Viable but non-culturable

Cools et al. (2005) used SPC with ChemChrome V6 labelhng to demonstrate the existence of a viable but non-culturable (VBNC) form of Campylobacter jejuni in water. For freshly cultured viable cells, an excellent correspondence was noticed between culture and SPC. Therefore, if discrepancies between the two results occur in older cultures, they can be attributed to the transition of culturable Campylobacter jejuni cells into the VBNC form. 

Catala P, Parthuisot N, Bernard L, Baudart J, Lemarchand K, Lebaron P (1999) Effectiveness of CSE to counterstain particles and dead bacterial cells with permeabilised membranes application to viability assessment in waters. FEMS Microb Lett 178 219-226 Cools 1, D Haese E, Uyttendaele M, Storms E, NeUs HJ, Debevere J (2005) Solid phase cytometry as a tool to detect viable but non-culturable cells of Campylobacter jejuni. J Microbiol Meth 63 107-114... 

Traditionally methods of viable counting and microscopy have been used to study phagocytosis of bacteria and subsequent survival or destruction." However, such indirect methods give an underestimate of bacterial survival within cells. It has been shown that bioluminescence acts as a convenient real-time method for monitoring survival of Bordetella bronchiseptica in vitro, whilst a dual gfp-luxABCDE operon has been used to monitor real-time replication of Staphylococcus aureus. The use of clinically important bacteria transformed with the lux cassette overcomes the problems with viable but non-culturable bacteria, as the expression of lux genes depends on the functional biochemistry of the bacteria. ""... 

McKay. A. M. <1992). Viable but non-culturable forms of poientially pathogenic bacteria in water. Letters inAf lied Microbiology 14 129-135. 

The specific dyes used in fluorescence microscopy are directly taken up by the cells and may react indiscriminately with organic material or be incorporated and concentrated in specific subcellular organelles. Still other techniques utilize immunochemical methodology (Section 16.3.2) whereby a fluorescent dye react with specific moieties of the viable cell membrane. In these cases, the dye can be used to distinguish between viable and dead cells. These methods have been applied toward detection of viable-but-non-culturable cells because these cells cannot be cultured using standard microbiological media (Section 6.1). 

Splittstoesser (1992) described a method using fluorescent dye (acridine orange) known as direct epifluorescence filter technique (DEFT), a method also applied by Divol and Lonvaud-Funel (2005). Divol and Lonvaud-Funel (2005) used a different substrate, fluoresceine diacetate, which is hydrolyzed by viable cells to form a fluorescent product, fluoresceine. However, Atlas and Bartha (1981) observed that cell population values can differ substantially between (epifluorescence and direct plating) methods (10 vs. 10 CFU), possibly due to the presence of viable-but-non-culturable cells. In addition, Meidell (1987) reported interference... 

Millet, V. and A. Lonvaud-Funel. 2000. The viable but non-culturable state of wine micro-organisms during storage. Lett. Appl. Microbiol. 30 136-141. 

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