Growth and enumeration of the meat spoilage bacterium Brochothrix thermosphacta

School of Chemical Engineering, University of Birmingham, Edgbaston, UK.
International Journal of Food Microbiology (Impact Factor: 3.08). 11/1999; 51(2-3):145-58. DOI: 10.1016/S0168-1605(99)00129-4
Source: PubMed


Brochothrix thermosphacta is a common meat spoilage bacterium. The morphology of this bacterium changes from coccobacilli and short rods to chains during growth, which may give a false estimation in numbers using some enumeration techniques. Methods for the quantification of this bacterium have been compared. Turbidimetric readings showed good agreement with cell dry weight indicating that the former provides a good measure of the change in cell mass during growth. The turbidimetric method also correlated well with bacterial numbers determined by plate counts, flow cytometry and manual counts (by microscope) over a limited range of 10(7)-10(9) cells/ml. Flow cytometry and manual counts gave a linear relationship over a wider range of 10(5)-10(9) cells/ml. The sensitivity of analysis, growth rates and lag time attained using these methods were also compared. As a consequence of changes in bacterial cell size during growth, turbidimetry over-estimated the growth rate. The plate count method proved unable to detect the difference between bacteria existing as chains or single cells. The sensitivity of analysis and the calculated growth related parameters were similar for flow cytometry and manual counts. This suggests that flow cytometry is capable of counting individual cells in a chain. Further investigation showed that passage of B. thermosphacta cells through the flow cytometer resulted in the breakage of chains into single cells. The reliability, low error and rapidity of this technique make it attractive for bacterial enumeration, something which has been demonstrated using B. thermosphacta, a bacterium which exhibits complex morphologies.

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Available from: Mohamed Al-Rubeai, Jan 16, 2014
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    • "Otherwise, after 24 h in culture, total counts determined by FC with the discriminator set on FL1 were slightly higher than CFU values; however, these were not statistically different. Similar differences between FC and other traditional methods have been reported with the bacterium Brochothrix thermosphacta and attributed to the breakage of chains into single cells when passed through the flow cytometer (Rattanasomboon et al. 1999). When considered together, the results presented in this study demonstrate that MmmLC growth in broth medium can be determined by FC with a similar sensitivity and reproducibility as the traditional plate count method. "
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    ABSTRACT: The potential of using flow cytometry (FC) in combination with a fluorescent dye (SYBR green-I) for rapidly estimating Mycoplasma mycoides subSPS. mycoides large-colony type (MmmLC) in broth culture was examined. The FC analysis was performed by staining the MmmLC cells with a fluorescent dye, SYBR green-I (SYBR), and the results were compared with plate count method (colony forming units, - CFUs). There was a good correlation (linear regression, r(2) = 0.93) between mycoplasma counts determined by FC (cells ml(-1)) and by traditional plate count method (CFU ml(-1)). The lowest bacterial concentration detected by FC and traditional plate count was of the order of 10(4) cells ml(-1) and 10(3) CFU ml(-1), respectively. FC method allowed results in 20-30 min, whereas at least 24 h were necessary to obtain results with the traditional plate count method (CFU). Growth rates of MmmLC in broth medium determined by FC were highly reproducible and correlated well with mycoplasma counts assessed by the plate count method. These findings suggest that FC could be a good alternative to replace other time-consuming techniques that are currently used to enumerate mycoplasma in broth medium, such as plate count method (CFU).
    Journal of Applied Microbiology 05/2006; 100(4):878-84. DOI:10.1111/j.1365-2672.2005.02858.x · 2.48 Impact Factor
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    • "Flow cytometry (FC) is as a sensitive, specific and rapid technique that can be used to analyse at the same time a large number of cellular parameters (Orfao and González 1995). Albeit initially designed to study different aspects related to eucaryotic cell growth and physiology, FC has been successfully applied to the study of polymorphic cells of Brochothrix thermosphacta (Rattanasomboom et al. 1999), the analysis of bacterial populations of different sizes and physiological statuses in sea water samples (Troussellier et al. 1999; Gregory et al. 2001; Lebaron et al. 2001) and the growth dynamic and cellular viability of different micro-organisms (Henningson et al. 1998; Bunthof et al. 2001). Despite the different applications in microbiology (for a review see A ´ lvarez-Barrientos et al. 2000) the FC technique has not been used to evaluate the growth of any species of Mollicutes. "
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    Journal of Applied Microbiology 02/2005; 98(5):1047-54. DOI:10.1111/j.1365-2672.2005.02536.x · 2.48 Impact Factor
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    • "For enumeration of marine samples this is sometimes implemented along with membrane filtration (Delille and Delille, 2000; Leecaster and Weisberg, 2001; Lipp et al., 2002). Flow cytometry is a useful tool for counting and identifying microorganisms (Marie et al., 1997; Connil et al., 1998; Davey et al., 1999; Rattanasomboon et al., 1999; Shvalov et al., 2000), assessing their viability (L o opez-Amor o os et al., 1997), and cell sorting (Nebe- Von-Caron et al., 2000). Cells are usually stained or labelled with fluorescent tags that enable them to be electronically identified when passing through a beam of laser light. "
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