Specific detection of Pseudomonas spp. in milk by fluorescence in situ hybridization using ribosomal RNA directed probes

Thusitha S. Gunasekera*, M. R. Dorsch, M. B. Slade, D. A. Veal

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    66 Citations (Scopus)

    Abstract

    Aims: Pseudomonas spp. are considered the most important milk spoilage organisms. Here we describe development of a fluorescence in situ hybridization (FISH) probe specific for detection and enumeration of Pseudomonas spp. in milk. Methods and Results: 16S rRNA sequences were analysed to develop specific oligonucleotide probe for the genus Pseudomonas. Twenty different Pseudomonas spp. and 23 bacterial species from genera other than Pseudomonas (as negative controls) were tested. All tested Pseudomonas spp. yielded a positive FISH reaction, whereas negative controls showed no FISH reaction except for Burkholderia cepacia that showed a relatively weak FISH reaction. The FISH assay specifically stains Pseudomonas in milk when the milk contains a mixture of other bacterial species. The FISH assay takes 2 h and compares favourably with current culturing methods, which take a minimum of 48 h. Specificity of the probe was validated using polymerase chain reaction to selectively amplifying the Pseudomonas rDNA gene and sequencing the gene products. Conclusions: The method presented in this study allows simultaneously detection, identification and enumeration of Pseudomonas spp. in milk. Significance and Impact of the Study: Rapid and accurate enumeration of Pseudomonas facilitates the identification of specific contamination sources in dairy plants, the accurate validation of pasteurization treatments and the prediction of shelf life of processed milk.

    Original languageEnglish
    Pages (from-to)936-945
    Number of pages10
    JournalJournal of Applied Microbiology
    Volume94
    Issue number5
    DOIs
    Publication statusPublished - 2003

    Keywords

    • 16S rRNA
    • Fluorescence in situ hybridization
    • Milk spoilage
    • PCR
    • Pseudomonas spp.

    Fingerprint

    Dive into the research topics of 'Specific detection of Pseudomonas spp. in milk by fluorescence in situ hybridization using ribosomal RNA directed probes'. Together they form a unique fingerprint.

    Cite this