Publications by authors named "Pramukh N Jayasekera"

4 Publications

  • Page 1 of 1

The cell membrane as a major site of damage during aerosolization of Escherichia coli.

Appl Environ Microbiol 2011 Feb 10;77(3):920-5. Epub 2010 Dec 10.

Defence Science & Technology Laboratory, Porton Down, Salisbury, Wiltshire SP4 0JQ, United Kingdom.

This study aimed to provide data on the survival and site of damage of Escherichia coli cells following aerosolization using two different techniques, nebulization and flow focusing. Four metabolic stains were assessed for their ability to detect respiratory activities and membrane homeostasis in aerosolized E. coli cells. The degree of sublethal injury increased significantly over the 10-min period of aerosolization in E. coli cells aerosolized by using the Collison nebulizer, reaching up to 99.9% of the population. In contrast, a significantly lower proportion of the population was sublethally damaged during aerosolization using the flow-focusing aerosol generator (FFAG). Concomitantly, loss of membrane homeostasis increased at a higher rate in nebulized cells (68 to 71%) than in those aerosolized by using the FFAG (32 to 34%). The activities of respiratory enzymes decreased at increased rates in nebulized cells (27 to 37%) compared to the rates of decrease in cells aerosolized by using the FFAG (59 to 61%). The results indicate that the physiology of an aerosolized bacterium is linked to the method of aerosol generation and may affect the interpretation of a range of aerobiological phenomenon.
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http://dx.doi.org/10.1128/AEM.01116-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3028717PMC
February 2011

Different pathologies but equal levels of responsiveness to the recombinant F1 and V antigen vaccine and ciprofloxacin in a murine model of plague caused by small- and large-particle aerosols.

Infect Immun 2009 Apr 2;77(4):1315-23. Epub 2009 Feb 2.

Defence Science & Technology Laboratory (Dstl), Porton Down, Salisbury, Wiltshire SP40JQ, United Kingdom.

Presently there is a significant effort to develop and evaluate vaccines and antibiotics against the potential bioterrorism agent Yersinia pestis. The animal models used to test these countermeasures involve the deposition of small particles within the lung. However, deliberate aerosol release of Y. pestis will generate both small and large inhalable particles. We report in this study that the pathogenesis patterns of plague infections caused by the deposition of 1- and 12-microm-particle aerosols of Y. pestis in the lower and upper respiratory tracts (URTs) of mice are different. The median lethal dose for 12-mum particles was 4.9-fold greater than that for 1-microm particles. The 12-microm-particle infection resulted in the degradation of the nasal mucosa and nasal-associated lymphoid tissue (NALT) plus cervical lymphadenopathy prior to bacteremic dissemination. Lung involvement was limited to secondary pneumonia. In contrast, the 1-microm-particle infection resulted in primary pneumonia; in 40% of mice, the involvement of NALT and cervical lymphadenopathy were observed, indicating entry via both URT lymphoid tissues and lungs. Despite bacterial deposition in the gastrointestinal tract, the involvement of Peyer's patches was not observed in either infection. Although there were major differences in pathogenesis, the recombinant F1 and V antigen vaccine and ciprofloxacin protected against plague infections caused by small- and large-particle aerosols.
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http://dx.doi.org/10.1128/IAI.01473-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2663150PMC
April 2009

Characterization and deposition of respirable large- and small-particle bioaerosols.

Appl Environ Microbiol 2008 Oct 22;74(20):6437-43. Epub 2008 Aug 22.

Defence Science and Technology Laboratory, Salisbury, Wiltshire, United Kingdom.

The deposition patterns of large-particle microbiological aerosols within the respiratory tract are not well characterized. A novel system (the flow-focusing aerosol generator [FFAG]) which enables the generation of large (>10-microm) aerosol particles containing microorganisms under laboratory conditions was characterized to permit determination of deposition profiles within the murine respiratory tract. Unlike other systems for generating large aerosol particles, the FFAG is compatible with microbiological containment and the inhalational challenge of animals. By use of entrapped Escherichia coli cells, Bacillus atrophaeus spores, or FluoSphere beads, the properties of aerosols generated by the FFAG were compared with the properties of aerosols generated using the commonly available Collison nebulizer, which preferentially generates small (1- to 3-microm) aerosol particles. More entrapped particulates (15.9- to 19.2-fold) were incorporated into 9- to 17-microm particles generated by the FFAG than by the Collison nebulizer. The 1- to 3-microm particles generated by the Collison nebulizer were more likely to contain a particulate than those generated by the FFAG. E. coli cells aerosolized using the FFAG survived better than those aerosolized using the Collison nebulizer. Aerosols generated by the Collison nebulizer and the FFAG preferentially deposited in the lungs and nasal passages of the murine respiratory tract, respectively. However, significant deposition of material also occurred in the gastrointestinal tract after inhalation of both the small (89.7%)- and large (61.5%)-particle aerosols. The aerosols generated by the Collison nebulizer and the FFAG differ with respect to mass distribution, distribution of the entrapped particulates, bacterial survival, and deposition within the murine respiratory tract.
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http://dx.doi.org/10.1128/AEM.01194-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2570277PMC
October 2008

Clostridium absonum alpha-toxin: new insights into clostridial phospholipase C substrate binding and specificity.

J Mol Biol 2003 Oct;333(4):759-69

School of Crystallography, Birkbeck College, Malet Street, London WC1E 7HX, UK.

Clostridium absonum phospholipase C (Caa) is a 42.7 kDa protein, which shows 60% amino acid sequence identity with the Clostridium perfringens phospholipase C, or alpha-toxin (Cpa), and has been isolated from patients suffering from gas gangrene. We report the cloning and sequencing, purification, characterisation and crystal structure of the Caa enzyme. Caa had twice the phospholipid-hydrolysing (lecithinase) activity, 1.5 times the haemolytic activity and over seven times the activity towards phosphatidylcholine-based liposomes when compared with Cpa. However, the Caa enzyme had a lower activity than Cpa to the free (i.e. not in lipid bilayer) substrate para-nitrophenylphosphorylcholine, towards sphingomyelin-based liposomes and showed half the cytotoxicity. The lethal dose (LD(50)) of Caa in mice was approximately twice that of Cpa. The crystal structure of Caa shows that the 72-93 residue loop is in a conformation different from those of previously determined open-form alpha-toxin structures. This conformational change suggests a role for W84 in membrane binding and a possible route of entry into the active site along a hydrophobic channel created by the re-arrangement of this loop. Overall, the properties of Caa are compatible with a role as a virulence-determinant in gas gangrene caused by C.absonum.
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http://dx.doi.org/10.1016/j.jmb.2003.07.016DOI Listing
October 2003
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