The study evaluates the survivability and storage stability of seven Trichoderma strains belonging to the species: T. harzianum (1), T. atroviride (4), and T. virens (2) after the lyophilization of their solid state cultures on wheat straw. Biomass of Trichoderma strains was freeze-dried with and without the addition of maltodextrin. Furthermore, in order to determine the ability of tested Trichoderma strains to preserve selected technological features, the biosynthesis of extracellular hydrolases (cellulases, xylanases, and polygalacturonases) after a 3-month storage of lyophilizates was investigated. Strains of T. atroviride (except TRS40) and T. harzianum TRS85 showed the highest viability after lyophilization process (up to 100%). After 3 months of storage, T. atroviride TRS14 exhibited the highest stability (95.23%); however, the number of active conidia remained at high level of 106-107 cfu/g for all tested T. atroviride strains and T. harzianum TRS85. Interestingly, after a 3-month storage of lyophilized formulations, most of the tested Trichoderma strains exhibited higher cellulolytic and xylanolytic activities compared to the control, i.e., before freeze-drying process. The highest activities of these enzymes exhibited the following: T. atroviride TRS14-2.37 U/g and T. atroviride TRS25-21.47 U/g, respectively, whereas pectinolytic activity was weak for all tested strains, with the highest value of 0.64 U/g registered for T. virens TRS109.

Department of Biotechnology and Food Microbiology Wrocław University of Environmental and Life Sciences ul Chełmońskiego 37 51 630 Wrocław Poland

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Bai Z, Jin B, Li Y, Chen J, Li Z. Utilization of winery wastes for Trichoderma viride biocontrol agent production by solid state fermentation. J Environ Sci. 2008;20(3):353–358. doi: 10.1016/S1001-0742(08)60055-8. PubMed DOI

Benítez T, Rincón AM, Limón MC, Codón AC. Biocontrol mechanisms of Trichoderma strains. Int Microbiol. 2004;7(4):249–260. PubMed

Bhat KA, Anwar A, Lone GM, Hussain K, Nazir G. Shelf life of liquid fermented product of Trichoderma harzianum in talc. J Mycol Pl Pathol. 2009;39(2):263–265.

Daryaei A, Jones EE, Glare TR, Falloon RE. Biological fitness of Trichoderma atroviride during long-term storage, after production in different culture conditions. Biocontrol Sci Techn. 2016;26(1):86–103. doi: 10.1080/09583157.2015.1077929. DOI

Day J, Stacey G. Cryopreservation and freeze-drying protocols. Totowa: Humana Press Inc.; 2007.

Druzhinina IS, Komoń-Zelazowska M, Atanasova L, Seidl V, Kubicek CP. Evolution and ecophysiology of the industrial producer Hypocrea jecorina (anamorph Trichoderma reesei) and a new sympatric agamospecies related to it. PLoS One. 2010;5(2):e9191. doi: 10.1371/journal.pone.0009191. PubMed DOI PMC

Fernández-Sandoval MT, Ortiz-Garcia M, Galindo E, Serrano-Correon L. Cellular damage during drying and storage of Trichoderma harzianum spores. Process Biochem. 2012;47(2):186–194. doi: 10.1016/j.procbio.2011.10.006. DOI

Gade RM, Wardhe SR, Armarker SV. Shelf life study of Trichoderma spp. in different carrier materials. Journal of Maharashtra Agricultural Universities. 2009;34(2):181–182.

Ganga A, Gonzalez-Candelas L, Ramon D, Perez-Gonzalez JA. Glucose-tolerant expression of Trichoderma longibrachiatum endoglucanase I, an enzyme suitable for use in wine production. J Agric Food Chem. 1997;45(6):2359–2362. doi: 10.1021/jf960904x. DOI

Grzegorczyk M, Szalewicz A, Żarowska B, Połomska X, Wątorek W, Wojtatowicz M. Microorganisms in biological control of phytopathogenic fungi. Acta Sci Pol Biotechnol. 2015;14(2):19–42.

Gusakov AV. Alternatives to Trichoderma reesei in biofuel production. Trends Biotechnol. 2011;29(9):419–425. doi: 10.1016/j.tibtech.2011.04.004. PubMed DOI

Juhasz T, Szengyel Z, Reczey K, Siika-Aho M, Viikari L. Characterization of cellulases and hemicellulases produced by Trichoderma reesei on various carbon sources. Process Biochem. 2004;40:3519–3525. doi: 10.1016/j.procbio.2005.03.057. DOI

Kancelista A, Witkowska D. Biosynthesis of some lytic enzymes in medium containing waste corn cobs by filamentous fungi from Trichoderma genus. Acta Sci Pol Biotechnol. 2008;7(1):17–25.

Kancelista A, Tril U, Stempniewicz R, Piegza M, Szczech M, Witkowska D. Application of lignocellulosic waste materials for the production and stabilization of Trichoderma biomass. Pol J Environ Stud. 2013;4:1083–1090.

Kovacs K, Macrelli S, Szakacs G, Zacchi G. Enzymatic hydrolysis of steam-pretreated linocellulosic materials with Trichoderma atroviride enzymes produced in-house. Biotechnol Biofuels. 2009;2(14):1–11. PubMed PMC

Kuhad RC, Gupta R, Singh A. Microbial cellulases and their industrial applications. Enzyme Res. 2011;2011:1–10. doi: 10.4061/2011/280696. PubMed DOI PMC

Kumar S, Thakur M, Rani A. Trichoderma: mass production, formulation, quality control, delivery and its scope in commercialization in India for the management of plant diseases. Afr J Agric Res. 2014;9(53):3838–3852.

Longa CMO, Pertot I, Tosi S. Ecophysiological requirements and survival of Trichoderma atroviride isolate with biocontrol potential. J Basic Microbiol. 2008;48(4):269–277. doi: 10.1002/jobm.200700396. PubMed DOI

Magan N. Physiological approaches to improving the ecological fitness of fungal biocontrol agents, fungi as biocontrol agents. Bristol: CABI Book; 2001.

Masutti DC, Borgognone A, Setti L (2012) Production of enzymes from rice husks and wheat straw in solid state fermentation. 3rd International Conference on Industrial Biotechnology (IBIC), Vol: 27. doi: 10.3303/CET1227023

Miller GL. Use dinitrosalicylic acid reagent for determination of reducing sugars. Anal Chem. 1959;31(3):426–428. doi: 10.1021/ac60147a030. DOI

Mitchell DA, Berovic M, Krieger N. Overview of solid state bioprocessing. Biotechnol Annu Rev. 2002;8:183–225. doi: 10.1016/S1387-2656(02)08009-2. PubMed DOI

Mohamed S, Christensen T, Mikkelsen J. New polygalacturonases from Trichoderma reesei: characterization and their specificities to partially methylated and acetylated pectins. Carbohydr Res. 2003;338(6):515–524. doi: 10.1016/S0008-6215(02)00398-1. PubMed DOI

Mohamed SA, Al-Malki AL, Khan JA, Kabli SA, Al-Garni SM. Solid state production of polygalacturonase and xylanase by Trichoderma species using cantaloupe and watermelon rinds. J Microbiol. 2013;51(5):605–611. doi: 10.1007/s12275-013-3016-x. PubMed DOI

Olsson L, Christensen T, Hansen K, Palmqvisit E. Influence of the carbon source on production of cellulases, hemicellulases and pectinases by Trichoderma reesei Rut C-30. Enzyme Microb Tech. 2003;33(5):612–619. doi: 10.1016/S0141-0229(03)00181-9. DOI

Panahian R, Rahnama K, Jafari M. Mass production of Trichoderma ssp. and application. Intl Res J Appl Basic Sci. 2012;3(2):292–298.

Pandey A. Aspects of fermenter design for solid-state fermentations. Process Biochem. 1991;26(6):355–361. doi: 10.1016/0032-9592(91)85026-K. DOI

Pandya JR, Sabalpara AN, Chawda SK (2011) Trichoderma: a particular weapon for biological control of pathogens. J Agric Technol 7(5):1187–1191

Phitsuwan P, Laohakunjit N, Kerdchoechuen O, Kyu KL, Ratanakhanokchai K. Present and potential applications of cellulases in agriculture, biotechnology, and bioenergy. Folia Microbiol. 2013;58(2):163–176. doi: 10.1007/s12223-012-0184-8. PubMed DOI

Piegza M, Stolaś J, Kancelista A, Witkowska D. Influence of Trichoderma strains on the growth of pathogenic moulds in biotic test on untypical carbon sources. Acta Sci Pol Biotechnol. 2009;8(1):4–14.

Sanghvi GV, Koyani RD, Rajput KS. Thermostable xylanase production and partial purification by solid-state fermentation using agricultural waste wheat straw. Mycology. 2010;1(2):106–112. doi: 10.1080/21501203.2010.484029. DOI

Sargin S, Gezgin Y, Eltem R, Vardar F. Micropropagule production from Trichoderma harzianum EGE-K38 using solid-state fermentation and a comparative study for drying methods. Turk J Biotechnol. 2013;37:139–146.

Shuster A, Shmoll M. Biology and biotechnology of Trichoderma. Appl Microbiol Biot. 2010;87(3):787–799. doi: 10.1007/s00253-010-2632-1. PubMed DOI PMC

Singh A, Srivastava S, Singh HB. Effect of substrates on growth and shelf life of Trichoderma harzianum and its use in biocontrol of diseases. Bioresour Technol. 2007;98(2):470–473. doi: 10.1016/j.biortech.2006.01.002. PubMed DOI

Skoneczny D, Oskiera M, Szczech M, Bartoszewski G. Genetic diversity of Trichoderma atroviride strains collected in Poland and identification of loci useful in detection of within species diversity. Folia Microbiol. 2015;60(4):297–307. doi: 10.1007/s12223-015-0385-z. PubMed DOI PMC

Smolińska U, Kowalska B, Kowalczyk W, Szczech M. The use of agro-industrial wastes as carriers of Trichoderma fungi in the parsley cultivation. Sci Hortic. 2014;179:1–8. doi: 10.1016/j.scienta.2014.08.023. DOI

Soliman HM, Abdel-Dayem A, El-Tanash AB, El-Tanash SA. Production of xylanase by Aspergillus niger and Trichoderma viride using some agricultural residues. Int J Agric Res. 2012;7(1):46–57. doi: 10.3923/ijar.2012.46.57. DOI

Sukumaran RK, Singhania RR, Pandey A (2005) Microbial cellulases - Production, applications and challenges. J Sci Ind Res 64:832–844

Sukumaran RK, Singhania RR, Mathew GM, Pandey A. Cellulase production using biomass feed stock and its application in lignocellulose saccharification for bio-ethanol production. Renew Energy. 2009;34(2):421–424. doi: 10.1016/j.renene.2008.05.008. DOI

Szczech M, Staniaszek M, Habdas H, Uliński Z, Szymański J. Trichoderma spp.—the cause of green mold on Polish mushroom farms. Vegetable Crops Research Bulletin. 2008;69:105–114. doi: 10.2478/v10032-008-0025-0. DOI

Szczech M, Witkowska D, Kancelista A, Piegza M, Gajewska E, Małolepsza U (2011) Method for selection of active fungal isolates of the genus Trichoderma. Patent Application No. PL397659. Dec 30, 2011

Tan CS, van Ingen CW, Stalpers JA. Freeze-drying fungi using a shelf freeze-drier. Methods Mol Biol. 2007;368:119–125. doi: 10.1007/978-1-59745-362-2_8. PubMed DOI

Tewari L, Bhanu C. Evaluation of agro-industrial wastes for conidia based inoculums production of biocontrol agent: Trichoderma harzianum. J Sci Ind Res. 2004;63:807–812.

Texido N, Canamas TP, Usall J, Torres R, Magan N, Vinas I. Accumulation of the compatible solutes, glycine-betaine and ectoine, in osmotic stress adaptation and heat shock cross-protection in the biocontrol agent Pantoea agglomerans CPA-2. Lett App Microbiol. 2005;41(3):248–252. doi: 10.1111/j.1472-765X.2005.01757.x. PubMed DOI

Tokarzewska-Zadora J, Rogalski J, Szczodrak J. Xylan-degrading enzymes—characterization and application in biotechnology. Biotechnologia. 2005;2(69):163–182.

Valmaseda M, Martínez MJ, Martínez AT. Kinetics of wheat straw solid-state fermentation with Trametes versicolor and Pleurotus ostreatus—lignin and polysaccharide alteration and production of related enzymatic activities. Appl Microbiol Biotechnol. 1991;35:817. doi: 10.1007/BF00169902. DOI

Witkowska D, Bień M. Activity in biosynthesis of extracellular hydrolases of Trichoderma viride mutants obtained in two-stage mutation. Acta Alient Polon. 1991;XVII/XLI(2):127–135.

Witkowska D, Maj A. Production of lytic enzymes by Trichoderma spp. and their effect on the growth of phytopathogenic fungi. Folia Microbiol. 2002;47(3):279–282. doi: 10.1007/BF02817652. PubMed DOI

Witkowska D, Wróblewska A, Jurgielewicz W. Degradation of cellulose and lignocellulose by Trichoderma reesei M7-1 hydrolases. Pol J Food Nutr Sci. 1997;6/47(2):57–62.

Witkowska D, Kancelista A, Wilczak A, Stempniewicz R, Pasławska M, Piegza M, Łaba W, Szczech M. Survivability and storage stability of Trichoderma atroviride TRS40 preserved by fluidised bed drying on various agriculture by-products. Biocontrol Sci Tech. 2016;26(12):1591–1604. doi: 10.1080/09583157.2016.1201457. DOI

Zuoxing Z, Kalidas S. Solid state production of polygalacturonase by Lentinusedodes using fruit processing wastes. Process Biochem. 2000;35:825–830. doi: 10.1016/S0032-9592(99)00143-0. DOI