Document Detail

Rapid differentiation and enumeration of the total, viable vegetative cell and spore content of thermophilic bacilli in milk powders with reference to Anoxybacillus flavithermus.
MedLine Citation:
PMID:  16238756     Owner:  NLM     Status:  MEDLINE    
AIMS: The development of a rapid method for the selective detection and enumeration of the total and viable vegetative cell and spore content of thermophilic bacilli in milk powder by PCR. METHODS AND RESULTS: Quantitative PCR and microscopy indicate the presence of up to 2.9 log units more cells in milk powder than accounted for by plate counting due to the majority of cells being killed during milk processing. Two approaches for viable and dead cell differentiation of thermophilic bacilli by quantitative PCR were evaluated, these being the nucleic binding dye ethidium monoazide (EMA) and DNase I digestion. The former agent exposed to a viable culture of Anoxybacillus flavithermus caused considerable cell inactivation. In contrast, DNase I treatment had no effect on cell viability and was utilized to develop DNA extraction methods for the differential enumeration of total, viable vegetative cells and spores in milk powder. Moreover, the methods were further applied and evaluated to 41 factory powder samples taken throughout eight process runs to assess changes in numbers of vegetative cells and spores with time. DNase I treatment reduced vegetative cell numbers enumerated with PCR by up to 2.6 log units. The quantification of spores in the factory milk powders investigated indicates on average the presence of 1.2 log units more spores than determined by plate counting. CONCLUSIONS: The method presented in this study provides the ability to selectively enumerate the total and viable cell and spore content of reconstituted milk. SIGNIFICANCE AND IMPACT OF THE STUDY: The current study provides a tool to monitor the extent of thermophilic contamination during milk powder manufacturing 60-90 min after sampling.
A Rueckert; R S Ronimus; H W Morgan
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of applied microbiology     Volume:  99     ISSN:  1364-5072     ISO Abbreviation:  J. Appl. Microbiol.     Publication Date:  2005  
Date Detail:
Created Date:  2005-10-21     Completed Date:  2006-02-16     Revised Date:  2009-11-19    
Medline Journal Info:
Nlm Unique ID:  9706280     Medline TA:  J Appl Microbiol     Country:  England    
Other Details:
Languages:  eng     Pagination:  1246-55     Citation Subset:  IM    
Thermophile Research Unit, Department of Biological Sciences, University of Waikato, Hamilton, New Zealand.
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MeSH Terms
Azides / pharmacology
Bacillaceae / drug effects,  growth & development*,  isolation & purification
Colony Count, Microbial
Cultured Milk Products / microbiology*
DNA, Bacterial / analysis,  drug effects
Deoxyribonuclease I / pharmacology
Food Contamination / analysis
Food Microbiology*
Geobacillus stearothermophilus / isolation & purification
Intercalating Agents / pharmacology
Microscopy, Fluorescence / methods
Microscopy, Phase-Contrast / methods
Polymerase Chain Reaction / methods
Spores, Bacterial / drug effects,  growth & development,  isolation & purification
Reg. No./Substance:
0/Azides; 0/DNA, Bacterial; 0/Intercalating Agents; 69498-50-6/8-azidoethidium; EC I

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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