Abstract
Fluorescence in situ hybridization (FISH) is a molecular method used to identify and quantify microorganisms in a wide range of samples. This technique combines the simplicity of microscopic observation and the specificity of DNA/rRNA hybridization, allowing detection of selected bacterial species and morphologic visualization. Here, we describe a quantitative molecular diagnosis of bacterial vaginosis, based on the classical Nugent score. Our probes are able to differentiate Lactobacillus spp. and Gardnerella vaginalis from the other undefined bacterial species considered in the Nugent score.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Bretelle F, Rozenberg P, Pascal A, Favre R, Bohec C, Loundou A, Senat MV, Aissi G, Lesavre N, Brunet J, Heckenroth H, Luton D, Raoult D, Fenollar F, Groupe de Recherche en Obstetrique Gynecologie (2015) High Atopobium vaginae and Gardnerella vaginalis vaginal loads are associated with preterm birth. Clin Infect Dis 60:860–867
Tibaldi C, Cappello N, Latino MA, Masuelli G, Marini S, Benedetto C (2009) Vaginal and endocervical microorganisms in symptomatic and asymptomatic non-pregnant females: risk factors and rates of occurrence. Clin Microbiol Infect 15:670–679
Verstraelen H, Swidsinski A (2013) The biofilm in bacterial vaginosis: implications for epidemiology, diagnosis and treatment. Curr Opin Infect Dis 26:86–89
Machado A, Cerca N (2015) Influence of biofilm formation by Gardnerella vaginalis and other anaerobes on bacterial vaginosis. J Infect Dis 212:1856–1861
Forsum U, Hallén A, Larsson P (2005) Bacterial vaginosis-a laboratory and clinical diagnostics enigma. Acta Pathol Microbiol Immunol Scand 113:153–161
Money D (2005) The laboratory diagnosis of bacterial vaginosis. Can J Infect Dis Med Microbiol 16:77–79
Nugent R, Krohn M, Hillier S (1991) Reliability of diagnosing bacterial vaginosis is improved by a standardized method of gram stain interpretation. J Clin Microbiol 29:297–301
Sha BE, Chen HY, Wang QJ, Zariffard MR, Cohen MH, Spear GT (2005) Utility of Amsel criteria, Nugent score, and quantitative PCR for Gardnerella vaginalis, Mycoplasma hominis, and Lactobacillus spp. for diagnosis of bacterial vaginosis in human immunodeficiency virus-infected women. J Clin Microbiol 43:4607–4612
Justé A, Thomma BP, Lievens B (2008) Recent advances in molecular techniques to study microbial communities in food-associated matrices and processes. Food Microbiol 25:745–761
Peleg AY, Tilahun Y, Fiandaca MJ, D’Agata EMC, Venkataraman L, Moellering RC, Eliopoulos GM (2009) Utility of peptide nucleic acid fluorescence in situ hybridization for rapid detection of Acinetobacter spp. and Pseudomonas aeruginosa. J Clin Microbiol 47:830–832
Stender H, Fiandaca M, Hyldig-Nielsen JJ, Coull J (2002) PNA for rapid microbiology. J Microbiol Methods 48:1–17
Amann R, Fuchs BM (2008) Single-cell identification in microbial communities by improved fluorescence in situ hybridization techniques. Nat Rev Microbiol 6:339–348
Almeida C, Azevedo NF, Iversen C, Fanning S, Keevil CW, Vieira MJ (2009) Development and application of a novel peptide nucleic acid probe for the specific detection of Cronobacter genomospecies (Enterobacter sakazakii) in powdered infant formula. Appl Environ Microbiol 75:2925–2930
Shepard JR, Addison RM, Alexander BD, Della-Latta P, Gherna M, Haase G, Hall G, Johnson JK, Merz WG, Peltroche-Llacsahuanga H, Stender H, Venezia RA, Wilson D, Procop GW, Wu F, Fiandaca MJ (2008) Multicenter evaluation of the Candida albicans/Candida glabrata peptide nucleic acid fluorescent in situ hybridization method for simultaneous dual-color identification of C. albicans and C. glabrata directly from blood culture bottles. J Clin Microbiol 46:50–55
Trnovsky J, Merz W, Della-Latta P, Wu F, Arendrup MC, Stender H (2008) Rapid and accurate identification of Candida albicans isolates by use of PNA FISH flow. J Clin Microbiol 46:1537–1540
Machado A, Almeida C, Salgueiro D, Henriques A, Vaneechoutte M, Haesebrouck F, Vieira MJ, Rodrigues L, Azevedo NF, Cerca N (2013) Fluorescence in situ hybridization method using peptide nucleic acid probes for rapid detection of Lactobacillus and Gardnerella spp. BMC Microbiol 13:82
Machado A, Castro J, Cereija T, Almeida C, Cerca N (2015) Diagnosis of bacterial vaginosis by a new multiplex peptide nucleic acid fluorescence in situ hybridization method. Peer J 3:e780
Acknowledgment
Research on BV biofilms in NC laboratory is supported by funding from the Fundação para a Ciência e a Tecnologia (FCT) strategic project of unit UID/BIO/04469/2013.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2017 Springer Science+Business Media LLC
About this protocol
Cite this protocol
Machado, A., Cerca, N. (2017). Multiplex Peptide Nucleic Acid Fluorescence In Situ Hybridization (PNA-FISH) for Diagnosis of Bacterial Vaginosis. In: Bishop-Lilly, K. (eds) Diagnostic Bacteriology. Methods in Molecular Biology, vol 1616. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7037-7_13
Download citation
DOI: https://doi.org/10.1007/978-1-4939-7037-7_13
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-7035-3
Online ISBN: 978-1-4939-7037-7
eBook Packages: Springer Protocols