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dc.contributor.authorMinogue, Elizabeth
dc.contributor.authorTuite, Nina
dc.contributor.authorSmith, Cindy J.
dc.contributor.authorReddington, Kate
dc.contributor.authorBarry, Thomas
dc.identifier.citationMinogue E., Tuite N.L., Smith C.J., Reddington K., Barry T. (2015) 'A rapid culture independent methodology to quantitatively detect and identify common human bacterial pathogens associated with contaminated high purity water'. BMC Biotechnol, 16 (6).en_IE
dc.description.abstractWater and High Purity Water (HPW) distribution systems can be contaminated with human pathogenic microorganisms. This biocontamination may pose a risk to human health as HPW is commonly used in the industrial, pharmaceutical and clinical sectors. Currently, routine microbiological testing of HPW is performed using slow and labour intensive traditional microbiological based techniques. There is a need to develop a rapid culture independent methodology to quantitatively detect and identify biocontamination associated with HPW. Results: A novel internally controlled 5-plex real-time PCR Nucleic Acid Diagnostics assay (NAD), was designed and optimised in accordance with Minimum Information for Publication of Quantitative Real-Time PCR Experiments guidelines, to rapidly detect, identify and quantify the human pathogenic bacteria Stenotrophomonas maltophilia, Burkholderia species, Pseudomonas aeruginosa and Serratia marcescens which are commonly associated with the biocontamination of water and water distribution systems. The specificity of the 5-plex assay was tested against genomic DNA isolated from a panel of 95 microorganisms with no cross reactivity observed. The analytical sensitivities of the S. maltophilia, B. cepacia, P. aeruginosa and the S. marcescens assays are 8.5, 5.7, 3.2 and 7.4 genome equivalents respectively. Subsequently, an analysis of HPW supplied by a Millipore Elix 35 water purification unit performed using standard microbiological methods revealed high levels of naturally occurring microbiological contamination. Five litre water samples from this HPW delivery system were also filtered and genomic DNA was purified directly from these filters. These DNA samples were then tested using the developed multiplex real-time PCR NAD assay and despite the high background microbiological contamination observed, both S. maltophilia and Burkholderia species were quantitatively detected and identified. At both sampling points the levels of both S. maltophilia and Burkholderia species present was above the threshold of 10 cfu/100 ml recommended by both EU and US guidelines. Conclusions: The novel culture independent methodology described in this study allows for rapid (<5 h), quantitative detection and identification of these four human pathogens from biocontaminated water and HPW distribution systems. We propose that the described NAD assay and associated methodology could be applied to routine testing of water and HPW distribution systems to assure microbiological safety and high water quality standards.en_IE
dc.description.sponsorshipCollege of Science and the Thomas Crawford Hayes Fund, National University of Ireland, Galway who have supported EM with a postgraduate scholarship. This publication, in part, has also emanated from research conducted with the financial support of Science Foundation Ireland under Grant SFI 13/TIDA/I2597. The authors also wish to acknowledge the European Union’s Seventh Programme for research.en_IE
dc.publisherBioMed Centralen_IE
dc.relation.ispartofBMC Biotechnolen
dc.rightsAttribution-NonCommercial-NoDerivs 3.0 Ireland
dc.subjectHuman microbial pathogensen_IE
dc.subjectBiocontaminated HPW distribution systemsen_IE
dc.subjectMultiplex real-time PCR NAD assaysen_IE
dc.subjectBiological water quality assuranceen_IE
dc.titleA rapid culture independent methodology to quantitatively detect and identify common human bacterial pathogens associated with contaminated high purity wateren_IE
dc.local.contactThomas Barry, Dept. Of Microbiology, Arts/Science Building, Nui Galway. 3189 Email:

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