Abstract
The anaerobic gut fungi (Neocallimastigomycota), first described almost 50 years ago, hold extraordinary potential for biotechnology. Anaerobic fungi could contribute to bioenergy and bio-based chemical production via their ability to degrade lignocellulose, may enhance animal health and production, and are now being revealed to have interesting biosynthetic potential for development. In this chapter, we briefly describe the biology of these species, discuss recent efforts to identify and exploit anaerobic fungal enzymes, and survey challenges and opportunities for their adoption in a variety of biotechnology applications.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Aditiya HB, Mahlia TMI, Chong WT, Nur H, Sebayang AH (2016) Second generation bioethanol production: a critical review. Renew Sustain Energy Rev 66:631–653. https://doi.org/10.1016/j.rser.2016.07.015
Ali BRS, Zhou L, Graves FM, Freedman RB, Black GW, Gilbert HJ, Hazlewood GP (1995) Cellulases and hemicellulases of the anaerobic fungus Piromyces constitute a multiprotein cellulose-binding complex and are encoded by multigene families. FEMS Microbiol Lett 125(1):15–22. https://doi.org/10.1111/j.1574-6968.1995.tb07329.x
Aydin S, Yildirim E, Ince O, Ince B (2017) Rumen anaerobic fungi create new opportunities for enhanced methane production from microalgae biomass. Algal Res 23:150–160. https://doi.org/10.1016/j.algal.2016.12.016
Balat M, Balat H (2009) Recent trends in global production and utilization of bio-ethanol fuel. Appl Energy 86:2273–2282. https://doi.org/10.1016/j.apenergy.2009.03.015
Bauchop T, Mountfort DO (1981) Cellulose fermentation by a rumen anaerobic fungus in both the absence and the presence of rumen methanogens. Appl Environ Microbiol 42(6):1103–1110. https://www.ncbi.nlm.nih.gov/pubmed/16345902
Bayer EA, Belaich JP, Shoham Y, Lamed R (2004) The cellulosomes: multienzyme machines for degradation of plant cell wall polysaccharides. Annu Rev Microbiol 58:521–554. https://doi.org/10.1146/annurev.micro.57.030502.091022
Blum DL, Li X-L, Chen H, Ljungdahl LG (1999) Characterization of an acetyl xylan esterase from the anaerobic fungus Orpinomyces sp. strain PC-2. Appl Environ Microbiol 65(9):3990–3995. https://aem.asm.org/content/aem/65/9/3990.full.pdf
Boxma B, Voncken F, Jannink S, Van Alen T, Akhmanova A, Van Weelden SWH, Van Hellemond JJ, Ricard G, Huynen M, Tielens AGM, Hackstein JHP (2004) The anaerobic chytridiomycete fungus Piromyces sp. E2 produces ethanol via pyruvate:formate lyase and an alcohol dehydrogenase E. Mol Microbiol 51(5):1389–1399. https://doi.org/10.1046/j.1365-2958.2003.03912.x
Caldwell DR, Bryant MP (1966) Medium without rumen fluid for nonselective enumeration and isolation of rumen bacteria. Appl Microbiol 14(5):794–801. https://www.ncbi.nlm.nih.gov/pubmed/5970467
Calkins S, Elledge NC, Hanafy RA, Elshahed MS, Youssef N (2016) A fast and reliable procedure for spore collection from anaerobic fungi: application for RNA uptake and long-term storage of isolates. J Microbiol Methods 127:206–213. https://doi.org/10.1016/j.mimet.2016.05.019
Calkins SS, Elledge NC, Mueller KE, Marek SM, Couger MB, Elshahed MS, Youssef NH (2018) Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungus Pecoramyces ruminantium strain C1A. PeerJ 6:e4276. https://doi.org/10.7717/peerj.4276
Callaghan TM, Podmirseg SM, Hohlweck D, Edwards JE, Puniya AK, Dagar SS, Griffith GW (2015) Buwchfawromyces eastonii gen. nov., sp. nov.: a new anaerobic fungus (Neocallimastigomycota) isolated from buffalo faeces. MycoKeys 9:11–28. https://doi.org/10.3897/mycokeys.9.9032
Catalano V, Vergara M, Hauzenberger JR, Seiboth B, Sarrocco S, Vannacci G, Kubicek CP, Seidl-Seiboth V (2011) Use of a non-homologous end-joining-deficient strain (delta-ku70) of the biocontrol fungus Trichoderma virens to investigate the function of the laccase gene lcc1 in sclerotia degradation. Curr Genet 57(1):13–23. https://doi.org/10.1007/s00294-010-0322-2
Chen H, Li XL, Ljungdahl LG (1995) A cyclophilin from the polycentric anaerobic rumen fungus Orpinomyces sp. strain PC-2 is highly homologous to vertebrate cyclophilin B. Proc Natl Acad Sci USA 92(7):2587–2591. https://doi.org/10.1073/pnas.92.7.2587
Chen H, Li XL, Ljungdahl LG (1997) Sequencing of a 1,3-1,4-beta-D-glucanase (lichenase) from the anaerobic fungus Orpinomyces strain PC-2: properties of the enzyme expressed in Escherichia coli and evidence that the gene has a bacterial origin. J Bacteriol 179(19):6028–6034. https://doi.org/10.1128/jb.179.19.6028-6034.1997
Chen H, Li X-L, Blum DL, Ljungdahl LG (1998) Two genes of the anaerobic fungus Orpinomyces sp. strain PC-2 encoding cellulases with endoglucanase activities may have arisen by gene duplication. FEMS Microbiol Lett 159(1):63–68. https://doi.org/10.1111/j.1574-6968.1998.tb12842.x
Chen H, Li XL, Xu H, Ljungdahl LG, Cerniglia CE (2006) High level expression and characterization of the cyclophilin B gene from the anaerobic fungus Orpinomyces sp. strain PC-2. Protein Pept Lett 13(7):727–732. https://doi.org/10.2174/092986606777790511
Chen YC, Chen WT, Liu JC, Tsai LC, Cheng HL (2014) A highly active beta-glucanase from a new strain of rumen fungus Orpinomyces sp.Y102 exhibits cellobiohydrolase and cellotriohydrolase activities. Bioresour Technol 170:513–521. https://doi.org/10.1016/j.biortech.2014.08.016
Cheng YF, Edwards JE, Allison GG, Zhu WY, Theodorou MK (2009) Diversity and activity of enriched ruminal cultures of anaerobic fungi and methanogens grown together on lignocellulose in consecutive batch culture. Bioresour Technol 100(20):4821–4828. https://doi.org/10.1016/j.biortech.2009.04.031
Conrad R (1999) Contribution of hydrogen to methane production and control of hydrogen concentrations in methanogenic soils and sediments. FEMS Microbiol Ecol 28(3):193–202. https://doi.org/10.1111/j.1574-6941.1999.tb00575.x
Couger MB, Youssef NH, Struchtemeyer CG, Liggenstoffer AS, Elshahed MS (2015) Transcriptomic analysis of lignocellulosic biomass degradation by the anaerobic fungal isolate Orpinomyces sp. strain C1A. Biotechnol Biofuels 8(1):208. https://doi.org/10.1186/s13068-015-0390-0
Dagar SS, Kumar S, Mudgil P, Singh R, Puniya AK (2011) D1/D2 domain of large-subunit ribosomal DNA for differentiation of Orpinomyces spp. Appl Environ Microbiol 77(18):6722–6725. https://doi.org/10.1128/AEM.05441-11
Dagar SS, Kumar S, Griffith GW, Edwards JE, Callaghan TM, Singh R, Nagpal AK, Puniya AK (2015) A new anaerobic fungus (Oontomyces anksri gen. nov., sp. nov.) from the digestive tract of the Indian camel (Camelus dromedarius). Fungal Biol 119(8):731–737. https://doi.org/10.1016/j.funbio.2015.04.005
Dalrymple BP, Cybinski DH, Layton I, McSweeney CS, Xue G-P, Swadling YJ, Lowry JB (1997) Three Neocallimastix patriciarum esterases associated with the degradation of complex polysaccharides are members of a new family of hydrolases. Microbiology 143(8):2605–2614. https://doi.org/10.1099/00221287-143-8-2605
Dashtban M, Schraft H, Qin W (2009) Fungal bioconversion of lignocellulosic residues; opportunities & perspectives. Int J Biol Sci 5(6):578–595. https://doi.org/10.7150/ijbs.5.578
Davies DR, Theodorou MK, Lawrence MI, Trinci AP (1993) Distribution of anaerobic fungi in the digestive tract of cattle and their survival in faeces. J General Microbiol 139:1395–1400. https://doi.org/10.1099/00221287-139-6-1395
den Haan R, van Rensburg E, Rose SH, Görgens JF, van Zyl WH (2015) Progress and challenges in the engineering of non-cellulolytic microorganisms for consolidated bioprocessing. Curr Opin Biotechnol 33:32–38.. https://doi.org/10.1016/j.copbio.2014.10.003
Dey A, Sehgal JP, Puniya AK, Singh K (2004) Influence of an anaerobic fungal culture (Orpinomyces sp.) administration on growth rate, ruminal fermentation and nutrient digestion in calves. Asian Australas J Anim Sci 17(6):820–824. https://doi.org/10.5713/ajas.2004.820
Dollhofer V, Callaghan TM, Griffith GW, Lebuhn M, Bauer J (2017) Presence and transcriptional activity of anaerobic fungi in agricultural biogas plants. Bioresour Technol 235:131–139. https://doi.org/10.1016/j.biortech.2017.03.116
Dollhofer V, Dandikas V, Dorn-In S, Bauer C, Lebuhn M, Bauer J (2018) Accelerated biogas production from lignocellulosic biomass after pre-treatment with Neocallimastix frontalis. Bioresour Technol 264:219–227. https://doi.org/10.1016/j.biortech.2018.05.068
Durand R, Rascle C, Fischer M, Fèvre M (1997) Transient expression of the β-glucuronidase gene after biolistic transformation of the anaerobic fungus Neocallimastix frontalis. Curr Genet 31(2):158–161. https://doi.org/10.1007/s002940050190
Durand R, Rascle C, Fèvre M (1999) Expression of a catalytic domain of a Neocallimastix frontalis endoxylanase gene (xyn3) in Kluyveromyces lactis and Penicillium roqueforti. Appl Microbiol Biotechnol 52(2):208–214. https://doi.org/10.1007/s002530051510
Eberhardt RY, Gilbert HJ, Hazlewood GP (2000) Primary sequence and enzymic properties of two modular endoglucanases, Cel5A and Cel45A, from the anaerobic fungus Piromyces equi. Microbiology 146(8):1999–2008. https://doi.org/10.1099/00221287-146-8-1999
Emerling CA, Delsuc F, Nachman MW (2018) Chitinase genes (CHIAs) provide genomic footprints of a post-Cretaceous dietary radiation in placental mammals. Sci Adv 4(5):eaar6478. https://doi.org/10.1126/sciadv.aar6478
Ferraro A, Dottorini G, Massini G, Mazzurco Miritana V, Signorini A, Lembo G, Fabbricino M (2018) Combined bioaugmentation with anaerobic ruminal fungi and fermentative bacteria to enhance biogas production from wheat straw and mushroom spent straw. Bioresour Technol 260:364–373.. https://doi.org/10.1016/j.biortech.2018.03.128
Filya I (2004) Nutritive value and aerobic stability of whole crop maize silage harvested at four stages of maturity. Anim Feed Sci Technol 116(1/2):141–150. https://doi.org/10.1016/j.anifeedsci.2004.06.003
Gilmore SP, Henske JK, O’Malley MA (2015) Driving biomass breakdown through engineered cellulosomes. Bioengineered 6(4):204–208. https://doi.org/10.1080/21655979.2015.1060379
Gordon GL, Phillips MW (1998) The role of anaerobic gut fungi in ruminants. Nutr Res Rev 11(1):133–168. https://doi.org/10.1079/NRR19980009
Griffith GW, Ozkose E, Theodorou MK, Davies DR (2009) Diversity of anaerobic fungal populations in cattle revealed by selective enrichment culture using different carbon sources. Fungal Ecol 2(2):87–97. https://doi.org/10.1016/j.funeco.2009.01.005
Griffith GW, Baker S, Fliegerová K, Liggenstoffer A, van der Giezen M, Voigt K, Beakes G (2010) Anaerobic fungi: Neocallimastigomycota. IMA Fungus 1(2):181–185. http://www.ncbi.nlm.nih.gov/pubmed/22679578
Grigoriev IV, Cullen D, Goodwin SB, Hibbett D, Jeffries TW, Kubicek CP, Kuske C, Magnuson JK, Martin F, Spatafora JW, Tsang A, Baker SE (2011) Fueling the future with fungal genomics. Mycology 2(3):192–209. https://doi.org/10.1080/21501203.2011.584577
Grigoriev IV, Nikitin R, Haridas S, Kuo A, Ohm R, Otillar R, Riley R, Salamov A, Zhao X, Korzeniewski F, Smirnova T, Nordberg H, Dubchak I, Shabalov I (2014) MycoCosm portal: gearing up for 1000 fungal genomes. Nucleic Acids Res 42(D1):D699–D704. https://doi.org/10.1093/nar/gkt1183
Gruninger RJ, Puniya AK, Callaghan TM, Edwards JE, Youssef N, Dagar SS, Fliegerova K, Griffith GW, Forster R, Tsang A, McAllister T, Elshahed MS (2014) Anaerobic fungi (phylum Neocallimastigomycota): advances in understanding their taxonomy, life cycle, ecology, role and biotechnological potential. FEMS Microbiol Ecol 90(1):1–17. https://doi.org/10.1111/1574-6941.12383
Gruninger RJ, Cote C, McAllister TA, Abbott DW (2016) Contributions of a unique β-clamp to substrate recognition illuminates the molecular basis of exolysis in ferulic acid esterases. Biochem J 473(7):839–849. https://doi.org/10.1042/bj20151153
Hackstein JHP, Baker SE, van Hellemond JJ, Tielens AGM (2019) Hydrogenosomes of anaerobic fungi: an alternative way to adapt to anaerobic environments. In: Tachezy J (ed) Hydrogenosomes and mitosomes: mitochondria of anaerobic eukaryotes. Springer, Cham, pp 159–175. https://doi.org/10.1007/978-3-030-17941-0_7
Haferkamp I, Hackstein JH, Voncken FG, Schmit G, Tjaden J (2002) Functional integration of mitochondrial and hydrogenosomal ADP/ATP carriers in the Escherichia coli membrane reveals different biochemical characteristics for plants, mammals and anaerobic chytrids. Eur J Biochem 269(13):3172–3181
Haitjema CH, Solomon KV, Henske JK, Theodorou MK, O’Malley MA (2014) Anaerobic gut fungi: advances in isolation, culture, and cellulolytic enzyme discovery for biofuel production. Biotechnol Bioeng 111(8):1471–1482. https://doi.org/10.1002/bit.25264
Haitjema CH, Gilmore SP, Henske JK, Solomon KV, de Groot R, Kuo A, Mondo SJ, Salamov AA, LaButti K, Zhao Z, Chiniquy J, Barry K, Brewer HM, Purvine SO, Wright AT, Hainaut M, Boxma B, van Alen T, Hackstein JHP, Henrissat B, Baker SE, Grigoriev IV, O’Malley MA (2017) A parts list for fungal cellulosomes revealed by comparative genomics. Nat Microbiol 2:17087. https://doi.org/10.1038/nmicrobiol.2017.87
Hanafy RA, Elshahed MS, Liggenstoffer AS, Griffith GW, Youssef NH (2017) Pecoramyces ruminantium, gen. nov., sp. nov., an anaerobic gut fungus from the feces of cattle and sheep. Mycologia 109(2):231–243. https://doi.org/10.1080/00275514.2017.1317190
Hanafy RA, Elshahed MS, Youssef NH (2018) Feramyces austinii, gen. nov., sp. nov., an anaerobic gut fungus from rumen and fecal samples of wild Barbary sheep and fallow deer. Mycologia 110(3):513–525. https://doi.org/10.1080/00275514.2018.1466610
Hanafy RA, Lanjekar VB, Dhakephalkar PK, Callaghan TM, Dagar SS, Griffith GW, Elshahed MS, Youssef NH (2020) Seven new Neocallimastigomycota genera from wild, zoo-housed, and domesticated herbivores greatly expand the taxonomic diversity of the phylum. Mycologia. https://doi.org/10.1080/00275514.2019.1696619
Hara KY, Kobayashi J, Yamada R, Sasaki D, Kuriya Y, Hirono-Hara Y, Ishii J, Araki M, Kondo A (2017) Transporter engineering in biomass utilization by yeast. FEMS Yeast Res 17(7). https://doi.org/10.1093/femsyr/fox061
Harhangi HR, Steenbakkers PJM, Akhmanova A, Jetten MSM, van der Drift C, Op den Camp HJM (2002) A highly expressed family 1 β-glucosidase with transglycosylation capacity from the anaerobic fungus Piromyces sp. E2. Biochimica et Biophysica Acta 1574(3):293–303. https://doi.org/10.1016/S0167-4781(01)00380-3
Hassan SS, Williams GA, Jaiswal AK (2018) Emerging technologies for the pretreatment of lignocellulosic biomass. Bioresour Technol 262:310–318. https://doi.org/10.1016/j.biortech.2018.04.099
Hazlewood GP, Gilbert HJ (1993) Recombinant xylanases. International patent number: WO9325693. https://patents.google.com/patent/WO1993025693A1/en
Henske JK, Gilmore SP, Knop D, Cunningham FJ, Sexton JA, Smallwood CR, Shutthanandan V, Evans JE, Theodorou MK, O’Malley MA (2017) Transcriptomic characterization of Caecomyces churrovis: a novel, non-rhizoid-forming lignocellulolytic anaerobic fungus. Biotechnol Biofuels 10(1):305. https://doi.org/10.1186/s13068-017-0997-4
Henske JK, Gilmore SP, Haitjema CH, Solomon KV, O’Malley MA (2018a) Biomass-degrading enzymes are catabolite repressed in anaerobic gut fungi. AIChE J. https://doi.org/10.1002/aic.16395
Henske JK, Wilken SE, Solomon KV, Smallwood CR, Shutthanandan V, Evans JE, Theodorou MK, O’Malley MA (2018b) Metabolic characterization of anaerobic fungi provides a path forward for bioprocessing of crude lignocellulose. Biotechnol Bioeng 115(4):874–884. https://doi.org/10.1002/bit.26515
Hillman ET, Readnour LR, Solomon KV (2017) Exploiting the natural product potential of fungi with integrated -omics and synthetic biology approaches. Curr Opin Syst Biol 5:50–56. https://doi.org/10.1016/j.coisb.2017.07.010
Hooker CA, Hillman ET, Overton JC, Ortiz-Velez A, Schacht M, Hunnicutt A, Mosier NS, Solomon KV (2018) Hydrolysis of untreated lignocellulosic feedstock is independent of S-lignin composition in newly classified anaerobic fungal isolate, Piromyces sp. UH3-1. Biotechnol Biofuels 11:293. https://doi.org/10.1186/s13068-018-1292-8
Irvine HL, Stewart CS (1991) Interactions between anaerobic cellulolytic bacteria and fungi in the presence of Methanobrevibacter smithii. Lett Appl Microbiol 12(2):62–64. https://doi.org/10.1111/j.1472-765X.1991.tb00504.x
James TY, Kauff F, Schoch CL, Matheny PB, Hofstetter V, Cox CJ, Celio G, Gueidan C, Fraker E, Miadlikowska J, Lumbsch HT, Rauhut A, Reeb V, Arnold AE, Amtoft A, Stajich JE, Hosaka K, Sung G-H, Johnson D, O’Rourke B, Crockett M, Binder M, Curtis JM, Slot JC, Wang Z, Wilson AW, Schüßler A, Longcore JE, O’Donnell K, Mozley-Standridge S, Porter D, Letcher PM, Powell MJ, Taylor JW, White MM, Griffith GW, Davies DR, Humber RA, Morton JB, Sugiyama J, Rossman AY, Rogers JD, Pfister DH, Hewitt D, Hansen K, Hambleton S, Shoemaker RA, Kohlmeyer J, Volkmann-Kohlmeyer B, Spotts RA, Serdani M, Crous PW, Hughes KW, Matsuura K, Langer E, Langer G, Untereiner WA, Lücking R, Büdel B, Geiser DM, Aptroot A, Diederich P, Schmitt I, Schultz M, Yahr R, Hibbett DS, Lutzoni F, McLaughlin DJ, Spatafora JW, Vilgalys R (2006) Reconstructing the early evolution of fungi using a six-gene phylogeny. Nature 443(7113):818–822. https://doi.org/10.1038/nature05110
Jin W, Cheng YF, Mao SY, Zhu WY (2011) Isolation of natural cultures of anaerobic fungi and indigenously associated methanogens from herbivores and their bioconversion of lignocellulosic materials to methane. Bioresour Technol 102(17):7925–7931. https://doi.org/10.1016/j.biortech.2011.06.026
Joblin K (1981) Isolation, enumeration, and maintenance of rumen anaerobic fungi in roll tubes. Appl Environ Microbiol 42(6):1119-1122. http://aem.asm.org/content/42/6/1119.full.pdf
Joblin KN, Naylor GE (1989) Fermentation of woods by rumen anaerobic fungi. FEMS Microbiol Lett 65(1–2):119–122. https://doi.org/10.1111/j.1574-6968.1989.tb03608.x
Joblin KN, Campbell GP, Richardson AJ, Stewart CS (1989) Fermentation of barley straw by anaerobic rumen bacteria and fungi in axenic culture and in co-culture with methanogens. Lett Appl Microbiol 9(5):195–197. https://doi.org/10.1111/j.1472-765X.1989.tb00323.x
Jones DR, Uddin MS, Gruninger RJ, Pham TTM, Thomas D, Boraston AB, Briggs J, Pluvinage B, McAllister TA, Forster RJ, Tsang A, Selinger LB, Abbott DW (2017) Discovery and characterization of family 39 glycoside hydrolases from rumen anaerobic fungi with polyspecific activity on rare arabinosyl substrates. J Biol Chem 292(30):12606–12620. https://doi.org/10.1074/jbc.M117.789008
Joshi A, Lanjekar VB, Dhakephalkar PK, Callaghan TM, Griffith GW, Dagar SS (2018) Liebetanzomyces polymorphus gen. et sp. nov., a new anaerobic fungus (Neocallimastigomycota) isolated from the rumen of a goat. MycoKeys 40(40):89–110. https://doi.org/10.3897/mycokeys.40.28337
Kameshwar AKS, Qin W (2018) Genome wide analysis reveals the extrinsic cellulolytic and biohydrogen generating abilities of Neocallimastigomycota fungi. J Genomics 6:74–87. https://doi.org/10.7150/jgen.25648
Kuyper M, Harhangi HR, Stave AK, Winkler AA, Jetten MSM, Laat WTAM, Ridder JJJ, Op den Camp HJM, Dijken JP, Pronk JT (2003) High-level functional expression of a fungal xylose isomerase: the key to efficient ethanolic fermentation of xylose by Saccharomyces cerevisiae? FEMS Yeast Res 4(1):69–78. https://doi.org/10.1016/S1567-1356(03)00141-7
Kuyper M, Hartog MM, Toirkens MJ, Almering MJ, Winkler AA, Dijken JP, Pronk JT (2005) Metabolic engineering of a xylose-isomerase-expressing Saccharomyces cerevisiae strain for rapid anaerobic xylose fermentation. FEMS Yeast Res 5(4-5):399–409. https://doi.org/10.1016/j.femsyr.2004.09.010
la Grange DC, den Haan R, van Zyl WH (2010) Engineering cellulolytic ability into bioprocessing organisms. Appl Microbiol Biotechnol 87(4):1195–1208. https://doi.org/10.1007/s00253-010-2660-x
Lamed R, Setter E, Kenig R, Edward B (1983) The cellulosome: a discrete cell surface organelle of Clostridium thermocellum which exhibits separate antigenic, cellulose-binding and various cellulolytic activities. In: Biotechnology bioengineering symposium, vol 13. https://www.researchgate.net/profile/Bayer_Edward/publication/236400605_The_cellulosome_A_discrete_cell_surface_organelle_of_Clostridium_thermocellum_which_exhibits_separate_antigenic_cellulose-binding_and_various_cellulolytic_activities/links/545524ca0cf2cf51647dd28c.pdf
Lee SS, Ha JK, Cheng K-J (2000) Relative contributions of bacteria, protozoa, and fungi to in vitro degradation of orchard grass cell walls and their interactions. Appl Environ Microbiol 66(9):3807–3813. https://doi.org/10.1128/AEM.66.9.3807-3813.2000
Leis S, Dresch P, Peintner U, Fliegerová K, Sandbichler AM, Insam H, Podmirseg SM (2014) Finding a robust strain for biomethanation: anaerobic fungi (Neocallimastigomycota) from the Alpine ibex (Capra ibex) and their associated methanogens. Anaerobe 29:34–43. https://doi.org/10.1016/j.anaerobe.2013.12.002
Li X-L, Ljungdahl LG, Ximenes EA, Chen H, Felix CR, Cotta MA, Dien BS (2004) Properties of a recombinant β-glucosidase from polycentric anaerobic fungus Orpinomyces PC-2 and its application for cellulose hydrolysis. Appl Biochem Biotechnol 113–116:233–250. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5890932/
Li XL, Skory CD, Ximenes EA, Jordan DB, Dien BS, Hughes SR, Cotta MA (2007) Expression of an AT-rich xylanase gene from the anaerobic fungus Orpinomyces sp. strain PC-2 in and secretion of the heterologous enzyme by Hypocrea jecorina. Appl Microbiol Biotechnol 74(6):1264–1275. https://doi.org/10.1007/s00253-006-0787-6
Li Y, Jin W, Cheng Y, Zhu W (2016) Effect of the associated methanogen methanobrevibacter thaueri on the dynamic profile of end and intermediate metabolites of anaerobic fungus Piromyces sp. F1. Curr Microbiol 73(3):434–441. https://doi.org/10.1007/s00284-016-1078-9
Li Y, Li Y, Jin W, Sharpton TJ, Mackie RI, Cann I, Cheng Y, Zhu W (2019) Combined genomic, transcriptomic, proteomic, and physiological characterization of the growth of Pecoramyces sp. F1 in monoculture and co-culture with a syntrophic methanogen. Front Microbiol 10(435):435. https://doi.org/10.3389/fmicb.2019.00435
Liggenstoffer AS, Youssef NH, Couger MB, Elshahed MS (2010) Phylogenetic diversity and community structure of anaerobic gut fungi (phylum Neocallimastigomycota) in ruminant and non-ruminant herbivores. ISME J 4(10):1225–1235. https://doi.org/10.1038/ismej.2010.49
Liu JH, Selinger LB, Hu YJ, Moloney MM, Cheng KJ, Beauchemin KA (1997) An endoglucanase from the anaerobic fungus Orpinomyces joyonii: characterization of the gene and its product. Can J Microbiol 43(5):477–485. https://www.ncbi.nlm.nih.gov/pubmed/9165703
Ljungdahl LG (2008) The cellulase/hemicellulase system of the anaerobic fungus Orpinomyces PC-2 and aspects of its applied use. Ann N Y Acad Sci 1125(1):308–321. https://doi.org/10.1196/annals.1419.030
Lombard V, Ramulu HG, Drula E, Coutinho PM, Henrissat B (2014) The carbohydrate-active enzymes database (CAZy) in 2013. Nucleic Acids Res 42(Database issue):D490–D495. https://doi.org/10.1093/nar/gkt1178
López-Contreras AM, Smidt H, van der Oost J, Claassen PAM, Mooibroek H, de Vos WM (2001) Clostridium beijerinckii cells expressing Neocallimastix patriciarum glycoside hydrolases show enhanced lichenan utilization and solvent production. Appl Environ Microbiol 67(11):5127–5133. https://doi.org/10.1128/aem.67.11.5127-5133.2001
Lowe SE, Theodorou MK, Trinci AP, Hespell RB (1985) Growth of anaerobic rumen fungi on defined and semi-defined media lacking rumen fluid. J Gen Microbiol 131(9):2225–2229. http://mic.sgmjournals.org/content/131/9/2225.full.pdf
Marvin-Sikkema FD, Pedro Gomes TM, Gottschal JC, Prins RA, Marvin-Sikkema FD (1993) Characterization of hydrogenosomes and their role in glucose metabolism of Neocallimastix sp. L2. Arch Microbiol 160(5). https://doi.org/10.1007/bf00252226
Milne A, Theodorou MK, Jordan MGC, King-Spooner C, Trinci APJ (1989) Survival of anaerobic fungi in feces, in saliva, and in pure culture. Exp Mycol 13(1):27–37. https://doi.org/10.1016/0147-5975(89)90005-4
Mondo SJ, Dannebaum RO, Kuo RC, Louie KB, Bewick AJ, LaButti K, Haridas S, Kuo A, Salamov A, Ahrendt SR, Lau R, Bowen BP, Lipzen A, Sullivan W, Andreopoulos BB, Clum A, Lindquist E, Daum C, Northen TR, Kunde-Ramamoorthy G, Schmitz RJ, Gryganskyi A, Culley D, Magnuson J, James TY, O’Malley MA, Stajich JE, Spatafora JW, Visel A, Grigoriev IV (2017) Widespread adenine N6-methylation of active genes in fungi. Nat Genet 49(6):964–968. https://doi.org/10.1038/ng.3859. http://www.nature.com/ng/journal/v49/n6/abs/ng.3859.html#supplementary-information
Montenecourt BS, Eveleigh DE (1977) Preparation of mutants of Trichoderma reesei with enhanced cellulase production. Appl Environ Microbiol 34(6):777–782. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC242747/
Mood SH, Golfeshan AH, Tabatabaei M, Jouzani GS, Najafi GH, Gholami M, Ardjmand M (2013) Lignocellulosic biomass to bioethanol, a comprehensive review with a focus on pretreatment. Renew Sustain Energy Rev 27:77–93. https://doi.org/10.1016/j.rser.2013.06.033
Morrison JM, Elshahed MS, Youssef N (2016a) A multifunctional GH39 glycoside hydrolase from the anaerobic gut fungus Orpinomyces sp. strain C1A. PeerJ 4:e2289. https://doi.org/10.7717/peerj.2289
Morrison JM, Elshahed MS, Youssef NH (2016b) Defined enzyme cocktail from the anaerobic fungus Orpinomyces sp. strain C1A effectively releases sugars from pretreated corn stover and switchgrass. Sci Rep 6:29217. https://doi.org/10.1038/srep29217
Mountfort D, Orpin C (1994) Anaerobic fungi: biology, ecology, and function. CRC, Boca Raton, FL
Müller M, Mentel M, van Hellemond JJ, Henze K, Woehle C, Gould SB, Yu RY, van der Giezen M, Tielens AG, Martin WF (2012) Biochemistry and evolution of anaerobic energy metabolism in eukaryotes. Microbiol Mol Biol Rev 76(2):444–495. https://doi.org/10.1128/mmbr.05024-11
Murphy CL, Youssef NH, Hanafy RA, Couger MB, Stajich JE, Wang Y, Baker K, Dagar SS, Griffith GW, Farag IF, Callaghan TM, Elshahed MS (2019) Horizontal gene transfer as an indispensable driver for Neocallimastigomycota evolution into a distinct gut-dwelling fungal lineage. Appl Environ Microbiol. https://doi.org/10.1128/aem.00988-19
Nagler M, Kozjek K, Etemadi M, Insam H, Podmirseg SM (2019) Simple yet effective: microbial and biotechnological benefits of rumen liquid addition to lignocellulose-degrading biogas plants. J Biotechnol 300:1–10. https://doi.org/10.1016/j.jbiotec.2019.05.004
Nielsen BB, Zhu W-Y, Trinci APJ, Theodorou MK (1995) Demonstration of zoosporangia of anaerobic fungi on plant residues recovered from faeces of cattle. Mycol Res 99(4):471–474. https://doi.org/10.1016/S0953-7562(09)80648-5
Nkemka VN, Gilroyed B, Yanke J, Gruninger R, Vedres D, McAllister T, Hao X (2015) Bioaugmentation with an anaerobic fungus in a two-stage process for biohydrogen and biogas production using corn silage and cattail. Bioresour Technol 185:79–88. https://doi.org/10.1016/j.biortech.2015.02.100
Obembe OO, Jacobsen E, Timmers J, Gilbert H, Blake AW, Knox JP, Visser RGF, Vincken J-P (2007) Promiscuous, non-catalytic, tandem carbohydrate-binding modules modulate the cell-wall structure and development of transgenic tobacco (Nicotiana tabacum) plants. J Plant Res 120(5):605–617. https://doi.org/10.1007/s10265-007-0099-7
O’Malley MA, Theodorou MK, Kaiser CA (2012) Evaluating expression and catalytic activity of anaerobic fungal fibrolytic enzymes native topiromyces sp E2 in Saccharomyces cerevisiae. Environ Progr Sustain Energy 31(1):37–46. http://onlinelibrary.wiley.com/doi/10.1002/ep.10614/abstract
Orpin CG (1975) Studies on the rumen flagellate Neocallimastix frontalis. J Gen Microbiol 91(2):249–262. https://doi.org/10.1099/00221287-91-2-249
Orpin CG (1977) On the induction of zoosporogenesis in the rumen phycomycetes Neocallimastix frontalis, Piromonas communis and Sphaeromonas communis. J Gen Microbiol 101(2):181–189. https://doi.org/10.1099/00221287-101-2-181
Paul SS, Kamra DN, Sastry VR, Sahu NP, Agarwal N (2004) Effect of anaerobic fungi on in vitro feed digestion by mixed rumen microflora of buffalo. Reprod Nutr Dev 44 (4):313–319.. http://www.ncbi.nlm.nih.gov/pubmed/15535463
Paul SS, Deb SM, Punia BS, Das KS, Singh G, Ashar MN, Kumar R (2011) Effect of feeding isolates of anaerobic fungus Neocallimastix sp. CF 17 on growth rate and fibre digestion in buffalo calves. Arch Anim Nutr 65(3):215–228. https://doi.org/10.1080/1745039X.2011.559722
Peng XN, Gilmore SP, O’Malley MA (2016) Microbial communities for bioprocessing: lessons learned from nature. Curr Opin Chem Eng 14:103–109. https://doi.org/10.1016/j.coche.2016.09.003
Peng X, Swift CL, Theodorou MK, O’Malley MA (2018) Methods for genomic characterization and maintenance of anaerobic fungi. In: de Vries RP, Tsang A, Grigoriev I (eds) Fungal genomics. Methods in molecular biology, vol 1775. Humana, New York
Podolsky IA, Seppälä S, Lankiewicz TS, Brown JL, Swift CL, O’Malley MA (2019) Harnessing nature’s anaerobes for biotechnology and bioprocessing. 10 (1). https://doi.org/10.1146/annurev-chembioeng-060718-030340
Poidevin L, Levasseur A, Paës G, Navarro D, Heiss-Blanquet S, Asther M, Record E (2009) Heterologous production of the Piromyces equi cinnamoyl esterase in Trichoderma reesei for biotechnological applications. Lett Appl Microbiol 49(6):673–678. https://doi.org/10.1111/j.1472-765X.2009.02734.x
Prasad V, Strömberg CAE, Alimohammadian H, Sahni A (2005) Dinosaur coprolites and the early evolution of grasses and grazers. Science 310(5751):1177–1180. https://doi.org/10.1126/science.1118806
Procházka J, Mrázek J, Štrosová L, Fliegerová K, Zábranská J, Dohányos M (2012) Enhanced biogas yield from energy crops with rumen anaerobic fungi. Eng Life Sci 12(3):343–351. https://doi.org/10.1002/elsc.201100076
Qiu X, Selinger B, Yanke LJ, Cheng KJ (2000) Isolation and analysis of two cellulase cDNAs from Orpinomyces joyonii. Gene 245 (1):119–126. https://doi.org/10.1016/S0378-1119(00)00028-7
Ranganathan A, Smith OP, Youssef NH, Struchtemeyer CG, Atiyeh HK, Elshahed MS (2017) Utilizing anaerobic fungi for two-stage sugar extraction and biofuel production from lignocellulosic biomass. Front Microbiol 8(635):635. https://doi.org/10.3389/fmicb.2017.00635
Ribeiro GO, Oss DB, He Z, Gruninger RJ, Elekwachi C, Forster RJ, Yang W, Beauchemin KA, McAllister TA (2017) Repeated inoculation of cattle rumen with bison rumen contents alters the rumen microbiome and improves nitrogen digestibility in cattle. Sci Rep 7(1):1276. https://doi.org/10.1038/s41598-017-01269-3
Sauer M, Porro D, Mattanovich D, Branduardi P (2008) Microbial production of organic acids: expanding the markets. Trends Biotechnol 26(2):100–108. https://doi.org/10.1016/j.tibtech.2007.11.006
Saxena S, Sehgal JP, Puniya AK, Singh K (2010) Effect of administration of rumen fungi on production performance of lactating buffaloes. Benef Microbes 1(2):183–188. https://doi.org/10.3920/bm2009.0018
Schnürer A (2016) Biogas production: microbiology and technology. In: Hatti-Kaul R, Mamo G, Mattiasson B (eds) Anaerobes in biotechnology. Advances in biochemical engineering/biotechnology, vol 156. Springer, Cham. https://doi.org/10.1007/10_2016_5
Schnürer A (2018) Microbiology of the biogas process. Swedish University of Agricultural Sciences
Seppälä S, Solomon KS, Gilmore SP, Henske JK, Rieth MD, O’Malley MA (2016) Mapping the membrane proteome of anaerobic gut fungi using RNA-Seq. Biophys J 110 (3, Suppl 1):58a–59a. https://doi.org/10.1016/j.bpj.2015.11.382
Seppälä S, Wilken SE, Knop D, Solomon KV, O’Malley MA (2017) The importance of sourcing enzymes from non-conventional fungi for metabolic engineering and biomass breakdown. Metab Eng 44:45–59. https://doi.org/10.1016/j.ymben.2017.09.008
Seppälä S, Yoo JI, Yur D, O’Malley MA (2019) Heterologous transporters from anaerobic fungi bolster fluoride tolerance in Saccharomyces cerevisiae. Metab Eng Commun 9:e00091. https://doi.org/10.1016/j.mec.2019.e00091
Sharma HK, Xu C, Qin W (2017) Biological pretreatment of lignocellulosic biomass for biofuels and bioproducts: an overview. Waste Biomass Valorization 10(2):235–251. https://doi.org/10.1007/s12649-017-0059-y
Skyba O, Douglas CJ, Mansfield SD (2013) Syringyl-rich lignin renders poplars more resistant to degradation by wood decay fungi. Appl Environ Microbiol 79(8):2560–2571. https://doi.org/10.1128/aem.03182-12
Solomon KV, Haitjema CH, Henske JK, Gilmore SP, Borges-Rivera D, Lipzen A, Brewer HM, Purvine SO, Wright AT, Theodorou MK, Grigoriev IV, Regev A, Thompson DA, O’Malley MA (2016a) Early-branching gut fungi possess a large, comprehensive array of biomass-degrading enzymes. Science 351(6278):1192–1195. https://doi.org/10.1126/science.aad1431
Solomon KV, Henske JK, Theodorou MK, O’Malley MA (2016b) Robust and effective methodologies for cryopreservation and DNA extraction from anaerobic gut fungi. Anaerobe 38:39–46. https://doi.org/10.1016/j.anaerobe.2015.11.008
Solomon KV, Henske JK, Gilmore SP, Lipzen A, Grigoriev IV, Thompson D, O’Malley MA (2018) Catabolic repression in early-diverging anaerobic fungi is partially mediated by natural antisense transcripts. Fungal Genet Biol 121:1–9. https://doi.org/10.1016/j.fgb.2018.09.004
Strauss J, Mach RL, Zeilinger S, Hartler G, Stöffler G, Wolschek M, Kubicek CP (1995) Crel, the carbon catabolite repressor protein from Trichoderma reesei. FEBS Lett 376 (1):103–107. https://doi.org/10.1016/0014-5793(95)01255-5
Strzalka R, Schneider D, Eicker U (2017) Current status of bioenergy technologies in Germany. Renew Sustain Energy Rev 72:801–820. https://doi.org/10.1016/j.rser.2017.01.091
Swift CL, Brown JL, Seppälä S, O’Malley MA (2019) Co-cultivation of the anaerobic fungus Anaeromyces robustus with Methanobacterium bryantii enhances transcription of carbohydrate active enzymes. J Ind Microbiol Biotechnol. https://doi.org/10.1007/s10295-019-02188-0
Ter Beek J, Guskov A, Slotboom DJ (2014) Structural diversity of ABC transporters. J Gen Physiol 143(4):419–435. https://doi.org/10.1085/jgp.201411164
Teunissen MJ, Baerends RJS, Knelissen RAG, Op den Camp HJM, Vogels GD (1992) A semi-continuous culture system for production of cellulolytic and xylanolytic enzymes by the anaerobic fungus Piromyces sp. strain E2. Appl Microbiol Biotechnol 38(1):28–33. https://doi.org/10.1007/bf00169414
Theodorou MK, Gill M, King-Spooner C, Beever DE (1990) Enumeration of anaerobic chytridiomycetes as thallus-forming units: novel method for quantification of fibrolytic fungal populations from the digestive tract ecosystem. Appl Environ Microbiol 56(4):1073–1078. http://aem.asm.org/content/56/4/1073.full.pdf
Theodorou MK, Mennim G, Davies DR, Zhu WY, Trinci AP, Brookman JL (1996a) Anaerobic fungi in the digestive tract of mammalian herbivores and their potential for exploitation. Proc Nutr Soc 55(3):913–926. https://doi.org/10.1079/pns19960088
Theodorou MK, Zhu W-Y, Rickers A, Nielsen BB, Gull K, Trinci APJ (1996b) Biochemistry and ecology of anaerobic fungi. In: Howard DH, Miller JD (eds) Human and animal relationships. Springer, Berlin, pp 265–295. https://doi.org/10.1007/978-3-662-10373-9_14
Trinci APG, Davies DR, Gull K, Lawrence MI, Nielsen BB, Rickers A (1994) Anaerobic fungi in herbivorous animals. Mycol Res 98. https://doi.org/10.1016/s0953-7562(09)80178-0
Tripathi VK, Sehgal JP, Puniya AK, Singh K (2007) Effect of administration of anaerobic fungi isolated from cattle and wild blue bull (Boselaphus tragocamelus) on growth rate and fibre utilization in buffalo calves AU - Tripathi, Vimal K. Arch Anim Nutr 61(5):416–423. https://doi.org/10.1080/17450390701556759
Tsai C-F, Qiu X, Liu J-H (2003) A comparative analysis of two cDNA clones of the cellulase gene family from anaerobic fungus Piromyces rhizinflata. Anaerobe 9(3):131–140. https://doi.org/10.1016/S1075-9964(03)00087-8
Ushida K, Newbold CJ, Jouany JP (1997) Interspecies hydrogen transfer between the rumen ciliate Polyplastron multivesiculatum and Methanosarcina barkeri. J Gen Appl Microbiol 43(2):129–131. https://www.ncbi.nlm.nih.gov/pubmed/12501346
van der Giezen M, Kiel JAKW, Sjollema KA, Prins RA (1998) The hydrogenosomal malic enzyme from the anaerobic fungus Neocallimastix frontalis is targeted to mitochondria of the methylotrophic yeast Hansenula polymorpha. Curr Genet 33(2):131–135. https://doi.org/10.1007/s002940050318
Wang T-Y, Chen H-L, Lu M-Y, Chen Y-C, Sung H-M, Mao C-T, Cho H-Y, Ke H-M, Hwa T-Y, Ruan S-K, Hung K-Y, Chen C-K, Li J-Y, Wu Y-C, Chen Y-H, Chou S-P, Tsai Y-W, Chu T-C, Shih C-C, Li W-H, Shih M-C (2011) Functional characterization of cellulases identified from the cow rumen fungus Neocallimastix patriciarum W5 by transcriptomic and secretomic analyses. Biotechnol Biofuels 4(1):24. https://doi.org/10.1186/1754-6834-4-24
Wang X, Liu X, Groenewald JZ (2017) Phylogeny of anaerobic fungi (phylum Neocallimastigomycota), with contributions from yak in China. Antonie Van Leeuwenhoek 110(1):87–103. https://doi.org/10.1007/s10482-016-0779-1
Wang Y, Youssef N, Couger M, Hanafy R, Elshahed M, Stajich JE (2019) Comparative genomics and divergence time estimation of the anaerobic fungi in herbivorous mammals. bioRxiv 401869 [Preprint]. 31 Aug 2018 [cited 30 Mar 2019]. Available from https://doi.org/10.1101/401869
Weimer PJ, Russell JB, Muck RE (2009) Lessons from the cow: what the ruminant animal can teach us about consolidated bioprocessing of cellulosic biomass. Bioresour Technol 100(21):5323–5331. https://doi.org/10.1016/j.biortech.2009.04.075
Wilken SE, Swift CL, Podolsky IA, Lankiewicz TS, Seppälä S, O’Malley MA (2019) Linking ‘omics’ to function unlocks the biotech potential of non-model fungi. Curr Opin Syst Biol 14:9–17.. https://doi.org/10.1016/j.coisb.2019.02.001
Wilson CA, Wood TM (1992) The anaerobic fungus Neocallimastix frontalis: isolation and properties of a cellulosome-type enzyme fraction with the capacity to solubilize hydrogen-bond-ordered cellulose. Appl Microbiol Biotechnol 37(1):125–129. https://link.springer.com/article/10.1007/BF00174216
Wubah DA, Kim DS (1996) Chemoattraction of anaerobic ruminal fungi zoospores to selected phenolic acids. Microbiol Res 151(3):257–262. https://doi.org/10.1016/S0944-5013(96)80022-X
Wubah DA, Fuller MS, Akin DE (1991) Resistant body formation in Neocallimastix Sp., an anaerobic fungus from the rumen of a cow. Mycologia 83(1):40–47. https://doi.org/10.1080/00275514.1991.12025977
Yang S-T, Liu X, Zhang Y (2007) Chapter 4—Metabolic engineering—applications, methods, and challenges. In: Yang S-T (ed) Bioprocessing for value-added products from renewable resources. Elsevier, Amsterdam, pp 73–118. https://doi.org/10.1016/B978-044452114-9/50005-0
Ye XY, Ng TB, Cheng KJ (2001) Purification and characterization of a cellulase from the ruminal fungus Orpinomyces joyonii cloned in Escherichia coli. Int J Biochem Cell Biol 33(1):87–94. https://doi.org/10.1016/S1357-2725(00)00068-6
Yildirim E, Ince O, Aydin S, Ince B (2017) Improvement of biogas potential of anaerobic digesters using rumen fungi. Renew Energy 109:346–353. https://doi.org/10.1016/j.renene.2017.03.021
Young E, Poucher A, Comer A, Bailey A, Alper H (2011) Functional survey for heterologous sugar transport proteins, using Saccharomyces cerevisiae as a host. Appl Environ Microbiol 77(10):3311–3319. https://doi.org/10.1128/AEM.02651-10
Young D, Dollhofer V, Callaghan TM, Reitberger S, Lebuhn M, Benz JP (2018) Isolation, identification and characterization of lignocellulolytic aerobic and anaerobic fungi in one- and two-phase biogas plants. Bioresour Technol 268:470–479. https://doi.org/10.1016/j.biortech.2018.07.103
Youssef NH, Couger MB, Struchtemeyer CG, Liggenstoffer AS, Prade RA, Najar FZ, Atiyeh HK, Wilkins MR, Elshahed MS (2013) The genome of the anaerobic fungus Orpinomyces sp. strain C1A reveals the unique evolutionary history of a remarkable plant biomass degrader. Appl Environ Microbiol 79(15):4620–4634. https://doi.org/10.1128/AEM.00821-13
Zetsche B, Gootenberg JS, Abudayyeh OO, Slaymaker IM, Makarova KS, Essletzbichler P, Volz SE, Joung J, van der Oost J, Regev A, Koonin EV, Zhang F (2015) Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system. Cell 163(3):759–771. https://doi.org/10.1016/j.cell.2015.09.038
Zhu W-Y, Theodorou MK, Longland AC, Nielsen BB, Dijkstra J, Trinci APJ (1996) Growth and survival of anaerobic fungi in batch and continuous-flow cultures: ecology. Anaerobe 2(1):29–37. https://doi.org/10.1006/anae.1996.0004
Zhu W-Y, Theodorou MK, Nielsen BB, Trinci APJ (1997) Dilution rate increases production of plant cell-wall degrading enzymes by anaerobic fungi in continuous-flow culture. Anaerobe 3(1):49–59. https://doi.org/10.1006/anae.1997.0070
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2020 Springer Nature Switzerland AG
About this chapter
Cite this chapter
Flad, V. et al. (2020). The Biotechnological Potential of Anaerobic Gut Fungi. In: Benz, J.P., Schipper, K. (eds) Genetics and Biotechnology. The Mycota, vol 2. Springer, Cham. https://doi.org/10.1007/978-3-030-49924-2_17
Download citation
DOI: https://doi.org/10.1007/978-3-030-49924-2_17
Published:
Publisher Name: Springer, Cham
Print ISBN: 978-3-030-49923-5
Online ISBN: 978-3-030-49924-2
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)