Beilei Wu*, Mei Li, Chenchen Liu and Xiliang Jiang*
Institute of Plant Protection, Chinese Academy of Agricultural Sciences, China
Submission: April 01, 2020; Published: April 09, 2020
*Corresponding author: Beilei Wu, Xiliang Jiang, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, No.2 West Yuanmingyuan Rd., Haidian District, Beijing, 100193, PR China
How to cite this article: Beilei Wu, Mei Li, Chenchen Liu, Xiliang Jiang. The Insight of Mycovirus from Trichoderma spp. Agri Res & Tech: Open Access
J. 2020; 24(2): 556258. DOI:10.19080/ARTOAJ.2020.24.556258
Trichoderma spp. are used extensively in agriculture as a biological control agent to prevent soil-borne plant diseases. In recent years, mycoviruses from fungi have attracted increasing attention due to their effects on their hosts, but Trichoderma mycoviruses is in the beginning stage as the subject of extensive study. At present, eight researches were on the mycoviruses from Trichoderma spp. techniques of genome sequencing, elimination of dsRNA, detection of dsRNA, transmission of mycovirus were elaborated. With the deep research on the mycovirus, more and more effective methods for these basic researches should be applied. The topics about antagonism and biocontrol function of mycovirus will better push the deep exploration on the interaction among Trichoderma-mycovirus-plant (or pathogen), which also will have the driving role on seeking and screening more resources of Trichoderm spp. possessing biocontrol capabilities with mycoviruses.
Viruses are popular small organism, which are distributing in human, animals, plants and insects to microorganisms, including bacteria, archaea and fungi and induced the obvious disease or symbiosis with the hosts without any symptoms [1-4]. Mycovirus is a group of viruses that can infect and replicate in filamentous fungi, yeasts and oomycetes and is widespread [3,4] which was first described by Hollings et al. (1962), who found three kinds of spherical or short rod-shaped viruses related to diseases in cultivated mushrooms . Lampson et al. (1967) discovered a mycovirus in Penicillium funiculosum (Eurotiales, Ascomycota) and showed that it can induce an interferon-mediated response in the host [1,2,5-7]. In the same year, Ellis et al. (1967) observed the presence of virus particles in the culture fluid of P. stoloniferum by electron microscopy . Based on previous studies, Banks et al. (1968) found that both P. stoloniferum virus (PsV) and P. funiculosum virus (PfV) had a dsRNA genome . Since then, various types of mycoviruses have been reported. Until now, the classification of mycoviruses was based on the mode of replication and the type of the genome, which was divided all currently known mycoviruses into 16 families and an unclassified group by the International Committee on Taxonomy of Viruses . The 16 families are consisted with seven dsRNA virus families, five positive-sense ssRNA virus families, two reverse-transcribing
virus families (+ssRNA), one negative-sense ssRNA virus family, and one positive-sense ssDNA virus family . The taxonomic status of roughly 20% of fungal viruses still need to be determined in the future [11,12].
The transmission and function of mycovirus: The transmission of mycoviruses have two ways, vertical and horizontal transmission. Vertical transmission is through spores of the fungi, including both sexual and asexual spores. In case of the mycelial asexual spores, the virus can be transmitted through the cytoplasm. This mode of transmission is relatively easy and especially common for dsRNA virus . Horizontal transmission is accomplished by the fusion between hyphae, but this mode of transmission is limited by the incompatibility between the vegetative forms . In some cases, few mycoviruses from fungi and fungi-like protozoans could not be virulent for the hosts with effecting the host fitness, including improving mycelia growth or reducing growth, abnormal pigmentation or deficient sporulation [4,6], and most mycovirus infections are asymptomatic . But some mycoviruses had virulence, there were two main affections to plant pathogenic fungi: first, they can cause the host to become a low-virulence strain; second, the metabolites induced by the mycovirus can increase the pathogenicity of the host [16-18]. The most successful mycovirus biocontrol agent to date has been The transmission and function of mycovirus: The transmission of mycoviruses have two ways, vertical and horizontal transmission. Vertical transmission is through spores of the fungi, including both sexual and asexual spores. In case of the mycelial asexual spores, the virus can be transmitted through the cytoplasm. This mode of transmission is relatively easy and especially common for dsRNA virus . Horizontal transmission is accomplished by the fusion between hyphae, but this mode of transmission is limited by the incompatibility between the vegetative forms . In some cases, few mycoviruses from fungi and fungi-like protozoans could not be virulent for the hosts with effecting the host fitness, including improving mycelia growth or reducing growth, abnormal pigmentation or deficient sporulation [4,6], and most mycovirus infections are asymptomatic . But some mycoviruses had virulence, there were two main affections to plant pathogenic fungi: first, they can cause the host to become a low-virulence strain; second, the metabolites induced by the mycovirus can increase the pathogenicity of the host [16-18]. The most successful mycovirus biocontrol agent to date has been
The researches of mycovirus from Trichoderma: Despite
Trichoderma spp. is researched popularly in the world for
the function of biocontrol agent, and for producing important
industrial enzymes [20-23], mycoviruses from Trichoderma spp.
have been poorly studied and characterized. So far, there are
eight descriptions of researches about Trichoderma mycoviruses
The genome sequences of Trichoderma mycovirus: The
first report for Trichoderma mycovirus was from the research of
Jom-in in 2009 , however the signs of mycoviruses existing in
Trichoderma spp. were only explored by checking the dsRNA by
extraction methods, the genome sequences was not get anymore.
Until 2016, Yun et al. still used the dsRNA extraction method to
get 32 strains with dsRNA- mycoviruses from 315 strains of
Trichoderma spp. from Lentinula edodes in Korea . According
to the diversification of number and size of dsRNAs among isolates,
the band patterns of the dsRNA were categorized into 15 groups.
The genome sequence was also not get yet.
The first whole genome sequences of the Trichoderma
mycovirus was obtained from Lee’s research in 2017 . The
complete genome is consisted by 8566bp, which contains two
open reading frames (ORF), encoding structural proteins and RNA
dependent RNA polymerases (RdRP), respectively. Phylogenetic
analysis classified it belong to the family Fusagraviridae and
named Trichoderma atroviride mycovirus 1 (TaMV1) . In this
research, the detection method for dsRNA was the electrophoresis,
and then subjected to reverse transcription and cDNA library
synthesis by using random hexanucleotide primers and reverse
transcriptase, then RACE Analysis was used for 5’- and 3’-terminal
sequences. This method was also used in the later following
researches for sequences.
From then on, the five genome sequences of Trichoderma
mycovirus were come out one after another. In 2018, Chun
et al. obtained two genome sequences from mycovirus of
Trichoderma, Trichoderma atroviride partitivirus 1 (TaPV1) 
and Trichoderma harzianum partitivirus 1(ThPV1) . TaPV1
was from T. atroviride and had two segments. The bigger one
(dsRNA1) is consisted with 2023bp with one open reading frame
(ORF) encoding RdRP. The smaller one (dsRNA2) has a total length
of 2012bp with a single ORF encoding CP. Phylogenetic analysis
indicated that the virus was a new member of Alphapartitivirus in
the Partitividae family . Moreover, the electron micrographs
of purified viral particles of TaPV1 was shown as an isometric
structure approximately of 30 nm in diameter. It was the first
successful extraction of mycovirus particles from Trichoderma.
ThPV1 was from T. harzianum, which is consisted of two dsRNA
with similar sequence size. The larger dsRNA1 is 2289 bp with
a single open reading frame encoding RdRP. The smaller dsRNA2
with 2245 bp contains an ORF encoding capsid protein (CP).
Phylogenetic analysis indicated the virus was a new type of fungal
virus which was not specifically classified into species in the genus
Betartitivirus, family Partitividae, named Trichoderma harzianum
partitivirus 1(ThPV1) . All of these two mycoviruses possessed
two segment, belonging to family Partitividae.
At the same time, a new fungal virus isolated from T. asperellum
was reported in the laboratory of Guizhou Medical University,
China, which was named Trichoderma asperellum dsRNA virus
1 (TaRV1) with two ORF on its genomic plus strand. ORF1 is a
hypothetical protein, ORF2 encodes an RdRP. Based on RdRP
sequence, phylogenetic analysis TaRV1 belongs to unclassified
virus , which was the first report about Trichoderma
In 2019, Liu et al isolated two unclassified dsRNA mycovirus
isolates harzianum mycovirus 1 (ThMBV1)  and harzianum
mycovirus 1 (ThMV1)  from the 155 Trichoderma spp. strains,
which were collected in the soil from Xinjiang and Inner Mongolia,
China in 2019 [3,4]. The metagenetics as new method was used
for checking the sign of mycovirus in strains, then electrophoresis,
RT-PCR, 5’ RACE and 3’ RACE were used for whole genome
sequence [3,4]. ThMBV1 was a new type of virus with bipartite
segments mycovirus. one was 2088 bp encoding the RNAdependent
RNA polymerase (RdRP), and another segment was
1634 bp encoding a hypothetical protein. phylogenetic analysis
suggested it was identified as unclassified mycovirus, named
Trichoderma harzianum bipartite mycovirus 1. The phylogenetic
analysis indicated it belonged to an unclassified family of dsRNA
mycoviruses . ThMV1 had two ORFs on the negative strand,
ORF-A (residues 1857-109) encoded RdRP, ORF-B encoded a
putative protein. On the positive strand, there was an ORF C
(residues 1076-1370), presumed to be a hypothetical protein
containing 94 amino acids, with the poly(A) structure on the 3’
terminal. This was the first report about the RdRP and CP encoding
on the negative strand of mycovirus genome sequence.
For the elimination of dsRNA is very important step for
exploring the function of the Trichoderma strains with and
without dsRNA. The basic method should be single-spore isolation
followed by hyphal tipping, the auxiliary measures were always
variated. Some used heat therapy, some used cycloheximide or
ribavirin, and some also have been helped by the protoplasting/
regeneration. In the research of Jom-in , the method of
elimination of dsRNA was heat therapy, though not successful.
The details were alternately heating at 37°C for 3 hours and
room temperature (28-30°C) interval for 24 hours, and then like
this for 10 times, but he did not use single-spore isolation. In the
research of Yun et al , the elimination of dsRNA from strains
is different with Jom-in, the auxiliary measures were depended
on the cycloheximide (5 and 10μg/ml) or ribavirin (10 or 20mg/
ml), which were used to eliminate the dsRNA incubated at 25°C,
transferred agar plugs from the margin of each colony to the fresh
V8 agar plates (100×15mm)  with the same concentration ORF either cycloheximide or ribavirin for 3-4 times, and then grown
for 3 or 4 days in each time, and then hyphal tipped and grown
in 50 ml of potato dextrose broth (PDB) (Difco Laboratories,
Detroit) at 25°C for 2 weeks. The mycelium was used for further
analysis of dsRNA presence . Until 2018, the method of singlespore
isolation followed by hyphal tipping from protoplast was
successfully single used to eliminate dsRNA by Chun et al [27,28].
In our research, the ribavirin, protoplasting/regeneration and
single-spore isolation followed by hyphal tipping were together
used to successfully eliminate ThMV1 from Trichoderma strain
525 , but for ThMBV1, this method was not successful .
Moreover, RT-PCR and northern blotting should be a good method
for detecting dsRNA, in the researches of Chun, Jem-in and Liu, it
was good use of checking existing of dsRNA  and elimination
of dsRNA [3,4,27,28].
The researches on the Transmission of Trichoderma mycovirus
were limited. Normally, most vertical transmission of dsRNA into
asexual spores of ascomycetes occurs at a markedly higher rate,
for instance, the TaMV1 has a very high transmission rate 33/38
, but ThPV1 had low transmission rate into conidia with
exceptional considering, the low transmission rate of ThPV1 into
conidia is attributable to the intrinsic characteristics of the virus–
fungus (ThPV1–T. harzianum) interaction . We also found the
transmission between same species also had the difficulties for
ThMV1 (not published).
The antagonism and biocontrol researches were involved invitro
and in-vivo researches, some differences were found between
with and without strains, and some not. Jom-in comparing with
the free isolates without the dsRNA, the function of the isolates
of TM10 and TM20 with dsRNA reduced the host growth rate,
sporulation and biological control efficacy . But in the research
of TaPV1, no apparent difference in colony morphology was
observed between TaPV1-containing and the three virus-cured
strains, moreover, β-1,3-glucanase and chitinase, as the two
representative antifungal enzymes, no obvious alterations of
enzymatic activities were observed between the infected and viruscured
isogenic strains [2018a], Moreover, the enzymatic activities
of β-1,3-glucanase and chitinase were no changes in viruscured
strains, which was the first report of an Alphapartitivirus
in T. atroviride . In our research, antagonism characteristics
of ThMV1 was explored though in vitro and in vivo. In vitro
experiment, There were no obvious differences, when we tested
the antagonism of T525 with ThMV1and T525-F without ThMV1
to three pathonogenic fungi (F. oxysporum f.sp.cucumebrium
Owen, B. cinerea and F. oxysporum f. sp. vasinfectum). but in vivo,
the removal of ThMV1 from host strain 525 reduced host biomass
production and improved the biocontrol capability of the host on
Fusarium oxysporum f. sp. Cucumerinum. Moreover, the presence
of ThMV1 functioned to improve the growth of the cucumber .
It was the first research on detecting the changes of biocontrol in
vivo of the Trichoderma trigged by mycovirus currency.
At present, the researches of Trichoderma mycoviruses are
limited, the more molecular technique for extraction of dsRNA, the
genome sequences, eliminating methods, Transmission methods,
the antagonism and biocontrol aceessment of Trichoderma
mycovirus will be improved and abundant. In the next, the
research goals of Trichoderma mycoviruses should be focus on
a. For the resources of Trichoderma mycovirus should be
rich in the nature, there should be more dsRNA mycovirus need to
be explored in the future;
b. It is the total tendency to find more strains of Trichoderma
spp. with mycoviruses with biocontrol function, which was
hoped to the find more mycoviruses mediating the capabilities of
Trichoderma to control plant disease, promoting plant growth and
going through the environment stress;
c. The interval mechanism among Trichodermamycoviruses-
plant-pathogen need to be discovered, for the
complicated factors, the true interaction between or among the
factors will be discovered with the progress of molecular biological
informatics, transcriptome, proteomics and metabonomics;
d. Until now the resources of Trichoderma mycoviruses
are limited, with the richness of them, the origin and phylogenetic
research of the Trichoderma mycoviruses need to be more focuses
for the growing enormous taxonomy in the future.
All in all, according to an applied agronomical perspective,
discovering more mycoviruses infecting Trichoderma spp.
populations and characterizing the nature of their host-parasite
interactions would be of special interest for identifying new
agents with potential biotechnological interest and also to
better understand the ecology and temporal dynamics of fungal
communities in natural and agronomic ecosystems.
The information presented in this article is derived from projects
funded by National key research and development plan (Chemical
fertilizer and pesticide reducing efficiency synergistic technology
research and development): Research and demonstration of a new
high efficiency biocide (2017YFD0201100-2017YFD0201102)
granted to Xiliang Jiang. Natural Science Foundation of Beijing,
China: Exploration of mycovirus of Trichoderma and their effects
on the host biology (No. 6192022) granted to Beilei Wu; Survey of
basic resources of science and technology: A comprehensive survey
of biodiversity in the Mongolian Plateau (2019FY102000) granted
to Beilei Wu; Demonstration of comprehensive prevention and
control technology of non-point source pollution in main vegetable
producing areas of Huang Huai Hai (SQ2018YFD080026) granted
to Beilei Wu.
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