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Use of a polyphasic approach including MALDI-TOF MS for identification of Aspergillus section Flavi strains isolated from food commodities in Brazil

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Abstract

Brazil is one the largest producers and exporters of food commodities in the world. The evaluation of fungi capable of spoilage and the production mycotoxins in these commodities is an important issue that can be of help in bioeconomic development. The present work aimed to identify fungi of the genus Aspergillus section Flavi isolated from different food commodities in Brazil. Thirty-five fungal isolates belonging to the section Flavi were identified and characterised. Different classic phenotypic and genotypic methodologies were used, as well as a novel approach based on proteomic profiles produced by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). Type or reference strains for each taxonomic group were included in this study. Three isolates that presented discordant identification patterns were further analysed using the internal transcribed spacer (ITS) region and calmodulin gene sequences. The data obtained from the phenotypic and spectral analyses divide the isolates into three groups, corresponding to taxa closely related to Aspergillus flavus, Aspergillus parasiticus, and Aspergillus tamarii. Final polyphasic fungal identification was achieved by joining data from molecular analyses, classical morphology, and biochemical and proteomic profiles generated by MALDI-TOF MS.

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References

  • Batista PP, Santos JF, Oliveira NT, Pires APD, Motta CMS, Luna-Alves LEA (2008) Genetic characterization of Brazilian strains of Aspergillus flavus using DNA markers. Gen Mol Res 7:706–717

    Article  CAS  Google Scholar 

  • Bennett JW, Klich M (2003) Mycotoxins. Clin Microbiol Rev 16:497–516

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Chang PK, Bennett JW, Cotty PJ (2001) Association of aflatoxins biosynthesis and sclerotial development in Aspergillus parasiticus. Mycopathologia 153:41–48

    Article  Google Scholar 

  • Chang PK, Ehrlich KC, Hua SST (2006) Cladal relatedness among Aspergillus oryzae isolates and Aspergillus flavus S and L morphotype isolates. Int J Food Microbiol 108:172–177

    Article  CAS  PubMed  Google Scholar 

  • Dias N, Santos C, Portela M, Lima N (2011) Toenail onychomycosis in a Portuguese geriatric population. Mycopathologia 172:55–61

    Article  CAS  PubMed  Google Scholar 

  • Giorni P, Magan N, Pietri A, Bertuzzi T, Battilani P (2007) Studies on Aspergillus section Flavi isolated from maize in northern Italy. Int J Food Microbiol 113:330–338

  • Godet M, Munaut F (2010) Molecular strategy for identification in Aspergillus section Flavi. FEMS Microbiol Lett 304:157–168

    Article  CAS  PubMed  Google Scholar 

  • Goto T, Wicklow DT, Ito Y (1996) Aflatoxin and cyclopiazonic acid production by a sclerotium-producing Aspergillus tamarii strain. Appl Environ Microb 62:4036–4038

    CAS  Google Scholar 

  • Hedayati MT, Pasqualotto AC, Warn PA, Bower P, Denning DW (2007) Aspergillus flavus: Human pathogen, allergen and mycotoxin producer. Microbiology 153:1677–1692

    Article  CAS  PubMed  Google Scholar 

  • Ito Y, Peterson SW, Wicklow DT, Goto T (2001) Aspergillus pseudotamarii, a new aflatoxin-producing species in Aspergillus section Flavi. Mycol Res 105:233–239

    Article  Google Scholar 

  • Kimura M (1980) A simple model for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120

    Article  CAS  PubMed  Google Scholar 

  • Klich MA (2002) Identification of common Aspergillus species. CBS, Netherlands

    Google Scholar 

  • Klich MA (2007) Aspergillus flavus: the major producer of aflatoxin. Mol Plant Pathol 8:713–722

  • Klich MA, Pitt JL (1988) Differentiation of Aspergilus flavus and A. parasiticus and other closely related species. T Brit Mycol Soc 91:99–108

    Article  Google Scholar 

  • Kozakiewicz Z (1989) Aspergillus species on stored products. CAB International, Wallingford

    Google Scholar 

  • Kumeda Y, Asao T (2001) Heteroduplex panel analysis, a novel method for genetic identification of Aspergillus section Flavi strains. Appl Environ Microb 67:4084–4090

    Article  CAS  Google Scholar 

  • Kurtzman CP, Horn BW, Hesseltine CW (1987) Aspergillus nomius, a new aflatoxin-producing species related to Aspergillus flavus and Aspergillus tamari. A van Leeuw 53:147–158

    Article  CAS  Google Scholar 

  • Li T-Y, Liu B-H, Chen Y-C (2000) Characterization of Aspergillus spores by matrix assisted laser desorption⁄ionization time-of-flight mass spectrometry. Rapid Commun Mass Sp 14:2393–2400

    Article  CAS  Google Scholar 

  • Lima-Neto R, Santos C, Lima N, Sampaio P, Pais C, Neves RP (2014) Application of MALDI-TOF MS for requalification of a Candida clinical isolates culture collection. Braz J Microbiol 45:515–522

  • Midorikawa GEO, Sousa MLM, Silva OF, Dias JSA, Kanzaki LIB, Hanada RE, Mesquita RMLC, Goncalves RC, Alvares VS, Bittencourt DMC, Mille RNG (2014) Characterization of Aspergillus species on Brazil nut from the Brazilian Amazonian region and development of a PCR assay for identification at the genus level. BMC Microbiol 14:138

    Article  PubMed Central  PubMed  Google Scholar 

  • Nicolau A, Sequeira L, Santos C, Mota M (2014) Matrix-assisted laser desorption /ionisation time-of-flight mass spectrometry (MALDI-TOF MS) applied to diatom identification: influence of culturing age. Aquat Biol 20:139–144

    Article  Google Scholar 

  • Novas MV, Cabral D (2002) Association of mycotoxin and sclerotia production with compatibility groups in Aspergillus flavus from peanut in Argentina. Plant Dis 86:215–219

    Article  CAS  Google Scholar 

  • O’Donnell K, Kistler HC, Tacke BK, Casper HH (2000) Gene genealogies reveal global phylogeographic structure and reproductive isolation among lineages of Fusarium graminearum, the fungus causing wheat scab. Proc Natl Acad Sci U S A 97:7905–7910

    Article  PubMed Central  PubMed  Google Scholar 

  • Passarini MRZ, Santos C, Lima N, Berlinck RGS, Sette LD (2013) Filamentous fungi from the Atlantic marine sponge Dragmacidon reticulatum. Arch Microbiol 195:99–111

    Article  CAS  PubMed  Google Scholar 

  • Pereira L, Dias D, Santos C, Lima N (2014) The use of MALDI-TOF ICMS as an alternative tool for Trichophyton rubrum identification and typing. Enferm Infecc Microbiol Clin 32:11–17

    Article  PubMed  Google Scholar 

  • Pildain MB, Vaamonde G, Cabral D (2004) Analysis of population structure of Aspergillus flavus from peanut based on vegetative compatibility, geographic origin, mycotoxin and sclerotia production. Int J Food Microbiol 93:31–40

    Article  CAS  PubMed  Google Scholar 

  • Raper KB, Fennel DI (1965) The genus Aspergillus. Williams & Wilkins, Baltimore

    Google Scholar 

  • Razzaghi-Abyaneh M, Shams-Ghahfarokhi M, Allameh A, Kazeroon-Shiri A, Ranjbar-Bahadori S, Mirzahoseini H, Rezaee MB (2006) A survey on distribution of Aspergillus Section Flavi in corn field soils in Iran: population patterns based on aflatoxins, cyclopiazonic acid and sclerotia production. Mycopathologia 161:183–192

    Article  CAS  PubMed  Google Scholar 

  • Rodrigues P, Venancio A, Kozakiewicz Z, Lima N (2009) A polyphasic approach to the identification of aflatoxigenic and non-aflatoxigenic strains of Aspergillus section Flavi isolated from Portuguese almonds. Int J Food Microbiol 129:187–193

    Article  CAS  PubMed  Google Scholar 

  • Rodrigues P, Santos C, Venancio A, Lima N (2011) Species identification of Aspergillus section Flavi isolates from Portuguese almonds using phenotypic, including MALDI-TOF ICMS, and molecular approaches. J Appl Microbiol 111:877–892

    Article  CAS  PubMed  Google Scholar 

  • Rokas A, Payne G, Fedorova ND, Baker SE, Machida M, YU J, Georgianna DR, Dean RA, Bhatnagar D, Cleveland TE, Wortman JR, Maiti R, Joardar V, Amedeo P, Denning DW, Nierman WC (2007) What can comparative genomics tell us about species concepts in the genus Aspergillus? Stud Mycol 59:11–17

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Samson RA, Hoekstra ES, Frisvad JC, Filtenborg O (2000) Identification of the common food and airborne fung. In: Samson RA, Hoekstra ES, Frisvad JC (eds) Introduction to food and airborne fungi, 6th edn. Centraalbureau voor Schimmekultures, Utrecht, pp 64–97

    Google Scholar 

  • Samson RA, Hoekstra ES, Frisvad JC (2004) Introduction to food and airborne fungi, 7th edn. Wageningen, CBS

    Google Scholar 

  • Santos C, Paterson RRM, Venancio A, Lima N (2010) Filamentous fungal characterizations by matrix assisted laser desorption⁄ionization time-of-flight mass spectrometry. J Appl Microbiol 108:375–385

    Article  CAS  PubMed  Google Scholar 

  • Santos C, Lima N, Sampaio P, Pais C (2011) Matrix-assisted laser desorption/ionization time-of-flight intact cell mass spectrometry (MALDI-TOFICMS) to detect emerging pathogenic Candida species. Diag Microbiol Infec Dis 71:304–308

    Article  CAS  Google Scholar 

  • Simões MF, Pereira L, Santos C, Lima N (2013) Polyphasic identification and preservation of fungal diversity: concepts and applications. In: Malik A, Grohmann E, AlvesM (eds) Management of microbial resources in the environment, Chapter 5, ISBN 978-94-007-5930-5, 410. Springer, The Netherlands, pp 91–116

  • Tamura K, Dudley J, Nei M, Kumar S (2007) MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599

    Article  CAS  PubMed  Google Scholar 

  • Tanaka K, Waki H, Ido Y, Akita S, Yoshida Y (1988) Protein and polymer analyses up to m⁄z 100000 by laser ionization time-of-flight mass spectrometry. Rapid Commun Mass Sp 2:151–153

    Article  CAS  Google Scholar 

  • Varga J, Rigo K, Toth B, Teren J, Kozakiewicz Z (2003) Evolutionary relationships among Aspergillus species producing economically important mycotoxins. Food Technol Biotech 41:29–36

    CAS  Google Scholar 

  • White TJ, Bruns T, Lee S, Tailor S (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innins MA, Gelfand DH, Sninsky JJ, White TJ (eds) PCR protocols: a guide to methods and applications. Academic, San Diego, pp 315–322

    Google Scholar 

  • Yu J, Whitelaw CA, Nierman WC, Bhatnagar D, Cleveland TE (2004) Aspergillus flavus expressed sequence tags for identification of genes with putative roles in aflatoxin contamination of crops. FEMS Microbiol Lett 237:333–340

    PubMed  Google Scholar 

Download references

Acknowledgments

Acknowledgments are due to FAPEMIG - Fundacao de Amparo a Pesquisa do Estado de Minas Gerais (Brazil) for financial support. F. C. da Silva extends thanks to CAPES - Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (Brazil) for the PhD grant. C. Santos and N. Lima thank CAPES for the financial support as international visiting professors in the Post-Graduate Programme in Agricultural Microbiology, Federal University of Lavras, Lavras (MG), Brazil.

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Authors and Affiliations

  1. Post-Graduate Programme in Agricultural Microbiology, Federal University of Lavras, Lavras, MG, Brazil

    Fernanda Chaves da Silva, Cledir Santos & Nelson Lima

  2. Company of Agricultural Research of Minas Gerais (EPAMIG), Federal University of Lavras, Lavras, MG, Brazil

    Sara Maria Chalfoun

  3. Department of Food Sciences, Federal University of Lavras, Lavras, MG, Brazil

    Luis Roberto Batista

  4. Instituto Federal do Espirito Santo, Campus Barra de São Francisco, Barra de São Francisco, ES, Brazil

    Fernanda Chaves da Silva

  5. CEB – Centre of Biological Engineering, Micoteca da Universidade do Minho, Universidade do Minho, 4710-057, Braga, Portugal

    Fernanda Chaves da Silva, Cledir Santos & Nelson Lima

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  1. Fernanda Chaves da Silva
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  2. Sara Maria Chalfoun
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  3. Luis Roberto Batista
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  4. Cledir Santos
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  5. Nelson Lima
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Correspondence to Cledir Santos.

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da Silva, F.C., Chalfoun, S.M., Batista, L.R. et al. Use of a polyphasic approach including MALDI-TOF MS for identification of Aspergillus section Flavi strains isolated from food commodities in Brazil. Ann Microbiol 65, 2119–2129 (2015). https://doi.org/10.1007/s13213-015-1050-0

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  • DOI: https://doi.org/10.1007/s13213-015-1050-0

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