Antagonistic activity and hyphal interactions of Trichoderma spp. against Fusarium proliferatum and F. oxysporum in vitro

Trichoderma is a well-known antagonist against soilborne plant pathogens. However, the species and even various isolates have different biocontrol potential. To evaluate the antagonistic activities of Trichoderma harzianum, T. harzianum strain T100 (T100), T. viride and T. haematum against Fusarium oxysporum and F. proliferatum, we used dual culture and productions of volatile and non-volatile metabolites in three different phases in vitro. An analysis of the data in dual culture tests represented T. viride, T. haematum and T100 as effective antagonists of Fusarium while T100 was the only fungus being able to lyse the confronting mycelia. Similar results were obtained in the volatile metabolites tests also. In contrast with the two previous tests, the non-volatile metabolites produced by T. harzianum inhibited Fusarium mycelial growth the most, and T100 acted moderately. It was also clearly showed that the antagonistic effect of Trichoderma spp. was more on F. proliferatum than on F. oxysporum. Finally, because Trichoderma spp. was most effective in the second phase, we recommend to use T100 against F. proliferatum at the initial stages of infection as its mycoparasitism on F. oxysporum was observed microscopically through forming apressoria structures without any coiling around the pathogen.     

Inhibitory effect of Trichoderma isolates on growth of Alternaria alternata,the causal agent of leaf spot disease on sunflower,under laboratory conditions

Several species of the genus Alternaria are involved in leaf spot disease of sunflower, with Alternaria alternata being the dominant species responsible for this disease in Iran and many other sunflower-producing areas, worldwide. The disease causes a progressive destruction of the photosynthetic apparatus, resulting in annual yield loss. The routine disease management strategies are not effective for disease control; hence, alternative measures for disease management are of great interest. In the present study, the efficacy of Trichoderma harzianum T22 and Trichoderma sp. on biological control of the causal agent was evaluated under laboratory conditions. The effect of Trichoderma isolates on dry weight (DW) and radial growth (RG) rate of A. alternata was evaluated using dual culture, volatile and non-volatile cellular metabolites. The results obtained in this study revealed that in both Trichoderma isolates, non-volatile cellular metabolites had the highest inhibitory effect on DW and RG rate of the causal agent. Owing to explicit inhibitory effect of non-volatile cellular metabolites on A. alternata, the inhibitory effects of different concentrations of non-volatile cellular metabolites were evaluated on DW and RG rate of the A. alternata. The obtained results showed that non-autoclaved 75 and 50% concentrations and undiluted (100%) autoclaved non-volatile cellular metabolites from Trichoderma sp. had the highest inhibitory effect on DW and RG rate of the causal agent. The overall results of this study reveal that Trichoderma spp. have excellent efficacy on biological control of A. alternata under laboratory condition; such that, further studies on the potential of Trichoderma spp. in biological control of Alternaria leaf spot disease of sunflower under green house and field conditions are necessary.     

Biocontrol potential of Trichoderma species against mango malformation pathogens

Malformation disease of Mango (Mangifera indica L.) caused by Fusarium moniliforme var. subglutinans is one of the most destructive diseases, which is a major production constraint in the mango-growing regions of India. In this study, The bioagents Trichoderma viride (Tr1), Trichoderma virens (Tr2) and Trichoderma harzianum (Tr3) were evaluated in culture with the pathogens to monitor the antagonistic effect and their volatile compound and culture filtrates (non-volatile compound). It was found that all the three isolates of bioagents significantly checked the growth of F. moniliforme var. subglutinans. In dual culture, the best result was obtained with T. harzianum followed by T. virens and T. viride. A similar result was also observed in the case of culture filtrates ofTrichoderma spp. The results clearly showed that inhibition of the growth of the fusaria isolates by T. harzianum was significantly superior to T. viride andT.virens. In case of antifungal activity of volatile compounds released by Trichoderma isolates, it was also observed that T. virens was more superior to T.harzianum and T. viride.     

Control of fusarium wilt of <Emphasis Type="Italic">Solanum melongena</Emphasis> by <Emphasis Type="Italic">Trichoderma</Emphasis> spp.

Biological control of wilt of egg plant (Solanum melongena L.) caused by Fusarium solani was made with the application of five Trichoderma species, T. harzianum, T. viride, T. lignorum, T. hamatum and T. reesei. The effect of volatile and non-volatile antibiotics of Trichoderma origin on growth inhibition of the wilt pathogen was studied. T. harzianum showed maximum growth inhibition (86.44 %) of the pathogen through mycoparasitism. The non-volatiles produced by the Trichoderma species exhibited 100 % growth inhibition of the pathogen under in vitro condition. Production of siderophores and fungal cell wall degrading enzymes, chitinase and β-1,3-glucanase were found. Treatments with two most efficient Trichoderma species, T. harzianum and T. viride resulted in the decreasing population of Fusarium solani in soil thereby deterring disease incidence in field condition.     

Efficacy ofTrichoderma chitinases againstRhizoctonia solani, the rice sheath blight pathogen

Thirty-five strains ofTrichoderma viride andT. harzianum were screened for their antagonistic ability against the rice sheath blight pathogen,Rhizoctonia solani. The strains that inhibited/overgrew the phytopathogenic fungus were considered effective. Light microscopic studies showed the antagonism of the hyphae of effectiveTrichoderma strains towards their host hyphae. Chitinase activity ofTrichoderma culture filtrates was enhanced, when colloidal chitin was used as the sole carbon source, instead of glucose. Chitinase pattern differed among the four select strains. The chitinase isoforms are induced differentially by carbon sources. The chitin affinity column fraction ofTrichoderma culture filtrate inhibited,in vitro, the growth ofR. solani.     

Antagonistic effect of two isolates of Trichoderma harzianum against postharvest pathogens of tomato (Lycopersicon esculentum)

Pathogenicity test of all fungi (14 different isolates) isolated from both infected tomato fruits and the surface wash of other healthy fruits had different pathogenicity rates. The genus Rhizopus sp. was the most pathogenic one followed by Fusarium sp. Trichoderma harzianum isolates (T3 and T4) had a different pattern of antagonism against the tested pathogens. In dual plate test of the antagonistic action of T3 and T24 against the postharvest pathogens, clear zone size ranged between 1 and 4 and 3 and 6 mm by T3 and T24, respectively. The antagonists (T3 and T24) didn't show inhibition zone against Rhizopus sp., but they could overgrow it by 100% after 9 days of incubation. Mostly, all the other postharvest isolates showed high degree of overgrowth by T3 than T24. The two antagonists failed to overgrow Aspergillus species except for A. niger (3) which was overgrown by T3. Volatile and non-volatile metabolite tests indicated that mycelial growth of Penicillium stekii was significantly inhibited by T3 and T24 more than the other tested pathogens. The inhibition of A. niger (1) was 12% by non-volatile metabolites of T24 produced after 1 day incubation, and reached to 97% inhibition by the metabolites of 3 days. Interestingly, inhibition of Aspergillus sp. by volatile compounds of T3 and T24 was 2% and 20%, respectively, whereas the inhibition of the same pathogen by non-volatile compounds reached 75% and 87%, respectively. The results of slice assay clearly indicate that T. harzianum (T3 and T24) could provide a complete protection to tomato slices from the infection of the tested pathogens. After 3 days of incubation, Trichoderma suppressed the linear growth of these pathogens on tomato slices and the percentage of suppression was significant and ranged between 80 and 100%, except with Rhizopus sp. the suppression reached 33% only.     

Tunisian isolates of Trichoderma spp. and Bacillus subtilis can control Botrytis fabae on faba bean

Faba bean crops worldwide are often attacked by different diseases, particularly chocolate spot caused by Botrytis fabae Sard. Fungal and bacterial isolates collected from faba bean and barley leaves in Tunisia were evaluated for their antagonistic potential against B. fabae. In a test on detached leaves, the highest rate of decrease in disease severity was scored by Trichoderma viride, followed by T. harzianum, the fungicide Carbendazim then Bacillus subtilis. Under glasshouse conditions, all tested fungi resulted in significant disease severity reduction. T. viride reduced the rate of chocolate spot infestation on leaves and stems by 35% and 31.5%, respectively, when the rate on the control was 100%. For T. harzianum, Carbendazim and B. Subtilis, the rates of infestation on the leaves were 41.7%, 43.1% and 59.7%, respectively. On the stems, T. harzianum scored the lowest rate of 54.2% followed by B. subtilis with 79.2% then Carbendazim with 87.5%. Two consecutive seasons of field trials using the Trichoderma species, B. subtilis and Carbendazim showed significant and consistent reduction in the severity of chocolate spot infestation rates. The highest protection against the disease was obtained from T. viride. Based on these results, Tunisian isolates of Trichoderma spp. can be recommended for developing commercial bio-fungicides for integrated management of chocolate spot.     

Genetic and physiological relatedness of antagonistic Trichoderma isolates against soil borne plant pathogenic fungi

In this study, the in vitro potential of 42 Trichoderma spp. were evaluated against four isolates of soil borne phytopathogenic fungi viz., Rhizoctonia solani, Macrophomina sp., Sclerotium rolfsii and Pythium aphanidermatum in dual culture techniques and through production of volatile and non-volatile inhibitors. In vitro screening results showed that the proportion of isolates with antagonistic activities was highest for the S. rolfsii followed by R. solani, Macrophomina sp. and P. aphanidermatum, respectively. The isolates TNT1, TNP2 and TWP1 showed consistent results in volatile and non-volatile activity in vitro against any of the two pathogens tested. Based on genomic finger prints, potential isolates showed no particular correlation between the origin of the isolates and the Random Amplified Polymorphic DNA (RAPD) groups could not be established. However, the polymorphism shown by the isolates did not correlate to their level of antagonism. Whereas, in physiology studies using BIOLOG (microbial identification system), three groups were formed, one group consists with 14 different Trichoderma species and two groups with two isolates each comprised of only T. koningii and T. viride.     

<i>Trichoderma</i>-Induced Improvement in Growth,Photosynthetic Pigments,Proline, and Glutathione Levels in <i>Cucurbita pepo</i> Seedlings under Salt Stress

Salt stress is one of the major abiotic stress in plants. However, traditional approaches are not always efficient in conferring salt tolerance. Experiments were conducted to understand the role of Trichoderma spp. (T. harzianum and T. viride) in growth, chlorophyll (Chl) synthesis, and proline accumulation of C. pepo exposed to salinity stress. There were three salt stress (50, 100, and 150 mM NaCl) lavels and three different Trichoderma inoculation viz. T. harzianum, T. viride, and T. harzianum + T. viride. Salt stress significantly declined the growth in terms of the shoot and root lengths; however, it was improved by the inoculation of Trichoderma spp. C. pepo inoculated with Trichoderma exhibited increased synthesis of pigments like chl a, chl b, carotenoids, and anthocyanins under normal conditions. It was interesting to observe that such positive effects were maintained under salt-stressed conditions, as reflected by the amelioration of the salinity-mediated decline in growth, physiology and antioxidant defense. The inoculation of Trichoderma spp. enhanced the synthesis of proline, glutathione, proteins and increased the relative water content. In addition, Trichoderma inoculation increased membrane stability and reduced the generation of hydrogen peroxide. Therefore, Trichoderma spp. can be exploited either individually or in combination to enhance the growth and physiology of C. pepo under saline conditions.     

Fusant Trichoderma HF9 with enhanced extracellular chitinase and protein content

Strain improvement was carried out to obtain higher chitinase and protein by inter-specific protoplast fusion between Trichoderma harzianum and Trichoderma viride. Fusant HF9 and parental strains of Trichoderma were compared for chitinase and protein production. 1% of glucose, sucrose and fungal cell wall (Rhizoctonia solani), were used as carbon source for cultivation of Trichoderma and fungal cell wall was the best to induce chitinase and protein. Usage of 0.5% colloidal chitin for the fungal growth under aerated conditions at pH 6.5 and 28°C led to higher chitinase and protein production. In these conditions fusant Trichoderma HF9 in comparison with parent strains had 3-, 2.5- and 1.5-fold increase of total chitinase, specific chitinase and protein, respectively. SDS-PAGE analysis revealed that it had 9 major protein bands with up-regulation compared to parent strains. Amino acid analysis showed that protein of culture filtrate of T. harzianum, T. viride and fusant Trichoderma HF9 had 8, 6 and 10 amino acids, respectively. The results obtained suggested that fusant HF9 could be an integration of T. harzianum and T. viride through protoplast fusion.     

Biocontrol efficacy of Trichoderma spp. against sesame wilt caused by Fusarium oxysporum f. sp. sesami

Sesame (Sesamum indicum L.) is one of the most important oilseed crops in Egypt and worldwide. It is being infected with many pathogens, among these pathogens Fusarium oxysporum f.sp. sesami (Zap.) Cast is causing severe economic losses on sesame. In this study, antagonistic capability of 24 isolates of Trichoderma spp. was assessed in vitro against F. oxysporum f.sp. sesami. Two strains; T. harzianum (T9) and T. viride (T21) were revealed to have high antagonistic effect against F. oxysporum f.sp. sesami in vitro with inhibition percentage about 70 and 67%, respectively. These two isolates proved to have high ability to control Fusarium wilt disease under greenhouse conditions. The highest reduction in disease severity was achieved with T. viride followed by T. harzianum with reduction in disease severity about 77 and 74%, respectively. This study revealed that the time of application of bioagents is a decisive factor in determining the efficacy of Trichoderma isolates to control Fusarium wilt of sesame. It was revealed that the highest reduction in the disease severity was achieved when either Trichoderma viride or T. harzianum were applied 7 days before challenging with the F. oxysporum f.sp. sesami.     

Potential for Biocontrol of Lasiodiplodia theobromae (Pat.) Griff. & Maubl. in Banana Fruits by Trichoderma Species

Fifteen Trichoderma isolates were tested for their antagonistic ability against Lasiodiplodia theobromae. Trichoderma harzianum exhibited the greatest inhibition in dual culture. Microscopic investigation demonstrated direct parasitism and coiling of T. harzianum and T. viride around hyphae of L. theobromae, causing swollen, deformed, shortened, or rounded cells of the pathogen. Granulation of cytoplasm and disintegration of the hyphal walls of L. theobromae also were noted in dual culture. Trichoderma viride reduced rotting by 29.07 to 65.06% in artificially inoculated banana fruits. Treatment of banana fruits with T. viride 4 h prior to inoculation with L. theobromae provided better protection than simultaneous application or treatment 4 h after inoculation.     

Isolation and Characterization of Trichoderma spp. for Antagonistic Activity Against Root Rot and Foliar Pathogens

Trichoderma, soil-borne filamentous fungi, are capable of parasitising several plant pathogenic fungi. Twelve isolates of Trichoderma spp. isolated from different locations of South Andaman were characterized for their cultural, morphological and antagonistic activity against soil borne and foliar borne pathogens. The sequencing of these isolates showed seven different species. The isolates revealed differential reaction patterns against the test pathogens viz., Sclerotium rolfsii, Colletotrichum gloeosporioides and C. capsici. However, the isolates, TND1, TWN1, TWC1, TGD1 and TSD1 were most effective in percentage inhibition of mycelial growth of test pathogens. Significant chitinase and β-1,3-glucanase activities of all Trichoderma isolates has been recorded in growth medium. T. viride was found with highest chitinase whereas T. harzianum was recorded with highest β-1,3-glucanase activities.     

Mycoparasitism studies of Trichoderma species against three phytopathogenic fungi: evaluation of antagonism and hydrolytic enzyme production

Trichoderma spp. are used for biocontrol of several plant pathogens. However, their efficient interaction with the host needs to be accompanied by production of secondary metabolites and cell wall-degrading enzymes. Three parameters were evaluated after interaction between four Trichoderma species and plant-pathogenic fungi: Fusarium solani, Rhizoctonia solani and Sclerotinia sclerotiorum. Trichoderma harzianum and T. asperellum were the most effective antagonists against the pathogens. Most of the Trichoderma species produced toxic volatile metabolites, having significant effects on growth and development of the plant pathogens. When these species were grown in liquid cultures with cell walls from these plant pathogens, they produced and secreted β-1,3-glucanase, NAGAse, chitinase, acid phosphatase, acid proteases and alginate lyase.     

Biocontrol of onion white rot by application of Trichoderma species formulated on wheat bran powder

In vitro, Trichoderma album, Trichoderma harzianum, Trichoderma koningii, Trichoderma viride and Trichoderma virens showed antagonistic effect against the most pathogenic isolate (Sc2) of Sclerotium cepivorum, the cause of onion white rot disease. Five Trichoderma preparations of each Trichoderma sp. were prepared on wheat bran powder to be used for controlling white rot disease of onion. Greenhouse and field experiments followed the same trend where T. harzianum and T. koningii were the most effective in reducing the incidence and severity of white rot disease compared with the control. Trichoderma species preparations caused promotion to vegetative parameters of onion plants in pots and increase bulb productivity in filed. In this regard, T. harzianum and T. koningii were the most effective. A positive correlation was found between the biocontrol activity of Trichoderma species preparations and enhancement of peroxidase, polyphenoloxidase and chitinase enzymes in onion plants to resist infection with S. cepivorum.     

Ecological management of tropical sugar beet (TSB) root rot (Sclerotium rolfsii (Sacc.) by rhizosphere Trichoderma species

Trichoderma species are collected from different location of sugarbeet growing areas of Tamil Nadu and it is effective against Sclerotium rolfsii pathogen caused by sugarbeet ecosystems. Out of thirty-one isolates of Trichoderma viride and four isolates of Trichoderma harzianum collected and tested for their antagonistic activity against S. rolfsii by dual culture technique, one isolate was found to be effective T. viride (TVB1) that recorded the maximum (73.03%) inhibition on the mycelial growth recording only 2.40 cm growth as against 8.90 cm in the control. The isolates of T. harzianum THB-1 recorded 71.19% mycelial growth reduction over control. The colonisation behaviour of T. viride (TVB1) revealed that it completely over grew on pathogen within 48 h after interaction with the pathogen, and speed of growth on pathogen was also high and it possesses a higher level of competitive saprophytic ability. The best four isolates of TVB1, TVB-2, TVB-3 and TVB31 and two isolates of T. harzianum THB-1 and THB-2 were compared with other species of Trichoderma longibrachiatum, Trichoderma reesei, Trichoderma koningii and Chaetomium globosum and tested under in vitro condition. BA of neem cake at 150 kg ha^?1 + T. viride isolate (TVB1) at 2.5 kg/ha recorded least root rot disease incidence of 17.05% which accounted for 75.37% disease reduction over control and highest recorded maximum root yield 65.73 t ha^?1 and increasing sugar content.     

拮抗北细辛菌核病木霉菌的分离、鉴定及生防效果

[背景] 菌核病是北细辛根部主要病害之一,木霉菌作为目前应用最广泛的生物防治真菌,利用木霉菌防治北细辛菌核病是目前研究的热点。[目的] 通过稀释分离法对健康北细辛植株根际土壤进行菌株分离,以期筛选出有效拮抗北细辛菌核病的生防木霉菌。[方法] 以北细辛菌核病菌为靶标菌,采用平板对峙培养、挥发性与非挥发性物质抑菌的方法对分离得到的木霉菌进行筛选,采用生长速率法对筛选出的木霉菌的发酵液进行抑菌效果测定,并采用硫代巴比妥酸法测定筛选出的木霉对北细辛菌核病菌的丙二醛（Malondialdehyde,MDA）含量、紫外吸收法测定过氧化氢酶（Catalase,CAT）活性、氮蓝四唑法测定超氧化物歧化酶（Superoxide Dismutase,SOD）活性、愈创木酚法测定过氧化物酶（Peroxidase,POD）活性的影响。[结果] 从土壤中分离出木霉菌共14株,通过形态学和ITS-RPB2双基因联合构建系统发育树,鉴定其为哈茨木霉（Trichoderma harzianum）、钩状木霉（Trichoderma hamatum）、拟康氏木霉（Trichoderma koningiopsis）、深绿木霉（Trichoderma atroviride）、短密木霉（Trichoderma brevicompactum）和装絮木霉（Trichoderma tomentosum）。对峙培养试验表明,钩状木霉A26、拟康氏木霉B30、钩状木霉C6、哈茨木霉A17对北细辛菌核病菌抑制率均在90%以上,挥发性物质抑制测定结果显示钩状木霉C6抑制率最高,为53.73%±0.07%,木霉菌的非挥发性物质抑菌作用较强,哈茨木霉A17、钩状木霉A26、钩状木霉C6的非挥发性物质对细辛菌核病菌的抑制率均在75%以上,而拟康氏木霉B30抑制率可达100%。因此,筛选出的哈茨木霉A17、钩状木霉A26、拟康氏木霉B30、钩状木霉C6为拮抗效果较强的生防木霉菌,这4株木霉菌的发酵液对北细辛菌核病菌的抑制率分别为56.33%±0.12%、77.22%±0.06%、82.28%±0.03%、46.20%±0.04%。经这4株木霉菌的非挥发性物质处理7 d后,菌核病菌MDA含量显著增加,钩状木霉A26是对照组的7.7倍,最为显著;菌核病菌抗氧化酶活性均降低,与对照组相比,CAT、SOD、POD活性分别下降了19.67%-75.84%、4.71%-68.71%和3.57%-67.86%。[结论] 从北细辛健康植株根际土壤中分离的木霉菌株哈茨木霉A17、钩状木霉A26、拟康氏木霉B30、钩状木霉C6对北细辛菌核病菌均有较好的抑制效果,可用于北细辛菌核病的生物防治。     

Biological control ofRhizoctonia solani in strawberry fields byTrichoderma harzianum

Summary Trichoderma harzianum preparations was used in two successive field experiments in commercial strawberry nurseries and fruiting fields. Disease severity ofRhizoctonia solani in daughter plants was reduced by 18–46 % in the treated nursery plots. Infestation of nursery soil with the pathogen, as tested by planting beans in soil samples was reduced by the Trichoderma treatment by up to 92% as compared to the untreated control. A rapid decline of the disease was observed in soil fromT. harzianum treated plots, successively planted with bean seedlings. More isolates ofTrichoderma sp. antagonistic toR. solani, were found in the infested field as compared to the non infested one.Trichoderma harzianum treated plants, transferred to the commercial field gave a 21–37% increase in early yield of strawberries. A combined treatment in the nursery and in the fruiting field resulted in a 20% yield increase as compared to control plots.     

Interaction between Colletotrichum falcatum pathotypes and biocontrol agents

Abstract

To select efficient antagonistic strain(s) of biocontrol agents against most of the existing pathotypes of Colletotrichum falcatum, an in vitro interaction study was carried out with 13 pathotypes, 12 isolates of Pseudomonas spp. and 6 isolates of Trichoderma spp. Antagonistic pseudomonad strains exhibited greater variation in their activity depending on the virulence of the pathotype. The lower the pathogen virulence, the higher was the antagonistic activity noticed. In general, sub-tropical pathotypes were suppressed at a comparatively higher level than the tropical pathotypes. Among the four efficient P. fluorescens strains selected based on their inhibitory effect against various pathotypes, ARR1G and VPT4 were effective against tropical pathotypes and FP7 showed moderate effect against all the pathotypes. The strain KKM2 was effective against sub-tropical and weaker tropical pathotypes. Strains of Trichoderma spp. did not show much variation in antagonism, but varied in their mode of action in suppressing the pathogen growth. However, based on higher rate of hyperparasitism, T. harzianum strains T5 and T62 were selected against all the pathotypes.     

Trichoderma viride as a mycoparasite ofAspergillus spp.

Summary Trichoderma viride was found to be parasitic on three species of Aspergillus. The mycoparasitism was characterized by frequent coiling, penetration and hyphal growth of the parasite inside the conidiophores of Aspergillus. The volatile and non-volatile metabolites ofT. viride, more or less, inhibited radial growth of all the testAspergillus spp.