Antagonistic Isolates Research Articles

Antagonistic Potential Marine Bacteria Against Luminescent Vibriosis Pathogen Vibrio Harveyi Obtained from Shrimp Farm, Marakkanam, Tamil Nadu, India

Aquaculture farming practices in India and other countries are facing a lot of economic loss due to luminescent vibriosis.Ecological and cost-effective practices may have been warranted to combat this tricky disease. A total of 57 morphologicallydifferent marine bacterial isolates were isolated from samples collected at different aquaculture ponds in Marakkanam, Tamil Nadu,India and screened for their antibacterial potential against luminescent vibriosis causing bacteria Vibrio harveyi. All marine bacterialisolates were screened for antibacterial activity by cross streak method against two different isolates V. harveyi GNC01 and V.harveyi GNC03. Secondary screening of antagonistic isolates by dual culture plate method and agar well diffusion method leads tothe identification of antagonistic isolates. The antagonistic isolates were characterized by molecular taxonomy (16S rRNA genesequence) and phylogeny and identified as Neobacillus niacini (GNC04), Bacillus halotolerans (GNC06), Bacillus amyloliquefaciens(GNC16), Lacticaseibacillus paracasei (GNC23) and Lactiplantibacillus plantarum (GNC24). The minimum inhibitory concentrationof all these selected bacterial crude secondary metabolites against both Vibrio harveyi isolates GNC01 and GNC03 ranged between0.062 to 0.50 mg/mL. The GC-MS profiling of the ethyl acetate extract revealed the presence of antibacterial compounds such asOxime-,methoxy-phenyl (26.63%), 2,3-Dihydroxybutane (9.52%), Phosphite, menthyldimethyl- (2.83%), Pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl)- (2.07%). The results of this study suggest that Lactiplantibacillus plantarum (GNC24) is apotential source for antagonistic secondary metabolites against luminescent vibriosis causing bacteria Vibrio harveyi. Further,biological control formulation with these isolates and their metabolites will be a reward for the integrated pest management inthe aquaculture sector.

Selection and Characterization of Antagonistic Microorganisms for Biological Control of Acidovorax citrulli Causing Fruit Rot in Watermelon

This study was performed to screen the efficacy of antagonistic bacterial isolates from various sources against the bacterial fruit blotch (BFB) causing pathogen (Acidovorax citrulli) in cucurbit crops. In addition, plant growth promoting traits of these antagonistic bacterial isolates were characterized. Two thousand seven hundred ninety-four microorganisms were isolated from the collected samples. Molecular identification revealed two A. citrulli out of 2,794 isolates. In vitro antagonistic results showed that, among the 28 antagonistic bacterial isolates, 24 and 14 bacterial isolates exhibited antagonism against HPP-3-3B and HPP-9-4B, respectively. Antagonistic and growth promotion characterization of the antagonistic bacterial isolates were further studied. Results suggested that, 4 antagonistic bacteria commonly showed both antagonism and growth promotion phenotypes. Moreover, 3 isolates possessed growth promoting activities. Overall results from this study suggests that BFB causing bacterial pathogen (A. citrulli) was suppressed in in vitro antagonism assay by antagonistic bacterial isolates. Furthermore, these antagonistic bacterial isolates possessed growth promotion and antagonistic enzyme production ability. Therefore, data from this study can provide useful basic data for the in vivo experiments which ultimately helps to develop the eco-friendly agricultural materials to control fruit rot disease in cucurbit crops in near future.

Biocontrol effects of endophytic and rhizospheric bacteria against the root pathogen Fusarium oxysporum of hot pepper (Capsicum annum L.)

Plant disease needs to be controlled to keep the quality ofproducts and the abundance of food produced by farmersall over the world. Hot pepper production in Ethiopia hasbeen reduced from time to time. This is mainly due to theoutbreak of different diseases especially fusarium wilt inthe hot pepper growing areas. The objective of this studywas to isolate and characterize Plant Growth PromotingBacteria from the hot pepper rhizosphere and internal tissueswith potential biocontrol activity against Fusariumoxysporum. Thirty healthy and vigorous hot pepper plantswith intact roots and soil from the rhizospheric region werecollected randomly from hot pepper growing areas of YimaliKebele. A total of 23 endophytic and rhizospheric bacteriawere isolated from the hot pepper root and rhizosphericsoil. The combined endophytic and rhizospheric bacteriashowed a significant effect on the growth of F. oxysporumthan single isolates (P d”0.05). The bacteria isolated fromMarkofana hot pepper varieties of black clay soil showed abetter antagonistic effect on F. oxysporum than bacteriaisolated from local hot pepper varieties of reddish sandysoil. A significant percentage of inhibition between combinedendophytic and rhizospheric bacteria [endophytic (GE) +rhizospheric (GR) isolates] compared to GE+GE and GR+GR(P =0.002) was obtained. Antagonistic bacterial isolates wereable to produce different hydrolytic enzymes such aschitinase, protease, and cellulase. Bacillus species showedbetter antagonistic performance and decreased radial growthof F. oxysporum than Pseudomonas species.

The effect of Trichoderma spp. isolates on some morphological traits of canola inoculated with Sclerotinia sclerotiorum and evaluation of their efficacy in biological control of pathogen

In this study, to evaluate the effect of isolates of different species of Trichoderma on some morphological traits of canola and to obtain suitable antagonist isolates for biological control of canola crown rot disease, the effect of 71 Trichoderma isolates on Sclerotinia sclerotiorum was investigated in the laboratory by volatile compound and non-volatile metabolite methods. In the volatile metabolite assay, the isolate ARCTr281 ( T. harzianum ) was more efficient than the other isolates with a 20% reduction in hyphae density and non-formation of S. sclerotiorum sclerotia. In the non-volatile metabolite assay, ARCTr272 ( T. harzianum ) with 73.7 and 62.22 at 25 and 15% concentrations and T. viride isolate ARCTr432 (20.37%) at 5% concentration were successful in inhibiting S. sclerotiorum mycelia growth. Six superior isolates were selected for greenhouse studies. Using two-stage Trichoderma isolates in the greenhouse, height, fresh and dry weight of shoots and roots, number of pods, pod length, pod diameter and number of seeds per pod in the presence of pathogen all increased. The results of principal component analysis showed that the first two components accounted for 86% of the total variation among the data. The results showed that for the first component, the traits of total grain weight, pod diameter, branch number, shoot dry weight and fresh weight of root were the most important traits in grouping the isolates. In the trait second component, plant height and number of seeds per pod were important traits. Mean comparisons were performed using an LSD test and showed significant differences between the fungi used in non-volatile metabolite and greenhouse studies. According to the results of this research, the fungal isolates of T. harzianum (ARCTr281, ARCTr272 and ARCTr418) were the most effective antagonists in improving plant traits and controlling canola rot disease in both the laboratory and the greenhouse.

Isolation of Antimicrobial Producing Actinomycetes from Indigenous Microhabitats.

Unusual microhabitats in different locations present opportunities for the bio-prospecting of actinomycetes with high antimicrobial producing potentials. A total of 113 actinomycetes from specific indigenous microhabitats were isolated and screened for their antimicrobial activity against selected test organisms. The earthworm castings, forest litter, and domestic dumpsite sites, respectively, accounted for approximately 24.78%, 19.47%, and 18.5% of the total number of isolates recovered. The preliminary antagonism assay using the perpendicular streak method yielded four isolates that showed high antimicrobial activity and broad spectrum activity against test bacteria or fungi. These isolates showed excellent growth and sporulation on milk agar and starch casein agar, and the micro-morphological characterization of the isolates revealed the presence of spore-bearing aerial hyphae. Bioactive extracts from DS15 and EC5, at 125 mg/mL, had respective zones of inhibition measuring 20.7 ± 0.6 and 33.0 ± 1.0 against multidrug resistant Escherichia coli (clinical isolate) and Salmonella typhi (ATCC 2923) respectively. Bioactive extracts from EC1 and EC9, at 125 mg/mL, show respective zones of inhibition measuring 18.7 ± 1.2 and 16.7 ± 1.2 against Candida albicans (ATCC 10231) and C. glabrata (ATCC 22018) respectively. This study reinforces the evidence that unique and unusual microhabitats, in different geographical locations, present an excellent opportunity for the bio prospecting of actinomycetes with high antimicrobial producing potentials. Furthermore, it shows that the indigenous antagonistic actinomycete isolates exhibited potent antimicrobial activity against test bacteria and fungi employed in the in-vitro antimicrobial assays.

Inoculations of soil by antagonistic strains modify tuberosphere bacterial communities and suppress common scab of potatoes

Common scab of potatoes is a soil-borne disease causing considerable economic losses worldwide. Although many management practices to control this disease have been introduced, the lack of efficiency increases an interest in effective and environmentally friendly alternatives. One of the attractive approaches to manage the disease is an application of antagonistic strains into soil. The aim of this study was to develop a selection approach for suitable biocontrol agents based on their in vitro properties and demonstrate their effect on soil microbial communities and host plant nutrition. Out of 56 isolates of Actinobacteria, three isolates and a mixture of six isolates with strong inhibitory activities against common scab pathogens and high growth rates were selected for the pot experiment to assess their biocontrol activity. The microbial community composition of the tuberosphere and the nutrient content in potato leaves was determined. One isolate significantly reduced the severity of the common scab. This isolate showed strong inhibitory activity against pathogens and reached the highest number of CFU/g of vermiculite. In the pot experiment, all the isolates affected the microbial community composition, yet the most substantial changes occurred in the treatment of the most efficient isolate. Moreover, it was determined that inoculation with isolates led to alterations in host plant nutrition suggesting plant-microbe interactions. We provided insights into relationships between antagonistic isolates, common scab pathogens, host plant and microbial communities, and their role in the disease control.

Suppression of Pepper Root Rot and Wilt Diseases Caused by Rhizoctonia solani and Fusarium oxysporum.

Pepper is vulnerable to soil-borne fungal pathogens such as Rhizoctonia solani and Fusarium oxysporum. The potential of beneficial rhizosphere microorganisms to control R. solani and F. oxysporum f.sp. capsici was evaluated in pepper plants. Paenibacillus polymyxa and Trichoderma longibrachiatum were isolated from rhizospheric soil samples of healthy pepper plants. In vitro, both isolates caused clear reductions in the radial growth of root rot and wilt pathogens. Scanning electron microscopy displayed lysis and abnormal shape of the pathogens in dual cultures with P. polymyxa and T. longibrachiatum. The incidence and severity of root rot and wilt diseases were significantly reduced in pepper plants treated with the growth-promoting fungi (PGPF isolates; Fusarium equiseti GF19-1, Fusarium equiseti GF18-3, and Phoma sp. GS8-3), P. polymyxa, or T. longibrachiatum in comparison to the control. Moreover, the induction treatments led to increased pepper growth compared with their control. The defense related gene (CaPR4) expression was shown to be significantly higher in the treated plants than in the control plants. In conclusion, the antagonistic isolates (P. polymyxa and T. longibrachiatum) and PGPF isolates have a clear impact on the prevention of root rot and wilt diseases in pepper plants incited by R. solani and F. oxysporum f.sp. capsici. The expression of the CaPR4 gene added to the evidence that PGPF isolates generate systemic resistance to pathogen infections.

Biocontrol properties from phyllospheric bacteria isolated from Solanum lycopersicum and Lactuca sativa and genome mining of antimicrobial gene clusters

BackgroundBiocontrol agents are sustainable eco-friendly alternatives for chemical pesticides that cause adverse effects in the environment and toxicity in animals including humans. An improved understanding of the phyllosphere microbiology is of vital importance for biocontrol development. Most studies have been directed towards beneficial plant-microbe interactions and ignore the pathogens that might affect humans when consuming vegetables. In this study we extended this perspective and investigated potential biocontrol strains isolated from tomato and lettuce phyllosphere that can promote plant growth and potentially antagonize human pathogens as well as plant pathogens. Subsequently, we mined into their genomes for discovery of antimicrobial biosynthetic gene clusters (BGCs), that will be further characterized.ResultsThe antimicrobial activity of 69 newly isolated strains from a healthy tomato and lettuce phyllosphere against several plant and human pathogens was screened. Three strains with the highest antimicrobial activity were selected and characterized (Bacillus subtilis STRP31, Bacillus velezensis SPL51, and Paenibacillus sp. PL91). All three strains showed a plant growth promotion effect on tomato and lettuce. In addition, genome mining of the selected isolates showed the presence of a large variety of biosynthetic gene clusters. A total of 35 BGCs were identified, of which several are already known, but also some putative novel ones were identified. Further analysis revealed that among the novel BGCs, one previously unidentified NRPS and two bacteriocins are encoded, the gene clusters of which were analyzed in more depth.ConclusionsThree recently isolated strains of the Bacillus genus were identified that have high antagonistic activity against lettuce and tomato plant pathogens. Known and unknown antimicrobial BGCs were identified in these antagonistic bacterial isolates, indicating their potential to be used as biocontrol agents. Our study serves as a strong incentive for subsequent purification and characterization of novel antimicrobial compounds that are important for biocontrol.

Potential of Trichoderma spp. for Biocontrol of Aflatoxin-Producing Aspergillus flavus.

The inhibitory action of 20 antagonistic Trichoderma isolates against the aflatoxigenic isolate A. flavus ITEM 9 (Af-9) and their efficacy in reducing aflatoxin formation in vitro were examined. Production of metabolites with inhibitory effect by the Trichoderma isolates was also investigated. Antagonistic effect against Af-9 was assessed by inhibition of radial growth of the colonies and by fungal interactions in dual confrontation tests. A total of 8 out of 20 isolates resulted in a significant growth inhibition of 3-day-old cultures of Af-9, ranging from 13% to 65%. A total of 14 isolates reduced significantly the aflatoxin B1 (AfB1) content of 15-day-old Af-9 cultures; 4 were ineffective, and 2 increased AfB1. Reduction of AfB1 content was up to 84.9% and 71.1% in 7- and 15-day-old cultures, respectively. Since the inhibition of Af-9 growth by metabolites of Trichoderma was not necessarily associated with inhibition of AfB1 production and vice versa, we investigated the mechanism of reduction of AfB1 content at the molecular level by examining two strains: one (T60) that reduced both growth and mycotoxin content; and the other (T44) that reduced mycotoxin content but not Af-9 growth. The expression analyses for the two regulatory genes aflR and aflS, and the structural genes aflA, aflD, aflO and aflQ of the aflatoxin biosynthesis cluster indicated that neither strain was able to downregulate the aflatoxin synthesis, leading to the conclusion that the AfB1 content reduction by these Trichoderma strains was based on other mechanisms, such as enzyme degradation or complexation. Although further studies are envisaged to identify the metabolites involved in the biocontrol of A. flavus and prevention of aflatoxin accumulation, as well as for assessment of the efficacy under controlled and field conditions, Trichoderma spp. qualify as promising agents and possible alternative options to other biocontrol agents already in use.

Exploring Citrus Nematodes and its Antagonists in Citrus Growing Areas of Punjab, Pakistan

Citrus nematode (Tylenchulus semipenetrans) is a major threat to the citrus industry in Pakistan. During 2019-20, a survey was carried out for citrus nematode and its antagonists in citrus orchards (Both healthy and declining trees were samples) of Punjab, Pakistan. Bhalwal, Shahpur, Sillanwali, Kotmomin, Sargodha, Quaidabad, Jhang, and Rahim Yar Khan were surveyed for disease incidence, prevalence, and association of T. semipenetrans. Maximum disease prevalence (100%) was recorded in Rahim Yar Khan while minimum (20%) in Bhalwal and Kotmomin. Maximum slow decline with 80% field incidence was observed in Rahim Yar Khan while a minimum of 20% in Quaidabad. Soil and root samples were used for nematode extraction using Baerman's funnel method and the Whitehead Hemming tray method. A maximum number of J2s/100ml of soil were observed in soil samples taken from Rahim Yar Khan (1674) followed by Shahpur (1534), Qaidabad (1432), Sargodha (1347), and Bhalwal (1172). The highest number of nematode females per gram of root were recorded in Rahim Yar Khan (652) followed by Quaidabad (611), Bhalwal (490), Kotmomin (421), Sillanwali (387). Isolation of different fungal and bacterial antagonist isolates was also done by soil dilution plate technique using nutrient agar media and potato dextrose agar media. Identification of fungal antagonists was made on colony growth, color, and spore structure. Bacterial identification was also done on morphological characters and gram tests. Trichoderma harzianum, Trichoderma viride, Trichoderma koningii Trichoderma atroviride, Pseudomonas fluorescens, Pseudomonas putida, Bacillus cereus and Bacillus subtilis were found associated.

In vitro evaluation of grapevine endophytes, epiphytes and sap micro-organisms for potential use to control grapevine trunk disease pathogens

Grapevine trunk diseases (GTDs) threaten the economic sustainability of viticulture, causing reductions of yield and quality of grapes. Biological control is a promising sustainable alternative to cultural and chemical methods to mitigate the effects of pathogens causing GTDs, including Botryosphaeria dieback, Eutypa dieback and Esca. This study aimed to identify naturally occurring potential biological control agents from grapevine sap, cane and pith tissues, and evaluate their in vitro antagonistic activity against selected fungal GTD pathogens. Bacterial and fungal isolates were preliminarily screened in dual culture assays to determine their antifungal activity against Neofusicoccum parvum and Eutypa lata. Among the fungal isolates, Trichoderma spp. inhibited mycelium growth of E. lata by up to 64% and of N. parvum by up to 73%, with overgrowth and growth cessation being the likely antagonistic mechanisms. Among the bacterial isolates, Bacillus spp. inhibited mycelium growth of E. lata by up to 20% and of N. parvum by up to 40%. Selected antagonistic isolates of Trichoderma, Bacillus and Aureobasidium spp. were subjected to further dual culture antifungal analyses against Diplodia seriata and Diaporthe ampelina, with Trichoderma isolates consistently causing the greatest inhibition. Volatile organic compound antifungal analyses showed that these Trichoderma isolates inhibited mycelium growth of N. parvum (20% inhibition), E. lata (61% inhibition) and Dia. ampelina (71% inhibition). Multilocus sequence analyses revealed that the Trichoderma isolates were most closely related to Trichoderma asperellum and Trichoderma hamatum. This study had identified grapevine sap as a novel source of potential biological control agents for control of GTDs. Further testing will be necessary to fully characterize modes of antagonism of these microorganisms, and assess their efficacy for pruning wound protection in planta.

Morphological and molecular characterization of a new autochthonous Trichoderma sp. isolate and its biocontrol efficacy against Alternaria sp.

The development of agriculture requires the use of microorganisms in the management of phytopathogens as a way to compensate for the use of chemical pesticides, in order to produce healthy crops. The objective of this study was to characterize a new isolate of Trichoderma sp. based on morphological and molecular features, and its potential ability to control the pathogen Alternaria sp. The antagonistic isolate was isolated from soil samples of potato fields in Guasave Sinaloa, Mexico, whereas the pathogen was collected from infected apple leaves in the orchard “La Escondida” in Guerrero County, Chihuahua, Mexico. For morphological characterization both fungi were grown on solid PDA medium. DNA of Trichoderma sp. was isolated using the CTAB method and PCR analyses were done using ITS1, ITS4 primers resulting in amplified products of 600 bp. These were sequenced, submitted to Genbank (acc. no. MN950427) and used for further phylogenetic analysis through Bayesian inference approach. Five clades were identified and the polytome topography recovered from clade 4 indicates a high genetic similarity with T. asperellum. A BLAST examination of the resulting sequence in GenBank showed 98.11% similarity with T. asperellum. This result together with the morphological and the phylogenetic analyses indicates that the isolate belongs to Trichoderma asperellum Samuels, Lieckfeldt & Nirenberg. Biocontrol tests of this isolate showed inhibition of Alternaria sp. between 50% and 93%. These results are essential for biodiversity research and give some new possibilities for pest management.

Biological control of fungal pathogens of tomato (Lycopersicon esculentum) by chitinolytic bacterial strains

The need to increase food production and to reduce the pollution caused by synthetic chemicals has led to a search for biocontrol agents against plant pathogens. In the present study, a total of 37 chitinolytic bacteria were isolated from the rhizospheric soil of tomatoes using a chitin agar medium. In vitro bacterial isolates, that is, TD9, TD11, TD15, and TD24 showed strong antagonistic and enzymatic activities against Rhizoctonia (8%-55%), Fusarium (31%-48%), Colletotrichum (24%-49%), and Aspergillus on a dual culture plate and enzyme assay. Furthermore, these putative antagonistic bacterial isolates were identified as Pantoea agglomerans (TD9), Bacillus subtilis (TD11), Bacillus cereus (TD15 and TD24) using 16S rRNA sequence analysis. Additionally, in culture filtrate in vivo assay, the isolates TD11 and TD15 inhibited the growth of Rhizoctonia solani about 40% and Fusarium oxysporum about 80%. However,in the pot trials, these two bacterial isolates (TD11 and TD15) considerably suppressed the disease rate in tomatoes caused by Fusarium and Rhizoctonia fungal species. Moreover, it was concluded that B. subtilis (TD11) was found to be the most promising putative biocontrol agent, inhibiting the fungal diseases of tomatoes by 50% and showing versatile antagonistic potential.

Characterisation of rice-associated antagonistic pseudomonads and their application in combination with plant resistance inducer molecules for the control of sheath blight disease of rice

ABSTRACT Rice sheath blight caused by Rhizoctonia solani AG1-IA is one of the most important diseases of high-yielding rice (Oryza sativa L.) cultivars in rice-growing areas of the world including Iran. This study aimed to determine the efficacy of native fluorescent pseudomonads in combination with plant defence activators in controlling rice sheath blight disease. Sampling was done from local and high-yielding rice cultivars grown in Mazandaran province, Iran. A total of 291 fluorescent strains were isolated and screened for antagonism against R. solani. The isolates Ps-275, Ps-128, Ps-287, Ps-240 and Ps-243 reduced the disease symptoms up to 53.1­%, 49.3%, 44.76%, 40.83% and 35.7­%, respectively, only when applied in combination with potassium silicate at the booting stage of rice growth. The isolate Ps-3 reduced the disease symptoms when applied alone and in combination with oxalic acid by 60.5% and 58.7%, respectively. Multilocus sequence analysis of house-keeping genes, rpoD, oprI and oprL identified the most effective antagonistic isolates, Ps-275, Ps-3 and Ps-287, as endemic Pseudomonas strains affiliated with the P. protegens, P. mosselii and P. aeruginosa (sub) groups, respectively. These strains significantly inhibited the growth of R. solani AG1-IA in vitro and reduced the disease symptoms at the booting stage of the rice plant as well.

Screening of indigenous rhizobacteria as potential biological control against faba bean (Vicia faba L.) gall disease caused by Olpidium viciae

BackgroundEmerged faba bean gall disease attacks the stem and leaves of the plant and results in complete crop losses. This study was initiated to screen multi-trait rhizobacteria for their antagonistic efficacy under in-vitro and in-vivo conditions against Olpidium viciae to control the effect of gall disease on faba bean. Sixty antagonistic isolates were first examined for their morphological, biochemical, and phenotypic traits.ResultsPseudomonas fluorescens AAUPF62, P. aeruginosa AAUS31, Bacillus AAUMF42, and Bacillus AAUAm28 showed greater than 68, 62, 57, and 54% suppression of O. viciae in dual culture, volatile metabolites, culture filtrate assay, and detached leaves experiments, respectively. The in-vivo study revealed that early treatment of the crop with P. aeruginosa AAUS31 reduced severity by 63% (in FB-Obse) and 54% (in FB-26869) faba bean varieties. The co-inoculation of P. fluorescens AAUPF62 and P. aeruginosa AAUS31 significantly enhanced the shoot (P = 0.003; mean = 122 cm) and root (P = 0.018; mean = 94 cm) length, increased shoot dry weight by 8 factors (83 g pot −1), and reduced final disease severity by 92% in FB-Obse variety.ConclusionsThe results revealed that P. fluorescens AAUPF62 and P. aeruginosa AAUS31 strains could be the potential antagonistic agents of gall disease. The use and early treatment of moderately resistant faba bean varieties by co-inoculation of synergistic potential bioagents were recommended.

Biocontrol Potential of Trichoderma afroharzianum TM24 Against Grey Mould on Tomato Plants.

Grey mould caused by Botrytis cinerea leads to severe economic loss on commercial tomato production. Application of beneficial microorganism offers an eco-friendly alternative for mitigation of tomato fungal disease damage, considering negative influences of fungicides. In the present study, an antagonistic Trichoderma afroharzianum isolate TM24 was evaluated for its biocontrol potential on tomato grey mould. The isolate TM24 showed obviously antagonistic effect on B. cinerea mycelium growth and production of glucanase and chitinase. Leaf spraying with spore suspension of isolate TM24 showed a biocontrol efficiency of over 54% against tomato grey mould in greenhouse pot experiment. The activities of plant defense-related enzymes including polyphenol oxidase, phenylalanine ammonialyase, superoxide dismutase, and peroxidase were all increased to varying degrees in tomato leaves after isolate TM24 treatment. Transcriptome analysis showed that, a total of 1941, 1753 and 38 differentially expressed genes (DEGs) were obtained at 24, 48 and 72 hpi, respectively, in tomato leaves pretreated with T. afroharzianum TM24, and then challenged with B. cinerea inoculation. The DEGs were mainly enriched in MAPK signaling pathway and plant hormones signal transduction pathway. Multiple genes that regulated crucial nodes of defense-related pathways, like flavonoid, phenylpropanoid, jasmonic acid and ethylene metabolisms were also identified, which may have positive correlations with the biocontrol potential of isolate TM24 in tomato plants. These promising results provided valuable information on using T. afroharzianum TM24 as a beneficial biocontrol agent in tomato grey mould management.

Inhibition of mycelial growth of Fusarium oxysporum f. sp. lycopersici by isolates of Trichoderma spp.

Tomato crops have great economic importance in Brazil and worldwide; however, its economic return is dependent on the control of several diseases, including Fusarium wilt, caused by Fusarium oxysporum f. sp. lycopersici. The control of this disease is usually based on biological products, which are commonly manufactured with antagonist isolates of Trichoderma sp. Therefore, the antagonistic potential of Trichoderma sp. should be better studied against other phytopathogens, thus expanding the biofungicide options on the market. The objective of the present work was to evaluate the antagonistic potential of different isolates of Trichoderma against the in vitro mycelial growth of F. oxysporum f. sp. lycopersici. Four isolates of Trichoderma spp. were tested, three collected in litterfall and one commercial isolate (T. asperellum). The Trichoderma spp. isolates were evaluated for antagonistic potential against F. oxysporum f. sp. lycopersici using the culture pairing technique. The variables evaluated were: mean colony size (MCS), mycelial growth inhibition percentage (MGIP), mycelial growth rate index (MGRI) of phytopathogen, and antagonism of isolates of Trichoderma spp. through a scale of grades. All isolates of Trichoderma spp. tested reduced the in vitro mycelial growth of F. oxysporum f. sp. lycopersici.

Selection and control efficiency of Trichoderma isolates against Fusarium oxysporum f. sp. ciceris in Iran

The antagonistic potential of forty isolates of Trichoderma spp. obtained from major chickpea growing areas in western part of Iran, were evaluated against Fusarium oxysporum f. sp. ciceris, the most important soil-born pathogen of chickpea. Among those, six species of Trichoderma harzianum, T. longibrachiatum, T. pleurotum, T. crassum, T. guizhouense,and Trichoderma sp. were identified based on morphological, molecular and phylogenetic analysis of concatenated DNA sequences of tef1 and rpb2 genes. All isolates were tested in vitro using dual culture method for their inhibition of the mycelial growth of the pathogen. Eight most antagonistic isolates, which suppressed colony growth of pathogen by 61.1–65.5%, were tested in vitro for production of volatile metabolites. All eight selected isolates produced volatile metabolites which prevented radial growth of pathogen from 17.5% to 31.0%. Antagonistic isolates provided significant protection against pathogen when mixed with soil or treated on seeds. Wilt control efficiency was greater for T. longibrachiatum KT8 (64.4%), T. harzianum KT9 (66.1%), and KT10 (69.5%) after 60 days of sowing highly susceptible chickpea cultivar JG-62 in inoculated soils. Greater wilt control efficiencies were detected for seed treatments with KT8 (69.1%), KT9 (72.7%), and KT10 (74.5%) isolates. Higher plant height, dry weight of root and shoot were recorded with these three superior isolates in seed treatments, inoculated and non-inoculated soils. Such superior isolates could be considered as promising antagonistic agents in biocontrol of chickpea Fusarium wilt for future studies. To the best of our knowledge, T. pleurotum and T. guizhouense are reported for the first time from Iran.

Suppression of Fusarium Wilt Caused by Fusarium oxysporum f. sp. lactucae and Growth Promotion on Lettuce Using Bacterial Isolates.

This study was carried out to explore a non-chemical strategy for enhancing productivity by employing some antagonistic rhizobacteria. One hundred eighteen bacterial isolates were obtained from the rhizospheric zone of various crop fields of Gangwon-do, Korea, and screened for antifungal activity against Fusarium wilt (Fusarium oxysporum f. sp. lactucae) in lettuce crop under in vitro and in vivo conditions. In broth-based dual culture assay, fourteen bacterial isolates showed significant inhibition of mycelial growth of F. oxysporium f. sp. lactucae. All of the antagonistic isolates were further characterized for the antagonistic traits under in vitro conditions. The isolates were identified on the basis of biochemical characteristics and confirmed at their species level by 16S rRNA gene sequencing analysis. Arthrobacter sulfonivorans, Bacillus siamensis, Bacillus amyloliquefaciens, Pseudomonas proteolytica, four Paenibacillus peoriae strains, and Bacillus subtilis were identified from the biochemical characterization and 16S rRNA gene sequencing analysis. The isolates EN21 and EN23 showed significant decrease in disease severity on lettuce compared to infected control and other bacterial treatments under greenhouse conditions. Two bacterial isolates, EN4 and EN21, were evaluated to assess their disease reduction and growth promotion in lettuce in field conditions. The consortium of EN4 and EN21 showed significant enhancement of growth on lettuce by suppressing disease caused by F. oxysporum f. sp. lactucae respectively. This study clearly indicates that the promising isolates, EN4 (P. proteolytica) and EN21 (Bacillus siamensis), can be commercialized and used as biofertilizer and/or biopesticide for sustainable crop production.

Characterization of Streptomyces spp. from Rice Fields as a Potential Biocontrol Agent against Burkholderia glumae and Rice Plant Growth Promoter

The usage of plant growth-promoting (PGP) Streptomyces to improve plant growth is an alternative to existing strategies of chemical fertilizers that commonly caused environmental pollution. The aim of this study was to isolate and characterize PGP Streptomyces from Malaysian rice fields with antagonistic activity against Bacterial Panicle Blight disease in rice caused by Burkholderia glumae. A total of 50 bacterial isolates were recovered from the soil, rhizosphere, and endosphere; 22% showed antagonistic activity against B. glumae. Molecular identification using 16S rRNA amplification and phylogenetic tree analyses revealed that the antagonistic isolates belonged to the genus Streptomyces. Among the antagonistic isolates, Streptomyces sp. TBS5 (26.35 ± 0.14 mm) demonstrated the most antagonistic activity (p < 0.05) against B. glumae. Through API® ZYM analysis, the antagonistic isolates were revealed to have phosphoric hydrolase, ester hydrolase, peptide hydrolase, and glycosidase activities that play a crucial role in plant growth promotion. The antagonistic isolates demonstrated the highest (80%) PGP traits including able to fix nitrogen and solubilize phosphate, as well as produce siderophore and indole-3-acetic acid. Plant growth promotion assay under laboratory and greenhouse conditions were analyzed with the treatment of rice, Oryza sativa, seeds with the antagonistic isolates. Seeds treated with Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 showed improvement in vigor index by more than 100% compared to water-treated control plants under both conditions. Augmentation of root length showed an overall increment of more than 101.5% under laboratory condition and 151.9% under greenhouse condition using Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 treatments. Results suggest that Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 are a good candidate to be developed as both biocontrol agent against B. glumae and PGP agent in rice.