Comparative analysis of Chlamydia bacteriophages reveals variation localized to a putative receptor binding domain

Three recently discovered ssDNA Chlamydia-infecting microviruses, phiCPG1, phiAR39, and Chp2, were compared with the previously characterized phage from avian C. psittaci, Chp1. Although the four bacteriophages share an identical arrangement of their five main genes, Chpl has diverged significantly in its nucleotide and protein sequences from the other three, which form a closely related group. The VP1 major viral capsid proteins of phiCPG1 and phiAR39 (from guinea pig-infecting C. psittaci and C. pneumoniae, respectively) are almost identical. However, VP1 of ovine C. psittaci phage Chp2 shows a high rate of nucleotide sequence change localized to a region encoding the "IN5" loop of the protein, thought to be a potential receptor-binding site. Phylogenetic analysis suggests that the ORF4 replication initiation protein is evolving faster than the other phage proteins. phiCPG1, phiAR39, and Chp2 are closely related to an ORF4 homolog inserted in the C. pneumoniae chromosome. This sequence analysis opens the way toward understanding the host-range and evolutionary history of these phages.     

Defective bacteriophages

Naturally occurring defective phage particles, which do not form plaques on any known host, but have a restricted host killing range, appear to be widely distributed. The defective phages are produced spontaneously but can be induced, at much higher levels, by chemical and physical agents which interfere with metabolism or structure of DNA. The defective phages discussed in this article have been divided into various categories on the basis of their structural complexity, which ranges from what appears to be phage tail components through to intact phage particles, and the source of the DNA packaged into the heads of the phage-like particles. The evolution of the defective phages is discussed and the possibility is entertained that they may have originated from temperate phages.     

Packaging of genomes in bacteriophages: a comparison of ssRNA bacteriophages and dsDNA bacteriophages.

In complex DNA bacteriophages like lambda, T4, T7, P22, P2, the DNA is packaged into a preformed precursor particle which sometimes has a smaller size and often a shape different from that of the phage head. This packaging mechanism is different from the one suggested for the RNA phages, according to which RNA nucleates the shell formation. The different mechanisms could be understood by comparing the genomes to be packaged: single stranded fII RNA has a very compact structure with high helix content. It might easily form quasispherical structures in solution (as seen in the electron microscope by Thach & Thach (1973)) around which the capsid could assemble. Double stranded phage DNA, on the other hand, is a rigid molecule which occupies a large volume in solution and has to be concentrated 15-fold during packaging into the preformed capsid, and the change in the capsid structure observed hereby might provide the necessary DNA condensation energy.     

Native marine bacteriophages

Abstract Isolation and cultivation of marine bacteriophages have shown that they are ubiquitous in seawater, and direct counting has shown that the total numbers of viruses frequently exceed the bacterial concentration by a factor of 10. About 150 different isolates of phages from marine environments have been characterized in the literature reviewed in the present report. Knoblike projections on phage heads seem to be a morphological property more common in marine phages than among phages from other sources. The cultured phages were generally much larger than the majority of viruses observed by direct transmission electron microscopy of seawater samples, indicating that culturing methods are not providing unbiased samples of environmental viruses. Cultured marine viruses frequently are more sensitive to organic solvents than the more intensively studied phages from other sources. Burst sizes from recent in situ studies are 50% lower than the average from culture studies. Phages in the marine environment may have half lives lasting less than a day, with consequent high turnover. Host ranges varies, and cross species host ranges have not been demonstrated. More information and further development of methods are needed, both from culture and from in situ studies.     

Single-stranded DNA-containing bacteriophages

Roots presents articles on major discoveries that laid the basis for contemporary molecular and cellular biology. In this article, Norton D. Zinder reviews the first findings about the single-stranded DNA-containing bacteriophages and what is known today about the genetics and molecular biology of these phages.     

Lytic bacteriophages ofStreptococcus mutans

Three phages ofStreptococcus mutans were obtained and partially characterized. The three phages, designated M102, e10, and f1, were found to be strictly lytic, with host ranges restricted to only serotype c, e, and f strains of this species, respectively. Phage sensitivity was not correlated with the presence of plasmids, at least in host strains of serotypes c and e. Each phage produced clear plaques in a number of standard media, even in the presence of sucrose, indicating that the extracellular glucan polysaccharides (mutan) produced by the hosts from this substrate do not prevent phage adsorption and growth. The phages were similar in size and morphology, having icosahedral heads and long (283–287 nm), flexible, noncontractile tails. The genome of each phage was found to consist of linear, double-stranded DNA, 31–35 kb in length, with a base composition of 37–38% G+C. Restricting phage DNAs with four enzymes produced fragment patterns unique to each phage, but common bands between M102 and e10 and between e10 and f1 were produced byBamHI. Labeled e10 and M102 DNAs hybridized strongly with all three phage DNAs, indicating that they share some common sequences. The three phages appear to be more similar than expected and probably evolved from a common ancestor.     

Characterization of Paenibacillus larvae bacteriophages and their genomic relationships to firmicute bacteriophages

Background

Paenibacillus larvae is a Firmicute bacterium that causes American Foulbrood, a lethal disease in honeybees and is a major source of global agricultural losses. Although P. larvae phages were isolated prior to 2013, no full genome sequences of P. larvae bacteriophages were published or analyzed. This report includes an in-depth analysis of the structure, genomes, and relatedness of P. larvae myoviruses Abouo, Davis, Emery, Jimmer1, Jimmer2, and siphovirus phiIBB_Pl23 to each other and to other known phages.

Results

P. larvae phages Abouo, Davies, Emery, Jimmer1, and Jimmer2 are myoviruses with ~50 kbp genomes. The six P. larvae phages form three distinct groups by dotplot analysis. An annotated linear genome map of these six phages displays important identifiable genes and demonstrates the relationship between phages. Sixty phage assembly or structural protein genes and 133 regulatory or other non-structural protein genes were identifiable among the six P. larvae phages. Jimmer1, Jimmer2, and Davies formed stable lysogens resistant to superinfection by genetically similar phages. The correlation between tape measure protein gene length and phage tail length allowed identification of co-isolated phages Emery and Abouo in electron micrographs. A Phamerator database was assembled with the P. larvae phage genomes and 107 genomes of Firmicute-infecting phages, including 71 Bacillus phages. Phamerator identified conserved domains in 1,501 of 6,181 phamilies (only 24.3%) encoded by genes in the database and revealed that P. larvae phage genomes shared at least one phamily with 72 of the 107 other phages. The phamily relationship of large terminase proteins was used to indicate putative DNA packaging strategies. Analyses from CoreGenes, Phamerator, and electron micrograph measurements indicated Jimmer1, Jimmer2, Abouo and Davies were related to phages phiC2, EJ-1, KC5a, and AQ113, which are small-genome myoviruses that infect Streptococcus, Lactobacillus, and Clostridium, respectively.

Conclusions

This paper represents the first comparison of phage genomes in the Paenibacillus genus and the first organization of P. larvae phages based on sequence and structure. This analysis provides an important contribution to the field of bacteriophage genomics by serving as a foundation on which to build an understanding of the natural predators of P. larvae.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-745) contains supplementary material, which is available to authorized users.     

Preparation of endotoxin-free bacteriophages

Bacteriophages (phages) are bacterial viruses that interact with bacterial walls and invade bacterial cells. Moreover, they disturb bacterial metabolism and lead to bacteria lysis. In the case of Gram-negative bacteria crude phage cultures, apart from the phages themselves, the bacterial debris, bacterial proteins and nucleic acids contain endotoxins. These endotoxins (lipopolysaccharides) posses a high degree of toxicity in vitro and in vivo, and their removal is essential for safety in antibacterial bacteriophage therapy. An effective, scaleable purification of bacteriophages from endotoxins was accomplished by sequential ultrafiltration through polysulfone membrane (30 nm) followed by chromatography on sepharose 4B and Matrex Cellulofine Sulfate. The phage fraction after gel filtration chromatography routinely contained endotoxins in the 150-2500 EU/ml range. The procedure yielded bacteriophages contaminated with as little as 0.4-7 EU/ml (Limulus assay). This value lies within the permitted level for intravenous applications (5 EU/kg/h by European Pharmacopoeia, 1997).