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.
- MeSH
- Biomass MeSH
- Time Factors MeSH
- Fermentation MeSH
- Hydrolases metabolism MeSH
- Freeze Drying * MeSH
- Microbial Viability * MeSH
- Triticum metabolism MeSH
- Drug Storage MeSH
- Spores, Fungal growth & development MeSH
- Trichoderma classification growth & development physiology MeSH
- Publication type
- Journal Article MeSH
Molecular markers that enable monitoring of fungi in their natural environment or assist in the identification of specific strains would facilitate Trichoderma utilization, particularly as an agricultural biocontrol agent (BCA). In this study, sequence analysis of internal transcribed spacer regions 1 and 2 (ITS1 and ITS2) of the ribosomal RNA (rRNA) gene cluster, a fragment of the translation elongation factor 1-alpha (tef1) gene, and random amplified polymorphic DNA (RAPD) markers were applied to determine the genetic diversity of Trichoderma atroviride strains collected in Poland, and also in order to identify loci and PCR-based molecular markers useful in genetic variation assessment of that fungus. Although tef1 and RAPD analysis showed limited genetic diversity among T. atroviride strains collected in Poland, it was possible to distinguish major groups that clustered most of the analyzed strains. Polymorphic RAPD amplicons were cloned and sequenced, yielding sequences representing 13 T. atroviride loci. Based on these sequences, a set of PCR-based markers specific to T. atroviride was developed and examined. Three cleaved amplified polymorphic sequence (CAPS) markers could assist in distinguishing T. atroviride strains. The genomic regions identified may be useful for further exploration and development of more precise markers suitable for T. atroviride identification and monitoring, especially in environmental samples.
- MeSH
- DNA, Fungal chemistry genetics MeSH
- Peptide Elongation Factor 1 genetics MeSH
- Phylogeny MeSH
- Genetic Variation * MeSH
- Genetic Loci * MeSH
- Genetic Markers MeSH
- DNA, Ribosomal Spacer chemistry genetics MeSH
- Molecular Sequence Data MeSH
- Molecular Typing MeSH
- Mycological Typing Techniques MeSH
- Sequence Analysis, DNA MeSH
- Cluster Analysis MeSH
- Random Amplified Polymorphic DNA Technique MeSH
- Trichoderma classification genetics isolation & purification MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Poland MeSH
The present study includes the molecular characteristics of Trichoderma pleurotum and Trichoderma pleuroticola isolates collected from green moulded cereal straw substrates at 47 oyster mushroom farms in Poland. The screening of the 80 Trichoderma isolates was performed by morphological observation and by using the multiplex PCR assay. This approach enabled specific detection of 47 strains of T. pleurotum and 2 strains of T. pleuroticola. Initial identifications were confirmed by sequencing the fragment of internal transcribed spacer regions 1 and 2 (ITS1 and ITS2) of the rRNA gene cluster and the fragment including the fourth and fifth introns and the last long exon of the translation-elongation factor 1-alpha (tef1) gene. ITS and tef1 sequence information was also used to establish the intra- and interspecies relationship of T. pleurotum and T. pleuroticola originating from the oyster mushroom farms in Poland and from other countries. Comparative analysis of the ITS sequences showed that all T. pleurotum isolates from Poland represent one haplotype, identical to that of T. pleurotum strains from Hungary and Romania. Sequence analysis of the tef1 locus revealed two haplotypes ("T" and "N") of Polish T. pleurotum isolates. The "T" type isolates of T. pleurotum were identical to those of strains from Hungary and Romania. The "N" type isolates possessed a unique tef1 allele. Detailed analysis of the ITS and tef1 sequences of two T. pleuroticola isolates showed their identicalness to Italian strain C.P.K. 1540.
- MeSH
- DNA, Fungal chemistry genetics MeSH
- Peptide Elongation Factor 1 genetics MeSH
- Genetic Variation * MeSH
- Haplotypes MeSH
- DNA, Ribosomal Spacer chemistry genetics MeSH
- Microscopy MeSH
- Molecular Sequence Data MeSH
- Multiplex Polymerase Chain Reaction MeSH
- Pleurotus * MeSH
- Base Sequence MeSH
- Sequence Analysis, DNA MeSH
- Trichoderma classification cytology genetics isolation & purification MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Poland MeSH
Twenty-eight isolates of Trichoderma belonging to four different species were screened in vitro for their antagonistic ability against Fusarium oxysporum f.sp. dianthi causing carnation wilt. Three different levels of antagonism observed in dual plate assay were further confirmed by cell-free culture filtrate experiments. Isolates showing class I level of antagonism produced maximum lytic enzymes, chitinases and beta-1,3-glucanases. Genetic variability of 25 selected isolates was assessed by random amplified polymorphic DNA technique and the amplified products were correlated for their level of antagonism. Unweighed pair-group method with arithmetical averages cluster analysis revealed prominent inter-and intraspecific genetic variation among the isolates. Based on their genetic relationship, the isolates were mainly distributed into 3 major groups representing T. atroviride, T. pseudokoningii and T. harzianum, with 20-35% interspecific dissimilarity. However, the polymorphism shown by the isolates did not correlate to their level of antagonism.
- MeSH
- Antibiosis MeSH
- Chitinases genetics metabolism MeSH
- Fungal Proteins genetics metabolism MeSH
- Fusarium physiology MeSH
- Phylogeny MeSH
- Genetic Variation MeSH
- Plant Diseases microbiology MeSH
- Polymorphism, Restriction Fragment Length MeSH
- Soil Microbiology MeSH
- Trichoderma physiology genetics isolation & purification classification MeSH
