Virologica Sinica special issue on “Phages and Therapy”

Note: Open access to this special issue no longer appears to be available…

2015, Volume 30, Issue 1


Bacteriophages, revitalized after 100 years in the shadow of antibiotics [pubmed]
Hongping Wei
In this issue, readers will not only find that bacteriophage research is a booming field but also learn about the diverse applications currently being explored for bacteriophages. The biggest driving force behind these applications is the serious threat of bacterial antibiotic resistance that is emerging in the current era.


Bacteriophage secondary infection [pubmed]
Stephen T Abedon
Phages are credited with having been first described in what we now, officially, are commemorating as the 100th. anniversary of their discovery. Those one-hundred years of phage history have not been lacking in excitement, controversy, and occasional convolution. One such complication is the concept of secondary infection, which can take on multiple forms with myriad consequences. The terms secondary infection and secondary adsorption, for example, can be used almost synonymously to describe virion interaction with already phage-infected bacteria, and which can result in what are described as superinfection exclusion or superinfection immunity. The phrase secondary infection also may be used equivalently to superinfection or coinfection, with each of these terms borrowed from medical microbiology, and can result in genetic exchange between phages, phage-on-phage parasitism, and various partial reductions in phage productivity that have been termed mutual exclusion, partial exclusion, or the depressor effect. Alternatively, and drawing from epidemiology, secondary infection has been used to describe phage population growth as that can occur during active phage therapy as well as upon phage contamination of industrial ferments. Here primary infections represent initial bacterial population exposure to phages while consequent phage replication can lead to additional, that is, secondary infections of what otherwise are not yet phage-infected bacteria. Here I explore the varying meanings and resultant ambiguity that has been associated with the term secondary infection. I suggest in particular that secondary infection, as distinctly different phenomena, can in multiple ways infl uence the success of phage-mediated biocontrol of bacteria, also known as, phage therapy.

Bacteriophage therapy against Enterobacteriaceae [pubmed]
Youqiang Xu, Yong Liu, Yang Liu, Jiangsen Pei, Su Yao, Chi Cheng
The Enterobacteriaceae are a class of gram-negative facultative anaerobic rods, which can cause a variety of diseases, such as bacteremia, septic arthritis, endocarditis, osteomyelitis, lower respiratory tract infections, skin and soft-tissue infections, urinary tract infections, intra-abdominal infections and ophthalmic infections, in humans, poultry, animals and fi sh. Disease caused by Enterobacteriaceae cause the deaths of millions of people every year, resulting in enormous economic loss. Drug treatment is a useful and effi cient way to control Enterobacteriaceae infections. However, with the abuse of antibiotics, drug resistance has been found in growing number of Enterobacteriaceae infections and, as such, there is an urgent need to find new methods of control. Bacteriophage therapy is an efficient alternative to antibiotics as it employs a different antibacterial mechanism. This paper summarizes the history of bacteriophage therapy, its bacterial lytic mechanisms, and the studies that have focused on Enterobacteriaceae and bacteriophage therapy.

Survival and proliferation of the lysogenic bacteriophage CTXΦ in Vibrio cholerae [pubmed]
Fenxia Fan, Biao Kan
The lysogenic phage CTXΦ of Vibrio cholerae can transfer the cholera toxin gene both horizontally (inter-strain) and vertically (cell proliferation). Due to its diversity in form and species, the complexity of regulatory mechanisms, and the important role of the infection mechanism in the production of new virulent strains of V. cholerae, the study of the lysogenic phage CTXΦ has attracted much attention. Based on the progress of current research, the genomic features and their arrangement, the host-dependent regulatory mechanisms of CTXΦ phage survival, proliferation and propagation were reviewed to further understand the phage’s role in the evolutionary and epidemiological mechanisms of V. cholerae.

Phage lytic enzymes: a history [pubmed]
David Trudil
There are many recent studies regarding the efficacy of bacteriophage-related lytic enzymes: the enzymes of ‘bacteria-eaters’ or viruses that infect bacteria. By degrading the cell wall of the targeted bacteria, these lytic enzymes have been shown to efficiently lyse Gram-positive bacteria without affecting normal fl ora and non-related bacteria. Recent studies have suggested approaches for lysing Gram-negative bacteria as well (Briersa Y, et al., 2014). These enzymes include: phage-lysozyme, endolysin, lysozyme, lysin, phage lysin, phage lytic enzymes, phageassociated enzymes, enzybiotics, muralysin, muramidase, virolysin and designations such as Ply, PAE and others. Bacteriophages are viruses that kill bacteria, do not contribute to antimicrobial resistance, are easy to develop, inexpensive to manufacture and safe for humans, animals and the environment. The current focus on lytic enzymes has been on their use as anti-infectives in humans and more recently in agricultural research models. The initial translational application of lytic enzymes, however, was not associated with treating or preventing a specifi c disease but rather as an extraction method to be incorporated in a rapid bacterial detection assay (Bernstein D, 1997).The current review traces the translational history of phage lytic enzymes-from their initial discovery in 1986 for the rapid detection of group A streptococcus in clinical specimens to evolving applications in the detection and prevention of disease in humans and in agriculture.


Selection of phages and conditions for the safe phage therapy against Pseudomonas aeruginosa infections [pubmed]
Victor Krylov, Olga Shaburova, Elena Pleteneva, Sergey Krylov, Alla Kaplan, Maria Burkaltseva, Olga Polygach, Elena Chesnokova
The emergence of multidrug-resistant bacterial pathogens forced us to consider the phage therapy as one of the possible alternative approaches to treatment. The purpose of this paper is to consider the conditions for the safe, long-term use of phage therapy against various infections caused by Pseudomonas aeruginosa. We describe the selection of the most suitable phages, their most effective combinations and some approaches for the rapid recognition of phages unsuitable for use in therapy. The benefi ts and disadvantages of the various different approaches to the preparation of phage mixtures are considered, together with the specifi c conditions that are required for the safe application of phage therapy in general hospitals and the possibilities for the development of personalized phage therapy.

Molecular dissection of phage lysin PlySs2: integrity of the catalytic and cell wall binding domains is essential for its broad lytic activity [pubmed]
Yanling Huang, Hang Yang, Junping Yu, Hongping Wei
The novel phage lysin PlySs2, is reported to be highly active against various bacteria, including staphylococci, streptococci and Listeria. However, the molecular mechanisms underlying its broad lytic spectrum remain to be established. In the present study, the lytic activity of the catalytic domain (CD, PlySc) and binding specificity of the cell wall binding domain (CBD, PlySb) of PlySs2 were examined. Our results showed that PlySc alone maintains very limited lytic activity. Enhanced green fluorescent protein (EGFP)-fused PlySb displayed high binding affinity to the streptococcal strains tested, including S. suis, S. dysgalactiae, and S. agalactiae, but not staphylococci, supporting its utility as a good CBD donor for streptococcal-targeted lysin engineering. EGFP-fused intact PlySs2 similarly displayed high affinity for streptococci, but not staphylococci. Notably, four truncated PlySb fragments showed no binding capacity. These fi ndings collectively indicate that integrity of the PlySc and PlySb domains is an essential determinant of the broad lytic activity of PlySs2.

Isolation and characterization of glacier VMY22, a novel lytic cold-active bacteriophage of Bacillus cereus [pubmed]
Xiuling Ji, Chunjing Zhang, Yuan Fang, Qi Zhang, Lianbing Lin, Bing Tang, Yunlin Wei
As a unique ecological system with low temperature and low nutrient levels, glaciers are considered a “living fossil” for the research of evolution. In this work, a lytic cold-active bacteriophage designated VMY22 against Bacillus cereus MYB41-22 was isolated from Mingyong Glacier in China, and its characteristics were studied. Electron microscopy revealed that VMY22 has an icosahedral head (59.2 nm in length, 31.9 nm in width) and a tail (43.2 nm in length). Bacteriophage VMY22 was classifi ed as a Podoviridae with an approximate genome size of 18 to 20 kb. A one-step growth curve revealed that the latent and the burst periods were 70 and 70 min, respectively, with an average burst size of 78 bacteriophage particles per infected cell. The pH and thermal stability of bacteriophage VMY22 were also investigated. The maximum stability of the bacteriophage was observed to be at pH 8.0 and it was comparatively stable at pH 5.0-9.0. As VMY22 is a cold-active bacteriophage with low production temperature, its characterization and the relationship between MYB41-22 and Bacillus cereus bacteriophage deserve further study.


Variation of resistance and infectivity between Pseudomonas fluorescens SBW25 and bacteriophage Ф2 and its therapeutic implications [pubmed]
Hanchen Chen, Guohua Chen
Studies of the coevolutionary dynamics between Pseudomonas fluorescens SBW25 and bacteriophage Ф can explore host resistance and parasite infectivity with applications in the ecological and therapeutic fields.Coevolutionary dynamics determine the efficacy of phage-based therapy. In the study described here, bacterial resistance and phage infectivity fluctuated with culturetime, perhaps resulting from random mutation and temporaladaptation, which reminds us of the necessity toconsider evolutionary mechanisms when applying phageto treat bacterial infections.

A novel transposable Mu-like prophage in Bacillus alcalophilus CGMCC 1.3604 (ATCC 27647) [pubmed]
Junjie Yang, Yimeng Kong, Xuan Li, Sheng Yang
In this letter, we provide evidence for the first transposable prophage BalMu-1 in Bacilli. The transposable prophage (BalMu-1, Genbank No. KP063902 and KP063903) was identified in Bacillus alcalophilus CGMCC 1.3604(ATCC 27647) through high throughput genome sequencing and PCR-dideoxy chain-termination(Sanger) sequencing.

Isolation and characterization of a lytic bacteriophage φKp-lyy15 of Klebsiella pneumoniae [pubmed]
Yinyin Lu, Hongyan Shi, Zhe Zhang, Fang Han, Jinghua Li, Yanbo Sun
In conclusion, the lytic bacteriophage φKp-lyy belonging to the Siphoviridae family specific for K. pneumonia was isolated and characterized. φKp-lyy displayed a short latent period, stability to a wide pH rang, high thermal resistance, and lytic activity toward a relatively broad range of K. pneumonia isolates. Thus, phage φKplyy should be considered as a candidate for inclusion in phage cocktails to control K. pneumoniae-associated nosocomial infections.

Expression and purification of recombinant lyase gp17 from the LSB-1 phage in Escherichia coli [pubmed]
Taiwu Wang, Hui Lin, Lu Zhang, Guorong Huang, Long Wu, Lei Yu, Hongyan Xiong
In this study, we successfully expressed and purified the recombinant gp17 protein from the LSB-1 phage and also confirmed its bacteriostatic effect. Assays also showed that the recombinant enzyme was soluble and had signifi cant lyase effects on the host bacterium, EIEC8401. A preliminary study demonstrated that the enzyme did not have inhibitory effects on other strains (unpublished data), which might indicate that the exclusive antibacterial effect of gp17 on EIEC8401 could have a special significance in practical application in bacterial therapy.

T4-like coliphage φKAZ14 virulent to pathogenic and extended spectrum β-lactamase-producing Escherichia coli of poultry origin [pubmed]
Kaikabo Adamu Ahmad, Abdulkarim Sabo Mohanmmed, Faridah Abas, Sieo Chin Chin
The aim of the present study was to isolate bacteriophages for the pre-harvest biocontrol of APEC 01 and ESBL-producing E. coli in chicken, in order to mitigate the risk of these pathogens to the food chain. Isolation and characterization of the T4-like coliphage KAZ14, lytic to APEC 01 and ESBL-producing E. coli, is reported and discussed.

Isolation and complete genome sequence of a novel virulent mycobacteriophage, CASbig [pubmed]
Tieshan Teng, Junping Yu, Hang Yang, Hongping Wei
In this study, we report the isolation and the complete genome of a novel mycobacteriophage, CASbig, which has an icosahedral head (diameter 50 ± 2 nm) and a long, non-contractile tail (length 160 ± 5 nm) with transverse striations, ending in a small knob. The length of the tail includes the middle of the baseplate, and the head measurements were taken between opposite apices. These characteristics indicate that the phage belongs to the family Siphoviridae morphotypes.


Experience of the Eliava Institute in bacteriophage therapy [pubmed]
Mzia Kutateladze
The rapid propagation of multidrug resistant bacterial strains is leading to renewed interest in bacteriophage therapy. With challenges in the treatment of bacterial infections, it is essential for people worldwide to understand how alternative approaches, such as bacteriophages, could be used to combat antibiotic resistant bacteria. The Eliava Institute of Bacteriophages, Microbiology and Virology (Tbilisi, Georgia) is arguably the most famous institution in the world focused on the isolation, study, and selection of phages active against a variety of bacterial pathogens.


Fossilized viruses found in the geological record

In the 90s there was a notion that was inexplicably popular in the literature (Folk, 1993; Folk and Lynch, 1997; Pedone and Folk, 1996; Sillitoe et al., 1996; McKay et al., 1996; Vasconcelos and McKenzie, 1997; Kajander and Çiftçioglu, 1998), and still stubbornly held by some, that the distinct structured particles under 150 nm in size commonly found in the geological record associated with putatively biogenic rock constituted a new form of “nanobacteria,” despite being too small to support living systems and based on often poorly labelled micrographs. How much the controversy this engendered unknowingly recalled the classical extended fights over Martinus Beijerinck‘s contagium vivum fluidum and the original discovery of viruses may seem obvious only in hindsight, but a new paper now convincingly argues that many of these particles may indeed be fossilized viruses of microbes,

Viruses as new agents of organomineralization in the geological record

Muriel Pacton, David Wacey, Cinzia Corinaldesi, et al. Published 2014 in Nat. Commun. doi:10.1038/ncomms5298
Viruses are the most abundant biological entities throughout marine and terrestrial ecosystems, but little is known about virus–mineral interactions or the potential for virus preservation in the geological record. Here we use contextual metagenomic data and microscopic analyses to show that viruses occur in high diversity within a modern lacustrine microbial mat, and vastly outnumber prokaryotes and other components of the microbial mat. Experimental data reveal that mineral precipitation takes place directly on free viruses and, as a result of viral infections, on cell debris resulting from cell lysis. Viruses are initially permineralized by amorphous magnesium silicates, which then alter to magnesium carbonate nanospheres of ~80–200 nm in diameter during diagenesis. Our findings open up the possibility to investigate the evolution and geological history of viruses and their role in organomineralization, as well as providing an alternative explanation for enigmatic carbonate nanospheres previously observed in the geological record.

While the authors seem primarily concerned with how viruses could affect the chemical formation of geological features, if real, this has the exciting potential to allow us to track the evolution of viral morphology through geological time.

Figure 2(a) Virus-like particle characterized by an icosahedral capsid-like structure (black arrow). Scale bar, 200 nm. (b) Virus-like particles. Black arrows point to the capsid-like structure in each case. Scale bar, 500 nm. (c) First stage of the amMg-Si mineralization process of the icosahedral capsid-like structure (black arrow); white arrow points to the viral DNA inside. Scale bar, 100 nm. (d) Second stage of the amMg-Si mineralization process of a virus-like particle (black arrow) showing its icosahedral capsid-like structure (white arrow). Scale bar, 100 nm. (e) Early mineralization of virus-like particles showing amMg-Si permineralized capsid-like structures (arrows). Scale bar, 200 nm. (f) amMg-Si permineralized virus-like particles occurring as single entities and chains (examples arrowed). Scale bar, 500 nm.



A Quote or Two from Hoeflmayr (1963): “Inhalation Therapy Using Bacteriophages in Therapy-Resistant Infections”

Stephen T. Abedon

Department of Microbiology – The Ohio State University – –


I just came across this “report”, which can be found here:

This dates from 1963, I believe as a translation, and the full citation is “Inhalation Therapy Using Bacteriophages in Therapy-Resistant Infections”. Fortschritte der Biologischen Aerosol-Forschung-Jahren 1957-1961 (Progress of the Biological Aerosol Research-Years 1957-1961), pp. 403-409. See under “Further reading” for what presumably is the original and/or complete citation.

At any rate, this paper/chapter/publication/translation/report has some interesting passages.

In view of the growing resistance against antibiotics, it is vitally important that we try to find ways to counteract this development. … Farsighted clinicians warned us as long as 10 years ago, when we were still students, that we should not hastily treat any little infection with penicillin.
If we should discover any-new possibilities for treating infections, then we should look at these possibilities only from the angle that such a therapy would have to preclude the formation of resistance as much as possible, therapy with bacteriophages fills the requirement. The fact that this therapy has so far met with skepticism is due to the results which, until a few years ago, did not ome up to expectations
[Schaefer, W., “Contribution on Epidemic Control” Vol. 3, Hippocrates, Stuttgart, 1948]. If we try to track down the reason for the failure of the earlier bacteriophage therapy, we will find that this was mostly due to the biological properties of the phages.
Now it is important to know that the bacteriophage has high specificity. Therefore, therapy can be effective
only If the administered phage encounters its homologous bacterium.
The disadvantage of our earlier phage preparations was to be found not only in the inadequate breeding methods but above all in the fact that only about 1-2 phage strains were available. If we consider the large number of pathogenic bacteria strains, which play a role even in a very simple infection or which at least at times might play a role, then we would have to set up two requirements. First of all, in order to have a wide range of effectiveness, such a therapeutic substance would have to contain a large number of various phage strains. Second, it is necessary that phages which would come into consideration for therapy should have sufficient virulence with regard to pathogenic viruses.
We used the preparation (Diriphagen ® Dr. Heinz Haury Chemical Plant, Munich) because we believed that this preparation met the requirements we just set up. According to Information received, this reparation contains 180-200 different phage strains and thus has a broad spectrum of effectiveness. In addition, it also contains so-called aimed antimicrobics which act against those bacteria that reveal primary phage resistance. We might note here that both the phage components and the added microbics in every ampule are standardized and meet the requirements for biological standardisation as regards phage effect [Penso, G., and Ortali, V., Arch. belges Med. Soc., 1, 1959]. If we mention the two therapeutic components, that is the bacteriphages and aimed (directed) antimicrobics, we are really not fully describing the effects mechanism as such. We have a third factor here. What we are dealing with here is the stimulation of the inherent defenses of the body which are bound to be aroused and which are based on the following: In breeding phages and antimicrobics, the pathogenic microbes used for this purpose give rise to lysates. But these lysates are not eliminated; instead they are also fed into the body. They act like antigens and lead to the formation of antibodies which in turn are specifically directed against the bacteria to which lyntes were added [Glauser, H. A., Med. achr., 13, 420, 1959.]. This reaction requires a latency period of about 8-10 days. The value of this antibody formation is hard to estimate in the individual case. We can get some specific figures on this only if we determine the phagocytosis capability; but this must be done in the clinic. Any new therapy is very often impaired by the fact that we do not employ it until other, more familiar measures have failed. We must admlt that we did not use Deriphagen until we had some patients in whom other preparations had not produced success. This is further by reports from other authors who achieved surprisingly good results with this preparation [Cevey, M., Schweiz. Z. Tuberk. (Swiss Tuberculosis Joural),
15, 34, 1958; Corbelli, G., Bologna Med., 6, 57, 1959; Delacoste, P., Rev. suisse Med., August 1959; Schaefer, W., “Contribution on Epidemic Control” Vol. 3, Hippoprates, Stuttgart, 1948].

Figure 1 shows the result of our treatment. The first column shows the total number of all patients treated; then we have the number of patients cured which abounted to 55.1%; then we cow to those who showed substantial improvement and on the right we have those patients who did not improve as a result of therapy [34.8%].

The author notes, however, that there is a discrepancy between microbiological and clinical results. That is, patients apparently reported a return to healthfulness but this did not coincide with elimination of pathogen, which the author seems to suggest is a consequence of phage- resistant forms not being pathogenic.

The text in the PDF then essentially fades away, though the main text of the paper continues on for two more pages!

Further reading:

Hoeflmayr, J. (1962). Inhalationstherapie mit Bakteriophagen bei therapieresistenten Infektionen [Inhalation Therapy with Bacteriophages for Treatment-Resistant Infections]. Fortschritte der biologischen Aerosol-Forschung in den Jahren 1957–1961 [Advances in Biological Aerosols Research in the Years 1957–1961].  403-409. 1962. (I believe this is the original reference)

Abedon, S. T., Kuhl, S., Blasdel, R., Kutter, E. M. (2011). Phage Treatment of Human Infections. Bacteriophage 1(2): 66-85. [PubMed link] (this article provides further historical context on European use of phage therapy, though note that description of a German tradition in that article is completely lacking and so far as I am aware was unknown to the authors at the time of its writing)