Zeynep CERİT, Mehmet Cengiz BALOĞLU, Remziye YILMAZ

BEYAZ PEYNİR MİKROBİYOTASINDA KÜLTÜROMİK VE SHOTGUN METAGENOMİK TEKNOLOJİLERİN DEĞERLENDİRİLMESİ

Omik teknolojiler; DNA, RNA, genler, proteinler ve metabolitlerin araştırılması için kullanılan araçlar ve metotlardan oluşan sistematik yöntemler bütünüdür. Son yıllarda mikroorganizmaların tanımlanmasında ve işlevlerinin belirlenmesinde genomik, transkriptomik, proteomik ve metabolomik alanlarda yapılan çalışmalar artış göstermektedir. Genomik ve transkritptomik çalışmalar kapsamında mikroorganizmaların genom dizilerinin belirlenmesinde ve gen ifade analizlerinde yeni nesil dizileme sistemleri ile biyoinformatik araçlar birlikte kullanılmaktadır. Bu çalışmada, beyaz peynirin taşıdığı toplam mikrobiyel yükün oluşturduğu beyaz peynir mikrobiyotasının belirlenmesinde, kültürden bağımsız bir yöntem olan shotgun metagenomik ile kültüre dayalı bir yöntem olan ve mikroorganizmaların tanımlanmasına olanak sağlayan kültüromik metotları üzerinde durulmuştur. Çalışma ile yakın gelecekte beyaz peynir gibi geleneksel gıda ürünlerinin yeni teknikler değerlendirilerek araştırılması gerekliliğinin önemi vurgulanmıştır. Kültüromik, metagenomik gibi yenilikçi teknikler, geleneksel gıda ürünlerinin mikrobiyota tanımlanması üzerinde daha az belirsizlik ile çalışılmasına olanak sağlayabilmektedir.

Anahtar Kelimeler:

beyaz peynir, mikrobiyota, omik teknolojiler, metagenomik, kültüromik, MALDI-TOF MS

EVALUATION OF CULTUROMICS AND SHOTGUN METAGENOMIC TECHNOLOGIES IN WHITE CHEESE MICROBIOTA

Omics technologies are a set of systematic methods consisting of tools and applications used to investigate genes, DNA, RNA, proteins and metabolites. In recent years, genomics, transcriptomics, proteomics and metabolomics have been mainly utilized in the identification and determination of microorganisms. Within the scope of genomic and transcriptomics studies, next generation sequencing systems and bioinformatics tools are used together to determine the genome sequences of microorganisms and gene expression analysis, respectively. In this study, shotgun metagenomics, the culture-independent method, and culture-dependent culturomics approach that allows identification of microorganisms are emphasized for the detection of white cheese microbiota. The study has been emphasized the importance of researching traditional food products such as white cheese by evaluating new techniques in the future. Innovative techniques such as culturomics and metagenomics can enable the study of traditional food products to be studied with less uncertainty on the microbiota identification.

Keywords:

white cheese, microbiota, omic technologies, metagenomics, culturomics, MALDI-TOF MS,

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Agyirifo, D.S., Wamalwa, M., Otwe, E.P., Galyuon, I., Runo, S., Takrama, J., Ngeranwa, J. (2019). Metagenomics analysis of cocoa bean fermentation microbiome identifying species diversity and putative functional capabilities, Heliyon 5(7): e02170, doi: 10.1016/j.heliyon.2019.e02170.
Anonymous, Türk Gıda Kodeksi Mikrobiyolojik Kriterler Tebliğinde Değişiklik Yapılması Hakkında Tebliğ (2009/68). Tarım ve Köyişleri Bakanlığı. 8 Ocak 2010 tarih ve 27456 sayılı Resmi Gazete, Ankara.
Anonymous, Türk Gıda Kodeksi Peynir Tebliği (2015/6). Tarım ve Köyişleri Bakanlığı, 8 Şubat 2015 tarih ve 29261 sayılı Resmi Gazete, Ankara.
Anonymous, Tarımsal Ekonomi ve Politika Geliştirme Enstitüsü Müdürlüğü (TEPGE) 2017-2018 Süt ve Süt Ürünleri Durum Tahmin Raporu, https://arastirma.tarimorman.gov.tr/tepge/Belgeler/PDF%20Durum-Tahmin%20Raporlar%C4%B1/2017-2018%20Durum-%20Tahmin%20Raporlar%C4%B1/S%C3%BCt%20Durum%20Tahmin%20Raporu%202017-2018-305.pdf (Erişim tarihi: 15 Mart 2020).
Anonymous, Ulusal Süt Konseyi 2018 Süt Raporu, https://ulusalsutkonseyi.org.tr/wp-content/uploads/Sut_Raporu_2018_Web_Kapakli.pdf (Erişim tarihi: 15 Mart 2020).
Arslan, S. (2017). Türkiye'nin farklı yörelerinden toplanan beyaz peynir örneklerinden laktik asit bakterilerinin izolasyonu, identifikasyonu ve moleküler karakterizasyonu Atatürk Üniversitesi Fen Bilimleri Enstitüsü Moleküler Biyoloji ve Genetik Anabilim Dalı Yüksek Lisans Tezi, Erzurum, Türkiye, 116 s.
Bora, S.S., Keot, J., Das, S. Sarma, K., Barooah, M. (2016). Metagenomics analysis of microbial communities associated with a traditional rice wine starter culture (Xaj-pitha) of Assam, India. 3 Biotech, 6:153, doi: 10.1007/s13205-016-0471-1.
Breitwieser, F. P., Lu, J., Salzberg, S. L. (2019). A review of methods and databases for metagenomic classification and assembly. Brief Bioinform, 20(4): 1125-1136, doi: 10.1093/bib/bbx120.
Cerutti, F., Cravero, D., Costantini, A., Pulcini, L., Modesto, P., Acutis, P.L., Vaudano, E., Peletto, S. (2019). Impact of DNA purification method and primerselection on 16S rRNA gene metabarcoding on wine. OENO One, 53. doi: 10.20870/oeno-one.2019.53.3.2368.
Cocolin, L., Ercolini, D. (2015). Zooming into food-associated microbial consortia: a ‘cultural’ evolution. Curr Opin Food Sci 2: 43-50, doi: 10.1016/j.cofs.2015.01.003.
Dalmasso, A., Río, M.D., Civera, T., Pattono, D., Cardazzo, B., Bottero, M.T. (2016). Characterization of microbiota in Plaisentif cheese by high-throughput sequencing. Lebensm Wiss Technol 69: 490-496, doi: 10.1016/j.lwt.2016.02.004.
Dec, M., Puchalski, A., Urban-Chmiel, R., Wernicki, A. (2016). 16S-ARDRA and MALDI-TOF mass spectrometry as tools for identification of Lactobacillus bacteria isolated from poultry. BMC Microbiol 16, doi: 10.1186/s12866-016-0732-5.
Deurenberg, R.H., Bathoorn, E., Chlebowicz, M.A., Couto, N., Ferdous, M., García- Cobos, S., Kooistra-Smid, A.M., Raangs, E.C., Rosema, S., Veloo, A.C., Zhou, K., Friedrich, A.W., Rossen, J.W. (2017). Application of next generation sequencing in clinical microbiology and infection prevention. J Biotechnol 243: 16-24, doi: 10.1016/j.jbiotec.2016.12.022.
Domingos-Lopesa, M.F.P., Stanton, C., Ross, P.R., Dapkevicius, M.L.E., Silva, C.C.G. (2017). Genetic diversity, safety and technological characterization of lactic acid bacteria isolated from artisanal Pico cheese. Food Microbiol 63:178-190, doi: 10.1016/j.fm.2016.11.014.
Doyle, C.J., Gleeson, D.E., O'Toole, P.W., Cotter, P.D. (2017). Impacts of Seasonal Housing and Teat Preparation on Raw Milk Microbiota: a High-Throughput Sequencing Study. Appl Environ Microbiol 83: 2, doi: 10.1128/AEM.02694-16.
Dugat-Bony, E., Garnier, L., Denonfoux, J., Ferreira, S., Sarthou, A., Bonnarme, P., Irlinger, F. (2016). Highlighting the microbial diversity of 12 French cheese varieties. Int J Food Microbiol 238: 265-273, doi: 10.1016/j.ijfoodmicro.2016.09.026.
Duru, I.C., Laine, P., Andreevskaya, M., Paulin, L., Kananen, S., Tynkkynen, S., Auvinen, P., Smolander, O.P. (2018). Metagenomic and metatranscriptomic analysis of the microbial community in Swiss-type Maasdam cheese during ripening. Int J Food Microbiol 281: 10-22 doi: 10.1016/j.ijfoodmicro.2018.05.017.
Ercolini, D., Mauriello, G., Blaiotta, G., Moschetti, G., Coppola, S. (2004). PCR–DGGE Fingerprints Of Microbial Succession During A Manufacture Of Traditional Water Buffalo Mozzarella Cheese. J Appl Microbiol 96(2): 263-70, doi: 10.1046/j.1365-2672.2003.02146.x.
Ercolini, D. (2013). High-Throughput Sequencing And Metagenomics: Moving Forward In The Culture-Independent Analysis Of Food Microbial Ecology. Appl Environ Microbiol 79 3148–3155, doi: 10.1128/AEM.00256-13.
Ertürkmen, P., Öner, Z. (2015) "Beyaz Peynir Örneklerinden İzole Edilen Laktik Asit Bakterilerinin Başlatıcı (Starter) Kültür Özelliklerinin Biyokimyasal Yöntemlerle Belirlenmesi". Süleyman Demirel Üniversitesi Fen Bilimleri Enstitüsü Dergisi 19: 9-16, doi: 10.19113/sdufbed.25545.
Escobar-Zepeda, A., Vera-Ponce De León, A., Sanchez-Flores, A. (2015). The Road to Metagenomics: From Microbiology to DNA Sequencing Technologies and Bioinformatics. Front Genet, 6, doi: 10.3389/fgene.2015.00348.
Escobar-Zepeda, A., Sanchez-Flores, A., Barucha, M. Q. (2016). Metagenomic analysis of a Mexican ripened cheese reveals a unique complex microbiota. Food Microbiol 57: 116-127, doi: 10.1016/j.fm.2016.02.004.
Escobar-Zepeda, A., Godoy-Lozano, E.E., Raggi, L. (2018). Analysis of sequencing strategies and tools for taxonomic annotation: Defining standards for progressive metagenomics. Sci Rep, 8, doi: 10.1038/s41598-018-30515-5.
Ferrocino, I., Bellio, A., Giordano, M., Macori, G., Romano, A., Rantsiou, K., Decastelli, L., Cocolin, L. (2018). Shotgun Metagenomics and Volatilome Profile of the Microbiota of Fermented Sausages. Appl Environ Microbiol 84(3): e02120-17. doi: 10.1128/AEM.02120-17.
Garofalo, C., Osimani, A., Milanović, V., Aquilanti, L., Filippis, F.D., Stellato, G., Mauro, S.D., Turchetti, B., Buzzini, P., Ercolini, D., Clementi, F. (2015). Bacteria and yeast microbiota in milk kefir grains from different Italian regions. Food Microbiol 49: 123-133, doi: 10.1016/j.fm.2015.01.017.
Guidone A., Zotta, T., Matera, A., Ricciardi, A., Filippis, F.D., Ercolini, D., Parente,E. (2016). The microbiota of high-moisture mozzarella cheese produced with different acidification methods. Int J Food Microbiol, 216: 9–17, doi: 10.1016/j.ijfoodmicro.2015.09.002.
Handelsman, J. (2004). Metagenomics: Application of Genomics to Uncultured Microorganisms. Microbiol Mol Biol Rev, 68(4): 669-685, doi: 10.1128/MMBR.68.4.669-685.2004.
Harnpicharnchai, P., Jaresitthikunchai, J., Seesang, M., Jindamorakot, S., Tanapongpipat, S., Ingsriswang, S. (2020). Characterization of Yeast and Bacterial Type Strains with Food and Agricultural Applications by MALDI-TOF Mass Spectrometry Biotypin. Hanguk Misaengmul Saengmyong Konghakhoe Chi, 48(2): 138-147. doi: 10.4014/mbl.1910.10008.
Hayaloğlu, A.A., Güven, M., P. F. Fox. (2002). Microbiological, Biochemical and Technological Properties Of Turkish White Cheese ‘Beyaz Peynir’. Int Dairy J, 12(8): 635-648. doi: 10.4172/2572-4134.1000117.
Jagadeesan, B., Gerner-Smidt, P., Allard, M.W., Leuillet, S., Winkler, A., Xiao, Y., Chaffron, S., Vossen, J.V., Tang, S., Katase, M., McClure, P., Kimura, B., Chai, L.C., Chapman, J., Grant, K. (2019). The use of next generation sequencing for improving food safety: Translation into practice. Food microbiol, 79: 96-115, doi: 10.1016/j.fm.2018.11.005.
Jonnala, B.R.Y., McSweeney, P.L.H., Sheehan, J.J., Cotter, P.D. (2018). Sequencing of the Cheese Microbiome and Its Relevance to Industry. Front Microbiol 23(9): 1020, doi: 10.3389/fmicb.2018.01020.
Jongman, M., Carmichael, P.C., Bill, M. (2020). Technological Advances in Phytopathogen Detection and Metagenome Profiling Techniques, Curr Microbiol 77: 675–681, doi: 10.1007/s00284-020-01881-z.
Kable, M.E., Srisengfa, Y., Laird, M., Zaragoza, J.L., Mcleod, J., Heidenreich, J.M., Marco, M.L. (2016). The Core and Seasonal Microbiota of Raw Bovine Milk in Tanker Trucks and the Impact of Transfer to a Milk Processing Facility. mBio, 7, doi: 10.1128/mBio.00836-16.
Kader, T., Goode, D. L., Wong, S. Q., Connaughton, J., Rowley, S. M., Devereux, L., Byrne, D., B. F., Stephen, Arnau, G. M., Tothill, R. W., Campbell I. G., Gorringe, K. L. (2016). Copy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue, Genome Med, 8(121), doi: 10.1186/s13073-016-0375-z.
KEGG GENOME Database, Organisms and ecosystems with genome sequence information. https://www.genome.jp/kegg/genome.html (Erişim tarihi: 20 Şubat 2020).
Knight R., Vrbanac, A., Taylor, B. C., Aksenov, A., Callewaert, C., Debelius, J., Gonzalez, A., Kosciolek, T., McCall, L. I., McDonald, D., Melnik, A. V., Morton, J. T., Navas, J., Quinn, R.A., Sanders, J. G., Swafford, A. D., Thompson, L. R., Tripathi, Xu, Z. Z., Zaneveld, J. R., Zhu, Q., Caporaso J. G., Dorrestein, P. C. (2018). Best practices for analysing microbiomes, Nat Rev Microbiol, 16: 410–422, doi: 10.1038/s41579-018-0029-9.
Lagier, J., Dubourg, G., Million, M., Cadoret, F., Bilen, M., Fenollar, F., Levasseur, A., Rolain, J.M., Fournier, P.E., Raoult, D. (2018). Culturing the human microbiota and culturomics. Nat Rev Microbiol 16, 540–550, doi:. 10.1038/s41579-018-0041-0.
Liu, X. (2016). Microbiome. Yale J Biol Med, 89(3): 275–276, doi: 10.3389/fmicb.2017.02224.
Nair, H.P., Bhat, S.G. (2020). Metagenomic data on bacterial diversity profiling of Arabian sea sediment by amplicon sequencing, Data Brief, 28, doi: 10.1016/j.dib.2019.104791.
Nacef, M., Chevalier, M., Chollet, S., Drider, D., Flahaut, C. (2017). MALDI-TOF mass spectrometry for the identification of lactic acid bacteria isolated from a French cheese: The Maroilles. Int J Food Microbiol, 247: 2-8, doi: 10.1016/j.ijfoodmicro.2016.07.005.
Nalbantoğlu, U., Cakar, A., Dogan, H., Abaci, N., Ustek, D., Sayood, K., Can, H., (2014). Metagenomic analysis of the microbial community in kefir grains. Food Microbiol, 41: 42-51, doi: 10.1016/j.fm.2014.01.014.
Parente, E., Ricciardi, A., Zotta, T. (2020). The microbiota of dairy milk: A review. Int Dairy J, 107: 104714, doi: 10.1016/j.idairyj.2020.104714.
Pothakos, V., Vuyst, L.D., Zhang, S.J., Bruyn, F.D., Verce, M., Torres, J., Callan, M., Moccand, C., Weckx, S. (2020). Temporal shotgun metagenomics of an Ecuadorian coffee fermentation process highlights the predominance of lactic acid bacteria. Curr Res Biotechnol, 2:1-15, doi: 10.1016/j.crbiot.2020.02.001.
Prescott, S.L. (2017). History of medicine: Origin of the term microbiome and why it matters. Hum Microb J, 4: 24-25, doi: 10.1016/j.humic.2017.05.004.
Ranjan, R., Rani, A., Metwally, A.A., McGee, H.S., Perkins, D.L. (2016). Analysis of the microbiome: Advantages of whole genome shotgun versus 16S ampliconsequencing. Biochem Biophys Res Commun, 469(4): 967-977, doi: 10.1016/j.bbrc.2015.12.083.
Santos, I.C., Hildenbrandbc, Z.L., Schug, K.A. (2016). Applications of MALDI-TOF MS in environmental microbiology. Analyst, 141: 2827-2837, doi: 10.1039/C6AN00131A.
Savaşan, S., Beyaz, D. (2019). Erken Olgunlaşma Dönemindeki Geleneksel Peynir Mikrobiyomunun Metagenomik Analizi. Vet Mikrobiyol, 30(1): 27-35, doi: 10.35864/EVMD.586548.
Serra, J.L., Moura, F.G., Pereira, G.V., Soccol, C.R., Rogez, H.L., Darnet, S. (2019). Determination of the microbial community in Amazonian cocoa bean fermentation by Illumina-based metagenomic sequencing. Lebensm Wiss Technol, 106: 229-239, doi: 10.1016/J.LWT.2019.02.038.
Sielaff, A.C., Urbaniak, C., Mohan, G.B., Stepanov, V.G., Tran, Q.H., Wood, J.M., Minich, J.J., McDonald, D., Mayer, T., Knight, R., Karouia, F., Fox, G.E., Venkateswaran, K. (2019). Characterization of the total and viable bacterial and fungal communities associated with the International Space Station surfaces. Microbiome, 7, doi: 10.1186/s40168-019-0666-x.
Stellato, G., Filippis, F.D., Storia, A.L., Ercolini, D. (2015). Coexistence of Lactic Acid Bacteria and Potential Spoilage Microbiota in a Dairy Processing Environment. Appl Environ Microbiol, 81(22): 7893-7904, doi: 10.1128/AEM.02294-15.
Sternes, P.R., Lee, D., Kutyna, D.R., Borneman, A.R. (2017). A combined meta- barcoding and shotgun metagenomic analysis of spontaneous wine fermentation. GigaScience, 6: 1 – 10, doi: 10.1093/gigascience/gix040.
Tilocca, B., Costanzo, N., Morittu, V.M., Spina, A.A., Soggiu, A., Britti, D., Roncada, P., Piras, C. (2019). Milk microbiota: Characterization methods and role in cheese production. J proteomics, 210: 103534, doi: 10.1016/j.jprot.2019.103534.
Togay, S.O., Capece, A., Siesto, G., Aksu, H., Altunatmaz, S.S., Aksu, F., Romano, P., Yuceer, Y.K. (2020). Molecular characterization of yeasts isolated from traditional Turkish cheeses. Food Sci Technol, doi: 10.1590/fst.24319.
Topçu, A., Saldamli, I. (2006). Proteolytical, Chemical, Textural and Sensorial Changes During the Ripening of Turkish White Cheese Made of Pasteurized Cows' Milk. Int J Food Prop, 9: 665-678, doi: 10.1080/10942910500542238.
TÜİK, Süt ve Süt Ürünleri Raporu Sayı: 33697, https://ulusalsutkonseyi.org.tr/wp-content/uploads/Haber_Bulteni_mart_2020.pdf (Erişim Tarihi: 10 Temmuz 2020).
Tunail, N. (2009). Mikrobiyoloji. Pelin Ofset Tipo Matbaacılık, Ankara, Türkiye, 434 s. ISBN: 978605-603-62-0-0.
Verce, M., Vuyst, L.D., Weckx, S. (2020). The metagenome-assembled genome of Candidatus Oenococcus aquikefiri from water kefir represents the species Oenococcus sicerae. Food microbiol 88: 103402, doi: 10.1016/j.fm.2019.103402.
Walsh, A. M., Crispie, F., Kilcawley, K., O'Sullivan, O., O'Sullivan, M. G., Claesson, M. J., Cotter, P. D. (2016). Microbial Succession and Flavor Production in the Fermented Dairy Beverage Kefir. mSystems 1(5): e00052-16, doi: 10.1128/mSystems.00052-16.
Wei, Y. J., Wu, Y., Yan, Y. Z., Zou, W., Xue, J., Ma, W. R., Wang, W., Tian, G., Wang, L. Y. (2018). High-throughput sequencing of microbial community diversity in soil, grapes, leaves, grape juice and wine of grapevine from China. PloS one 13(3):e0193097, doi:10.1371/journal.pone.0193097.
Weiss, H., Hertzberg, V.S., Dupont, C., Espinoza, J.L., Levy, S., Nelson, K., Norris, S., The FlyHealthy Research Team (2019). The Airplane Cabin Microbiome. Environ Microbiol 77(1):87-95, doi: 10.1007/s00248-018-1191-3.
Wolfe, B. E., Button, J. E., Santarelli, M., Dutton, R. J. (2014). Cheese rind communities provide tractable systems for in situ and in vitro studies of microbial diversity. Cell 158(2): 422-433, doi: 10.1016/j.cell.2014.05.041.
Yılmaz R., Temiz, A. Açık, L., Çelebi Keskin, A. (2015). Genetic Differentiation of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus Strains Isolated from Raw Milk Samples Collected from Different Regions of Turkey, Food Biotechnol, 29(4): 336-355, doi: 10.1080/08905436.2015.1092091.
Zhang, L., Loh, K., Lim, J.W., Zhang, J. (2019). Bioinformatics analysis of metagenomics data of biogas-producing microbial communities in anaerobic digesters: A review. Renew. Sustain. Energy Rev, 100: 110-126, doi: 10.1016/J.RSER.2018.10.021.
Zheng J., Wittouck S., Salvetti E., Franz C.M.A.P., Harris H.M.B., Mattarelli P., O'Toole, P.W., Pot, B., Vandamme, P., Walter, J., Watanabe, K., Wuyts, S., Felis, G.E.,Gänzle, M.G., Lebeer, S. (2020). A taxonomic note on the genus Lactobacillus: description of 23 novel genera, emended description of the genus Lactobacillus beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. Int J Syst Evol Microbiol, 70(4): 2782–2858, doi: 10.1099/ijsem.0.004107.
Zhou, L.M., Proctor, L.M., Creasy, H., Fettweis, J.M., Lloyd-Price, J., Mahurkar, A., Zhou, W., Buck, G.A., Snyder, M.P., Strauss, J.F., Weinstock, G.M., White, O.L., Huttenhower, C. (2019). The Integrative Human Microbiome Project. Nature, 569: 641-648, doi: 10.1038/s41586-019-1238-8.