Burkholderia cepacia X4降解油脂特性研究

从长期受油污染的土壤中分离筛选得到的Burkholderia cepaciaX4菌株能高效降解油脂。该菌株降解油脂的最适温度和pH分别为30℃和7.0,菌株降解油脂时适宜的氮源为硫酸铵,适宜碳氮比为4∶1。共基质碳源的添加有利于生物量的迅速增加和油脂降解率的提高,添加适量的葡萄糖能使油脂降解率提高8%~10%。50mg/L Ca2 对菌株生长和油脂降解更有利。在橄榄油浓度高达20g/L条件下最大油脂降解率仍可达83%。在油脂浓度≤2500mg/L时,该菌对油脂的降解符合抑制动力学Monod方程。     

革兰氏阴性菌锌摄取机制及抵抗宿主营养免疫的策略

锌是一种重要的金属元素,不仅充当许多蛋白质和酶的辅因子,还广泛参与糖类、脂质等的代谢过程。锌通常以二价离子的形式存在,在自然界主要分布在植物、土壤和水中,而在生物体内则是分散于肌肉和骨等组织中。对于大多数革兰氏阴性菌而言,锌离子也是其生长过程中必不可少的营养物质。正常情况下,细菌通过ZnuABC和ZIP锌转运系统从宿主体内夺取锌离子,用于体内蛋白质和酶的合成。当过多的锌离子被摄入时,细菌为了避免锌毒性则会启动特定的锌转录调节蛋白,以维持体内外的锌平衡。另一方面,当宿主察觉体内的锌离子被夺取,便会迅速采取锌限制性营养免疫等措施来制止锌离子的进一步流失。为了抵抗宿主的营养免疫,细菌进化出了相应的抵抗策略。较为典型的例子有鲍曼不动杆菌（Acinetobacter baumannii）的锌金属伴侣ZigA,其可在低锌环境中帮助细菌转运锌离子。本文将介绍革兰氏阴性菌锌摄取机制和抵抗宿主营养免疫的典型策略,为控制细菌感染途径和开发相关免疫疫苗等方面提供理论依据。     

Bioremediation of Vegetable Oil and Grease from Polluted Wastewater Using a Sand Biofilm System

Pseudomonas sp. (L1), P. diminuta(L2) were among eight bacterial strains isolated from vegetable grease and oil-contaminated industrial wastewater, four of which only were found to have the ability to degrade oil and grease. They were identified and investigated for oil and grease degradation either individually or in combinations in previous unpublished work by the authors. Since the combination M1 (Pseudomonas sp. andP. diminuta) produced the highest degradative activity, it was used in the present study in a biofilm sand filter system for vegetable oil and grease removal. This system was tested either as one unit or two units in sequence where different flow rates (30, 50, 100 ml/h) were applied compared to a control unit(s). Results showed that both biofilm systems reduced oily wastewater, even in cases of high degree of pollution (fat, oil & grease (FOG), 7535 ppm; biochemical oxygen demand (BOD₅), 525 ppm; chemical oxygen demand (COD), 1660 ppm). Results also showed a removal of FOG with efficiency at 100%; BOD₅ at 95.9% and COD at 96%, at 50 ml/h flow rate using one unit of biofilm system. On using two units in sequence, a complete removal of FOG, BOD₅ and COD with efficiency 100%, at flow rate 100 ml/h was achieved. In conclusion, the previous biofilm results indicated the efficiency of such a system in treating oily polluted wastewater (vegetable oil origin) on the basis of bacterial isolates being used, the optimum flow rate, and the number of biofilm units used in sequence to obtain the highest removal capacity of such a system. This revised version was published online in July 2006 with corrections to the Cover Date.     

革兰氏阴性菌TonB依赖性受体的功能与结构研究进展*

为维持生长所需,革兰氏阴性菌需要从外界摄取多种营养物质。分子量小于600 Da的分子可以通过自由扩散的方式通过革兰氏阴性菌的外膜,而大分子物质则需要特殊的转运系统才能将其转运至革兰氏阴性菌的胞内。革兰氏阴性菌对大分子营养物质的识别和转运主要由TonB依赖性受体负责完成。所有革兰氏阴性菌中均有TonB依赖性受体的存在,然而不同种类的革兰氏阴性菌拥有TonB依赖性受体的数量不同且功能各异。最近研究表明,TonB依赖性受体不仅参与了铁、血红素、锰、锌、镍、维生素、碳水化合物等多种营养物质的摄取,而且参与了蛋白酶的分泌。为对TonB依赖性受体提供更为深入和系统的理解,详细介绍了目前已知的TonB依赖性受体的功能及结构,以期为更进一步探知TonB依赖性受体未知功能提供可参考依据。     

革兰氏阴性菌TonB-ExbB-ExbD复合物的功能、作用机制、分布及进化

革兰氏阴性菌在生长繁殖过程中需要从外界摄取营养物质。一些小分子营养物质可以自由地通过革兰氏阴性菌的细胞膜,而一些大分子营养物质的转运需要特异性的TonB复合物依赖性的外膜受体进行转运。TonB复合物由TonB、ExbB、ExbD构成,是革兰氏阴性菌对外界营养物质主动转运过程的能量提供单位,在革兰氏阴性菌分布广泛。近年来,对TonB-ExbB-ExbD复合物的功能、结构及作用机制取得了重大研究进展,然而此复合物在不同的细菌也存在功能及作用机制上的差异。基于此背景,本文综述了TonB复合物的功能和结构研究进展,并分析了TonB复合物在革兰氏阴性菌中的分布、进化,比较了不同革兰氏阴性菌此复合物的差异,有助于进一步发现和揭示TonB复合物的新功能与作用机制。     

革兰氏阴性细菌外膜主要成分的跨膜转运机制研究进展

革兰氏阴性细菌的外膜由脂多糖、磷脂、外膜蛋白和脂蛋白等成分组成,是细菌抵御外界有害物质的首要物理屏障,与细菌致病性和耐药性密切相关.外膜各组分依赖特定的系统进行跨膜转运,包括脂多糖转运系统(lipopolysaccharide transport, Lpt)、脂质不对称维持系统(maintenance of lipid asymmetry, Mla)、β-桶状装配机器(β-barrel assembly machinery,Bam)以及脂蛋白定位系统(localization of lipoprotein,Lol).这些系统能够保证细菌外膜的完整与稳定,被视为维持细菌生命活动的"命门".因此,本文系统地综述革兰氏阴性细菌外膜主要成分的跨膜转运系统结构与功能,并对其未来研究方向进行展望,为新型靶向抗菌类药物研发提供新的思路.     

一株抗G- 菌和酵母菌的乳酸乳球菌的分离鉴定与抗菌活性

以G+ 菌金黄色葡萄球菌(Staphylococcus aureus)作为指示菌, 通过抑菌筛选法从生牛奶中初筛得到具有抑菌活性的14株细菌菌株, 然后通过个体形态与培养特征观测、部分生理生化反应、G + C mol%测定、16S rDNA序列比对分析、PCR扩增特异性N-乙酰胞壁酸水解酶基因和序列对比分析等鉴定, 确定其中的一株具有较高抑菌活性的分离株为乳酸乳球菌乳酸亚种(Lactococcus lactis subsp. lactis)菌株, 命名为MB191。对多种G+ 细菌、G- 细菌、酵母菌和丝状真菌的对峙培养抗性测定结果表明, MB191除对供试G+ 细菌具有较高的抑菌活性以外, 还对丁香假单胞菌(Pseudomonas syringae)、荧光假单胞菌(P. fluorescens)等G- 细菌和汉逊德巴利酵母(Debaryomyces hansenii)等具有明显的抑菌活性。乳酸乳球菌的这一特性目前尚未见文献报道。     

The Geomicrobiological Role of Sulphate-Reducing Bacteria in Environments Contaminated by Petroleum Products

This paper reviews the geomicrobiological role of sulphate-reducing bacteria (SRB) in environments contaminated with petroleum products and describes the habitats of SRB and their capacity for bioremediation in anaerobic conditions. Moreover, the participation of SRB in biocorrosion and formation of different minerals and sediments is discussed.     

高龄患者血液感染常见革兰阴性杆菌及其耐药性分析

目的分析本院80~100岁高龄患者血液感染常见革兰阴性杆菌的种类及其耐药状况,为本院合理使用抗生素提供依据。方法采用BacT/Alert 3D血培养仪对血液标本进行阳性鉴定;VITEK-2Compact全自动微生物鉴定仪进行鉴定;K-B纸片扩散法对抗菌药物进行敏感性测定;使用WHONET 5.4分析软件分析数据。结果本院高龄患者血液培养阳性标本中共分离出革兰阴性杆菌108株,以肠杆菌科细菌为主,其次为非发酵菌,前者主要为大肠埃希菌52株(48.15%)和肺炎克雷伯菌37株(34.26%),后者主要包括铜绿假单胞菌10株(9.26%)和鲍曼不动杆菌8株(7.41%)。其中大肠埃希菌和肺炎克雷伯菌对亚胺培南的耐药率分别为1.92%和13.51%,两者对氨苄西林、氨苄西林/舒巴坦、头孢唑林、头孢呋辛的耐药率均高于50.00%;铜绿假单胞菌和鲍曼不动杆菌对亚胺培南的耐药率分别为20.00%和25.00%,后者对其他抗菌药物的耐药率均高于前者。结论碳青酶烯类抗生素可作为本院高龄患者常见革兰阴性杆菌所致血液感染的首选药物;但在治疗中应考虑细菌的耐药特点及患者的代谢特点合理选择抗生素。     

Discriminative features of type I and type III secreted proteins from Gram-negative bacteria

The amino acid composition of sequences and structural attributes (α-helices, β-sheets) of C-and N-terminal fragments (50 amino acids) were compared to annotated (SWISS-PROT/ TrEMBL) type I (20 sequences) and type III (22 sequences) secreted proteins of Gram-negative bacteria. The discriminant analysis together with the stepwise forward and backward selection of variables revealed the frequencies of the residues Arg, Glu, Gly, Ile, Met, Pro, Ser, Tyr, Val as a set of strong (1-P < 0.001) predictor variables to discriminate between the sequences of type I and type III secreted proteins with a cross-validated accuracy of 98.6–100 %. The internal and external validity of discriminant analysis was confirmed by multiple (15 repeats) test-retest procedures using a randomly split original set of proteins; this validation method demonstrated an accuracy of 100 % for 191 non-selected (retest) sequences. The discriminant analysis was also applied using selected variables from the propensities for β-sheets and polarity of C-terminal fragments. This approach produced the next highest and comparable cross-validated classification accuracy for randomly selected and retest proteins (85.4–86.0 % and 82.4–84.5 %, respectively). The proposed sets of predictor variables could be used to assess the compatibility between secretion substrates and secretion pathways of Gram-negative bacteria by means of discriminant analysis.     

Purification and Visualization of Lipopolysaccharide from Gram-negative Bacteria by Hot Aqueous-phenol Extraction

Lipopolysaccharide (LPS) is a major component of Gram-negative bacterial outer membranes. It is a tripartite molecule consisting of lipid A, which is embedded in the outer membrane, a core oligosaccharide and repeating O-antigen units that extend outward from the surface of the cell^{1, 2}. LPS is an immunodominant molecule that is important for the virulence and pathogenesis of many bacterial species, including Pseudomonas aeruginosa, Salmonella species, and Escherichia coli^3-5, and differences in LPS O-antigen composition form the basis for serotyping of strains. LPS is involved in attachment to host cells at the initiation of infection and provides protection from complement-mediated killing; strains that lack LPS can be attenuated for virulence^6-8. For these reasons, it is important to visualize LPS, particularly from clinical isolates. Visualizing LPS banding patterns and recognition by specific antibodies can be useful tools to identify strain lineages and to characterize various mutants. In this report, we describe a hot aqueous-phenol method for the isolation and purification of LPS from Gram-negative bacterial cells. This protocol allows for the extraction of LPS away from nucleic acids and proteins that can interfere with visualization of LPS that occurs with shorter, less intensive extraction methods⁹. LPS prepared this way can be separated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and directly stained using carbohydrate/glycoprotein stains or standard silver staining methods. Many anti-sera to LPS contain antibodies that cross-react with outer membrane proteins or other antigenic targets that can hinder reactivity observed following Western immunoblot of SDS-PAGE-separated crude cell lysates. Protease treatment of crude cell lysates alone is not always an effective way of removing this background using this or other visualization methods. Further, extensive protease treatment in an attempt to remove this background can lead to poor quality LPS that is not well resolved by any of the aforementioned methods. For these reasons, we believe that the following protocol, adapted from Westpahl and Jann¹⁰, is ideal for LPS extraction.     

常见革兰阴性菌耐药性分析与合理用药研究

目的 分析芜湖市第一人民医院革兰阴性病原菌分布情况及对常见抗菌药物的耐药程度,为临床抗菌药物的选择及其合理使用提供参考。方法 收集2015年1月至2017年12月我院临床科室送检的细菌培养标本,采用法国生物梅里埃公司生产VITEK2 compact全自动微生物分析系统及其配套试剂盒进行细菌鉴定及药敏试验,数据采用WHONET 5.6软件进行统计分析。对年度耐药株数使用R语言3.4.0软件进行分析。结果 2015—2017年我院共检出病原菌3 728株,其中革兰阳性菌1 055株,占28.3%;革兰阴性菌2 673株,占71.7%。革兰阴性菌中主要致病菌为大肠埃希菌、肺炎克雷伯菌、铜绿假单胞杆菌、鲍曼不动杆菌,对青霉素类和第一代头孢菌素类抗菌药物高水平耐药。2017年这4种细菌对头孢唑林的耐药率均已达100.0%。鲍曼不动杆菌对头孢曲松的耐药率逐年上升,对头孢替坦连续3年耐药率均为100.0%,对氨苄西林/舒巴坦、氟喹诺酮类以及头孢吡肟耐药率未逐年上升,对亚胺培南耐药率变化显著,差异均有统计学意义（P<0.05）。铜绿假单胞杆菌对氨曲南耐药率逐年上升,对氨基糖苷类及头孢他啶耐药率逐年下降,差异均有统计学意义（P<0.05）。大肠埃希菌和肺炎克雷伯菌对头孢吡肟耐药率逐年下降,差异有统计学意义（P<0.05）。结论 我院住院患者病原菌以革兰阴性菌为主,不同病原菌对药物的敏感性各异,临床医生应结合患者情况和药敏结果综合分析,合理选用抗菌药物。     

The Phylogenetic Diversity of Aerobic Organotrophic Bacteria from the Dagang High-Temperature Oil Field

The distribution and species diversity of aerobic organotrophic bacteria in the Dagang high-temperature oil field (China), which is exploited with water-flooding, have been studied. Twenty-two strains of the most characteristic thermophilic and mesophilic aerobic organotrophic bacteria have been isolated from the oil stratum. It has been found that, in a laboratory, the mesophilic and thermophilic isolates grow in the temperature, pH, and salinity ranges characteristic of the injection well near-bottom zones or of the oil stratum, respectively, and assimilate a wide range of hydrocarbons, fatty acids, lower alcohols, and crude oil, thus exhibiting adaptation to the environment. Using comparative phylogenetic 16S rRNA analysis, the taxonomic affiliation of the isolates has been established. The aerobic microbial community includes gram-positive bacteria with a high and low G+C content of DNA, and γ and β subclasses of Proteobacteria. The thermophilic bacteria belong to the genera Geobacillus and Thermoactinomyces, and the mesophilic strains belong to the genera Bacillus, Micrococcus, Cellulomonas, Pseudomonas, and Acinetobacter. The microbial community of the oil stratum is dominated by known species of the genus Geobacillus (G. subterraneus, G. stearothermophilus, and G. thermoglucosidasius) and a novel species “Geobacillus jurassicus.” A number of novel thermophilic oil-oxidizing bacilli have been isolated.__________Translated from Mikrobiologiya, Vol. 74, No. 3, 2005, pp. 401–409.Original Russian Text Copyright © 2005 by Nazina, Sokolova, Shestakova, Grigoryan, Mikhailova, Babich, Lysenko, Tourova, Poltaraus, Qingxian Feng, Fangtian Ni, Belyaev.     

大连湾原油降解菌的分离和多样性分析

[目的]研究大连湾原油污染海域可培养原油降解菌的多样性,并获得新的原油降解菌.[方法]通过大连湾海水、海泥和海绵样品采集,以原油作为唯一碳源,培养、富集、分离筛选原油降解菌,根据16S rRNA基因序列确定其系统进化地位.[结果]通过形态观察和16S rRNA基因分析,共获得22个属的50株菌.其中,有6株菌的16S rRNA序列与最相近的菌株序列一致性仅为95％-97％,可能是潜在的新菌.单菌实验表明,45株菌具有石油降解能力.[结论]揭示了大连湾可培养原油降解菌的多样性,并获得了新的原油降解菌,为海洋石油污染的生物治理提供新资源.     

The comparison of susceptibility patterns of Gram-negative invasive and non-invasive pathogens in Estonian hospitals

A total of 560 invasive and 1062 non-invasive isolates were collected. The antimicrobial susceptibility of invasive versus non-invasive Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae isolates were evaluated using the E-tests. The equal domination of Gram-negative among both invasive and non-invasive pathogens was estimated in our study if contaminants were excluded. The emergence trend of Gram-positive microbes especially of coagulase negative staphylococci may be proved only after application of exclusive algorithms. Due to similar susceptibility, the data of non-invasive Gram-negative pathogens can be useful to predict resistance of invasive ones. Also, the surveillance of invasive pathogens provides useful information about the general susceptibility of pathogens.     

Enhancement of the antimicrobial activity and selectivity of GNU7 against Gram-negative bacteria by fusion with LPS-targeting peptide

Antimicrobial peptides (AMPs) provide a potential source of new antimicrobial therapeutics for the treatment of multidrug-resistant pathogens. To develop Gram-negative selective AMPs that can inhibit the effects of lipopolysaccharide (LPS)-induced sepsis, we added various rationally designed LPS-targeting peptides [amino acids 28–34 of lactoferrin (Lf28–34), amino acids 84–99 of bactericidal/permeability increasing protein (BPI84–99), and de novo peptide (Syn)] to the potent AMP, GNU7 (RLLRPLLQLLKQKLR). Compared to our original starting peptide GNU7, hybrid peptides had an 8- to 32-fold improvement in antimicrobial activity against Gram-negative bacteria, such as Escherichia coli and Salmonella typhimurium. Among them, Syn-GNU7 showed the strongest LPS-binding and -neutralizing activities, thus allowing it to selectively eliminate Gram-negative bacteria from within mixed cultures. Our results suggest that LPS-targeting peptides would be useful to increase the antimicrobial activity and selectivity of other AMPs against Gram-negative bacteria.     

临床分离4544株革兰阴性杆菌的耐药分布与变迁

目的了解玉溪市革兰阴性杆菌的耐药分布与变迁,为临床用药提供依据。方法对玉溪市人民医院1999—2004年临床各科送检的各类标本中培养分离出的4544株革兰阴性杆菌作回顾性分析。结果9种（属）细菌对25种药物的药敏结果耐药率〉50％者72种次（40．9％）,〈20％者44种次（25．0％）。总的耐药情况为不动杆菌和阴沟肠杆菌最高,铜绿假单胞菌和大肠埃希菌次之,甲型副伤寒沙门菌和福氏志贺菌最低。亚胺培南对多种细菌有较高的敏感覆盖率。有4种细菌对10种抗菌药物耐药率的上升具有临床意义（P〈0.01或P〈0．05）,其中以甲型副伤寒沙门菌对氟喹诺酮类上升最为显著。结论临床分离革兰阴性杆菌的耐药形势十分严峻,应定期监测区域内细菌的耐药变化．指导临床合坪用药。     

Isolation and Characterization of Arsenic-Resistant Bacteria from Contaminated Water-Bodies in West Bengal,India

Bengal Basin is known for severe arsenic contamination. In the present study, we have isolated six bacteria from the arsenic contaminated surface water of Bengal Basin. 16S rDNA sequence analysis identified them as Microbacterium oleivorans, Acinetobacter soli, Acinetobacter venetianus, Acinetobacter junii, Acinetobacter baumannii, Acinetobacter calcoaceticus. All the isolates possess arsenic accumulation potential and high molecular weight plasmid (>10 kb). PCR amplification indicated the presence of arsenic-resistance genes (arsB and aoxB) either in the genome or plasmid or in both in the isolated bacteria (except in Acinetobacter venetianus). Exposure to arsenic affected bacterial growth and induced alteration in cytoplasmic membrane integrity.     

The Bioremediation Potential of Hydrocarbonoclastic Bacteria Isolated From a Mangrove Contaminated by Petroleum Hydrocarbons on the Cilacap Coast,Indonesia

The main purpose of the study was to isolate strains of bacteria capable of degrading hydrocarbons from contaminated mangroves and to investigate the ability of the isolated bacteria to degrade total petroleum hydrocarbons (TPH) in a microcosm model of an oily sludge. The potential use of these bacteria strains as environmental clean-up agents was tested by culturing them with six different polyaromatic hydrocarbon (PAH) compounds (phenothiazine, fluorene, fluoranthene, dibenzothiophene, phenanthrene, and pyrene). Six viable and culturable bacteria were isolated, and the 16S rDNA sequence for each was amplified using the primers 9F and 1510R. Sequence results were compared using the National Center for Biotechnology Information (NCBI) BLAST program and, combined with phenotypic and phylogenetic data, were used to identify three strains that belonged to the Bacillus genus and were most closely related (98–99%) to Bacillus aquimaris, Bacillus megaterium, and Bacillus pumilus. The other three strains were closely related (98–100%) to Flexibacteraceae bacterium, Halobacilus trueperi, and Rhodobacteraceae bacterium. Two isolates, BA-PZN and BM-PFFP, which were related to Bacillus aquimaris and Bacillus megaterium, respectively, were further characterized and showed great potential for the removal of more complex hydrocarbon compounds in the oily microcosm model.     

某医院2014-2017年耐碳青霉烯类革兰阴性杆菌分布及耐药性

目的 了解2014-2017年玉溪市人民医院耐碳青霉烯类革兰阴性杆菌在临床标本及科室中的检出情况及耐药情况,为防治耐碳青霉烯类革兰阴性杆菌提供依据。 方法 采用VITEK2 COMPACT微生物分析仪对细菌进行鉴定和药敏试验。 结果 共检出952株耐碳青霉烯类革兰阴性杆菌,涉及17个菌属30个菌种,主要来自痰样本（669株,72.27%）、尿液（90株,9.45%）和体表分泌物（54株,5.67%）。科室分布显示重症医学科有309株（32.46%）,急诊内科121株（12.71%）,神经内科113株（11.87%）,呼吸内科83株（8.72%）。检出细菌中鲍曼不动杆菌仅对庆大霉素和左氧氟沙星的耐药率小于40.00%;大肠埃希菌仅对阿米卡星和呋喃妥因的耐药率小于40.00%;铜绿假单胞菌对哌拉西林、头孢吡肟、氨曲南、阿米卡星、庆大霉素、左氧氟沙星和环丙沙星的耐药率低于40.00%;肺炎克雷伯菌肺炎亚种对厄他培南、阿米卡星、四环素、复方新诺明的耐药率小于40.00%。 结论 耐碳青霉烯类革兰阴性杆菌分布较广,不同细菌对抗菌药物的耐药状况不同。鲍曼不动杆菌和大肠埃希菌的耐药性较高,应重视监测细菌耐药情况,指导临床合理使用抗菌药物,更好地控制耐药菌株的产生及传播。