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AFLP protocol comparison for microbial diversity fingerprinting

  • Microbial Genetics • Original Paper
  • Published:
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Abstract

Over the last decade, several methods based on genomic DNA have been developed for the identification and genotyping of prokaryotic and eukaryotic organisms. These genomic methods differ regarding taxonomic range, discriminatory power, reproducibility, and ease of interpretation and standardization. The amplified fragment length polymorphism (AFLP) technique is a very powerful DNA fingerprinting technique for DNA of any source or complexity, varying in both size and base composition. In addition, this method shows high discriminatory power and good reproducibility allowing it to be efficient in discriminating at both the species and strain levels. The development and application of AFLP have allowed significant progress in the study of biodiversity and taxonomy of microorganisms. In the last years, the Applied Biosystems AFLP Microbial Fingerprinting Kit, now out of production, was widely used in various studies to perform AFLP characterization of selected bacteria strains (described by Vos et al. (Nucleic Acids Res 23(21):4407–4414, 1995)). Its replacement gives the possibility for laboratories to continue the use of the previous AFLP data as a reference for bacteria genetic fingerprinting analysis in biodiversity studies. To overcome this issue a result comparison, by using an improved AFLP protocol and the AFLP commercial kit, was performed. In particular, previous results on different species (Listeria monocytogenes, Lactobacillus plantarum, and Streptococcus thermophilus) obtained with the commercial kit were compared with the improved AFLP procedure to validate the protocol. When compared with the AFLP Microbial Fingerprinting Kit, the improved protocol shows high reproducibility, resolution, and overall, is a faster method with lower costs.

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References

  • Bernini V, Bottari B, Dalzini E, Sgarbi E, Lazzi C, Neviani E, Gatti M (2013) The presence, genetic diversity and behaviour of Listeria monocytogenes in blue-veined cheese rinds during the shelf life. Food Control 34(2):323–330

    Article  CAS  Google Scholar 

  • Blears M, De Grandis S, Lee H, Trevors JT (1998) Amplified fragment length polymorphism (AFLP): a review of the procedure and its applications. J Ind Microbiol Biotechnol 21:99–114

    Article  CAS  Google Scholar 

  • Cappello MS, Stefani D, Grieco F, Logrieco A, Zapparoli G (2008) Genotyping by amplified fragment length polymorphism and malate metabolism performances of indigenous Oenococcus oeni strains isolated from Primitivo wine. Int J Food Microbiol 127(3):241–245

    Article  CAS  PubMed  Google Scholar 

  • Cocconcelli PS, Porro D, Galandini S, Senini L (1995) Development of RAPD protocol for typing of strains of lactic acid bacteria and enterococci. Lett Appl Microbiol 21(6):376–379

    Article  CAS  PubMed  Google Scholar 

  • Cocolin L, Ercolini D (2008) Molecular techniques in the microbial ecology of fermented foods. Food microbiology and food safety. Springer, New York, pp 1–30

    Book  Google Scholar 

  • Curtin CD, Bellon JR, Henschke PA, Godden PW, de Barros Lopes MA (2007) Genetic diversity of Dekkera bruxellensis yeasts isolated from Australian wineries. FEMS Yeast Res 7:471–481

    Article  CAS  PubMed  Google Scholar 

  • De Filippis F, Parente E, Ercolini D (2017) Metagenomics insights into food fermentations. Microb Biotechnol 10(1):91–102

    Article  PubMed  Google Scholar 

  • Di Cagno R, Minervini G, Sgarbi E, Lazzi C, Bernini V, Neviani E, Gobbetti M (2010) Comparison of phenotypic (biolog system) and genotypic (random amplified polymorphic DNA-polymerase chain reaction, RAPD-PCR, and amplified fragment length polymorphism, AFLP) methods for typing Lactobacillus plantarum isolates from raw vegetables and fruits. Int J Food Microbiol 143(3):246–253

    Article  CAS  PubMed  Google Scholar 

  • Garofalo C, Osimani A, Milanović V, Taccari M, Cardinali F, Aquilanti L, Riolo P, Ruschioni S, Isidoro N, Clementi F (2017) The microbiota of marketed processed edible insects as revealed by high-throughput sequencing. Food Microbiol 62:15–22

    Article  CAS  PubMed  Google Scholar 

  • Giraffa G, Paris A, Valcavi L, Gatti M, Neviani E (2001) Genotypic and phenotypic heterogeneity of Streptococcus thermophilus strains isolated from dairy products. J Appl Microbiol 91(5):937–943

    Article  CAS  PubMed  Google Scholar 

  • Gulitz A, Stadie J, Ehrmann MA, Ludwig W, Vogel RF (2013) Comparative phylobiomic analysis of the bacterial community of water kefir by 16S rRNA gene amplicon sequencing and ARDRA analysis. J Appl Microbiol 114(4):1082–1091

    Article  CAS  PubMed  Google Scholar 

  • Hamza AA, Robene-Soustrade I, Jouen E, Lefeuvre P, Chiroleu F, Fisher-Le Saux M, Gagnevin L, Pruvost O (2012) MultiLocus sequence analysis and amplified fragment length polymorphism-based characterization of xanthomonads associated with bacterial spot of tomato and pepper and their relatedness to Xanthomonas species. Syst Appl Microbiol 35(3):183–190

    Article  CAS  PubMed  Google Scholar 

  • Heir E, Lindstedt B-A, Vardund T, Wasteson Y, Kapperud G (2000) Genomic fingerprinting of shigatoxin-producing Escherichia coli (STEC) strains: comparison of pulsed-field gel electrophoresis (PFGE) and fluorescent amplified-fragment-length polymorphism (FAFLP). Epidemiol Infect 125:537–548

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Janssen P, Coopman R, Huys G, Swings J, Bleeker M, Vos P, Zabeau M, Kersters K (1996) Evaluation of the DNA fingerprinting method AFLP as a new tool in bacterial taxonomy. Microbiology 142:1881–1893

    Article  CAS  PubMed  Google Scholar 

  • Jarraud S, Mougel C, Thioulouse J, Lina G, Meugnier H, Forey F, Vandenesch F (2002) Relationships between Staphylococcus aureus genetic background, virulence factors, agr groups (alleles), and human disease. Infect Immun 70(2):631–641

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jérôme M, Macé S, Dousset X, Pot B, Joffraud JJ (2016) Genetic diversity analysis of isolates belonging to the Photobacterium phosphoreum species group collected from salmon products using AFLP fingerprinting. Int J Food Microbiol 217:101–109

    Article  CAS  PubMed  Google Scholar 

  • Kovacs A, Yacoby K, Gophna U (2010) A systematic assessment of automated ribosomal intergenic spacer analysis (ARISA) as a tool for estimating bacterial richness. Res Microbiol 161(3):192–197

    Article  CAS  PubMed  Google Scholar 

  • Lazzi C, Rossetti L, Zago M, Neviani E, Giraffa G (2004) Evaluation of bacterial communities belonging to natural whey starters for Grana Padano cheese by length heterogeneity-PCR. J Appl Microbiol 96(3):481–490

    Article  CAS  PubMed  Google Scholar 

  • Lazzi C, Bove CG, Sgarbi E, Gatti M, La Gioia F, Torriani S, Neviani E (2009) Application of AFLP fingerprint analysis for studying the biodiversity of Streptococcus thermophilus. J Microbiol Methods 79(1):48–54

    Article  CAS  PubMed  Google Scholar 

  • Lévesque S, St-Pierre K, Frost E, Arbeit RD, Michaud S (2012) Use of amplified-fragment length polymorphism to study the ecology of Campylobacter jejuni in environmental water and to predict multilocus sequence typing clonal complexes. Appl Environ Microbiol 78(7):2470–2473

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nabhan S, Wydra K, Linde M, Debener T (2012) The use of two complementary DNA assays, AFLP and MLSA, for epidemic and phylogenetic studies of pectolytic enterobacterial strains with focus on the heterogeneous species Pectobacterium carotovorum. Plant Pathol 61:498–508

    Article  Google Scholar 

  • Perin LM, Savo Sardaro ML, Nero LA, Neviani E, Gatti M (2017) Bacterial ecology of artisanal Minas cheeses assessed by culture-dependent and -independent methods. Food Microbiol 65:160–169

    Article  CAS  PubMed  Google Scholar 

  • Savelkoul PH, Aarts HJ, De Haas J, Dijkshoorn L, Duim B, Otsen M, Rademaker JL, Schouls L, Lenstra JA (1999) Amplified-fragment length polymorphism analysis: the state of an art. J Clin Microbiol 37(10):3083–3091

    CAS  PubMed  PubMed Central  Google Scholar 

  • Savo Sardaro ML, Perin LM, Bancalari E, Neviani E, Gatti M (2018) Advancement in LH-PCR methodology for multiple microbial species detections in fermented foods. Food Microbiol 74:113–119

    Article  CAS  PubMed  Google Scholar 

  • Vaneechoutte M, Rossau R, De Vos P, Gillis M, Janssen D, Paepe N, De Rouck A, Fiers T, Claeys G, Kersters K (1992) Rapid identification of bacteria of the Comamonadaceae with amplified ribosomal DNA-restriction analysis (ARDRA). FEMS Microbiol Lett 93:227–234

    Article  CAS  Google Scholar 

  • Vos P, Hogers R, Bleeker M, Reijans M, Van de Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M (1995) AFLP: a new technique for DNA fingerprinting. Nucleic Acids Res 23(21):4407–4414

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zabeau M, Vos P (1993) Selective restriction fragment amplification: a general method for DNA fingerprinting. European Patent Office. Publication 0 534 858 Al

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Acknowledgements

Authors acknowledge the Department of Soil, Plant, and Food Science, University of Bari, Italy, for providing L. plantarum strains (POM1, POM31, POM43, POM40, POM8, C6, POM38). The authors are grateful to “Consorzio tutela Pecorino Toscano DOP” and “Consorzio tutela del formaggio Gorgonzola” for providing the samples.

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Authors and Affiliations

  1. Food and Drug Department, University of Parma, Parco Area delle Scienze 49/A, Parma, Italy

    Gaia Bertani, Maria Luisa Savo Sardaro, Erasmo Neviani & Camilla Lazzi

  2. Department of Human Science and Promotion of the Quality of Life, University San Raffaele Srl, Via Val Cannuta 247, Rome, Italy

    Maria Luisa Savo Sardaro

Authors
  1. Gaia Bertani
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  2. Maria Luisa Savo Sardaro
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  3. Erasmo Neviani
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  4. Camilla Lazzi
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Contributions

Bertani contributed to the analysis, acquisition, and interpretation of data; drafting of the manuscript; and critical revision. Savo Sardaro contributed to the study conception and designs; the work, analysis, and interpretation of data; drafting of the manuscript; and critical revision. Lazzi and Neviani contributed to design the work and in the critical revision.

Corresponding author

Correspondence to Maria Luisa Savo Sardaro.

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The authors declare that they have no conflict of interest.

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This article does not contain any studies with animals performed by any of the authors.

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Communicated by: Agnieszka Szalewska-Palasz

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Bertani, G., Savo Sardaro, M.L., Neviani, E. et al. AFLP protocol comparison for microbial diversity fingerprinting. J Appl Genetics 60, 217–223 (2019). https://doi.org/10.1007/s13353-019-00492-0

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  • DOI: https://doi.org/10.1007/s13353-019-00492-0

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