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Testing hypotheses for the presence of tRNA genes in mycobacteriophage genomes 验证分支噬菌体基因组中tRNA基因存在的假设
Pub Date : 2016-07-02 DOI: 10.1080/21597081.2016.1219441
V. Delesalle, Natalie T Tanke, Albert C. Vill, G. Krukonis
ABSTRACT The presence of tRNA genes in bacteriophages has been explained on the basis of codon usage (tRNA genes are retained in the phage genome if they correspond to codons more common in the phage than in its host) or amino acid usage (independent of codon, the amino acid corresponding to the retained tRNA gene is more common in the phage genome than in the bacterial host). The existence of a large database of sequenced mycobacteriophages, isolated on the common host Mycobacterium smegmatis, allows us to test the above hypotheses as well as explore other hypotheses for the presence of tRNA genes. Our analyses suggest that amino acid rather than codon usage better explains the presence of tRNA genes in mycobacteriophages. However, closely related phages that differ in the presence of tRNA genes in their genomes are capable of lysing the common bacterial host and do not differ in codon or amino acid usage. This suggests that the benefits of having tRNA genes may be associated with either growth in the host or the ability to infect more hosts (i.e., host range) rather than simply infecting a particular host.
tRNA基因在噬菌体中的存在是基于密码子的使用(如果tRNA基因与噬菌体中比宿主中更常见的密码子相对应,则tRNA基因保留在噬菌体基因组中)或氨基酸的使用(与密码子无关,与保留的tRNA基因相对应的氨基酸在噬菌体基因组中比在细菌宿主中更常见)来解释的。从耻垢分枝杆菌的共同宿主上分离出的已测序的分枝噬菌体的大型数据库的存在,使我们能够检验上述假设,并探索tRNA基因存在的其他假设。我们的分析表明,氨基酸的使用比密码子的使用更能解释分枝噬菌体中tRNA基因的存在。然而,在其基因组中存在tRNA基因的密切相关的噬菌体能够裂解共同的细菌宿主,并且在密码子或氨基酸的使用上没有差异。这表明,携带tRNA基因的好处可能与宿主的生长或感染更多宿主(即宿主范围)的能力有关,而不是简单地感染特定宿主。
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引用次数: 56
Characterization of novel Staphylococcus aureus lytic phage and defining their combinatorial virulence using the OmniLog® system 使用OmniLog®系统鉴定新型金黄色葡萄球菌裂解噬菌体并确定其组合毒力
Pub Date : 2016-07-02 DOI: 10.1080/21597081.2016.1219440
Luis A. Estrella, Javier Quiñones, M. Henry, R. M. Hannah, R. Pope, Theron Hamilton, Nimfa Teneza-mora, E. Hall, Biswas Biswajit
ABSTRACT Skin and soft tissue infections (SSTI) caused by methicillin resistant Staphylococcus aureus (MRSA) are difficult to treat. Bacteriophage (phage) represent a potential alternate treatment for antibiotic resistant bacterial infections. In this study, 7 novel phage with broad lytic activity for S. aureus were isolated and identified. Screening of a diverse collection of 170 clinical isolates by efficiency of plating (EOP) assays shows that the novel phage are virulent and effectively prevent growth of 70–91% of MRSA and methicillin sensitive S. aureus (MSSA) isolates. Phage K, which was previously identified as having lytic activity on S. aureus was tested on the S. aureus collection and shown to prevent growth of 82% of the isolates. These novel phage group were examined by electron microscopy, the results of which indicate that the phage belong to the Myoviridae family of viruses. The novel phage group requires β-N-acetyl glucosamine (GlcNac) moieties on cell wall teichoic acids for infection. The phage were distinct from, but closely related to, phage K as characterized by restriction endonuclease analysis. Furthermore, growth rate analysis via OmniLog® microplate assay indicates that a combination of phage K, with phage SA0420ᶲ1, SA0456ᶲ1 or SA0482ᶲ1 have a synergistic phage-mediated lytic effect on MRSA and suppress formation of phage resistance. These results indicate that a broad spectrum lytic phage mixture can suppress the emergence of resistant bacterial populations and hence have great potential for combating S. aureus wound infections.
耐甲氧西林金黄色葡萄球菌(MRSA)引起的皮肤和软组织感染(SSTI)很难治疗。噬菌体(噬菌体)是一种潜在的替代治疗抗生素耐药细菌感染。本研究分离鉴定了7个具有广泛裂解金黄色葡萄球菌活性的新型噬菌体。通过电泳效率(EOP)试验对170个临床分离株进行筛选,结果表明该新型噬菌体具有强毒力,可有效阻止70-91%的MRSA和甲氧西林敏感金黄色葡萄球菌(MSSA)分离株的生长。噬菌体K先前被确定对金黄色葡萄球菌具有裂解活性,在金黄色葡萄球菌收集物上进行了测试,结果显示可阻止82%的分离株的生长。电镜观察结果表明,该噬菌体属肌病毒科。新的噬菌体群需要细胞壁上的β- n -乙酰氨基葡萄糖(GlcNac)片段来感染。通过限制性内切酶分析,该噬菌体与噬菌体K不同,但与K密切相关。此外,通过OmniLog®微孔板分析,生长速率分析表明,噬菌体K与噬菌体SA0420 1、SA0456 1或SA0482 1结合对MRSA具有协同的噬菌体介导裂解作用,并抑制噬菌体耐药性的形成。这些结果表明,广谱溶噬菌体混合物可以抑制耐药菌群的出现,因此在对抗金黄色葡萄球菌伤口感染方面具有很大的潜力。
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引用次数: 42
The impact of orally administered phages on host immune response and surrounding microbial communities 口服噬菌体对宿主免疫反应和周围微生物群落的影响
Pub Date : 2016-07-02 DOI: 10.1080/21597081.2016.1211066
Yingying Hong, J. Thimmapuram, Jiayi Zhang, Clayton K. Collings, K. Bhide, Kyle Schmidt, P. Ebner
ABSTRACT Numerous studies have shown the efficacy of phage therapy in reducing foodborne pathogen carriage in food animals. Fewer studies have focused on host reactions, especially in terms of phage-mediated acute immune responses and effects on the gut microbiome. Here we administered E. coli O157:H7 phages in low (single dose of 105 PFU) or high (single dose of 107 PFU) quantities to mice. While there were time points at which cytokine levels in different treatment groups differed from one another, all cytokine levels remained within normal ranges for mice regardless of treatment. Similarly, the patterns of these differences were not dose related, indicating that phage treatment did not result in a strong acute immune response as measured here. In separate experiments, 3-week-old pigs received a diet containing an in-feed antibiotic or daily phage treatment. After two weeks, microbial DNA of ileal, cecal, and fecal contents was characterized using 16S rRNA sequencing. There were no statistical differences in performance among the different groups. Compared to control pigs (no antibiotic, no phage), antibiotic treatment significantly altered ileal microbiome composition (P < 0.05), with Bacilli being most affected (antibiotic treated: 22%; control: 76%; FDR = 0.0572). No significant differences were observed in cecal and fecal microbiome composition between antibiotic-treated and control pigs, and there were no differences in gut microbiome composition between phage treated and control pigs in any intestinal compartment. Significant abundance differences were observed at the OTU level, with OTUs belonging to genera such as Lactobacillus and Streptococcus being over- or under-represented in either antibiotic or phage treated groups compared to control pigs. Determining whether these changes are deleterious to host, however, requires further study.
大量研究表明噬菌体治疗在减少食用动物食源性病原体携带方面的有效性。很少有研究关注宿主反应,特别是噬菌体介导的急性免疫反应和对肠道微生物群的影响。我们给小鼠注射了低剂量(单剂量105 PFU)或高剂量(单剂量107 PFU)的大肠杆菌O157:H7噬菌体。虽然不同治疗组的细胞因子水平存在时间点差异,但无论治疗方式如何,小鼠的所有细胞因子水平均保持在正常范围内。同样,这些差异的模式与剂量无关,表明噬菌体治疗没有导致强烈的急性免疫反应。在不同的实验中,3周龄的猪在饲粮中添加抗生素或每日使用噬菌体。两周后,使用16S rRNA测序对回肠、盲肠和粪便内容物的微生物DNA进行表征。不同组之间的表现没有统计学差异。与对照组(无抗生素和无噬菌体)相比,抗生素处理显著改变了回肠微生物组组成(P < 0.05),其中杆菌受影响最大(抗生素处理占22%;控制:76%;FDR = 0.0572)。抗生素组与对照组之间盲肠和粪便微生物组组成无显著差异,噬菌体组与对照组之间各肠室肠道微生物组组成无显著差异。在OTU水平上观察到显著的丰度差异,与对照猪相比,抗生素或噬菌体处理组中属于乳酸杆菌和链球菌属的OTU过多或不足。然而,确定这些变化是否对宿主有害还需要进一步的研究。
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引用次数: 22
Grete Kellenberger-Gujer: Molecular biology research pioneer. Grete Kellenberger-Gujer:分子生物学研究先驱。
Pub Date : 2016-04-05 eCollection Date: 2016-04-01 DOI: 10.1080/21597081.2016.1173168
Sandra Citi, Douglas E Berg

Grete Kellenberger-Gujer was a Swiss molecular biologist who pioneered fundamental studies of bacteriophage in the mid-20(th) century at the University of Geneva. Her life and career stories are reviewed here, focusing on her fundamental contributions to our early understanding of phage biology via her insightful analyses of phenomena such as the lysogenic state of a temperate phage (λ), genetic recombination, radiation's in vivo consequences, and DNA restriction-modification; on her creative personality and interactions with peers; and how her academic advancement was affected by gender, societal conditions and cultural attitudes of the time. Her story is important scientifically, putting into perspective features of the scientific community from just before the molecular biology era started through its early years, and also sociologically, in illustrating the numerous "glass ceilings" that, especially then, often hampered the advancement of creative women.

Grete Kellenberger-Gujer是瑞士分子生物学家,20世纪中期在日内瓦大学开创了噬菌体的基础研究。本文回顾了她的生活和职业故事,重点介绍了她对噬菌体生物学早期理解的基本贡献,通过她对一些现象的深刻分析,如温和噬菌体的溶原状态(λ)、基因重组、辐射的体内后果和DNA限制性修饰;论她的创作个性和与同龄人的互动以及她的学术进步如何受到当时性别、社会条件和文化态度的影响。她的故事在科学上很重要,从分子生物学时代开始的早期开始,就把科学界的特征透视出来,从社会学上也很重要,说明了无数的“玻璃天花板”,特别是在那个时候,经常阻碍有创造力的女性的进步。
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引用次数: 1
Counteracting selection for antibiotic-resistant bacteria. 对耐抗生素细菌的对抗选择。
Pub Date : 2016-03-07 eCollection Date: 2016-01-01 DOI: 10.1080/21597081.2015.1096996
Ido Yosef, Miriam Manor, Udi Qimron

The occurrence of antibiotic-resistant bacterial pathogens is on the rise because antibiotics exert selection pressure that kills only the antibiotic-sensitive pathogens. Sanitation and cleansing of hospital surfaces and the skin of medical personnel do not counteract this selective pressure, but rather indiscriminately reduce total pathogens on treated surfaces. Here, we discuss two recently introduced genetic strategies, based on temperate bacteriophages as DNA-delivery vehicles, that aim to sensitize bacteria to antibiotics and selectively kill the antibiotic-resistant ones. Outlooks for rendering one such approach more efficient and applicable are proposed. We believe that using an end product designed according to the provided principles on hospital surfaces and in hand-sanitizers will facilitate substitution of antibiotic-resistant pathogens with sensitive ones.

耐抗生素细菌病原体的发生正在上升,因为抗生素施加选择压力,只杀死对抗生素敏感的病原体。卫生和清洁医院表面和医务人员的皮肤不能抵消这种选择性压力,而是不加选择地减少治疗表面上的病原体总数。在这里,我们讨论了最近引入的两种遗传策略,基于温和噬菌体作为dna递送载体,旨在使细菌对抗生素敏感并选择性地杀死耐抗生素的细菌。提出了使这种方法更加有效和适用的展望。我们认为,在医院表面和洗手液中使用根据提供的原则设计的最终产品,将有助于用敏感病原体替代抗生素耐药病原体。
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引用次数: 13
Bacteriophage P2. 噬菌体P2。
Pub Date : 2016-02-18 eCollection Date: 2016-01-01 DOI: 10.1080/21597081.2016.1145782
Gail E Christie, Richard Calendar

P2 is the original member of a highly successful family of temperate phages that are frequently found in the genomes of gram-negative bacteria. This article focuses on the organization of the P2 genome and reviews current knowledge about the function of each open reading frame.

P2是一个非常成功的温带噬菌体家族的原始成员,该家族经常在革兰氏阴性菌的基因组中发现。本文主要介绍P2基因组的组织结构,并回顾了目前关于每个开放阅读框功能的知识。
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引用次数: 35
T7 ejectosome assembly: A story unfolds. T7弹射体组装:一个故事展开。
Pub Date : 2016-02-18 eCollection Date: 2016-01-01 DOI: 10.1080/21597081.2015.1128513
Sebastian Leptihn, Julia Gottschalk, Andreas Kuhn

T7 phage DNA is transported from the capsid into the host cytoplasm across the cell wall by an ejectosome comprised of the viral proteins gp14, gp15 and gp16. Prior to infection, these proteins form the so-called internal core in the mature virion. Gp16 was shown to associate with pure phospholipid bilayers while gp15 bound to DNA. A complex of both proteins appears as spiral-like rods in electron micrographs. It was also shown that the proteins gp15 and gp16 have the propensity to regain their full structure after thermal unfolding. From these observations it was concluded that (partial) unfolding of the proteins occurs during the translocation through the narrow portal of the phage capsid. After leaving the phage head, the proteins refold to form the ejectosome channel across the periplasm of the host. In this work, we analyzed the structure of gp15 and gp16 in presence of lipids and their stability toward chemical denaturants. A model to explain how the ejectosome might assemble in the host cell is discussed.

T7噬菌体DNA通过由病毒蛋白gp14、gp15和gp16组成的喷射体从衣壳穿过细胞壁进入宿主细胞质。在感染之前,这些蛋白质在成熟的病毒粒子中形成所谓的内部核心。结果显示,Gp16与纯磷脂双分子层结合,而gp15与DNA结合。这两种蛋白质的复合体在电子显微镜下呈螺旋状。研究还表明,gp15和gp16蛋白在热展开后具有恢复其完整结构的倾向。从这些观察结果可以得出结论,蛋白质的(部分)展开发生在通过噬菌体衣壳狭窄入口的转运过程中。离开噬菌体头后,蛋白质重新折叠形成穿过宿主外周质的喷射体通道。在这项工作中,我们分析了gp15和gp16在脂质存在下的结构及其对化学变性剂的稳定性。讨论了一个解释喷射体如何在宿主细胞中组装的模型。
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引用次数: 20
The diverse genetic switch of enterobacterial and marine telomere phages. 肠杆菌和海洋端粒噬菌体的多样性遗传开关。
Pub Date : 2016-02-18 eCollection Date: 2016-04-01 DOI: 10.1080/21597081.2016.1148805
Jens A Hammerl, Claudia Jäckel, Eugenia Funk, Sabrina Pinnau, Christin Mache, Stefan Hertwig

Temperate bacteriophages possess a genetic switch which regulates the lytic and lysogenic cycle. The genomes of the enterobacterial telomere phages N15, PY54 and ϕKO2 harbor a primary immunity region (immB) comprising genes for the prophage repressor, the lytic repressor and a putative antiterminator, similar to CI, Cro and Q of lambda, respectively. Moreover, N15 and ϕKO2 contain 3 related operator (OR) sites between cI and cro, while only one site (OR3) has been detected in PY54. Marine telomere phages possess a putative cI gene but not a cro-like gene. Instead, a gene is located at the position of cro, whose product shows some similarity to the PY54 ORF42 product, the function of which is unknown. We have determined the transcription start sites of the predicted repressor genes of N15, PY54, ϕKO2 and of the marine telomere phage VP58.5. The influence of the genes on phage propagation was analyzed in E. coli, Y. enterocolitica and V.parahaemolyticus. We show that the repressors and antiterminators of N15, ϕKO2 and PY54 exerted their predicted activities. However, while the proteins of both N15 and ϕKO2 affected lysis and lysogeny by N15, they did not affect PY54 propagation. On the other hand, the respective PY54 proteins exclusively influenced the propagation of this phage. The immB region of VP58.5 contains 2 genes that revealed prophage repressor activity, while a lytic repressor gene could not be identified. The results indicate an unexpected diversity of the growth regulation mechanisms in these temperate phages.

温带噬菌体具有调节裂解和溶原循环的遗传开关。肠杆菌端粒噬菌体N15、PY54和ko2的基因组含有一个初级免疫区(imb),由噬菌体抑制因子、裂解抑制因子和推测的抗菌素基因组成,分别类似于λ的CI、Cro和Q。此外,N15和ko2在cI和cro之间含有3个相关的算子(OR)位点,而在PY54中只检测到一个位点(OR3)。海洋端粒噬菌体具有假定的cI基因,但不具有克隆样基因。相反,一个位于cro位置的基因,其产物与PY54 ORF42产物有一定的相似性,其功能未知。我们已经确定了N15, PY54, ko2和海洋端粒噬菌体VP58.5的预测抑制基因的转录起始位点。分析了这些基因对大肠杆菌、小肠结肠炎杆菌和副溶血性弧菌噬菌体繁殖的影响。结果表明,N15、ko2和PY54的抑制因子和反终止因子发挥了预期的活性。然而,虽然N15和ko2蛋白都影响N15的裂解和溶原性,但它们不影响PY54的繁殖。另一方面,各自的PY54蛋白只影响该噬菌体的繁殖。VP58.5的imb区含有2个显示前噬菌体抑制活性的基因,而裂解抑制基因未被鉴定。结果表明,这些温带噬菌体的生长调节机制具有意想不到的多样性。
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引用次数: 7
CTXϕ: Exploring new alternatives in host factor-mediated filamentous phage replications. ctxφ:探索宿主因子介导的丝状噬菌体复制的新选择。
Pub Date : 2016-02-11 eCollection Date: 2016-04-01 DOI: 10.1080/21597081.2015.1128512
Eriel Martínez, Javier Campos-Gómez, François-Xavier Barre
ABSTRACT For a long time Ff phages from Escherichia coli provided the majority of the knowledge about the rolling circle replication mechanism of filamentous phages. Host factors involved in coliphages replication have been fully identified. Based on these studies, the function of Rep protein as the accessory helicase directly implicated in filamentous phage replication was considered a paradigm. We recently reported that the replication of some filamentous phages from Vibrio cholerae, including the cholera toxin phage CTXφ, depended on the accessory helicase UvrD instead of Rep. We also identified HU protein as one of the host factors involved in CTXφ and VGJφ replication. The requirement of UvrD and HU for rolling circle replication was previously reported in some family of plasmids but had no precedent in filamentous phages. Here, we enrich the discussion of our results and present new preliminary data highlighting remarkable divergence in the lifestyle of filamentous phages.
长期以来,来自大肠杆菌的Ff噬菌体提供了关于丝状噬菌体滚圈复制机制的大部分知识。参与噬菌体复制的宿主因子已经完全确定。基于这些研究,Rep蛋白作为辅助解旋酶直接参与丝状噬菌体复制的功能被认为是一个范例。我们最近报道了来自霍乱弧菌的一些丝状噬菌体的复制,包括霍乱毒素噬菌体ctxφ,依赖于辅助解旋酶UvrD而不是rep。我们还确定了HU蛋白是参与ctxφ和vgjφ复制的宿主因子之一。UvrD和HU对滚动圈复制的要求在一些质粒家族中已有报道,但在丝状噬菌体中没有先例。在这里,我们丰富了对我们的结果的讨论,并提出了新的初步数据,突出了丝状噬菌体生活方式的显着差异。
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引用次数: 5
Lactococcus lactis phage TP901-1 as a model for Siphoviridae virion assembly. 乳球菌噬菌体TP901-1作为虹膜病毒科病毒粒子组装的模型。
Pub Date : 2016-01-06 eCollection Date: 2016-01-01 DOI: 10.1080/21597081.2015.1123795
Jennifer Mahony, Stephen R Stockdale, Barry Collins, Silvia Spinelli, Francois P Douillard, Christian Cambillau, Douwe van Sinderen

Phages infecting Lactococcus lactis pose a serious threat to the dairy fermentation sector. Consequently, they are among the most thoroughly characterized Gram positive-infecting phages. The majority of lactococcal phages belong to the tailed family of phages named the Siphoviridae. The coliphage lambda and the Bacillus subtilis phage SPP1 have been the predominant comparators for emerging siphophages both genomically and structurally and both phages recognize a membrane protein receptor. In contrast, the lactococcal P335 group phage TP901-1 attaches to cell wall surface polysaccharides. It is a typical "lambdoid" siphophage possessing a long non-contractile tail and a genomic architecture reminiscent of lambda and SPP1 despite low or undetectable sequence homology in many of its encoded products, especially those involved in host recognition. A functional analysis of the structural components of TP901-1 was undertaken based on the characterization of a series of mutants in the region encoding the capsid and tail morphogenetic elements. Through this analysis, it was possible to deduce that, despite the lack of sequence homology, the overall genomic architecture of Siphoviridae phages typified by functional synteny is conserved. Furthermore, a model of the TP901-1 assembly pathway was developed with potential implications for many tailed phages.

感染乳酸乳球菌的噬菌体对乳制品发酵行业构成严重威胁。因此,它们是最彻底表征革兰氏阳性感染噬菌体之一。大多数乳球菌噬菌体属于尾状噬菌体科。噬菌体lambda和枯草芽孢杆菌噬菌体SPP1在基因组和结构上都是新兴噬菌体的主要比较物,两种噬菌体都识别膜蛋白受体。相反,乳球菌P335噬菌体TP901-1附着在细胞壁表面的多糖上。它是一种典型的“lambdoid”噬菌体,具有长而不收缩的尾巴,尽管其许多编码产物的序列同源性很低或无法检测到,但其基因组结构与lambda和SPP1相似,特别是那些参与宿主识别的产物。基于编码衣壳和尾部形态发生元件区域的一系列突变的特征,对TP901-1的结构成分进行了功能分析。通过这一分析,可以推断,尽管缺乏序列同源性,但以功能共生性为特征的Siphoviridae噬菌体的整体基因组结构是保守的。此外,建立了TP901-1组装途径模型,该模型对许多尾状噬菌体具有潜在的指导意义。
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引用次数: 12
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