Comparison between standard culture and peptide nucleic acid 16S rRNA hybridization quantification to study the influence of physico-chemical parameters on Legionella pneumophila survival in drinking water biofilms

• Published in: Biofouling (Chur, Switzerland) Switzerland), 2009, Vol.25 (4), p.335-343
• Main Authors: Gião, M. S., Wilks, S. A., Azevedo, N. F., Vieira, M. J., Keevil, C. W.
• Format: Article
• Language: English
• Subjects: Bacteriological Techniques; Biofilms - growth & development; carbon concentration; Colony Count, Microbial; Culture Media; drinking water biofilms; Fresh Water - chemistry; Fresh Water - microbiology; Humans; Legionella pneumophila; Legionella pneumophila - genetics; Legionella pneumophila - growth & development; Legionella pneumophila - isolation & purification; Nucleic Acid Hybridization - methods; Peptide Nucleic Acids - genetics; PNA hybridization; RNA, Ribosomal, 16S - genetics; shear stress; temperature; Water Microbiology; Water Supply
• ISSN: 0892-7014; 1029-2454
• DOI: 10.1080/08927010902802232

Legionella pneumophila is a waterborne pathogen that is mainly transmitted by the inhalation of contaminated aerosols. In this article, the influence of several physico-chemical parameters relating to the supply of potable water was studied using a L. pneumophila peptide nucleic acid (PNA) specific probe to quantify total L. pneumophila in addition to standard culture methods. A two-stage chemostat was used to form the heterotrophic biofilms, with biofilm generating vessels fed with naturally occurring L. pneumophila. The substratum was the commonly used potable water pipe material, uPVC. It proved impossible to recover cultivable L. pneumophila due to overgrowth by other microorganisms and/or the loss of cultivability of this pathogen. Nevertheless, results obtained for total L. pneumophila cells in biofilms using a specific PNA probe showed that for the two temperatures studied (15 and 20°C), there were no significant differences when shear stress was increased. However, when a source of carbon was added there was a significant increase in numbers at 20°C. A comparison of the two temperatures showed that at 15°C, the total cell numbers for L. pneumophila were generally higher compared with the total microbial flora, suggesting that lower temperatures support the inclusion of L. pneumophila in drinking water biofilms. The work reported in this article suggests that standard culture methods are not accurate for the evaluation of water quality in terms of L. pneumophila. This raises public health concerns since culture methods are still considered to be the gold standard for assessing the presence of this opportunistic pathogen in water.