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A bacteriolytic agent that detects and kills Bacillus anthracis

Abstract

The dormant and durable spore form of Bacillus anthracis is an ideal biological weapon of mass destruction1,2. Once inhaled, spores are transported by alveolar macrophages to lymph nodes surrounding the lungs, where they germinate; subsequent vegetative expansion causes an overwhelming flood of bacteria and toxins into the blood, killing up to 99% of untreated victims. Natural and genetically engineered antibiotic-resistant bacilli amplify the threat of spores being used as weapons, and heighten the need for improved treatments and spore-detection methods after an intentional release. We exploited the inherent binding specificity and lytic action of bacteriophage enzymes called lysins for the rapid detection and killing of B. anthracis. Here we show that the PlyG lysin, isolated from the γ phage of B. anthracis, specifically kills B. anthracis isolates and other members of the B. anthracis ‘cluster’ of bacilli in vitro and in vivo. Both vegetative cells and germinating spores are susceptible. The lytic specificity of PlyG was also exploited as part of a rapid method for the identification of B. anthracis. We conclude that PlyG is a tool for the treatment and detection of B. anthracis.

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Figure 1: Characterization of PlyG.
Figure 2: The specific and rapid killing action of PlyG.
Figure 3: PlyG causes profound morphological changes in, and ultimately lysis of, RSVF1.
Figure 4: Survival of PlyG-treated BALB/c mice infected with RSVF1.
Figure 5: PlyG-mediated spore killing and detection.

References

  1. Mock, M. & Fouet, A. Anthrax. Annu. Rev. Microbiol. 55, 647–671 (2001)

    CAS  Article  Google Scholar 

  2. Inglesby, T. V. et al. Anthrax as a biological weapon, 2002: Updated recommendations for management. J. Am. Med. Assoc. 287, 2236–2252 (2002)

    Article  Google Scholar 

  3. Wang, I. N., Smith, D. L. & Young, R. Holins: The protein clocks of bacteriophage infections. Annu. Rev. Microbiol. 54, 799–825 (2000)

    CAS  Article  Google Scholar 

  4. Lopez, R., Garcia, E., Garcia, P. & Garcia, J. L. The pneumococcal cell wall degrading enzymes: A modular design to create new lysins? Microb. Drug Resist. 3, 199–211 (1997)

    CAS  Article  Google Scholar 

  5. Loessner, M. J., Kramer, K., Ebel, F. & Scherer, S. C-terminal domains of Listeria monocytogenes bacteriophage murein hydrolases determine specific recognition and high-affinity binding to bacterial cell wall carbohydrates. Mol. Microbiol. 44, 335–349 (2002)

    CAS  Article  Google Scholar 

  6. Loeffler, J. M., Nelson, D. & Fischetti, V. A. Rapid killing of Streptococcus pneumoniae with a bacteriophage cell wall hydrolase. Science 294, 2170–2172 (2001)

    ADS  CAS  Article  Google Scholar 

  7. Nelson, D., Loomis, L. & Fischetti, V. A. Prevention and elimination of upper respiratory colonization of mice by group A streptococci by using a bacteriophage lytic enzyme. Proc. Natl Acad. Sci. USA 98, 4107–4112 (2001)

    ADS  CAS  Article  Google Scholar 

  8. Redmond, C., Henderson, I., Turnbull, P. C. B. & Bowen, J. Phage from different strains of Bacillus anthracis. Salisbury Med. Bull. Spec. Suppl. 87, 60–63 (1996)

    Google Scholar 

  9. Brown, E. R. & Cherry, W. B. Specific identification of Bacillus anthracis by means of a variant bacteriophage. J. Infect. Dis. 96, 34–39 (1955)

    CAS  Article  Google Scholar 

  10. Turnbull, P. C. B. Definitive identification of Bacillus anthracis—a review. J. Appl. Microbiol. 87, 237–240 (1999)

    CAS  Article  Google Scholar 

  11. Helgason, E. et al. Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis—one species on the basis of genetic evidence. Appl. Environ. Microbiol. 66, 2627–2630 (2000)

    CAS  Article  Google Scholar 

  12. Burdon, K. L., Davis, J. S. & Wende, R. D. Experimental infection of mice with Bacillus cereus: Studies of pathogenesis and pathologic changes. J. Infect. Dis. 117, 307–316 (1967)

    CAS  Article  Google Scholar 

  13. Lamanna, C. & Jones, L. Lethality for mice of vegetative and spore forms of Bacillus cereus and Bacillus cereus-like insect pathogens injected intraperitoneally and subcutaneously. J. Bacteriol. 85, 532–535 (1963)

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Fischetti, V. A. Phage antibacterials make a comeback. Nature Biotechnol. 19, 734–735 (2001)

    CAS  Article  Google Scholar 

  15. Makino, S., Ito, N., Inoue, T., Miyata, S. & Moriyama, R. A spore-lytic enzyme released from Bacillus cereus spores during germination. Microbiology 140, 1403–1410 (1994)

    CAS  Article  Google Scholar 

  16. Jenkinson, H. F., Kay, D. & Mandelstam, J. Temporal dissociation of late events in Bacillus subtilis sporulation from expression of genes that determine them. J. Bacteriol. 141, 793–805 (1980)

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Henriques, A. O. & Moran, C. P. Jr. Structure and assembly of the bacterial endospore coat. Methods 20, 95–110 (2000)

    CAS  Article  Google Scholar 

  18. Santo, L. Y. & Doi, R. H. Ultrastructural analysis during germination and outgrowth of Bacillus subtilis spores. J. Bacteriol. 120, 475–481 (1974)

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Brussow, H. & Hendrix, R. W. Phage genomics: Small is beautiful. Cell 108, 13–16 (2002)

    CAS  Article  Google Scholar 

  20. Helgason, E., Caugant, D. A., Olsen, I. & Kolsto, A. B. Genetic structure of population of Bacillus cereus and B. thuringiensis isolates associated with periodontitis and other human infections. J. Clin. Microbiol. 38, 1615–1622 (2000)

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Pannucci, J., Okinaka, R. T., Sabin, R. & Kuske, C. R. Bacillus anthracis pXO1 plasmid sequence conservation among closely related bacterial species. J. Bacteriol. 184, 134–141 (2002)

    CAS  Article  Google Scholar 

  22. Ticknor, L. O. et al. Fluorescent amplified fragment length polymorphism analysis of norwegian Bacillus cereus and Bacillus thuringiensis soil isolates. Appl. Environ. Microbiol. 67, 4863–4873 (2001)

    CAS  Article  Google Scholar 

  23. Schmitt, C. K. et al. Absence of all components of the flagellar export and synthesis machinery differentially alters virulence of Salmonella enterica serovar Typhimurium in models of typhoid fever, survival in macrophages, tissue culture invasiveness, and calf enterocolitis. Infect. Immun. 69, 5619–5625 (2001)

    CAS  Article  Google Scholar 

  24. Jackson, P. J. et al. Characterization of the variable-number tandem repeats in vrrA from different Bacillus anthracis isolates. Appl. Environ. Microbiol. 63, 1400–1405 (1997)

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Guzman, L. M., Belin, D., Carson, M. J. & Beckwith, J. Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J. Bacteriol. 177, 4121–4130 (1995)

    CAS  Article  Google Scholar 

  26. Loessner, M. J., Wendlinger, G. & Scherer, S. Heterogeneous endolysins in Listeria monocytogenes bacteriophages: A new class of enzymes and evidence for conserved holin genes within the siphoviral lysis cassettes. Mol. Microbiol. 16, 1231–1241 (1995)

    CAS  Article  Google Scholar 

  27. Stopa, P. J., Tieman, D., Coon, P. A., Milton, M. M. & Paterno, D. Detection of biological aerosols by luminescence techniques. Field Anal. Chem. Technol. 3, 283–290 (1999)

    CAS  Article  Google Scholar 

  28. Corran, J. The induction of supersuppressor mutants of Bacillus subtilis by ethyl methanesulphonate and the posttreatment modification of mutation yield. Mol. Gen. Genet. 103, 42–57 (1968)

    CAS  Article  Google Scholar 

  29. Mazas, M., Martinez, S., Lopez, M., Alvarez, A. B. & Martin, R. Thermal inactivation of Bacillus cereus spores affected by the solutes used to control water activity of the heating medium. Int. J. Food Microbiol. 53, 61–67 (1999)

    CAS  Article  Google Scholar 

  30. Keim, P. et al. Multiple-locus variable-number tandem repeat analysis reveals genetic relationships within Bacillus anthracis. J. Bacteriol. 182, 2928–2936 (2000)

    CAS  Article  Google Scholar 

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Acknowledgements

We thank P. J. Piggot, L. W. Mayer, A. L. Turetsky, A. Aronson, A. Keynan, H.-W. Ackerman, R. J. McNall and T. A. Kokjohn for their gifts of strains; E. Sphicas at the Bio-imaging Resource Center at The Rockefeller University for help with electron microscopy; R. L. Russell and S. Zhu for technical help; New Horizons Diagnostics for their luminometer and reagents and P. Model and members of the V.A.F. laboratory for reviewing this manuscript. We also thank A. Keynan for reviewing this manuscript and for advice regarding spore preparation and germination. This work was supported by a grant from the Defense Advanced Research Projects Agency (DARPA).

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Schuch, R., Nelson, D. & Fischetti, V. A bacteriolytic agent that detects and kills Bacillus anthracis. Nature 418, 884–889 (2002). https://doi.org/10.1038/nature01026

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