Abstract
Microscopic analysis is a well-accepted technique in microbiology to characterize a single colony and single cell morphotypes. Although colony morphotype can be imaged by light microscopy, morphotypes of microbial cells are normally viewed by transmission electron microscopy (TEM). The specific location of proteins on microbial cell surface or inside the cells is normally detected using immunoelectron microscopy (IEM). Here we describe the technique of detection of type-specific cellular appendages that are produced by several strains of Pseudomonas aeruginosa under conditions of phosphate limitation. The ability to produce the appendages in P. aeruginosa herein described is thus far attributable only to certain multidrug-resistant strains isolated from critically ill patients. The appearance of appendages is highly suppressed in phosphate rich media and enhanced during growth in phosphate depleted media. Under these dual conditions, the absence or presence of appendages correlates with an adhesive and virulent phenotype in P. aeruginosa strains. The proper technique for appendage visualization is critical to facilitate the necessary studies to further elucidate their structure and function.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Josenhans C, Suerbaum S (2002) The role of motility as a virulence factor in bacteria. Int J Med Microbiol 291:605–614
Jonson AB, Normark S, Rhen M (2005) Fimbriae, pili, flagella and bacterial virulence. Contrib Microbiol 12:67–89
Zaborina O, Holbrook C, Chen Y, Long J, Zaborin A et al (2008) Structure-function aspects of PstS in multi-drug-resistant Pseudomonas aeruginosa. PLoS Pathog 4:e43
Rouached H, Arpat AB, Poirier Y (2010) Regulation of phosphate starvation responses in plants: signaling players and cross-talks. Mol Plant 3:288–299
Yang XJ, Finnegan PM (2010) Regulation of phosphate starvation responses in higher plants. Ann Bot 105:513–526
Lin WY, Lin SI, Chiou TJ (2009) Molecular regulators of phosphate homeostasis in plants. J Exp Bot 60:1427–1438
Hanrahan G, Salmassi TM, Khachikian CS, Foster KL (2005) Reduced inorganic phosphorus in the natural environment: significance, speciation and determination. Talanta 66:435–444
Romanowski K, Zaborin A, Valuckaite V, Rolfes RJ, Babrowski T et al (2012) Candida albicans isolates from the gut of critically ill patients respond to phosphate limitation by expressing filaments and a lethal phenotype. PLoS One 7:e30119
Lamarche MG, Wanner BL, Crepin S, Harel J (2008) The phosphate regulon and bacterial virulence: a regulatory network connecting phosphate homeostasis and pathogenesis. FEMS Microbiol Rev 32:461–473
Rao NN, Gomez-Garcia MR, Kornberg A (2009) Inorganic polyphosphate: essential for growth and survival. Annu Rev Biochem 78:605–647
Alverdy J, Holbrook C, Rocha F, Seiden L, Wu RL et al (2000) Gut-derived sepsis occurs when the right pathogen with the right virulence genes meets the right host: evidence for in vivo virulence expression in Pseudomonas aeruginosa. Ann Surg 232:480–489
Alverdy JC, Chang EB (2008) The re-emerging role of the intestinal microflora in critical illness and inflammation: why the gut hypothesis of sepsis syndrome will not go away. J Leukoc Biol 83:461–466
Long J, Zaborina O, Holbrook C, Zaborin A, Alverdy J (2008) Depletion of intestinal phosphate after operative injury activates the virulence of P. aeruginosa causing lethal gut-derived sepsis. Surgery 144:189–197
Norris FA, Wilson MP, Wallis TS, Galyov EE, Majerus PW (1998) SopB, a protein required for virulence of Salmonella dublin, is an inositol phosphate phosphatase. Proc Natl Acad Sci U S A 95:14057–14059
Ruska E (1987) Nobel lecture. The development of the electron microscope and of electron microscopy. Biosci Rep 7:607–629
Ruska E (1980) The early development of electron lenses and electron microscopy. Microsc Acta (Suppl):1–140
Shah M et al (2014) Localization of DING proteins on PstS-containing outer-surface appendages of Pseudomonas aeruginosa. FEMS Microbiol Lett 352:54–61
Acknowledgments
This study was funded by the National Institute of Health (RO1-GM062344-12).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2014 Springer Science+Business Media New York
About this protocol
Cite this protocol
Zaborina, O., Alverdy, J., Shah, M., Chen, Y. (2014). Microscopic Analysis: Morphotypes and Cellular Appendages. In: Filloux, A., Ramos, JL. (eds) Pseudomonas Methods and Protocols. Methods in Molecular Biology, vol 1149. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-0473-0_11
Download citation
DOI: https://doi.org/10.1007/978-1-4939-0473-0_11
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-0472-3
Online ISBN: 978-1-4939-0473-0
eBook Packages: Springer Protocols