- 作者：C谩ssio do Nascimento ; Janine Navarro dos Santos ; Vin铆cius Pedrazzi ; Murillo Sucena Pita ; Nadia Monesi ; Ricardo Faria Ribeiro ; Rubens Ferreira de Albuquerque Junior
- 关键词：DNA storage ; Bacteria ; Candida ; Diagnosis ; Clinical assessment ; Checkerboard DNA&ndash ; DNA hybridization
- 刊名：Archives of Oral Biology
- 出版年：January, 2014
- 年：2014
- 卷：59
- 期：1
- 页码：12-21
- 全文大小：724 K
Purpose
Molecular diagnosis methods have been largely used in epidemiological or clinical studies to detect and quantify microbial species that may colonize the oral cavity in healthy or disease. The preservation of genetic material from samples remains the major challenge to ensure the feasibility of these methodologies. Long-term storage may compromise the final result. The aim of this study was to evaluate the effect of temperature and time storage on the microbial detection of oral samples by Checkerboard DNA-DNA hybridization.Methods
Saliva and supragingival biofilm were taken from 10 healthy subjects, aliquoted (n = 364) and processed according to proposed protocols: immediate processing and processed after 2 or 4 weeks, and 6 or 12 months of storage at 4 掳C, 鈭?0 掳C and 鈭?0 掳C.
Results
Either total or individual microbial counts were recorded in lower values for samples processed after 12 months of storage, irrespective of temperatures tested. Samples stored up to 6 months at cold temperatures showed similar counts to those immediately processed. The microbial incidence was also significantly reduced in samples stored during 12 months in all temperatures.
Conclusions
Temperature and time of oral samples storage have relevant impact in the detection and quantification of bacterial and fungal species by Checkerboard DNA-DNA hybridization method. Samples should be processed immediately after collection or up to 6 months if conserved at cold temperatures to avoid false-negative results.