Viruses are the most abundant biological entities on the planet, yet most classical principles of evolutionary biology and ecology were not developed with viruses in mind. Here, the concept of biological tradeoffs, a fundamental tenet of life history theory, is examined in the context of bacteriophage biology. Specifically, several important parameters of phage life histories—replication, persistence, host range, and adsorption—are evaluated for tradeoffs. Available data indicate that replication rate is strongly negatively correlated with both persistence and host range, suggesting that the well-documented tradeoff in macroorganisms between offspring production and offspring quality also applies to phages. The biological tradeoffs that appear to characterize viruses’ life histories have potential importance for viral evolution, ecology, and pathogenesis.
adsorption; bacteriophages; biological tradeoffs; ECOR; host range; life history theory; microbial ecology; replication rate; viral evolution
Designing lab exercises for introductory biology classes requires balancing the need for students to obtain results with a desire to provide unpredictable outcomes to better approximate actual research. Bacteriophage are particularly well suited for this as many species are well-understood but, with their hosts, represent a relatively complex interacting system. I have designed a seven week series of lab exercises that allow students to select bacteriophage resistant mutant hosts, isolate and sequence the corresponding receptor gene to identify the specific bacterial mutation from a large number of potential mutations. I also examined the possibility of collecting useful mutant strains for other studies. After two semesters, the lab series is working well with over 90% of students successfully isolating mutant bacteria and about half identifying the specific mutation. Here I discuss the advantages of using bacteriophage in an introductory class, the specific labs in this series and future plans.
teaching; laboratory teaching; bacteriophage T5; host range; receptor; receptor binding protein
Bacterial resistance to antibiotics poses a serious health threat. Since research into new antibiotics is not progressing at the same rate as the development of bacterial resistance, widespread calls for alternatives to antibiotics have been made. Phage therapy is an ideal alternative candidate to be investigated. However the success of phage therapy may be hampered by a lack of investment support from large pharmaceutical companies, due to their narrow spectrum of activity in antibiotics, very large costs associated with clinical trials of the variety of phages needed, and regulatory requirements remaining unclear. Intellectual property is difficult to secure for therapeutic phage products for a variety of reasons, and patenting procedures vary widely between the US and the EU. Consequently, companies are more likely to invest in phage products for decontamination or veterinary use, rather than clinical use in humans. Some still raise questions as to the safety of phage therapy overall, suggesting the possibility of cytotoxicity and immunogenicity, depending on the phage preparation and route. On the other hand, with patients dying because of infections untreatable with conventional antibiotics, the question arises as to whether it is ethical not to pursue phage therapy more diligently. A paradigm shift about how phage therapy is perceived is required, as well as more rigorous proof of efficacy in the form of clinical trials of existing medicinal phage products. Phage therapy potential may be fulfilled in the meantime by allowing individual preparations to be used on a named-patient basis, with extensive monitoring and multidisciplinary team input. The National Health Service and academia have a role in carrying out clinical phage research, which would be beneficial to public health, but not necessarily financially rewarding.
phage therapy; limitations; safety; ethics; paradigm changes; clinical use
Bacteriophage therapy, the use of viruses that infect bacteria as antimicrobials, has been championed as a promising alternative to conventional antibiotics. Although in the laboratory bacterial resistance against phages arises rapidly, resistance so far has been an only minor problem for the effectiveness of phage therapy. Resistance to antibiotics, however, has become a major issue after decades of extensive use. Should we expect similar problems after long-term use of phages as antimicrobials? Like antibiotics, phages are often noted to be drivers of bacterial evolution. Should we expect phage-treated pathogens to develop a general resistance to phages over time, a resistance against which only, for example, hypothetically co-evolved phages might be infective? Here we argue that the global infection patterns of phages suggest that this is not necessarily a concern as environmental phages often can infect bacteria with which those phages lack any recent co-evolutionary history.
antibiotic resistance; ecology; evolution; phage resistance; phage-therapy
The DNA packaging motors of double-stranded DNA phages are models for analysis of all multi-molecular motors and for analysis of several fundamental aspects of biology, including early evolution, relationship of in vivo to in vitro biochemistry and targets for anti-virals. Work on phage DNA packaging motors both has produced and is producing dualities in the interpretation of data obtained by use of both traditional techniques and the more recently developed procedures of single-molecule analysis. The dualities include (1) reductive vs. accretive evolution, (2) rotation vs. stasis of sub-assemblies of the motor, (3) thermal ratcheting vs. power stroking in generating force, (4) complete motor vs. spark plug role for the packaging ATPase, (5) use of previously isolated vs. new intermediates for analysis of the intermediate states of the motor and (6) a motor with one cycle vs. a motor with two cycles. We provide background for these dualities, some of which are under-emphasized in the literature. We suggest directions for future research.
ATPase; DNA packaging; bacteriophage genetics; bacteriophage structure; biological motor; cryo-electron microscopy; single-molecule analysis
The dawn of next generation sequencing technologies has opened up exciting possibilities for whole genome sequencing of a plethora of organisms. The 2nd and 3rd generation sequencing technologies, based on cloning-free, massively parallel sequencing, have enabled the generation of a deluge of genomic sequences of both prokaryotic and eukaryotic origin in the last seven years. However, whole genome sequencing of bacterial viruses has not kept pace with this revolution, despite the fact that their genomes are orders of magnitude smaller in size compared with bacteria and other organisms. Sequencing phage genomes poses several challenges; (1) obtaining pure phage genomic material, (2) PCR amplification biases and (3) complex nature of their genetic material due to features such as methylated bases and repeats that are inherently difficult to sequence and assemble. Here we describe conclusions drawn from our efforts in sequencing hundreds of bacteriophage genomes from a variety of Gram-positive and Gram-negative bacteria using Sanger, 454, Illumina and PacBio technologies. Based on our experience we propose several general considerations regarding sample quality, the choice of technology and a “blended approach” for generating reliable whole genome sequences of phages.
bacteriophage genome; Illumina HiSeq; Roche 454; PacBio; Sanger sequencing; Hybrid genome assembly; SISPA; Scaffolding
Phage targets for adsorption can include: (1) individual bacteria; (2) bacterial cellular arrangements such as streptococci; (3) microcolonies consisting of bacterial clones as can make up bacterial lawns and biofilms; and (4) bacterial biofilms themselves. While much effort has gone into considering category 1, and some into category 4, substantially less has been put into the question of how bacterial association into clonal arrangements or microcolonies might affect phage-bacterial interactions. Recently I have been exploring just this issue—within a single-authored monograph published in 2011 and a theoretical article published in 2012 as part of a special issue of the journal, Viruses. For this commentary, I have been invited to summarize my thinking on how bacterial association into either cellular arrangements or microcolonies might affect their susceptibility to phages along with related issues of bacterial resistance to phages and phage propagation in the context of both plaques and biofilms.
bacteriophages; biofilm; cellular arrangement; lysis inhibition; microcolony; phage; phage ecology; plaque formation; T-even phages
The quality of bacteriophage electron microscopy appears to be on a downward course since the 1980s. This coincides with the introduction of digital electron microscopes and a general lowering of standards, possibly due to the disappearance of several world-class electron microscopists The most important problem seems to be poor contrast. Positive staining is frequently not recognized as an undesirable artifact. Phage parts, bacterial debris, and aberrant or damaged phage particles may be misdiagnosed as bacterial viruses. Digital electron microscopes often seem to be operated without magnification control because this is difficult and inconvenient. In summary, most phage electron microscopy problems may be attributed to human failure. Journals are a last-ditch defense and have a heavy responsibility in selecting competent reviewers and rejecting, or not, unsatisfactory articles.
artifacts; contrast; crystals; digital electron microscopy; dimensions; fake viruses focus; misdiagnosis; monsters; positive staining; purification
Since the enlightenment, scientists have enjoyed a self-image as rational actors, guided only by reason, evidence and logic. When the Royal Society of London was founded in 1660 it chose as its motto “nullius in verba” (often translated as “on the word of no one”) a reference to Horace’s Epistles “Nullius addictus iurare in verba magistri…” (being not obliged swear allegiance to any master). Similar to our 21st century contemporaries who embrace the “new evidenced-based medicine,” the “virtuosi” of the Royal Society proclaimed a new era in science based only on observation and direct experience.
phage therapy; Félix d'Herelle; Gunther Stent; Frederick Twort; lytic principle
Vertebrate animals possess multiple anti-pathogen defenses. Individual mechanisms usually are differentiated into those that are immunologically adaptive vs. more “primitive” anti-pathogen phenomena described as innate responses. Here I frame defenses used by bacteria against bacteriophages as analogous to these animal immune functions. Included are numerous anti-phage defenses in addition to the adaptive immunity associated with CRISPR/cas systems. As these other anti-pathogen mechanisms are non-adaptive they can be described as making up an innate bacterial immunity. This exercise was undertaken in light of the recent excitement over the discovery that CRISPR/cas systems can serve, as noted, as a form of bacterial adaptive immunity. The broader goal, however, is to gain novel insight into bacterial defenses against phages by fitting these mechanisms into considerations of how multicellular organisms also defend themselves against pathogens. This commentary can be viewed in addition as a bid toward integrating these numerous bacterial anti-phage defenses into a more unified immunology.
abortive infection; adsorption resistance; antigen presentation; CRISPR; innate immunity; restriction-modification
A total of 30,000 phage papers, books, or book chapters, published between 1965 and 2010, were analyzed for the ethnic origins of 14,429 first authors. Their names represent 40 linguistic domains or geographic areas and at least 70 languages. British and German names predominate. Results broadly concur with statistics on the frequency of publications by country and show the growing role of Third-World countries in phage research. Irish and Jewish scientists are prominent. Historical and societal factors appear to be very important elements in the advancement of science.
bibliography; cultural theory; ethnic origin; family names; linguistics
The first phage electron micrographs were published in 1940 in Germany and proved the particulate nature of bacteriophages. Phages and infected bacteria were first examined raw and unstained. US American scientists introduced shadowing and freeze-drying. Phages appeared to be tailed and morphologically heterogeneous. Phage types identified by early electron microscopy include enterobacteriophages T4, T1, T7, T5, 7–11, ViI and Pseudomonas phage PB1. This paper retraces the development of early virus electron microscopy till the introduction of negative staining.
bacteriophage; electron microscopy; history
It can be difficult to appreciate just how small bacteria and phages are or how large, in comparison, the volumes that they occupy. A single milliliter, for example, can represent to a phage what would be, with proper scaling, an “ocean” to you and me. Here I illustrate, using more easily visualized macroscopic examples, the difficulties that a phage, as a randomly diffusing particle, can have in locating bacteria to infect. I conclude by restating the truism that the rate of phage adsorption to a given target bacterium is a function of phage density, that is, titer, in combination with the degree of bacterial susceptibility to adsorption by an encountering phage.
mean free path; phage adsorption; phage therapy