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juniperpublishers-agriculture · 5 years ago

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The Insight of Mycovirus from Trichoderma spp.-Juniper Publishers

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.

Keywords: Trichoderma spp., Mycovirus, dsRNA, Application

Introduction

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 [5]. 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 [8]. Based on previous studies, Banks et al. (1968) found that both P. stoloniferum virus (PsV) and P. funiculosum virus (PfV) had a dsRNA genome [9]. 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 [10]. 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 [10]. 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 [13]. Horizontal transmission is accomplished by the fusion between hyphae, but this mode of transmission is limited by the incompatibility between the vegetative forms [14]. 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 [15]. 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 [13]. Horizontal transmission is accomplished by the fusion between hyphae, but this mode of transmission is limited by the incompatibility between the vegetative forms [14]. 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 [15]. 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 [3,4,24-29].

The genome sequences of Trichoderma mycovirus: The first report for Trichoderma mycovirus was from the research of Jom-in in 2009 [24], 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 [25]. 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 [26]. 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) [26]. 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) [23] and Trichoderma harzianum partitivirus 1(ThPV1) [28]. 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 [27]. 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) [28]. 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 [29], which was the first report about Trichoderma mycovirus from

In 2019, Liu et al isolated two unclassified dsRNA mycovirus isolates harzianum mycovirus 1 (ThMBV1) [3] and harzianum mycovirus 1 (ThMV1) [4] 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 [3]. 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.

The Methods of Eliminating dsRNA

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 [24], 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 [25], 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) [30] 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 [30]. 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 [4], but for ThMBV1, this method was not successful [3]. 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 [20] and elimination of dsRNA [3,4,27,28].

Transmission of Trichoderma Mycovirus

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 [26], 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 [27]. We also found the transmission between same species also had the difficulties for ThMV1 (not published).

The Antagonism and Biocontrol Researches of Trichoderma Mycovirus

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 [24]. But in the research of TaPV1[27], 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 [27]. 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 [4]. It was the first research on detecting the changes of biocontrol in vivo of the Trichoderma trigged by mycovirus currency.

Prospect of the Study on Trichoderma Mycoviruses

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 these aspects:

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.

Acknowledgment

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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karanresearchlayer · 2 years ago

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Biofungicides Market :Advanced Technologies & Growth Opportunities Worldwide By 2024

Biofungicides are biological agents that are used to control fungal diseases in plants. They are made from naturally occurring microorganisms such as bacteria, fungi, and viruses that have been isolated from the environment and then formulated into products that can be applied to crops.

Biofungicides work by either inhibiting the growth of fungi or by causing the fungi to become inactive. They are generally considered safer for the environment and human health than chemical fungicides, as they are non-toxic and biodegradable.

Some common types of biofungicides include:

Bacillus subtilis: A bacterium that produces enzymes that can break down the cell walls of fungi.

Trichoderma harzianum: A fungus that can parasitize other fungi and suppress their growth.

Streptomyces lydicus: A bacterium that produces antifungal compounds.

Coniothyrium minitans: A fungus that parasitizes other fungi and can be used to control diseases such as Sclerotinia.

Biofungicides can be used in both organic and conventional agriculture, and are often used in combination with other management practices such as crop rotation and sanitation. However, they may not provide as rapid or complete control as chemical fungicides, and may require more frequent applications.

Overall, biofungicides are a promising tool for sustainable disease management in agriculture, as they offer a natural alternative to chemical fungicides that can reduce the impact of plant diseases on both the environment and human health.

#Biofungicides Market

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oaresearchpaper · 2 years ago

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Fighting Chestnut Blight: Antagonistic Microscopic Fungi

Abstract

The extent of cryphonecrosis among the chestnut populations of three Imeretian (west Georgia) villages: Darka, Eto, and Chala has been evaluated. 23 strains of Cryphonectria parasitica (Murrill) Barr (syn. Endothia parasitica (Murrill) were isolated and identified from the bark of sick trees. The collection of strains of the plant pathogen fungus has been created. The strategy of the struggle against the chestnut blight, based on the application of antagonistic to C. parasitica microscopic fungi, has been elaborated. For this purpose 50 strains of different microscopic fungi were isolated and identified (till genus) from the soil samples picked just under the stems of sick trees of above mentioned locations. The dominating genera of micromycetes in forest brown soils have been revealed. Strong biological antagonists of the plant pathogenic fungus, belonging to genera Penicillium, Trichoderma and Aspergillus have been selected on the base of the investigation of antagonistic activity of the “aboriginal” flora of studied soils. The collection of antagonistic to C. parasitica microscopic fungi, among them of new biological agents, has been created. The vegetative compatibility of the isolated strains of C. parasitica was investigated as well.

Introduction

Wood of excellent quality and high nutritional value fruits rise the chestnut plant - Castanea sativa Mill. in range of popular and economicaly significant trees in the world. (Heiniger and Rigling, 1994). Though, the plant is under the danger of extinction all over the world. The reason of it is so called “chestnut blight” disease caused by the plant pathogenic fungus from ascomycetes Cryphonectria parasitica (Murrill) Barr (syn. Endothia parasitica (Murrill)) (Anagnostakis, 1994; Rigling and Prospero, 2018).

It is more than half of a century the world scientists try efforts to fight the chestnut blight but in vain. Healing of sick trees with hypovirulent strains is regarded to be the most effective measure against C. parasitica today (Robin and Heiniger 2001; Puia et al., 2012). As it has established, hypovirulence is stipulated by the existence of Cryphonectria hypovirus (CHV) in the cytoplasm of C. parasitica. The genome of the virus is double chain RNA molecule (ds RNA). The virus lacks of capsid, which suppresses its active spreading in the environment. The virus is able “to enter” a new host organism only in case of formation of the hypae anastomoses between two stains of C. parasitica, or by means of the host’s asexual spores.

Though, wide scale integration of hypovirus under the natural conditions is not always possible because of the vegetative incompatibility of C. parasitica’s different stains (Anagnostakis, 1977). There exists another method of the biological inspection of chestnut blight: isolation of antagonistic to C. parasitica microorganisms from the natural sources and their application against the pathogen. According to literary data among the antagonistic microorganisms of C. parasitica are well known genera of microscopic fungi and bacteria (Trichoderma, Penicillium, Bacillus, Streptomyces) (Wilhelm et al., 1998; Groome et al., 2001; Akilli et al., 2011; Smith, 2013).

Combined application of antagonists and hypovirulent strains of C. parasitica appeared especially effective against the disease (Akilli et al., 2011). Those countries which are unable to control chestnut blight by hypovirulent strains, for the purpose to localize the pathogen, apply the easy way of in situ struggle against the pathogen – soil compressing or mud packing method (Anagnostakis, 1994; Groome et al., 2001).

The above mentioned botanical disaster of 20th century touched country of Geogia as well. Ten years ago 50% of Georgian chestnut forests were already dead (Prospero et al., 2013). There does not exist any effective fungicide against C. parasitica, according to the information of Georgian Ministery of Environment Protection. Cutting of sick trees is the only way of cryphonecrosis localization here. Except the joined scientific project of Swiss and Georgian investigators and attempts of the Turkish scientists of integration of hypovirulent strains in Ajara chestnut forest, practically no active measures have been performed for saving of the unique plant of Castanea sativa in Georgia, which is placed in “Georgian red list” and “Red book” (Prospero et al., 2013). Thus, elaboration and testing of the strategy of biological control against cryphonecrosis in Georgian chestnut forests is very urgent task today. Accordingly, the purpose of the presented work was to isolate the virulent strains of C. parasitica, spread in chestnut populations of one of the regions of west Georgia (Imereti), and to reveal the effective, antagonistic biological agents against them. The strategy of the fight against chestnut blight, spread in chestnut populations of west Georgian region Imereti, will be elaborated for the first time, and isolation of antagonistic to C. parasitica microscopic fungi directly from one of the hot spots of cryphonecrosis of Georgia will be performed for the first time, as well as creation of the collection of antagonists against the pathogen. All this may be regarded as the scientific novelty of the presented work.

Collection of this type may be considered as the material base for management and localization of chestnut blight epidemic in this region. We hope that creation of the antinecrotic bio-preparation and its in situ testing on the base of isolated particular antagonist microscopic fungi or their consortia will become possible in future.

Source : Microscopic fungi antagonistic to chestnut blight- Cryphonectria parasitica (Murrill) Barr.

#Chestnut blight #Cryphonectria parasitica #Antagonist micromycetes

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ijtsrd · 3 years ago

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Biodegradation Potentials of Aspergillus Flavipes Isolated from Uburu and Okposi Salt Lakes

by Kingsley C. Agu | Frederick J. C. Odibo "Biodegradation Potentials of Aspergillus Flavipes Isolated from Uburu and Okposi Salt Lakes"

Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-5 , August 2021,

URL: https://www.ijtsrd.com/papers/ijtsrd44949.pdf

Paper URL: https://www.ijtsrd.com/biological-science/microbiology/44949/biodegradation-potentials-of-aspergillus-flavipes-isolated-from-uburu-and-okposi-salt-lakes/kingsley-c-agu

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Saline lakes are water bodies with salinity greater than 3 g l 0.3 , while hypersaline lakes are water bodies that surpass the moderate 35 g l 3.5 salt of oceans. Hypersaline lakes, could either be thalassohaline which are creations of evaporation of seawater and as such contain sodium chloride as the major salt, with a salinity that surpasses that of seawater by a factor of 5 – 10, with a neutral or slightly alkaline pH . Whereas, athalassohaline lakes stem from non seawater sources and are made up of high concentrations of ions such as magnesium and calcium and sundry other ions such as potassium, or sodium in smaller amounts. This work has revealed the biodegradation potentials of some halophiles isolated from Uburu and Okposi salt lakes. The isolates recovered in descending order of salt tolerance were Aspergillus flavipes 13mm at 40 , Penicillium citrinum 10mm at 40 , Aspergillus ochraceus 9mm at 40 , Aspergillus nomius 15mm at 35 , Microsphaeropsis arundinis 12mm at 35 , Aspergillus sydowi 28mm at 30 , Penicillium janthinellum 26mm at 30 , Mucor sp 13mm at 30 , Aureobasidium sp 12mm at 30 , Trichoderma sp 9mm at 30 , Alternaria sp. 22mm at 25 , Aspergillus sp 18mm at 25 , Penicillium sp 20mm at 20 , Cladosporium sp. 7mm at 15 and identified using ITS rDNA Sequencing Macrogen, South Korea . They belonged to the borderline extreme halophiles and moderate halophiles respectively. The biodegradative potential of Aspergillus flavipes was ascertained by testing it against 2 , 4 and 6 crude oil and it grew only on 2 crude oil Bushnell Haas broth with a fungal count of 2.56x105 cfu ml. Crude oil degradation rate was evaluated biweekly gravimetrically with 22 degradation in 2 weeks, 36 in 4 weeks, 67 in 6 weeks and 89 in 8 weeks as well as by way of gas chromatography GC FID , which showed that fractions C10 C11 were significantly degraded, C12 C20, moderately degraded and C26 C34, insignificantly degraded.

#Biodegradation #Potentials #Aspergillus flavipes #Uburu

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crimson-biotechnology · 4 years ago

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Creation of Trichoderman: From an Idea to Realization _ Crimson Publishers

Creation of Trichoderman: From an Idea to Realization by Pakdaman BS in Journal of Biotechnology & Bioresearch

The current world of agriculture suffers from many limitations and limiting factors, and Trichoderma fungi as biologically beneficial agents can play considerable roles in soil and crop residue management, biological control of plant diseases and pests even in the niches hardly accessible by chemical pesticides, agricultural environment maintenance, as well as plant growth and yield promotion. Therefore, the creation of Trichoderma fungi via genetic transformation of superior isolates by the genetic constructs for the expression and secretion of insect-specific insecticidal peptides (Trichodermans) can improve their effectiveness in the control of insect pest targets.

The present world of agriculture suffers from multiple of problems. The increasing population of the world means increased demand for agricultural production [1]. Despite of this fact, only about 5% of current agricultural soils are disease suppressive and the remnants are disease conducive [2]. While agricultural soils are exposed to increased rate of erosion and gradual loss of fertility [3], the total area for agricultural activities comes down because of increased constructions and buildings. This situation is more aggravated where the cities are expanded over agricultural regions [1]. Plants are frequently affected by mostly fungal pathogens and insect pests that invade the internal tissues which are not accessible by the pesticides.

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juniperpublishers-ttsr · 4 years ago

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Evaluation of Antagonistic Effect of Trichoderma Harzianum against Fusarium oxysporum causal Agent of White Yam (Dioscorearotundata poir) Tuber Rot

Abstract

Studies were carried out on biological control using dual culture method to assess the potential of Trichoderma harzianum against Fusarium oxysporum, the causal agent of tuber dry rot disease of white yam in vitro. The rot-causing fungal organism was isolated from rotted yam tubers collected from different storage barns in Zaki-Biam, Benue State of Nigeria. The biological agent (T harzianum) was introduced at three different times (same time with the pathogen, 2 days before the inoculation of the pathogen and 2days after the inoculation of the pathogen). The experiment was conducted in the Advanced Plant Pathology Laboratory, Federal University of Agriculture, Makurdi, Nigeria. The plates were incubated for 192 hours and measurements of mycelial radial growths were done at 24 hours intervals beginning from 72hour. The result of the in vitro interactions showed that the highest percentage growth inhibition of mycelia of the pathogen (77.99%) was recorded when T. harzianum was introduced two days before the inoculation of F. oxysporum, followed by introduction of T. harzianum same time with F. oxysporum (45.69%) and the least percentage growth inhibition (13.72%) was recorded when the antagonist was introduced two days after inoculation of the pathogen. In all the treatments, T. harzianum was able to significantly (P<0.05) inhibits the growth of F. oxysporum throughout the period of incubation. T. harzianum was observed to be effective in reducing the mycelial growth of F. oxysporum in culture in all the treatments and therefore showed the potential for biological control of the pathogen. Minimum inhibition concentration recorded showed that the effectiveness levels of T. harzianum were slightly effective to effective across treatments. It is therefore concluded that the antagonistic potentials of T. harzianum could be used as a good biological control agent of yam tuber rot caused by F. oxysporum.

Keywords: Rot; In vitro; T. harzianum; Antagonistic; F. oxysporum; Yam; Inhibition

Abbreviations: PDA: Potato Dextrose Agar; PGI: Percent Growth Inhibition; MIC: Minimum Inhibition Concentration; CRD: Completely Randomized Design; LSD: Least Significance Difference

Introduction

Yam (Discorea) is a major staple food crop in West Africa, where it provides food for over 60 million people [1,2] estimated that the world production of yams is around 20 million tons per year. In spite of the high volume of yam production, rotting of yam in storage continue unabated. Fungal organisms commonly involved in yam rots includes: Aspergillus niger, Botryodiplodia theobromae, Fusarium solani, Fusarium oxysporum, Penicillium, Rhizopus stolonifer and Mucor [3-5]. Losses due to post-harvest rot significantly affect farmers' and traders' income, food security and seed yams stored for planting. Studies conducted by Okigbo RN [6] indicated that between 20 and 39.5% of stored tubers may be lost to rot organisms in storage. The use of synthetic chemicals such as sodium orthiophenylphenate, mancozeb and borax has been found to reduce storage rot in yam [4,7]. However, biological control is generally favoured as a method of plant disease management [8,9].This is because it is selective in action, produces no side effect and cost less. Resistance to biological control is rare and biological control agents are self- propagating and self-perpetuating [6]. The aim of this research was to study the antagonistic activities of a bio agent, T. harzianum introduced at different times in vitro to control Fusarium oxysporum, the causal agent of dry rot of yam tubers.

Materials and Methods

The study area

The experiment was conducted at the Advanced Plant Pathology Laboratory, Federal University of Agriculture, Makurdi, Nigeria.

Source of T. harzianum isolate

The biological control agent (T. harzianum) was obtained from yam Pathology Unit of University of Ibadan, Oyo State, Nigeria. Stock cultures of the isolate were maintained on slants of acidified Potato Dextrose Agar (PDA) in McCartney bottles for subsequent studies.

Collection of disease yam tubers

Rotten yam tubers of white yam varieties (Dioscorea rotundata) showing various diseased symptoms of dry rots were obtained from yam farmers from various storage barns in Zaki- Biam, Ukum local government area of Benue State, Nigeria which lies between longitudes 9° 25' and 9° 28'E, and latitude 7° 32' and 7° 35'N respectively The rotten yam tubers were packaged in sterile polyethylene bags. The tubers were protected using wire mesh to prevent rodent attack [10]. Potato Dextrose Agar (PDA) was the medium used. Test fungus for this study was F. oxysporum.

Isolation and identification of F. oxysporum

F. oxysporum was isolated from naturally infected yam tubers. Small sizes of approximately 2x2mm were cut out with sterile scalpel from yam tubers infected with rot at inter-phase between the healthy and rotted portions of the tubers. The cut tissues were soaked in 5% sodium hypochlorite for 2minutes for surface sterilization and then rinsed in four successive changes of sterile distilled water as reported by Ritchie B [11]. The infected tissues were later picked onto sterile filter paper using a sterile forceps and then wrapped with filter paper for 2-3minutes. The dried infected tissues were placed on several prepared sterile plates of acidified Potato Dextrose Agar (PDA) and the plates were incu-bated at ambient room temperature (30±5 °C) for 192 hours. The different fungi that grew from infected tissues were sub-cultured on separate sterile acidified PDA plates and incubated to obtain pure cultures of pathogens. Macroscopic and microscopic examination and morphological characteristics and identification were made and compared with already established authorities [12,13].

Antagonistic potential of T. harzianum against F. oxysporum in vitro

The assay for antagonism was performed on Potato Dextrose Agar (PDA) on Petri dishes by the dual culture method [14]. The mycelial plugs (5mm diameter) of 5 day old fungal antagonist and pathogen were placed on the same dish 6cm from each other. Isolate of test fungal antagonist was plated same time with pathogen, two days before the inoculation of the pathogen and two days after the inoculation of the pathogen. Paired cultures were incubated at room temperature (30±5 °C) for 192hours. Dishes inoculated only with test pathogens served as controls. Treatments comprised times of introduction of antagonist which were replicated three times for each treatment and arranged in completely randomized design [15].

Measurement of radial mycelial growth

Measurement of mycelia radial growths of the pathogen in dual culture and in control plates were done at 24 hour interval beginning from the 72nd hour up to the 192nd hour of incubation at ambient room temperature (30 ± 5 °C). Percent Growth Inhibition (PGI) of pathogen was also calculated as described by Korsten L [16].

PGI = Percent Growth Inhibition

R = the distance (measured in mm) from the point of inoculation to the colony margin in control plate,

R1 = the distance of fungal growth from the point of inoculation to the colony margin in treated plate in the direction of the antagonist.

The percent growth inhibition was calculated and values were used in selecting the Minimum Inhibition Concentration (MIC) that will be effective in controlling F. oxysporum. Antagonist was also rated for inhibitory effects using a scale by Sangoyomi T [17]:

≤ 0% inhibition (not effective),

>0-20% inhibition (slightly effective)

>20-50% inhibition (moderately effective),

>50-<100% inhibition (effective)

100% inhibition (highly effective)

Experimental design and data analysis

The experimental design was Completely Randomized Design (CRD) with three replicates as described by Gomez KA [15]. Test of variance was calculated using Analysis of variance (ANOVA) and statistical F-tests were evaluated at P≤0.05. Differences among treatment means for each measured parameter were further separated using fishers Least Significance Difference (LSD) to determine levels of significance according to Cochran GW [18].

Results

Isolation and identification of F. oxysporum

Fungal organisms including F. oxysporum were isolated and identified as one of the rot-causing pathogens of white yam tubers in storage in the study area. F. oxysporum was used as test pathogen in this study. Colony of F. oxysporum on PDA was moderately fast growing and produced a varying amount of aerial mycelium, initially white, and changing to peach, salmon, wine grey to purple or violet colour (Figure 1). Mycelia became thickly matted and sometimes wrinkled in older colonies. Spore masses were creamy white in colour. Macroconidia borne on short, branched conidiophores were generally abundant, hyaline, single-celled and variable, oval to kidney shaped. Macroconidia were hyaline, thin walled, only slightly curved and pointed at both ends, 3-7 septate, with a somewhat hooked apex and a foot-shaped basal cell (Figure 1).

Antagonistic potential of T. harzianum against F. oxysporum in vitro

The antagonistic potential of T. harzianum against F. Oxysporum show increased in inhibition with days of incubation in the dual culture method. The control plates that were not inoculated with T. harzianum grew much faster when compared with F. oxysporum in the dual culture plates. The dual culture plates showed initial rapid growth of F. oxysporum which terminated at the point of contact with T. harzianum (Figure 2 & 3). When T. harzianum was introduced 2 days before inoculation of the pathogen, the antagonist overgrew the pathogen and completely colonized it (Figure 4). The interaction between T. harzianum and F. oxysporum produced spores from both the antagonist and the pathogens at the point of contact when they were introduced same time. However, spores of F. oxysporum were not seen when the antagonist was introduced 2 days before inoculation of the pathogen. Chlamadosphore were observed when the antagonist was introduced two days after the inoculation of the pathogens (Figure 5).

Table 1 show that, T. harzianum assessed in this study was able to reduced mycelia growth of F. oxysporum when grown in dual culture irrespective of the time of introduction of the antagonist. The results of dual culture indicated that T. harzianum significantly (P≤0.05) inhibited the growth of F. oxysporum at varying degrees across duration of incubation (Table 1). The findings revealed that T. harzianum introduced 2days before inoculation of F. oxysporum has the highest mean percentage growth inhibition (77.99%), followed by the introduction of T. harzianum same time with the pathogen (45.69%) and the least mean percentage growth inhibition (13.72%) was recorded when the antagonist was introduced 2 days after inoculation of the pathogen (Table 1).

Means on the same column with the same superscript are not statistically significant (P≤0.05). Means on the same row (for Mean) with the same superscript are not statistically significant (P≤0.05) by time of introduction of T. harzianum. Thxpath = T.harzianum introduced same time with pathogen; Th2 dbipath = T.harzianum introduced 2 days before inoculation of pathogen; Th2daipath = T. harzianum introduced 2 days after inoculation of pathogen.

Radial mycelial growth and determination of inhibition

Figure 6 shows the radial mycelial growth of T. harzianum and F. oxysporum in dual culture and F. oxysporum in control plates for each of the treatments from 72 hours to 196 hours of incubation. The result revealed that growth of the pathogen was more rapid in the control plates than in the dual culture plates in all the treatments. It was also found that T. harzianum grew much faster than F. Oxysporum in all the treatments in dual culture there by inhibiting the growth of the fungus.

Minimum inhibition concentration recorded showed that the effectiveness levels of T. harzianum were slightly effective (introduction of T. harzianum 2days after inoculation of pathogen) to effective (introduction of T. harzianum 2 days before inoculation of pathogen) and was found to be significantly different (P≤0.05) across treatments (Table 2).

Thxpath = T.harzianum introduced same time with pathogen; Th2dbipath = T.harzianum introduced 2 days before inoculation of pathogen; Th2daipath = T. harzianum introduced 2 days after inoculation of pathogen; MIC = Minimum Inhibition Concentration (%); ≤0% inhibition (not effective); >0-20% inhibition (slightly effective); >20-50% inhibition (moderately effective); >50-x003C;100% inhibition (effective); 100% inhibition (highly effective).

Discussion

Fungal pathogenic organisms are the major cause of yam losses in storage in major yam producing areas of Nigeria. The rot-causing pathogens reduce output and productivity of yam annually. This has always placed the demand for yam tubers above its supply [19]. This result is similar to the work of [3,20,21] who had earlier on identified this organism in rotted yam tubers at different parts of Nigeria.

T. harzianum assessed in this work was able to reduced mycelial growth of F. oxysporum when grown in dual culture irrespective of the time of introduction of the antagonist. The inoculation of the test antagonists two days before the pathogen was done because there are no bio control agents that have enough competitive ability to displace an already established pathogen. The time lapse allows adequate increase in cell concentration and subsequent colonization by antagonist before the arrival of the pathogen [22]. The results of the dual culture showed that T. harzianum inhibited the growth of the target organisms through its ability to grow much faster than the pathogenic fungi thus competing efficiently for space and nutrients. Starvation was the most common cause of death for F. oxysporum, so that competition for limiting nutrients resulted in biological control of the test pathogen. This finding is similar to the work of [9,23]. Antagonists may also affect growth of pathogen either through antibiosis or mycoparasitism [24]. It was observed microscopically that the interaction region between F. oxysporum with T. harzianum showed mycelia of T. harzianum grew on the surface of the pathogen and later penetrated the cell walls directly without formation of appresorium structures. It also prevented the formation of both micro and macro conidia of the test fungus when the antagonist was introduced 2 days before inoculation of the pathogen. The pathogen mycelial then disintegrated suggesting enzyme action. This result agreed with the work done by Mokhtar H et al. [2 5] who reported the inhibition of the growth and spore formation of F. acuminatum by 66.66% using T. harzianum in vitro. Research conducted by Lorito M et al. [26] demonstrated possible role of chitinolytic and/or glucanases enzymes in bio-control by Trichoderma. They showed that the enzymes function by breaking down the polysaccharides, chitin, and glucans that are responsible for the rigidity of fungal cell walls, thereby destroying cell wall integrity limiting the growth of the pathogen.

Microscopic observation also showed that the antagonist, T. harzianum prevented the formation of spores of the test pathogen. It was observed in the study that a clear zone of interaction was formed in all T. harzianum and F. oxysporum interactions except in the interaction between T.harzianum and F. oxysporum; where T. harzianum was introduced 2 days before inoculation of F. Oxysporum where T. harzianum over grew the pathogen in the dual culture plates. The zones of inhibition produced might be due to the production of antifungal metabolites by the test antagonist as reported by Adejumo TO et al. [27].

Studies conducted by Ekefan EJ et al. [28] showed that T. harzianum isolates suppressed the growth of Colletotrichumcapsici eventually overgrowing it within seven days. Report by Ekefan EJ et al. [29] demonstrated that the Trichoderma isolates have a strong antagonism against wilt diseases caused by Fusarium sp, invitro, on potato dextrose agar medium, when growth was decreased by 88%, 86% and 80% for Trichoderma harzianum, T. hamatum and T. viride respectively. According to [30] the antagonism in vitro of Trichoderma harzianum against Fusarium oxysporium showed an inhibition on the pathogenic fungus growth with a ratio more than 65%, moreover the volatile metabolism substances of the antagonism reduced the pathogenic fungus growth by 63% compared with controls.

The inoculation of the test antagonists two days before the pathogen was done because there are no bio control agents that have enough competitive ability to displace an already established pathogen. The time lapse allows adequate increase in cell concentration and subsequent colonization by antagonist before the arrival of the pathogen [22].

Minimum Inhibition Concentration (MIC) showed that the introduction of T. harzianum 2 days before the inoculation of F. oxysporum was more effective in controlling the pathogen compared with the introduction of the antagonist same time with the pathogen as well as the introduction of the antagonist 2 days after inoculation of the fungus [31,32]. Introduction of T. harzianum 2 days before inoculation of F. oxysporum was therefore considered more effective in controlling the pathogenic fungus in vitro [33].

Conclusion

The study found out the antagonistic potentials of T. harzianum as good biological control agent of yam tuber rot caused by F. oxysporum. The study also revealed that the best time for the introduction of bio-agent is before the arrival of the pathogen. This is because the introduction of the antagonist 2days before inoculation of F. Oxysporum in vitro gave the highest inhibition of the mycelial growth of the pathogen.

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juniperpublishers-agriculture · 4 years ago

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The Exploration of Mycoviruses From Trichoderma Spp.- Juniper Publishers

Mycoviruses are a kind of widespread virus, which can infect filamentous fungi and yeasts [1-3]. Most viruses can infect the host without obvious infection symptoms [1-2]. Double-stranded RNA (dsRNA) genomes always the main nucleotide style of mycoviruses, which has been a marker of the existence of viruses [4]. The International Committee on Taxonomy of Viruses has divided all known branching viruses into 16 families and one unclassified group according to the mode of replication and genomics types of viruses [5]. At present, nine mycoviruses infecting Trichoderma spp. have been described [6-14]. 2009, Jom-in and Akarapisan provided the first description of two mycoviruses with sizes of 0.7kb and 1.1KB isolated from T. spp. [6]. Later, Yun et al. proposed that Lentinus edodes mycoviruses were widespread in Korea, and 32 different dsRNA-containing viruses were isolated from 315 strains of T. spp. [7]. Then, Lee et al. isolated an unclassified endovirus from Trichoderma atroviride, named Trichoderma atroviride mycovirus 1 (TaMV1) [8]. In 2018, Chun et al. obtained the complete genome sequences of two Trichoderma endoviruses, namely Trichoderma atroviride partitivirus 1 (TaPV1) and Trichoderma harzianum partitivirus 1(ThPV1) [9,10]. Meanwhile, a new mycovirus named Trichoderma asperellum dsRNA virus 1 (TaRV1) was reported in the laboratory of Guizhou Medical University in 2019 [11]. In 2019, Liu et al. isolated two unclassified dsRNA endoviruses, Trichoderma harzianum mycovirus 1 (ThMBV1) and Trichoderma harzianum mycovirus 1 (ThMV1), from 155 Trichoderma strains which were collected from the soils of Xinjiang and Inner Mongolia, China [12,13]. In 2019, You et al. from Huazhong Agricultural University isolated a new mycovirus from Trichoderma harzianum, named Trichoderma harzianum hypovirus 1 (ThHV1), which may be a member of Betahyprovirus [14]. In this paper, we chose 120 strains of T. spp. isolated from Xinjiang, Inner Mongolia, and Heilongjiang of China, to screen mycoviruses using molecular biology methods, and three mycoviruses strains were recovered.

Materials and methods

Fungal strains

One hundred and twenty T. spp. strains were isolated from the forests and grasslands of Xinjiang, Inner Mongolia and Heilongjiang of China, in 2017–2018. The species of all of these strains were identified by the lab experiment, the information about these strains were in the supplemental table S1. All fungal strains were cultured on potato dextrose agar (PDA) plates for 7 days at 28°C for the preparation of the next step.

Extraction of dsRNA

All fungal strains were transferred to in the potato dextrose liquid medium (PD), and placed on the shaker at 200 r/min and cultured at 28℃, after 2 days, collecting the mycelium for dsRNA extraction. DsRNA was extracted and purified from the mycelia by using CF-11 cellulose column chromatography [15]. Then, the samples were digested by RNase free DNase I (TaKaRa, Dalian, China) and S1 Nuclease (TaKaRa, Dalian, China) following the manufacturer’s instructions to remove any DNA and ssRNA contamination [13]. At last, electrophoresises of the samples were run in 1% agarose gel, and the dsRNA was detected by ultraviolet transillumination.

Results

After 2 days’ culture in PD, the mycelium of 120 strains of T. spp. were collected, then dsRNA of the 120 strains were extracted by the molecular biology methods. Though the existing of dsRNA in the electrophoresis, the three Trichoderma strains with mycoviruses have been screened out. T673 was T. harzianum, T703 and T827 were T. citrinoviride, all of which were from Xinjiang province (Figures 1A, B, C). After dsRNA was extracted from mycelium, two fragments of 1.5 and 1.7 KB were found in strain T673 (T. harzianum), then DNase I and S1 nuclease were used to confirm the type of nucleic acid, and dsRNAs were identified (Figure 2A). In the strain of T703 (T. citrinoviride), only one fragment of 9 KB was found, and the dsRNA was digested by DNase I and S1 nuclease to confirm the properties of nucleic acid (Figure 2 B). With the same method, one dsRNA fragment of 9 KB was found in strain T827 (T. citrinoviride) (Figure 2C).

Discussion

In this paper, we obtained three strains of mycoviruses from 144 strains of T. spp. isolated from the soil, which can preliminarily prove that mycovirus existed in T. spp., and provided insight into how to locate and isolate mycoviruses from T. spp.. In the obvious researches, five genome sequences of mycoviruses from T. spp. were obtained, from which three strains were from T. harzianum [6-14]. In this issue, one mycovirus strain T-673 weas explored from T. harzianum, another two strains from T. citrinoviride. At present, for the investigation on DNA mycovirus was few in the nature, the DNA mycovirus from T. spp. has not been recovered [3]. All in all, the investigated dsRNA and DNA mycoviruses from T. spp. were few until now, which need to be explored more and more. In the future, nucleic acid type, the elimination, complete genome sequence, transmission mode and antagonistic effect on Trichoderma and biocontrol function of the mycovirus, as well as the interaction between fungi and mycoviruses, need to be revealed.

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slowlyperfecttidalwave · 5 years ago

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Fungal Recombinant Proteins

The initial focus of the production of recombinant proteins by filamentous fungi associated with utilizing enzyme synthesis and secretion of extracellular incredible machine of industrial strains, including species of Aspergillus,

Trichoderma, Penicillium and Rhizopus, is to produce a recombinant protein products. An initial weakness fungal filaments recognized as the host of recombinant proteins is their common ability to produce protease homologues can degrade the heterologous protein products and strategies to prevent proteolysis has met with some limited success.

It was also recognized that the glycosylation patterns of proteins in filamentous fungi and mammals are very different, so it may not be a filamentous Fungal Recombinant Proteins host microbes that are most suitable for the production of recombinant human glycoproteins for therapeutic use.

By combining the experience gained from the production of recombinant proteins single new scientific information generated through genomics and proteomics research, biotechnology is now ready to expand the capabilities biomanufacturing filamentous fungi recombinant by allowing them to express a gene that encodes several proteins, including, for example, biosynthetic pathways new for the production of a new primary or secondary metabolites.

It is recognized that filamentous fungi, most species that have not been isolated, is a source of enormous variety of new biosynthetic pathways, and that the natural fungal host storing a valuable biosynthetic pathways organism may often not the most suitable for the purpose biomanufacture.

It is therefore expected that substantial efforts will be directed to change the other host fungal, microbial host non-fungal and indeed host non microbes to reveal some of the new biosynthetic pathways. But future applications of recombinant protein expression will not be limited to the biomanufacturing.

Opportunities to exploit recombinant technology to uncover the cause of the detrimental effects of fungi, such as humans, mammals and plant pathogens, and then to bring the solutions forward, is expected to represent a future focus very important technology of recombinant fungal proteins.

#Fungal Recombinant Proteins

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nuadox · 7 years ago

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Fungi can help concrete heal its own cracks

- By Congrui Jin , Binghamton University , The Conversation -

Infrastructure supports and facilitates our daily lives – think of the roads we drive on, the bridges and tunnels that help transport people and freight, the office buildings where we work and the dams that provide the water we drink. But it’s no secret that American infrastructure is aging and in desperate need of rehabilitation.

Concrete structures, in particular, suffer from serious deterioration. Cracks are very common due to various chemical and physical phenomena that occur during everyday use. Concrete shrinks as it dries, which can cause cracks. It can crack when there’s movement underneath or thanks to freeze/thaw cycles over the course of the seasons. Simply putting too much weight on it can cause fractures. Even worse, the steel bars embedded in concrete as reinforcement can corrode over time.

Very tiny cracks can be quite harmful because they provide an easy route in for liquids and gasses – and the harmful substances they might contain. For instance, micro-cracks can allow water and oxygen to infiltrate and then corrode the steel, leading to structural failure. Even a slender breach just the width of a hair can allow enough water in to undermine the concrete’s integrity.

But continuous maintenance and repair work is difficult because it usually requires an enormous amount of labor and investment.

So since 2013, I’ve been trying to figure out how these harmful cracks could heal themselves without human intervention. The idea was originally inspired by the amazing ability of the human body to heal itself of cuts, bruises and broken bones. A person takes in nutrients which the body uses to produce new substitutes to heal damaged tissues. In the same way, can we provide necessary products to concrete to fill in cracks when damage happens?

My Binghamton University colleagues Guangwen Zhou and David Davies, Ning Zhang from Rutgers University and I have found an unusual candidate to help concrete heal itself: a fungus called Trichoderma reesei.

Researchers screened a number of fungi looking for a candidate that could help fill concrete cracks. Congrui Jin, CC BY-ND

We initially screened about 20 different species of fungi in order to find one that could withstand the harsh conditions in concrete. Some we isolated from the roots of plants that grew in nutrient-poor soils, including from the New Jersey Pine Barrens and the Canadian Rocky Mountains in Alberta.

We found that as calcium hydroxide from concrete dissolved in water, the pH of our fungal growth medium increased from a close-to-neutral original value of 6.5 all the way to a very alkaline 13.0. Of all the fungi we tested, only T. reesei could survive this environment. Despite the drastic pH increase, its spores germinated into threadlike hyphal mycelium and grew equally well with or without concrete.

Once the spores (left) germinate with the addition of water, they grow into threadlike hyphal mycelium (right). Congrui Jin, CC BY-ND

We propose including fungal spores, together with nutrients, during the initial mixing process when building a new concrete structure. When the inevitable cracking occurs and water finds its way in, the dormant fungal spores will germinate.

As they grow, they’ll work as a catalyst within the calcium-rich conditions of the concrete to promote precipitation of calcium carbonate crystals. These mineral deposits can fill in the cracks. When the cracks are completely caulked and no more water can enter, the fungi will again form spores. If cracks form again and environmental conditions become favorable, the spores could wake up and repeat the process.

T. reesei is eco-friendly and nonpathogenic, posing no known risk to human health. Despite its widespread presence in tropical soils, there are no reports of adverse effects in aquatic or terrestrial plants or animals. In fact, T. reesei has a long history of safe use in industrial-scale production of carbohydrase enzymes, such as cellulase, which plays an important role in fermentation processes during winemaking. Of course, researchers will need to conduct a thorough assessment to investigate any possible immediate and long-term effects on the environment and human health prior to its use as a healing agent in concrete infrastructure.

Future cement recipes may include fungi. Midtown Crossing at Turner Park, CC BY

We still don’t fully understand this very young but promising biological repair technique. Concrete is a harsh environment for the fungus: very high pH values, relatively small pore sizes, severe moisture deficit, high temperatures in summer and low temperatures in winter, limited nutrient availability and possible exposure to ultraviolet rays from sunlight. All of these factors dramatically influence the fungi’s metabolic activities and make them vulnerable to death.

Our research is still in the initial stage and there’s a long way to go to make self-healing concrete practical and cost-effective. But the scope of American infrastructure’s challenges makes exploring creative solutions like this one worthwhile.

Congrui Jin, Assistant Professor of Mechanical Engineering, Binghamton University, State University of New York

This article was originally published on The Conversation. Read the original article.