BMB Reports

Korean Society for Biochemistry and Molecular Biology (생화학분자생물학회)
권호 표지
DOI QR코드

DOI QR Code

Detection of the cell wall-affecting antibiotics at sublethal concentrations using a reporter Staphylococcus aureus harboring drp35 promoter - lacZ transcriptional fusion

  • Mondal, Rajkrishna (Department of Biochemistry, Bose Institute) ;
  • Chanda, Palas K. (Department of Biochemistry, Bose Institute) ;
  • Bandhu, Amitava (Department of Biochemistry, Bose Institute) ;
  • Jana, Biswanath (Department of Biochemistry, Bose Institute) ;
  • Lee, Chia-Y. (Department of Microbiology and Immunology, University of Arkansas for Medical Sciences) ;
  • Sau, Subrata (Department of Biochemistry, Bose Institute)
  • Received : 2010.04.12
  • Accepted : 2010.05.26
  • Published : 2010.07.31
Download PDF
⟨ Previous Next ⟩

Abstract

Previously, various inhibitors of cell wall synthesis induced the drp35 gene of Staphylococcus aureus efficiently. To determine whether drp35 could be exploited in antistaphylococcal drug discovery, we cloned the promoter of drp35 ($P_d$) and developed different biological assay systems using an engineered S. aureus strain that harbors a chromosomally-integrated $P_d$ - lacZ transcriptional fusion. An agarose-based assay showed that $P_d$ is induced not only by the cell wall-affecting antibiotics but also by rifampicin and ciprofloxacin. In contrast, a liquid medium-based assay revealed the induction of $P_d$ specifically by the cell wall-affecting antibiotics. Induction of $P_d$ by sublethal concentrations of cell wall-affecting antibiotics was even assessable in a microtiter plate assay format, indicating that this assay system could be potentially used for high-throughput screening of new cell wall-inhibiting compounds.

Keywords

References

  1. Draghi, D. C., Sheehan, D. F., Hogan, P. and Sahm, D. F. (2006) Current antimicrobial resistance profiles among methicillin-resistant Staphylococcus aureus encountered in the outpatient setting. Diagn. Microbiol. Infect. Dis. 55, 129-133. https://doi.org/10.1016/j.diagmicrobio.2006.01.003
  2. Feiz, V. and Redline, D. E. (2007) Infectious scleritis after pars plana vitrectomy because of methicillin-resistant Staphylococcus aureus resistant to fourth-generation fluoroquinolones. Cornea. 26, 238-240.
  3. Howden, B. P., Davies, J. K., Johnson, P. D., Stinear, T. P. and Grayson, M. L. (2010) Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection, and clinical implications. Clin. Microbiol. Rev. 23, 99-139. https://doi.org/10.1128/CMR.00042-09
  4. Stryjewski, M. E. and Corey, G. R. (2009) New treatments for methicillin-resistant Staphylococcus aureus. Curr. Opin. Crit. Care. 15, 403-412. https://doi.org/10.1097/MCC.0b013e32832f0a74
  5. Murakami, H., Matsumaru, H., Kanamori, M., Hayashi, H. and Ohta, T. (1999). Cell wall-affecting antibiotics induce expression of a novel gene, drp35, in Staphylococcus aureus. Biochem. Biophys. Res. Commun. 264, 348-351. https://doi.org/10.1006/bbrc.1999.1388
  6. Tanaka, Y., Morikawa, K., Ohki, Y., Yao, M., Tsumoto, K., Watanabe, N., Ohta, T. and Tanaka, I. (2007) Structural and mutational analyses of drp35 from Staphylococcus aureus: a possible mechanism for its lactonase activity. J. Biol. Chem. 282, 5770-5780. https://doi.org/10.1074/jbc.M607340200
  7. Morikawa, K., Hidaka, T., Murakami, H., Hayashi, H. and Ohta. T. (2005) Staphylococcal drp35 is the functional counterpart of the eukaryotic PONs. FEMS. Microbiol. Lett. 249, 185-190. https://doi.org/10.1016/j.femsle.2005.06.038
  8. Osburne, M. S., Maiese, W. M. and Greenstein, M. (1993) An assay for the detection of bacterial DNA gyrase inhibitors. J. Antibiot. 46, 1764-1766. https://doi.org/10.7164/antibiotics.46.1764
  9. Ulijasz, A. T, Grenader, A. and Weisblum, B. (1996) A vancomycin-inducible lacZ reporter system in Bacillus subtilis: induction by antibiotics that inhibit cell wall synthesis and by lysozyme. J. Bacteriol. 178, 6305-6309. https://doi.org/10.1128/jb.178.21.6305-6309.1996
  10. Bianchi, A. A. and Baneyx, F. (1999) Stress responses as a tool to detect and characterize the mode of action of antibacterial agents. Appl. Environ. Microbiol. 65, 5023-5027.
  11. Tenhami, M., Hakkila, K. and Karp, M. (2001) Measurement of effects of antibiotics in bioluminescent Staphylococcus aureus RN4220. Antimicrob Agents Chemother. 45, 3456-3461. https://doi.org/10.1128/AAC.45.12.3456-3461.2001
  12. Shapiro, E. and Baneyx, F. (2002) Stress-based identification and classification of antibacterial agents: secondgeneration Escherichia coli reporter strains and optimization of detection. Antimicrob. Agents. Chemother. 46, 2490-2497. https://doi.org/10.1128/AAC.46.8.2490-2497.2002
  13. Chanda, P. K., Ganguly, T., Das, M., Lee, C. Y., Luong, T. T. and Sau, S. (2007) Detection of antistaphylococcal and toxic chemicals by biological assay systems developed with a reporter Staphylococcus aureus strain harboring a heat shock promoter - lacZ fusion. J. Biochem. Mol. Biol. 40, 936-943. https://doi.org/10.5483/BMBRep.2007.40.6.936
  14. Steidl, R., Pearson, S., Stephenson, R. E., Ledala, N., Sitthisak, S., Wilkinson, B. J. and Jayaswal, R. K. (2008) Staphylococcus aureus cell wall stress stimulon gene-lacZ fusion strains: potential for use in screening for cell wall-active antimicrobials. Antimicrob. Agents. Chemother. 52, 2923-2925. https://doi.org/10.1128/AAC.00273-08
  15. Mesak, L. R., Miao, V. and Davies, J. (2008) Effects of subinhibitory concentrations of antibiotics on SOS and DNA repair gene expression in Staphylococcus aureus. Antimicrob. Agents. Chemother. 52, 3394-3397. https://doi.org/10.1128/AAC.01599-07
  16. Chanda, P. K., Mondal, R., Sau, K. and Sau, S. (2009) Antibiotics, arsenate and $H_2O_2$ induce the promoter of Staphylococcus aureus cspC gene more strongly than cold. J. Basic Microbiol. 49, 205-211. https://doi.org/10.1002/jobm.200800065
  17. Sau, S., Chattoraj, P., Ganguly, T., Chanda, P. K. and. Mandal, N.C (2008) Inactivation of bacterial indispensable proteins by early/delayed early proteins of bacteriophages: implication in antibacterial drug discovery. Curr. Protein Pept Sci. 9, 284-290. https://doi.org/10.2174/138920308784533970
  18. Berg, J. D. and Fiksdal, L. (1988) Rapid detection of total and fecal coliforms in water by enzymatic hydrolysis of 4-methylumbelliferone-beta-D-galactoside. Appl. Environ. Microbiol. 54, 2118-2122.
  19. Miller, J. M. (1972) Experiments in Molecular Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor. N. Y.
  20. Sambrook, J. and Russell, D. W. (2001) Molecular Cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Laboratory Press, CSH, New York, USA.
  21. Ausubel, F. M., Brent, R., Kingston, R. E., Moore, D. D., Seidman, J. G., Smith, J. A. and Struhl, K. (1998) Current Protocols in Molecular Biology. John Wiley & Sons, Inc., USA.
  22. Lee, C. Y. and Iandolo, J. J. (1986) Integration of staphylococcal phage L54a occurs by site-specific recombination: structural analysis of the attachment sites. Proc. Natl. Acad. Sci. U.S.A. 83, 5474-5478. https://doi.org/10.1073/pnas.83.15.5474
  23. Kemp, E. H., Sammons, R. L., Moir, A., Sun, D. and Setlow, P. (1991) Analysis of transcriptional control of the gerD spore germination gene of Bacillus subtilis 168. J Bacteriol. 173, 4646-4652. https://doi.org/10.1128/jb.173.15.4646-4652.1991