Correlation of physical maps and some genetic functions in the genomes of the kappa-theta phage family of Bacillus licheniformis

Summary Natural recombinant genomes between several, phenotypically distinct forms of phages and were isolated and characterized by DNA restriction fragment mapping and electron microscopic heteroduplex analysis. The phenotypes of the recombinants were correlated with the physical maps of the genomes, and several genetic functions were therfore defined and mapped. All genes necessary for the assembly of infectious virus particles map in a contiguous tract of DNA comprising about 20 kb, or nearly one third of the genome length. No DNA homology occurs within these domains of the two genomes, so that homologous recombination does not take place here and phenotypic mixing of the phages is eo ipso excluded. Other regions of heterology contain regulatory genes responsible for the lytic or temperate character of the phages, and for exclusion of phage by .     

Bacterial host strains that support replication of somatic coliphages

Somatic coliphages detected by Escherichia coli strain WG5 have been proposed as potential indicators of water quality. Their potential replication in the water environment is considered a drawback for their use as indicators. However, the contribution of replication outside the gut to the total numbers has never been quantified. It has not been determined either the fraction of bacterial strains that might support replication of phages detected by strain WG5 in the water environment. We examined the sensitivity of 291 host strains to 25 phages by streaking slants of the presumptive host strain onto an agar layer that contains bacteriophages, which gives a total of 7275 combinations (sensitivity tests). Only a 3.02% of the tests showed sensitivity. Additionally, six environmental strains were used as hosts to count phages in sewage and seawater. Phages isolated on these strains were used to infect strain WG5. The environmental strains detected 1 log₁₀ fewer phages than strain WG5 in sewage and seawater. The fraction of phages that were detected by the six strains and that also infected strain WG5 ranged from < 0.07% to < 2.0% of the total amount of bacteriophages detected by strain WG5 in the same samples. Our results confirm that less than 3% of naturally occurring hosts support replication of phages infecting E. coli. We conclude that the contribution of replication to the number of somatic coliphages detected in the aquatic environment is negligible. This revised version was published online in June 2006 with corrections to the Cover Date.     

Phage release from biofilm and planktonic Staphylococcus aureus cells

The ability of pathogenic staphylococci to form biofilms facilitates colonization and the development of chronic infections. Therapy is hampered by the high tolerance of biofilms towards antibiotic treatment and the immune system. We found evidence that lysogenic Staphylococcus aureus cells in a biofilm and in planktonic cultures spontaneously release phages into their surroundings. Phages were detected over a much longer period in biofilm cultures than in planktonic supernatants because the latter were degraded by secreted proteases. Phage release in planktonic and biofilm cultures was artificially increased by adding mitomycin C. Two morphologically distinct phages in the S. aureus strain used in this work were observed by electron microscopy. We postulate that phage-release is a frequent event in biofilms. The resulting lysis of cells in a biofilm might promote the persistence and survival of the remaining cells, as they gain a nutrient reservoir from their dead and lysed neighboring cells. This might therefore be an early differentiation and apoptotic mechanism.     

Bacteriophages immunomodulate the response of monocytes

Bacteriophages are present in fluids from cirrhosis patients. However, their effect on the immune response is unknown. In this work, we explore the role of phages in the phenotype, function, and cytokine production of monocytes. We stimulated healthy monocytes with five different butanol-purified phage suspensions infective for Gram-negative and Gram-positive bacteria. We studied the expression of the monocyte markers involved in lipopolysaccharide recognition (LPS; CD14), antigen presentation (HLA-DR) and co-stimulation (CD86), and the concentration of induced cytokines (TNF-α, IFN-α, and IL-10) by phages. To confirm the direct role of phages without the interference of contaminating soluble LPS in phage suspensions, polymyxin B was added to the cell cultures. Phagocytosis experiments were assessed by flow cytometry using labeled phage suspensions. We observed that butanol-purified phages reduced the surface levels of CD14 and CD86 in monocytes and increased the secreted levels of TNF-α and IL-10 compared with the control sample containing only butanol buffer. All phage suspensions showed downregulation of HLA-DR expression but only Staphylococcus aureus phage contaminated with Escherichia coli reached statistical significance. The addition of polymyxin B did not restore the monocytic response induced by phages, suggesting that the effect was not caused by the presence of LPS. Monocytes were able to phagocyte phages in a dose- and time-dependent manner. To conclude, the phagocytosis of butanol-purified phages altered the phenotype and cytokine production of monocytes suggesting they become tolerogenic.     

限制和修饰系统LlaBIII在构建抗噬菌体菌株中的作用

限制和修饰 (ｒｅｓｔｒｉｃｔｉｏｎａｎｄｍｏｄｉｆｉｃａｔｉｏｎ ,Ｒ Ｍ)系统是指由限制性内切酶和甲基化酶组成的单亚基或多亚基复合酶系统 ,两者通常成对出现 ,具有相同的ＤＮＡ识别位点 ,其作用相反。Ｒ Ｍ系统在原核生物中普遍存在 ,在保护细胞免遭外源病毒侵害方面具有重要作用[1] 。作为发酵剂的乳酸乳球菌在乳制品发酵中具有重要作用 ,但这类菌株极易遭受噬菌体感染 ,导致菌株产酸力降低 ,甚至发酵失败 ,造成严重的经济损失。所以在乳制品发酵过程中防止噬菌体感染就成为十分重要的问题。通过自然筛选或诱变处理等手段筛选噬菌…     

Polysaccharases for microbial exopolysaccharides

Microbial exopolysaccharides (EPS) are the substrates for a wide range of enzymes most of which are highly specific. The enzymes are either endoglycanases or polysaccharide lyases and their specificity is determined by carbohydrate structure with uronic acids often playing a major role. The presence of various acyl substituents frequently has little effect on the action of many of the polysaccharases but markedly inhibits some of the polysaccharide lyases including alginate and gellan lyases. The commonest sources of such enzymes can be either microorganisms or bacteriophages. These specific polysaccharide-degrading enzymes can yield oligosaccharide fragments, which are amenable to NMR and other analytical techniques. They have thus proved to be extremely useful in providing information about microbial polysaccharide structures and were routinely used in many such studies. Complex systems containing various mixtures of enzymes may also be effective in the absence of single enzymes but may be difficult to obtain with reproducible activities. Such preparations may also cause extensive degradation of the polysaccharide structure and thus prove less useful in providing information. Commercially available enzyme preparations have seldom proved capable of degrading microbial heteropolysaccharides, although some are active against bacterial alginates and homopolysaccharides including bacterial cellulose and curdlan.     

Specific and Selective Bacteriophages in the Fight against Multidrug-resistant Acinetobacter baumannii

Acinetobacter baumannii causes serious infections especially in immunocompromised and/or hospitalized patients. Several A. baumannii strains are multidrug resistant and infect wounds, bones, and the respiratory tract. Current studies are focused on finding new effective agents against A. baumannii. Phage therapy is a promising means to fight this bacterium and many studies on procuring and applying new phages against A. baumannii are currently being conducted. As shown in animal models, phages against multidrug-resistant A. baumannii may control bacterial infections caused by this pathogen and may be a real hope to solve this dangerous health problem.     

Viruses in extreme environments

The tolerance limits of extremophiles in term of temperature, pH, salinity, desiccation, hydrostatic pressure, radiation, anaerobiosis far exceed what can support non-extremophilic organisms. Like all other organisms, extremophiles serve as hosts for viral replication. Many lines of evidence suggest that viruses could no more be regarded as simple infectious “fragments of life” but on the contrary as one of the major components of the biosphere. The exploration of niches with seemingly harsh life conditions as hypersaline and soda lakes, Sahara desert, polar environments or hot acid springs and deep sea hydrothermal vents, permitted to track successfully the presence of viruses. Substantial populations of double-stranded DNA virus that can reach 10⁹ particles per milliliter were recorded. All these viral communities, with genome size ranging from 14 kb to 80 kb, seem to be genetically distinct, suggesting specific niche adaptation. Nevertheless, at this stage of the knowledge, very little is known of their origin, activity, or importance to the in situ microbial dynamics. The continuous attempts to isolate and to study viruses that thrive in extreme environments will be needed to address such questions. However, this topic appears to open a new window on an unexplored part of the viral world. Marc Le Romancer and Mélusine Gaillard contributed equally to this work.     

噬菌体作为模式生物在蛋白表达和病毒学方面的应用

近年来,随着噬菌体研究的不断深入,人们逐渐意识到噬菌体巨大的应用潜力。噬菌体表面展示技术、噬菌体抗体库、噬菌体核酸疫苗是将噬菌体作为载体而实现基因型与表型的结合,在病毒学方面噬菌体常被作为模式生物应用于多个研究领域。本文就以上各点作一综述。