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Comprehensive analysis of the CRISPR-Cas systems in Streptococcus thermophilus strains isolated from traditional yogurts

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Abstract

Phage resistance is crucial for lactic acid bacteria in the dairy industry. However, identifying all phages affecting these bacteria is challenging. CRISPR-Cas systems offer a resistance mechanism developed by bacteria and archaea against phages and plasmids. In this study, 11 S. thermophilus strains from traditional yogurts underwent analysis using next-generation sequencing (NGS) and bioinformatics tools. Initial characterization involved molecular ribotyping. Bioinformatics analysis of the NGS raw data revealed that all 11 strains possessed at least one CRISPR type. A total of 21 CRISPR loci were identified, belonging to CRISPR types II-A, II-C, and III-A, including 13 Type II-A, 1 Type III-C, and 7 Type III-A CRISPR types. By analyzing spacer sequences in S. thermophilus bacterial genomes and matching them with phage/plasmid genomes, notable strains emerged. SY9 showed prominence with 132 phage matches and 30 plasmid matches, followed by SY12 with 35 phage matches and 25 plasmid matches, and SY18 with 49 phage matches and 13 plasmid matches. These findings indicate the potential of S. thermophilus strains in phage/plasmid resistance for selecting starter cultures, ultimately improving the quality and quantity of dairy products. Nevertheless, further research is required to validate these results and explore the practical applications of this approach.

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References

  • Achigar R, Magadán AH, Tremblay DM, Julia Pianzzola M, Moineau S (2017) Phage-host interactions in Streptococcus thermophilus: Genome analysis of phages isolated in Uruguay and ectopic spacer acquisition in CRISPR array. Sci Rep 7(1):43438

    Article  PubMed  PubMed Central  Google Scholar 

  • Ağagündüz D, Şahin TÖ, Ayten Ş, Yılmaz B, Güneşliol BE, Russo P, Spano G, Özogul F (2022) Lactic acid bacteria as pro-technological, bioprotective and health-promoting cultures in the dairy food industry. Food Biosci 47:101617

    Article  Google Scholar 

  • Barrangou R, Marraffini LA (2014) CRISPR-Cas systems: prokaryotes upgrade to adaptive immunity. Mol Cell 54(2):234–244

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P, Moineau S, Romero DA, Horvath P (2007) CRISPR provides acquired resistance against viruses in prokaryotes. Science 315(5819):1709–1712

    Article  CAS  PubMed  Google Scholar 

  • Chi H, Hoikkala V, Grüschow S, Graham S, Shirran S, White MF (2023) Antiviral type III CRISPR signalling via conjugation of ATP and SAM. Nature 622(7984):826–833

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chylinski K, Le Rhun A, Charpentier E (2013) The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems. RNA Biol 10(5):726–737

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Common J, Morley D, Westra ER, van Houte S (2019) CRISPR-Cas immunity leads to a coevolutionary arms race between Streptococcus thermophilus and lytic phage. Philos Trans R Soc B 374(1772):20180098

    Article  CAS  Google Scholar 

  • Datsenko KA, Pougach K, Tikhonov A, Wanner BL, Severinov K, Semenova E (2012) Molecular memory of prior infections activates the CRISPR/Cas adaptive bacterial immunity system. Nat Commun 3(1):945

    Article  PubMed  Google Scholar 

  • Deltcheva E, Chylinski K, Sharma CM, Gonzales K, Chao Y, Pirzada ZA, Eckert MR, Vogel J, Charpentier E (2011) CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III. Nature 471(7340):602–607

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Deveau H, Barrangou R, Garneau JE, Labonté J, Fremaux C, Boyaval P, Romero DA, Horvath P, Moineau S (2008) Phage response to CRISPR-encoded resistance in Streptococcus thermophilus. J Bacteriol 190(4):1390–1400

    Article  CAS  PubMed  Google Scholar 

  • Devi V, Harjai K, Chhibber S (2022) CRISPR-Cas systems: role in cellular processes beyond adaptive immunity. Folia Microbiol 67(6):837–850

    Article  CAS  Google Scholar 

  • Feng Z, Mao Y, Xu N, Zhang B, Wei P, Yang D-L, Wang Z, Zhang Z, Zheng R, Yang L (2014) Multigeneration analysis reveals the inheritance, specificity, and patterns of CRISPR/Cas-induced gene modifications in Arabidopsis. Proc Natl Acad Sci 111(12):4632–4637

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Garrett SC (2021) Pruning and tending immune memories: spacer dynamics in the CRISPR array. Front Microbiol 12:664299

    Article  PubMed  PubMed Central  Google Scholar 

  • Gophna U (2022) CRISPR‐Cas, horizontal gene transfer, and the flexible (Pan) genome. Crispr: Biology and Applications 121–130

  • Hao M, Cui Y, Qu X (2018) Analysis of CRISPR-Cas system in Streptococcus thermophilus and its application. Front Microbiol 9:288052

    Article  Google Scholar 

  • Hille F, Charpentier E (2016) CRISPR-Cas: biology, mechanisms and relevance. Philos Transact Royal Soc b: Biol Sci 371(1707):20150496

    Article  Google Scholar 

  • Horvath P, Romero DA, Coûté-Monvoisin A-C, Richards M, Deveau H, Moineau S, Boyaval P, Fremaux C, Barrangou R (2008) Diversity, activity, and evolution of CRISPR loci in Streptococcus thermophilus. J Bacteriol 190(4):1401–1412

    Article  CAS  PubMed  Google Scholar 

  • Horvath P, Coûté-Monvoisin A-C, Romero DA, Boyaval P, Fremaux C, Barrangou R (2009) Comparative analysis of CRISPR loci in lactic acid bacteria genomes. Int J Food Microbiol 131(1):62–70

    Article  CAS  PubMed  Google Scholar 

  • Hu T, Cui Y, Qu X (2020a) Characterization and comparison of CRISPR Loci in Streptococcus thermophilus. Arch Microbiol 202(4):695–710

    Article  CAS  PubMed  Google Scholar 

  • Hu T, Cui Y, Qu X (2020b) Characterization and comparison of CRISPR Loci in Streptococcus thermophilus. Arch Microbiol 202:695–710

    Article  CAS  PubMed  Google Scholar 

  • Hynes AP, Villion M, Moineau S (2014) Adaptation in bacterial CRISPR-Cas immunity can be driven by defective phages. Nat Commun 5(1):4399

    Article  CAS  PubMed  Google Scholar 

  • ILIKKAN OK (2021) Type III-A CRISPR/CAS systems and comparison of CAS1, CAS2, and CAS10 proteins of lactobacilli. Asian J Microbiol Biotechnol Environ Exp Sci 6(1):1–9

    Google Scholar 

  • Jackson SA, McKenzie RE, Fagerlund RD, Kieper SN, Fineran PC, Brouns SJ (2017) CRISPR-Cas: adapting to change. Science 356(6333):eaal5056

    Article  PubMed  Google Scholar 

  • Ka D, Jang DM, Han BW, Bae E (2018) Molecular organization of the type II-A CRISPR adaptation module and its interaction with Cas9 via Csn2. Nucleic Acids Res 46(18):9805–9815

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kahraman Ilıkkan Ö (2022) Analysis of Probiotic Bacteria Genomes: Comparison of CRISPR/Cas Systems and Spacer Acquisition Diversity. Ind J Microbiol 62(1):40–46

    Article  Google Scholar 

  • Kahraman-Ilıkkan Ö (2022) Comparison of Propionibacterium genomes: CRISPR-Cas systems, phage/plasmid diversity, and insertion sequences. Arch Microbiol 204(7):1–10

    Article  Google Scholar 

  • Kapse N, Pisu V, Dhakephalkar T, Margale P, Shetty D, Wagh S, Dagar S, Dhakephalkar PK (2024) Unveiling the Probiotic Potential of Streptococcus thermophilus MCC0200: Insights from In Vitro Studies Corroborated with Genome Analysis. Microorganisms 12(2):347

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Künne T, Kieper SN, Bannenberg JW, Vogel AI, Miellet WR, Klein M, Depken M, Suarez-Diez M, Brouns SJ (2016) Cas3-derived target DNA degradation fragments fuel primed CRISPR adaptation. Mol Cell 63(5):852–864

    Article  PubMed  Google Scholar 

  • Landsberger M, Gandon S, Meaden S, Rollie C, Chevallereau A, Chabas H, Buckling A, Westra ER, van Houte S (2018) Anti-CRISPR phages cooperate to overcome CRISPR-Cas immunity. Cell 174(4):908-916. e912

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Levin BR, Moineau S, Bushman M, Barrangou R (2013) The population and evolutionary dynamics of phage and bacteria with CRISPR–mediated immunity. PLoS Genet 9(3):e1003312

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Levy A, Goren MG, Yosef I, Auster O, Manor M, Amitai G, Edgar R, Qimron U, Sorek R (2015) CRISPR adaptation biases explain preference for acquisition of foreign DNA. Nature 520(7548):505–510

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu TY, Liu J-J, Aditham AJ, Nogales E, Doudna JA (2019) Target preference of Type III-A CRISPR-Cas complexes at the transcription bubble. Nat Commun 10(1):3001

    Article  PubMed  PubMed Central  Google Scholar 

  • Liu S, Liu H, Wang X, Shi L (2024) The immune system of prokaryotes: potential applications and implications for gene editing. Biotechnol J 19(2):2300352

    Article  CAS  Google Scholar 

  • Louwen R, Staals RH, Endtz HP, van Baarlen P, van der Oost J (2014) The role of CRISPR-Cas systems in virulence of pathogenic bacteria. Microbiol Mol Biol Rev 78(1):74–88

    Article  PubMed  PubMed Central  Google Scholar 

  • Makarova KS, Wolf YI, Koonin EV (2013) Comparative genomics of defense systems in archaea and bacteria. Nucleic Acids Res 41(8):4360–4377

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Maniv I, Jiang W, Bikard D, Marraffini LA (2016) Impact of different target sequences on type III CRISPR-Cas immunity. J Bacteriol 198(6):941–950

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Marchler-Bauer A, Derbyshire MK, Gonzales NR, Lu S, Chitsaz F, Geer LY, Geer RC, He J, Gwadz M, Hurwitz DI (2015) CDD: NCBI’s conserved domain database. Nucleic Acids Res 43(D1):D222–D226

    Article  CAS  PubMed  Google Scholar 

  • Marino ND, Pinilla-Redondo R, Csörgő B, Bondy-Denomy J (2020) Anti-CRISPR protein applications: natural brakes for CRISPR-Cas technologies. Nat Methods 17(5):471–479

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Marraffini LA (2015) CRISPR-Cas immunity in prokaryotes. Nature 526(7571):55–61

    Article  CAS  PubMed  Google Scholar 

  • McGinn J, Marraffini LA (2019) Molecular mechanisms of CRISPR–Cas spacer acquisition. Nat Rev Microbiol 17(1):7–12

    Article  CAS  PubMed  Google Scholar 

  • Mir A, Edraki A, Lee J, Sontheimer EJ (2018) Type II-C CRISPR-Cas9 biology, mechanism, and application. ACS Chem Biol 13(2):357–365

    Article  CAS  PubMed  Google Scholar 

  • Oh G-S, An S, Kim S (2024) Harnessing CRISPR-Cas adaptation for RNA recording and beyond. BMB Rep 57(1):40

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • O’Meara DM (2018) Examining the Role of Horizontal Gene Transfer on the Evolution of CRISPR-Cas. University of California, Riverside

    Google Scholar 

  • Paez-Espino D, Sharon I, Morovic W, Stahl B, Thomas BC, Barrangou R, Banfield JF (2015) CRISPR immunity drives rapid phage genome evolution in Streptococcus thermophilus. Mbio 6(2):e00262-e215

    Article  PubMed  PubMed Central  Google Scholar 

  • Philippe C, Moineau S (2021) The endless battle between phages and CRISPR–Cas systems in Streptococcus thermophilus. Biochem Cell Biol 99(4):397–402

    Article  CAS  PubMed  Google Scholar 

  • Philippe C, Morency C, Plante P-L, Zufferey E, Achigar R, Tremblay DM, Rousseau GM, Goulet A, Moineau S (2022) A truncated anti-CRISPR protein prevents spacer acquisition but not interference. Nat Commun 13(1):2802

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Richter C, Chang JT, Fineran PC (2012) Function and regulation of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated (Cas) systems. Viruses 4(10):2291–2311

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sampson TR, Weiss DS (2014) CRISPR-Cas systems: new players in gene regulation and bacterial physiology. Front Cell Infect Microbiol 4:37

    Article  PubMed  PubMed Central  Google Scholar 

  • Samson JE, Magadán AH, Sabri M, Moineau S (2013) Revenge of the phages: defeating bacterialdefences. Nat Rev Microbiol 11(10):675–687

    Article  CAS  PubMed  Google Scholar 

  • Scherz P (2017) The mechanism and applications of CRISPR-Cas9. The National Catholic Bioethics Quarterly 17(1):29–36

    Article  Google Scholar 

  • Semenova E, Jore MM, Datsenko KA, Semenova A, Westra ER, Wanner B, Van Der Oost J, Brouns SJ, Severinov K (2011) Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence. Proc Natl Acad Sci 108(25):10098–10103

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Severinov K, Ispolatov I, Semenova E (2016) The influence of copy-number of targeted extrachromosomal genetic elements on the outcome of CRISPR-Cas defense. Front Mol Biosci 3:45

    Article  PubMed  PubMed Central  Google Scholar 

  • Shabbir MAB, Shabbir MZ, Wu Q, Mahmood S, Sajid A, Maan MK, Ahmed S, Naveed U, Hao H, Yuan Z (2019) CRISPR-cas system: biological function in microbes and its use to treat antimicrobial resistant pathogens. Ann Clin Microbiol Antimicrob 18:1–9

    Article  CAS  Google Scholar 

  • Sun CL, Thomas BC, Barrangou R, Banfield JF (2016) Metagenomic reconstructions of bacterial CRISPR loci constrain population histories. ISME J 10(4):858–870

    Article  CAS  PubMed  Google Scholar 

  • Van Der Oost J, Westra ER, Jackson RN, Wiedenheft B (2014) Unravelling the structural and mechanistic basis of CRISPR–Cas systems. Nat Rev Microbiol 12(7):479–492

    Article  PubMed  PubMed Central  Google Scholar 

  • Vink JN, Baijens JH, Brouns SJ (2021) PAM-repeat associations and spacer selection preferences in single and co-occurring CRISPR-Cas systems. Genome Biol 22:1–25

    Article  Google Scholar 

  • Wang JY, Pausch P, Doudna JA (2022) Structural biology of CRISPR–Cas immunity and genome editing enzymes. Nat Rev Microbiol 20(11):641–656

    Article  CAS  PubMed  Google Scholar 

  • Waterhouse A, Bertoni M, Bienert S, Studer G, Tauriello G, Gumienny R, Heer FT, de Beer TAP, Rempfer C, Bordoli L (2018) SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res 46(W1):W296–W303

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Weinberger AD, Sun CL, Pluciński MM, Denef VJ, Thomas BC, Horvath P, Barrangou R, Gilmore MS, Getz WM, Banfield JF (2012) Persisting viral sequences shape microbial CRISPR-based immunity. PLoS Comput Biol 8(4):e1002475

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Westra ER, van Houte S, Oyesiku-Blakemore S, Makin B, Broniewski JM, Best A, Bondy-Denomy J, Davidson A, Boots M, Buckling A (2015) Parasite exposure drives selective evolution of constitutive versus inducible defense. Curr Biol 25(8):1043–1049

    Article  CAS  PubMed  Google Scholar 

  • Wright AV, Nuñez JK, Doudna JA (2016) Biology and applications of CRISPR systems: harnessing nature’s toolbox for genome engineering. Cell 164(1–2):29–44

    Article  CAS  PubMed  Google Scholar 

  • Wu Q, Cui L, Liu Y, Li R, Dai M, Xia Z, Wu M (2022) CRISPR-Cas systems target endogenous genes to impact bacterial physiology and alter mammalian immune responses. Molecular Biomedicine 3(1):22

    Article  PubMed  PubMed Central  Google Scholar 

  • Zhang Y, Showalter AM (2020) CRISPR/Cas9 genome editing technology: a valuable tool for understanding plant cell wall biosynthesis and function. Front Plant Sci 11:589517

    Article  PubMed  PubMed Central  Google Scholar 

  • Zhang Q, Ye Y (2017) Not all predicted CRISPR–Cas systems are equal: isolated cas genes and classes of CRISPR like elements. BMC Bioinformatics 18:1–12

    Article  Google Scholar 

  • Zheng Y, Li J, Wang B, Han J, Hao Y, Wang S, Ma X, Yang S, Ma L, Yi L (2020) Endogenous type I CRISPR-Cas: from foreign DNA defense to prokaryotic engineering. Frontiers Bioeng Biotechnol 8:62

    Article  Google Scholar 

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Authors and Affiliations

  1. Animal Originated Foodstuffs Department, Central Research Institute of Food and Feed Control, Bursa, Turkey

    Ali Özcan & Deniz Kiraz

  2. Food Hygiene and Technology Department, Faculty of Veterinary Medicine, Uludağ University, Bursa, Turkey

    Ali Özcan & Artun Yıbar

  3. Kahramankazan Vocational School, Food Quality Control and Analysis Program, Başkent University, Ankara, Turkey

    Özge Kahraman Ilıkkan

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  1. Ali Özcan
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  2. Artun Yıbar
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[Ali ÖZCAN]: Research design, data collection and analysis, writing and editing of the manuscript. [Deniz KİRAZ]: Literature review, data analysis, interpretation of results, revision of the manuscript. [Özge Kahraman ILIKKAN]: Statistical analysis, design of methods, proofreading of the manuscript. [Artun YIBAR]: Supervision, guiding the research process, final revision.

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Özcan, A., Yıbar, A., Kiraz, D. et al. Comprehensive analysis of the CRISPR-Cas systems in Streptococcus thermophilus strains isolated from traditional yogurts. Antonie van Leeuwenhoek 117, 63 (2024). https://doi.org/10.1007/s10482-024-01960-2

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