A simple and sensitive method to extract bacterial, yeast and fungal DNA from blood culture material

This study investigated the various commercially available kits and ‘in-house’ methods to extract DNA from Gram-negative and Gram-positive bacteria, yeast and fungal agents in commonly employed blood culture material. The main methods investigated were as follows; Qiagen QIAmp Blood kit, Roche high...

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Bibliographic Details
Published in:Journal of microbiological methods 2000-10, Vol.42 (2), p.139-147
Main Authors: Millar, Beverley C., Jiru, Xu, Moore, John E., Earle, John A.P.
Format: Article
Language:English
Subjects:
Bacillus coagulans
Bacteria
Bacteriological methods and techniques used in bacteriology
Bacteriology
Biological and medical sciences
Blood - microbiology
Blood-culture
Contaminant
Culture Media
DNA extraction
DNA, Bacterial - isolation & purification
DNA, Fungal - isolation & purification
DNA, Ribosomal - analysis
Fundamental and applied biological sciences. Psychology
Fungi
Lactococcus lactis
Microbiological Techniques - economics
Microbiology
Mycological methods and techniques used in mycology
Mycology
PCR inhibitor
Polymerase Chain Reaction
Reagent Kits, Diagnostic
RNA, Ribosomal, 16S - genetics
Sensitivity and Specificity
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Summary:This study investigated the various commercially available kits and ‘in-house’ methods to extract DNA from Gram-negative and Gram-positive bacteria, yeast and fungal agents in commonly employed blood culture material. The main methods investigated were as follows; Qiagen QIAmp Blood kit, Roche high PCR template preparation kit, Puregene DNA extraction kit, boiling, glass beads/sonication and wash/alkali/heat lysis. The results indicated that a simple wash/alkali/heat lysis method was the most sensitive, reproducible, simple and cost-effective extraction method. This was the only method which removed any PCR inhibitors and inherent DNA which existed in virgin BacT/Alert aerobic, anaerobic and paediatric blood culture material. Contaminating microbial DNA from Lactococcus lactis or Bacillus coagulans was identified in all batches of BacT/Alert ® FAN ® aerobic blood culture material examined.
ISSN:0167-7012
1872-8359
DOI:10.1016/S0167-7012(00)00174-3