Detection of Neuraminidase Activity in Pseudomonas aeruginosa PAO1

Document Type : Original Article

Authors

1 Department of Microbiology, Faculty of Veterinary, University of Urmia, Urmia, Iran

2 Department of Microbiology, Faculty of Veterinary Medicine, University of Urmia, Urmia, Iran

3 Department of Microbiology, Immunology and Genetics, Faculty of Medicine, Urmia University of Medical Sciences, Uremia, Iran

Abstract

Objective(s)
Some properties of neuraminidase produced by Pseudomonas aeruginosa PAO1 growth in a defined medium (BHI) were examined and evaluated for its features.
Materials and Methods
The obtained supernatant enzyme of P. aeruginosa PAO1 cultures was used in a sensitive fluorometric assay by using 2'-(4-methylumbelliferyl) a-D-N acetylneuraminic acid as substrate. As hydrolyzing MUN with neuraminidase; free N-acetylneuraminic acid and 4-methylumbelliferone were formed with a shift in the fluorescence spectra from 315/374 nm (substrate) to 365/450 nm (product). Enzyme activity was then measured by the fluorescence of 4-methylumbelliferone at 450 nm.
Results
Among the culture media to determine the enzyme production, the highest production of P. aeruginosa PAO 1 neuraminidase was found in BHI culture media. Neuraminidase production in P. aeruginosa PAO1 paralleled bacterial growth in defined medium (BHI) and was maximal in the late logarithmic phase of growth but decreased during the stationary phase, probably due to protease production or thermal instability. The neuraminidase of P. aeruginosa PAO1 possessed an optimum temperature of 56 °C and the activity was pH-dependent with maximal activity at pH 5. Heating the enzyme at 56 °C for 45 min in the presence of bovine serum albumin destroyed 33.1% of the activity while the addition of Ca+2, EDTA and N-acetyl neuraminic acid (NANA) decreased activity markedly.
Conclusion
Overall, the results indicated that neuraminidase of P. aeruginosa PAO1 is more an extracellular enzyme than K. pneumonia neuraminidase is.

Keywords

References

1.Cacalano G, Kays M, Saiman L, Prince A. Production of Pseudomonas aeruginosa neuraminidase is increased under hyperosmolar conditions and is regulated by genes involved in alginate expression. J Clin Invest 1992; 89:1866-1874.
2.Soong G, Muir A., Gomez Marisa I, Waks J, Reddy B, Planet P, et al. Bacterial neuraminidase facilitates mucosal infection by participating in biofilm production. J Clin Invest 2006; 116:2297-2305.
3.Camera M, Boulnois GJ, Andrew PW, Mitchell TJ. A neuraminidase from Streptococcus pneumoniae has the features of a surface protein. Infect Immun 1994; 62:3688-3695.
4.Shahjahan K, Nayeema A, Showkat A. Neuraminidase production by Vibrio cholerae o1 and other diarrheagenic bacteria. Infect Immun 1984; 49:747-749.
5.Straus DC, Portnoy-Duran C. Neuraminidase production by a Streptococcus sanguis strain associated with subacute bacterial endocarditis. Infect Immun 1983; 41:507-515.
6.Moncla BJ, Braham P. Detection of sialidase (neuraminidase) activity in actinomyces species by using 2'-(4- methylumbelliferyl) o1-d-n-acetylneuraminic acid in a filter paper spot test. J Clin Microbiol 1989; 27:182-184.
7.Winter AJ, Spiro DC, Osborne MP, Tarlow MJ, Stephen J, Andrew PE, et al. A role for pneumolysin but not neuraminidase in the hearing loss and cochlear damage induced by experimental pneumococcal meningitis in guinea pigs. Infect Immun 1997; 65:4411-4418.
8.Briselden A, Moncla BJ, Stevens CE, Sharon LH. Sialidases (neuraminidases) in bacterial vaginosis and bacterial vaginosis-associated microflora. J Clin Microbiol 1992; 30:663-666.
9.Anon.,               Protein determination-lowry      procedure.         Available              at:
http://wwwbiodavidsonedu/people/jowilliamson/Techniques/Protocolweek5html 2000.
10.Berg JO, Lindqvist L, Andersson G, Nord CE. Neuraminidase in Bacteroides fragilis. Appl Environ Microbiol 1983; 46:75-80.
11.Popoff MR, Dodin A. Survey of neuraminidase production by Clostridium butyricum, C. beijerinckii, and C. difficile strains from clinical and nonclinical sources. J Clin Microbiol 1985; 22:873-576.
12.White DJ, Jolley WL, Purdy CW, Straus DC.Extracellular neuraminidase production by a Pasteurella multocidaa: 3 strain associated with bovine pneumonia. Infect Immun 1995; 63: 1703-1709.
13.Burns B, Mendz G, Hazell S. Methods for the measurement of a bacterial enzyme activity in cell lysates and extracts. Biol Proced Online 1998; 1:17-26.
14.Ryan K, Ray CG. Sherris medical microbiology.4th ed. McGraw Hill; 2004.
15.Murray P, Rosenthal K, Pfaller MA. Medical microbiology. New York, NY: Elsevier Press; 2005.
16.Moncla BJ, Braham, P, Hillier SL. Sialidase (neuraminidase) activity among gram-negative anaerobic and capnophilic bacteria. J Clin Microbiol 1990; 28:422-425.
17.Flashner M, Kessler J, Tanenbaum SW. Structural requirements for neuraminidase induction in Arthrobacter sialophilus. J Bacteriol 1982; 151:1630-1632.
18.Vimr E, Kalivoda KA, Deszo EL, Steenbergen SM. Diversity of microbial sialic acid metabolism. Microbiol Mol Biol Rev 2004; 68:132-153.
19.Songer JG, Post k. W Veterinary microbiology. Elsevier Saunders; 2005.
20.Abrashev I, Dulguerova G. Neuraminidases (sialidases) from bacterial origin. Exp Pathol Parasitol 2001; 4:35-40.
21.Nikaido H. Molecular basis of bacterial outer membrane permeability revisited. Microbiol Mol Biol Rev 2003; 67:1092-2172.
22.Prithiviraj B, Bais H, Weir T, Suresh B, Najarro E, Dayakar B , et al. Down-regulation of virulence factors of Pseudomonas aeruginosa by salicylic acid attenuates its virulence on Arabidopsis thaliana and Caenorhabditis elegans. Infect Immun 2005; 73:5319-5320.
23.Naomi IA, Masami O, Hiroshi O, Sadahiko I, Misuzu K. Interactions between cations in modifying the binding of hexokinases i and ii to mitochondria. Mol Cell Biochem 1988; 81:37-41.

Files

Share

How to cite

Statistics