Pachnoda Marginata Larva'sının arka bağırsağından anaerobik lignoselülitik mikrobiyal kültür geliştirilmesi

Yıl 2020, Cilt: 26 Sayı: 6, 1117 - 1122, 13.11.2020

Emine Gözde Özbayram , Orhan İnce Sabine Kleınsteuber Marcell Nıkolausz

Öz

Tarım atıkları, yüksek enerji içerikleri ve üretim miktarları ile düşük maliyetleri nedeniyle anaerobik çürütücüler için en önemli biyokütle olarak değerlendirilmesine rağmen karmaşık lignoselülozik yapıları bu maddelerin hidroliz aşamalarını sınırlandırmaktadır. Hidroliz verimini artırmak için etkin lignoselüloz degradasyon stratejileri geliştirilmektedir. Bu çalışmanın amacı, Pachnoda marginata larvalarının arka bağırsağından lignoselüloz parçalayabilen anaerobik mikrobiyal topluluğun zenginleştirilmesidir. Bu kapsamda, Pachnoda marginata larvaları 3 hafta süre ile lignoselülozik substrat ile beslenmiş, sonrasında disekte edilmiştir. Arka bağırsak kültür şişelerine transfer edilmiştir. Bakteriyel topluluk çeşitliliği 16S rRNA amplikon dizileme yöntemi ile Illumina MiSeq platformunda analiz edilmiş, metanojenik arkeal topluluk ise T-RFLP yöntemi ile incelenmiştir. Kültür şişelerinde biyogaz üretimi zaman ile artış göstermiş, tüm şişelerde metan üretimi gözlenmiştir. Zenginleştirme prosedürü sonucunda, kültürün bakteriyel topluluk profili değişiklik göstermiş, üç transfer sonrasında alınan örneklerde Porphyromonadaceae (phylum: Bacteroidetes) bolluğunun artarak baskıladığı görülmüştür. Metanojenik topluluk ise Methanobrevibacter ile baskılanmıştır. Çalışmada elde edilen sonuçlar, Pachnoda marginata larvasının sindirim sisteminden üretilen zenginleştirilmiş kültürün, lignoselülozca zengin kompleks biyokütleyi etkin bir şekide parçalayabildiğini göstermiştir.

Anahtar Kelimeler

Bağırsak mikrobiyotası, Illumina Miseq, Lignoseüloz degredasyonu, Scarabaeidae, T-RFLP, Zenginleştirilmiş kültür

Kaynakça

• [1] Zhong W, Zhang Z, Luo Y, Sun S, Qiao W, Xiao M. “Effect of biological pretreatments in enhancing corn straw biogas production”. Bioresource Technology, 102, 11177-11182, 2011.
• [2] Ozbayram EG, Kleinsteuber S, Nikolausz M, Ince B, Ince O. “Effect of bioaugmentation by cellulolytic bacteria enriched from sheep rumen on methane production from wheat straw”. Anaerobe, 46, 122-130, 2017.
• [3] Akyol Ç, Ince O, Bozan M, Ozbayram EG, Ince B. “Biological pretreatment with Trametes versicolor to enhance methane T production from lignocellulosic biomass: A metagenomic approach”. Industrial Crops & Products, 140, 1-10, 2019.
• [4] Ozbayram EG, Kleinsteuber S, Nikolausz M, Ince B, Ince O. “Enrichment of lignocellulose-degrading microbial communities from natural and engineered methanogenic environments”. Applied Microbiology and Biotechnology, 102, 1035-1043, 2018.
• [5] Ozbayram EG, Akyol Ç, Ince B, Karakoc C, Ince O. “Rumen bacteria at work: bioaugmentation strategies to enhance biogas production from cow manure”. Journal of Applied Microbiology, 124, 491-502, 2018.
• [6] Egert M, Wagner B, Lemke T, Brune A, Friedrich MW. “Microbial community structure in midgut and hindgut of the humus-feeding larva of Pachnoda ephippiata (Coleoptera : Scarabaeidae)”. Applied Environmental Microbiology, 69, 6659-6668, 2003.
• [7] Takasuka TE, Book AJ, Lewin GR, Currie CR, Fox BG. “Aerobic deconstruction of cellulosic biomass by an insect-associated Streptomyces”. Scientific Reports, 3(1030), 1-10, 2013.
• [8] Lemke T, Stingl U, Egert M, Friedrich MW, Brune A. “Physicochemical Conditions and Microbial Activities in the Highly Alkaline Gut of the Humus-Feeding Larva of Pachnoda ephippiata (Coleoptera: Scarabaeidae)”. Applied Environmental Microbiology, 69, 6650-6658, 2003.
• [9] Morrison M, Pope PB, Denman SE, McSweeney CS. “Plant biomass degradation by gut microbiomes: more of the same or something new?” Current Opinion in Biotechnology, 20, 358-363, 2009.
• [10] Bayané A, Guiot SR. “Animal digestive strategies versus anaerobic digestion bioprocesses for biogas production from lignocellulosic biomass”. Reviews in Environmental Science and Biotechnology, 10, 43-62, 2011.
• [11] Huang SW, Zhang HY, Marshall S, Jackson TA. “The scarab gut: A potential bioreactor for bio-fuel production”. Insect Science, 17, 175-183, 2010.
• [12] Huang S, Sheng P, Zhang H. “Isolation and identification of cellulolytic bacteria from the gut of holotrichia parallela larvae (Coleoptera: Scarabaeidae)”. International Journal of Molecular Sciences, 13, 2563-2577, 2012.
• [13] Cazemier AE, Verdoes JC, Reubsaet FAG, Hackstein JHP, van der Drift C, Op den Camp HJM. “Promicromonospora pachnodae sp. nov., a member of the (hemi)cellulolytic hindgut flora of larvae of the scarab beetle Pachnoda marginata”. Antonie van Leeuwenhoek, International Journal of Molecular Medicine, 83, 135-148, 2003.
• [14] Watanabe H, Tokuda G. “Cellulolytic Systems in Insects”. Annual Review of Entomology, 55, 609-632, 2010.
• [15] Berasategui A, Shukla S, Salem H, Kaltenpoth M. “Potential applications of insect symbionts in biotechnology Targeting insect-microbe symbioses for biotechnological applications”. Applied Microbiology and Biotechnology, 1567-1577, 2016.
• [16] Sheng P, Huang J, Zhang Z, Wang D, Tian X, Ding J. “Construction and characterization of a cellulolytic consortium enriched from the hindgut of holotrichia parallela larvae”. International Journal of Molecular Sciences, 17(10), 2-12, 1646, 2016.
• [17] Ozbayram EG, Kleinsteuber S, Nikolausz M, Ince B, Ince O. “Bioaugmentation of anaerobic digesters treating lignocellulosic feedstock by enriched microbial consortia”. Engineering in Life Sciences, 18(7), 440-446, 2018.
• [18] Porsch K, Wirth B, Tóth EM, Schattenberg F, Nikolausz M. “Characterization of wheat straw-degrading anaerobic alkali-tolerant mixed cultures from soda lake sediments by molecular and cultivation techniques”. Microbial Biotechnology, 8, 801-814, 2015.
• [19] McDonald D, Price MN, Goodrich J, Nawrocki EP, DeSantis TZ, Probst A. “An improved Greengenes taxonomy with explicit ranks for ecological and evolutionary analyses of bacteria and archaea”. ISME Journal, 6, 610-618, 2012.
• [20] Wang Q, Garrity GM, Tiedje JM, Colen JR. “Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy”. Applied and Environmental Microbiology, 73, 5261-5267, 2007.
• [21] Ondov BD, Bergman NH, Phillippy AM. “Interactive metagenomic visualization in a Web browser”. BMC Bioinformatics, 12(385), 1-9, 2011.
• [22] Sträuber H, Bühligen F, Kleinsteuber S, Nikolausz M, Porsch K. “Improved anaerobic fermentation of wheat straw by alkaline pre-treatment and addition of alkali-tolerant microorganisms”. Bioengineering, 2, 66-93. 2015.
• [23] Depkat-Jakob PS, Hunger S, Schulz K, Brown GG, Tsai SM, Drake HL. “Emission of methane by Eudrilus eugeniae and other earthworms: From Brazil”. Applied and Environmental Microbiology, 78, 3014-3019, 2012.
• [24] Lory S. The Family Mycobacteriaceae. Editors: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F. The Prokaryotes, 571-575, Berlin, Heidelberg, Germany, Springer, 2014.
• [25] Stackebrandt E, Schumann P, Prauser H. The Family Cellulomonadaceae. Editors: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F. The Prokaryotes, 983-1001, Berlin, Heidelberg, Germany, Springer, 2006.
• [26] Octavia S, Lan R. The Family Enterobacteriaceae. Editors: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F. The Prokaryotes, 225-286, Berlin, Heidelberg, Germany, Springer, 2014.
• [27] Sakamoto M. The Family Porphyromonadaceae. Editors: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F. The Prokaryotes, 811-824, Berlin, Heidelberg, Germany, Springer, 2014.
• [28] Janssen PH, Kirs M. “Structure of the archaeal community of the rumen”. Applied Environmental Microbiology, 74, 3619-3625, 2008.