The threat of antibiotic-resistant bacteria is driving researchers to think up ever more clever ways to tackle infections. An enzyme from a bacterium-killing virus may prove effective against anthrax infections.
Following the deliberate dissemination of spores of Bacillus anthracis, the bacterium that causes anthrax, through the US mail last autumn, public-health officials were faced with two major problems: detecting spores in buildings and on exposed individuals, and treating those people thought to be exposed and the few who actually became infected. Testing for spores required samples to be sent to specialized labs to be cultured, and took several days — a painfully long time when the public was insisting on immediate information and action. And thousands of people who were thought to have been exposed were treated with antibiotics, usually ciprofloxacin. Fortunately, nobody treated preventatively in this way became infected; the intervention was effective because the particular strain used in the attack is wholly susceptible to the usual antibiotics. But the situation could have been much worse if the strain had been resistant to antibiotics.
On page 884 of this issue, Schuch and colleagues1 describe an innovative tool that might help to solve all these problems by rapidly detecting B. anthracis spores and curing an infection — even one resistant to standard antibiotics. The group's antimicrobial strategy originated in earlier work by the senior author, Vincent Fischetti, and defines a new class of antibiotics that is distinct from any previously available.
Recent years have seen a growing awareness of the problem of antibiotic-resistant bacteria. Even defined environments such as hospitals present difficulties in dealing with antibiotic-resistant variants of common bacterial infections, suggesting that deliberate, widespread dissemination of spores of an antibiotic-resistant B. anthracis strain could cause a far more serious public-health emergency than that which occurred last autumn (Fig. 1). These concerns — along with worries about the current anthrax vaccine that have led some in the US military to refuse immunization — have spurred intense research into alternative forms of preventing and treating B. anthracis infections.
Effective and more acceptable vaccines are being developed2. However, these, like many other vaccines, will require multiple immunizations, and time for protection to build up. To be effective, a vaccine would need to be administered well in advance of an attack. On another tack, recent breakthroughs in understanding the structure and function of anthrax toxin — one of the two major components the bacterium needs for its virulence — have led to candidate drugs that directly interfere with toxin assembly and function3,4,5,6. Such products may protect against the toxic effects of anthrax. But drugs that destroy the bacteria will also be important.
One technique for tackling bacterial infections is to use bacteriophage — viruses that infect and burst open (lyse) specific bacteria. Bacteriophage therapy has had a chequered history, but the recent successful application of a genetic means of selecting viral variants that resist elimination from the human blood circulation7 suggests that modern techniques might resurrect the practice. In an elegant variation on this classical approach, Schuch and colleagues1 have exploited a single key enzyme of bacteriophage γ, a virus often used in clinical labs to identify B. anthracis8,9. The authors first purified this enzyme, PlyG, which is a type of lysin; in the normal course of the bacteriophage life cycle, it destroys the bacterial cell wall so as to allow the release of progeny phage. Lysins are typically highly specific for particular species or strains of bacteria, because their binding regions are directed towards cell-wall carbohydrate structures that vary greatly among species and strains10,11. Accordingly, the PlyG lysin described by Schuch et al. was highly effective in killing B. anthracis, but did not affect most strains of the closely related species Bacillus cereus and Bacillus thuringiensis.
Although bacteriophage lysins are made inside bacteria and normally cause lysis from within, the authors showed that PlyG added externally had strong lytic activity towards all the B. anthracis strains tested when they were grown on solid or liquid media. Most importantly, Schuch et al. also found that when mice were injected with PlyG 15 minutes after being infected with a close relative of B. anthracis, about 80% of the animals were rescued from otherwise certain death. An obvious concern is that some of the mice might have died because PlyG-resistant mutants of B. anthracis had arisen. However, a controlled test showed that PlyG-resistant mutants were not generated in B. anthracis cultures, even when the cultures were treated chemically in a way that increased the percentage of mutants resistant to standard antibiotics.
Schuch et al. suggest that the absence of the development of resistance to PlyG is due to the fact that any mutational change to the cell-wall structure that prevents binding to PlyG would kill the bacterium. This targeting of phage lysins to an essential bacterial structure gives them an advantage over the small-molecule antibiotics to which bacteria can become resistant rather easily. One drawback, however, shared by many other candidate antibacterial tools, is that PlyG would have to be administered soon after a person became infected, before lethal levels of anthrax toxin could develop. As illustrated by recent human cases, it is often difficult to diagnose anthrax infections early enough for treatment to be effective.
It is here that PlyG has another possible application. Currently, environmental and clinical samples suspected of being contaminated by anthrax are usually sent to specialized labs for culture. One of the tests used at the Centers for Disease Control and Prevention in the United States involves checking cultured bacteria for sensitivity to bacteriophage γ (refs 1, 8, 9). But all culture methods are slow. Efforts are under way to develop quicker tests using, for example, molecular approaches that require DNA extraction and special instrumentation. However, such sophisticated methods are difficult to deploy outside the lab.
Schuch et al. have now shown that PlyG can be used in a simple system to rapidly detect B. anthracis spores. Spores are resistant to PlyG-induced lysis, so the authors added the amino acid l-alanine to trigger germination, and then treated the emerging bacteria with PlyG, causing lysis and the release of bacterial components. One of these components is ATP, the main cellular energy store, which the authors detected using a luciferase–luciferin system: the luciferase enzyme degrades luciferin in the presence of ATP, producing light, which can be detected with a hand-held luminometer. This type of system would readily lend itself to field detection of spore contamination, because the equipment could be easily incorporated into a relatively small and simple device. One limitation is that it would not distinguish between virulent and non-virulent strains of B. anthracis, because PlyG would lyse the bacteria regardless of whether or not they produced the two factors needed for full virulence — anthrax toxin and a polyglutamate capsule. Presumably, however, the test could be used outside the lab as a rapid first indicator of contamination by B. anthracis, and further characterization would follow.
There is still much to be done to develop PlyG into an effective drug. For example, it would probably need to be administered intravenously in a formulation that would give adequate concentrations in the blood, because that is where the bacteria grow rapidly during the dangerous final stage of infection. Nonetheless, Schuch et al.1 have introduced a potential treatment for anthrax that might be useful either alone or in combination with other therapies. Similarly, their means of detecting B. anthracis spores may prove valuable as the basis for an easily deployable test. Given these promising results, we expect to see rapid moves to test the use of bacteriophage lysins in detecting and treating other bacterial infections.
Schuch, R., Nelson, D. & Fischetti, V. A. Nature 418, 884–889 (2002).
Leppla, S. H., Robbins, J. B., Schneerson, R. & Shiloach, J. J. Clin. Invest. 110, 141–144 (2002).
Mourez, M. et al. Trends Microbiol. 10, 287–293 (2002).
Chaudry, G. J. et al. Trends Microbiol. 10, 58–62 (2002).
Pannifer, A. D. et al. Nature 414, 229–233 (2001).
Maynard, J. A. et al. Nature Biotechnol. 20, 597–601 (2002).
Biswas, B. et al. Infect. Immun. 70, 204–210 (2002).
Inglesby, T. V. et al. J. Am. Med. Assoc. 287, 2236–2252 (2002).
Brown, E. R. & Cherry, W. B. J. Infect. Dis. 96, 34–39 (1955).
Lopez, R., Garcia, E., Garcia, P. & Garcia, J. L. Microb. Drug Resist. 3, 199–211 (1997).
Loessner, M. J., Kramer, K., Ebel, F. & Scherer, S. Mol. Microbiol. 44, 335–349 (2002).
About this article
Investigations on the Interactions of λPhage-Derived Peptides Against the SrtA Mechanism in Bacillus anthracis
Applied Biochemistry and Biotechnology (2014)
Journal of Environmental Science and Technology (2014)
Phage lysin to control the overgrowth of normal flora in processed sputum samples for the rapid and sensitive detection of Mycobacterium tuberculosis by luciferase reporter phage assay
BMC Infectious Diseases (2013)
Complete Nucleotide Sequence and Molecular Characterization of Bacillus Phage TP21 and its Relatedness to Other Phages with the Same Name
Bioengineered Bugs (2010)