Impact of temperature and time storage on the microbial detection of oral samples by Checkerboard DNA-DNA hybridization method

Author(s)

do Nascimento, Cassio ^[1] ; dos Santos, Janine Navarro ^[1] ; Pedrazzi, Vinicius ^[1] ; Pita, Murillo Sucena ^[1] ; Monesi, Nadia ^[2] ; Ribeiro, Ricardo Faria ^[1] ; de Albuquerque Junior, Rubens Ferreira ^{[1, 3]}

Affiliation

^[1] Univ Sao Paulo, Mol Diag Lab, Dept Dent Mat & Prosthodont, Fac Dent Ribeirao Preto, BR-14040904 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Dept Clin Toxicol & Bromatol Anal, Fac Pharmaceut Sci Ribeirao Preto, BR-14040903 Ribeirao Preto, SP - Brazil

^[3] McGill Univ, Fac Dent, Strathcona Anat & Dent, Montreal, PQ H3A 2B2 - Canada

• Document type: Journal article
• Source: ARCHIVES OF ORAL BIOLOGY; v. 59, n. 1, p. 12-21, JAN 2014.
• Web of Science Citations: 2

Abstract

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 degrees C, -20 degrees C and -80 degrees 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. (C) 2013 Elsevier Ltd. All rights reserved. (AU)

• FAPESP's process: 10/17807-6 - Evaluation of the influence of temperature and storage time in the detection and quantification of oral cavity bacteria samples by the DNA Checkerboard technique.
• Grantee: Vinícius Pedrazzi
• Support Opportunities: Regular Research Grants
• FAPESP's process: 11/10008-3 - Effect of temperature and time sample storage on the bacterial detection of oral samples by DNA Checkerboard method
• Grantee: Janine Navarro Santos
• Support Opportunities: Scholarships in Brazil - Scientific Initiation