑龙江省大豆田大豆胞囊线虫胞囊可培养细菌的多样性。【方法】运用稀释平板法和16SrDNA基因序列的系统发育分析对胞囊可培养细菌多样性进行研究。【结果】用NA培养基从胞囊上分离90株具有不同菌落形态的细菌。16S rDNA序列分析结果表明:90株菌株分属于7个属22个种。46株属于变形菌门γ亚群(Gammaproteobacteria),32株属于厚壁菌门(Firmicutes),10株属于变形菌门β亚群(Betaproteobacteria),2株属于变形菌门ɑ亚群(Alphaproteobacteria)。假单胞菌属(Pseudomonas)和芽孢杆菌属(Bacillus)为优势菌属。【结果】黑龙江省大豆胞囊线虫胞囊中存在丰富的细菌物种多样性,这些细菌对大豆胞囊线虫可能具有一定的生理生态作用。     

Diversity of Culturable Bacteria Associated with Ascocarps of a Chinese White Truffle,Tuber panzhihuanense (Ascomycota)*

Culturable bacterial communities inhabiting ascocarps of Tuber panzhihuanense were investigated. Isolates obtained on tryptone soy agar (TSA) were screened with high performance capillary electrophoresis (HPCE) according to differences in size of 16S rDNA V3. Target isolates were identified by analysis of the whole length of 16S rDNA gene. The results revealed that the ascocarps of T. panzhihuanense harbored a great number of culturable bacteria which belonging to 20 species and 11 genera in 5 phyla. Most isolates (4968%) were affiliated to the γ Proteobacteria, dominated by Pseudomonas lurida. The second major subclass was α Proteobacteria (3742%), with Phyllobacterium and a nitrogen fixing bacterium Bradyrhizobium japonicum also occurring as dominant taxa. The remaining bacterial isolates contained members of Actinobacteria (322%) and Firmicutes (774%) of which Bacillus was the commonest bacterium. A novel Tuber associated culturable bacterium species, Terriglobus roseus, was isolated and detected for the first time in Tuber ascocarps.     

杂交水稻种子固有细菌群落多样性探究

采用传统的分离培养方法和分子生物学技术对我国高产杂交水稻(OryzasativaL.)金优611种子固有细菌进行研究,从而了解其中可培养细菌群落的多样性。对分离得到的91株细菌进行16SrDNA扩增、ARDRA分型和16SrDNA系统发育分析,结果表明,分离得到的91株细菌分属于10个属16个种。其中γ-变形杆菌(Gammaproteobacteria)(53.85%)占据优势地位,其次为α-变形杆菌(Alphaproteobacteria)(20.88%),其它分属放线菌门Actinobacteria(15.39%)及厚壁菌门Firmicutes(9.88%)。其中的泛菌属(Pantoea sp.)和鞘氨醇单胞菌属(Sphingomonas sp.)、假单胞菌属(Pseudomonas sp.)、微杆菌属(Microbacterium sp.)为分离到的优势种群,且在种子这一特殊的生存空间中有4株潜在的新种存在。首次报道了杂交水稻金优611种子具有丰富的微生物群落多样性,为进一步探索植物种子际微生态环境中微生物群落的形成和生态功能提供了基础信息。     

中国白块菌——攀枝花块菌子囊果内可培养细菌的多样性研究

对新近发现的块菌属一新种——攀枝花白块菌（Tuber panzhihuanense）子囊果中可培养细菌的多样性进行了研究。采用胰蛋白大豆培养基（TSA）对菌株进行分离。用毛细管电泳（HPCE）对所有获得的菌株的16S rDNA V3高变区进行筛选获得不同条带大小的菌株,对筛选出的菌株的16S rDNA进行测序,并进行细菌多样性分析和研究。结果显示,攀枝花块菌子囊果内可培养细菌在数量及种类上都表现出很高的多样性,所有细菌分属于5个门的11个属和20个种。在所分离到的变形菌门的细菌中,数量最多的菌株（4968%）属于γ Proteobacteria,其中假单胞菌属的Pseudomonas lurida为优势类群;其次为α Proteobacteria,占3742%,其中以固氮菌 Bradyrhizobium japonicum和Phyllobacterium spp.为优势类群。其余的菌株属于放线菌门（Actinobacteria） （322%）和厚壁菌门（Firmicutes） （774%）,厚壁菌门中以芽孢杆菌属（Bacillus）为代表菌群。酸杆菌门中的Terriglobus roseus（194%）首次从块菌中分离获得。     

16S rDNA metabarcoding of the bacterial community associated with workers of Pheidole rugaticeps Emery (Hymenoptera: Formicidae)

Insect microbiota are receiving increasing attention from researchers, particularly with the continued advances in next generation sequencing (NGS) techniques. However, there is a paucity of data on the microbiota of ants that scavenge around human settlements. In this study, we characterized the bacterial communities of Pheidole rugaticeps Emery that were collected scavenging on other household insects using Illumina MiSeq high-throughput sequencing of the bacterial 16S ribosomal DNA gene. P. rugaticeps DNA was extracted from the insect samples using a HiYield? Genomic DNA isolation kit according to the manufacturer’s protocols and amplified using polymerase chain reaction (PCR). The PCR products were sequenced with the Illumina MiSeq platform according to the standard protocols to amplify the V3–V4 of the 16S rDNA gene. The results for the 16S rDNA genes were analysed using QIIME 2 Core ? 2020.6, and a 16S rDNA metabarcoding dataset was presented. A total of 46,651 reads were obtained from three genomic samples. A total of 368 amplicon sequence variants (ASV) comprising 165 genera were revealed and classified into 17 phyla. Proteobacteria (57.47%) and Firmicutes (33.14%) were the most abundant taxa, while Acinetobacter (37.10%) was the most abundant genus in all three sampling groups. Pathogenic bacteria species, such as Acinetobacter baumannii (15%) and Pseudomonas aeruginosa (2.92%), were identified from P. rugaticeps samples collected from a hospital environment. However, this study recommends more studies on the microbiota of Pheidole ants with different feeding habits and habitats to establish their core microbiome.     

Application of molecular systematics to study of bacterial cultures consuming volatile organic compounds

A range of species of four mixed bacterial cultures was studied by molecular systematics methods with the use of 16S rRNA genes. The cultures had been developed for application in minireactors, to degrade volatile organic compounds (VOCs): ethyl benzene, m-xylene, styrene, and o-xylene. A sample of 30 plasmid rDNA clones was obtained for each of the mixed cultures. The clones were analyzed by RFLP according to two restriction sites. Major variants of the 16S-rDNA sequences, corresponding to the most abundant species, were determined for each association. Sequencing of four clones of predominant 16S-rDNAs showed that the culture consuming ethyl benzene was dominated by Pseudomonas fluorescens; o-xylene, by Achromobacter xylosoxydans; styrene, by Pseudomonas veronii; and m-xylene, by Delftia acidovorans. Minor components of all four cultures were generally similar. They included species of the genera Sphingobacter, Rhizobium, Mesorhizobium, Pedobacter, and Paenibacillus. Sampling sequencing of genes for 16S rRNA cloned from total genomic DNA allowed quantitative determination of the composition of actual bacterial associations consuming VOCs in minireactors.     

Plasmid transfer from Pseudomonas putida to the indigenous bacteria on alfalfa sprouts: characterization,direct quantification,and in situ location of transconjugant cells

The transfer of the plasmids pJKJ5 and TOL (pWWO) from Pseudomonas putida to the indigenous bacterial community on alfalfa sprouts was studied. Tagging with fluorescent protein markers allowed direct quantification of the introduced donor bacteria and of indigenous bacteria that had received the plasmids. The sprouts were observed for 9 days; during this time alfalfa seeds, inoculated with donor bacteria, developed to edible and subsequently decaying sprouts. The first transconjugants were detected on day 6 after donor inoculation and occurred at frequencies of 3.4 x 10(-4) and 2.0 x 10(-6) transconjugant cells per donor cell for pKJK5::gfp and TOL::gfp, respectively. Confocal laser scanning microscopy revealed that the sprouts were heavily colonized with donors and that most transconjugants were located around the hypocotyl and root areas. Randomly selected members of the indigenous bacterial community from both inoculated and uninoculated sprouts, as well as a representative part of the community that had received the plasmids, were characterized by polymorphisms of PCR-amplified ribosomal DNA (rDNA) spacer regions between the 16S and 23S genes, followed by partial 16S rDNA sequencing. This showed that the initially dominating genera Erwinia and Paenibacillus were gradually replaced by Pseudomonas on the fully developed sprouts. Transconjugants carrying either of the investigated plasmids mainly belonged to the genera Pseudomonas and ERWINIA: The numbers of transconjugant cells did not reach detectable levels until 6 days after the onset of germination, at which point these species constituted the majority of the indigenous bacteria. In conclusion, the alfalfa sprouts provided an environment that allowed noteworthy frequencies of plasmid transfer from P. putida in the absence of selective pressure that could favor the presence of the investigated plasmids.     

Rickettsial relative associated with male killing in the ladybird beetle (Adalia bipunctata).

A cytoplasmically inherited microorganism associated with male killing in the two-spot ladybird beetle, Adalia bipunctata, is shown to be closely related to bacteria in the genus Rickettsia. Sequencing of a PCR-amplified product of the 16S genes coding for rRNA (16S rDNA) shows the organism associated with male killing in ladybirds to share a common ancestry with the Rickettsias relative to other genera (e.g., Anaplasma, Ehrlichia, and Cowdria). The rickettsial 16S rDNA product is found in four strains of ladybird beetle showing male embryo lethality and is absent from two uninfected strains and an antibiotic-cured strain. In addition, a revertant strain that had naturally lost the male-killing trait failed to amplify the rickettsial 16S rDNA product. Use of PCR primers for a 17-kDa protein antigen which is found only in rickettsias also resulted in an amplified product from infected strains. Uninfected, cured, and revertant strains and insect species infected with related bacteria (cytoplasmic-incompatibility bacteria from Nasonia wasps) failed to amplify the product. Discovery of a close relative of rickettsias associated with sex ratio distortion in insects has implications for the evolution and population dynamics of this bacterial genus.